Preface

Chapter 8 **Health-Related Quality of Life in Maxillectomy Patients Rehabilitated with Obturator Prostheses: A**

**Literature Review 137**

Kadriye Peker

**VI** Contents

Head and neck cancer is an important health problem in all countries. The number of indi‐ viduals affected from the disease is increasing day by day. Important advances have occur‐ red in the diagnosis and management of cancer patients since the first reports. This also leads to an increase in the survival rate of these patients.

A multidisciplinary approach is required for the management of these patients. Dentists also are included in this team especially when the lesions are located in the oral cavity and jaws and when radiotherapy including the head and neck region is applied.

This book starts with a chapter including signs and symptoms and etiology and risk factors of head and neck cancer. It carries on with the epidemiology and recently identified micro‐ RNAs and the role of microRNAs in nasopharyngeal carcinoma and oral carcinogenesis. The readers can find current information regarding classification of salivary gland tumors, thyroid gland tumors, TNM staging, and preoperative assessment and management of these lesions. Recent updates on the diagnosis and management of medullary thyroid carcinoma are also included.  Oral side effects of head and neck irradiation, dentist's role in head and neck cancer team, dental management prior radiation therapy, and oral care of these patients during and after head and neck radiotherapy are described. The book finishes with a review including results of research about health-related quality of life in maxillectomy patients rehabilitated with obturator prostheses.

The contributions to this book have been made from researchers and specialists from differ‐ ent countries. I hope that this book will serve as a useful reference for the readers.

I want to thank Ms. Andrea Korić for the invitation of the book project and Ms. Martina Usljebrka for her help during the preparation of the book. I also want to thank my husband Veysel and my sons Utku and Uğur for their patience during the preparation of this book.

> **Associate Professor Zühre Akarslan** Department of Oral and Maxillofacial Radiology Gazi University Faculty of Dentistry Turkey

**Provisional chapter**

## **Introductory Chapter: Be Careful! It Can Be Cancer**

**Introductory Chapter: Be Careful! It Can Be Cancer**

DOI: 10.5772/intechopen.70045

#### Zühre Akarslan Additional information is available at the end of the chapter

Zühre Akarslan

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.70045

## **Introduction**

Head and neck cancer includes the cancers of oral cavity, salivary glands, nasal cavity, paranasal sinuses, oropharynx, hypopharynx, pharynx, and larynx. It has a high rate among all cancer types in the world, and it constitutes a major public health problem. Although there are different histopathological types, the majority of the lesions are squamous cell carcinomas which usually arise from mucosal lining [1]. Males are more commonly affected from the condition compared to females [2].

The signs and symptoms of head and neck cancer depend according to the localization of the lesion. The most common sign of laryngeal cancer is hoarseness, whereas difficulty in swallowing or sore throat is seen in pharyngeal cancers. Sometimes, a painless neck mass is present. The important point is that the clinician should be careful during the examination of patients with nonspecific signs and symptoms, or signs and symptoms mimicking benign pathologies [1]. Any of the following such as sore throat, hoarseness, stridor, difficulty in swallowing, lump in neck, unilateral ear pain, red or white patch in the oral cavity, oral ulcer, presence of an indurated or rolled border, loosened teeth over a short time, rapid swelling with no demonstrable cause, lateral neck mass, lymphadenopathy, rapidly growing thyroid mass, cranial nerve palsy, orbital mass, and unilateral ear effusion lasting for more than 3 weeks can be cancer [3, 4].

When we look at the etiology and risk factors of head and neck cancer, tobacco and alcohol are reported to be the major ones. If the patient is smoking and drinking alcohol at the same time, the risk is multiplied [5]. Genetic predisposition is related with some cases. Genetic polymorphisms in genes encoding the enzymes, which are responsible from the metabolism of tobacco and alcohol, increase the risk of development of cancer [1].  Human papilloma virus (HPV) is reported to be a risk factor, especially for oropharyngeal cancer [6].

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Premalignant lesions of the oral cavity are also suspected as risk factors. These lesions do not have a high-risk factor like the others addressed earlier but should be carefully examined and treated. Leukoplakia and erythroplakia are the common premalignant lesions. Oral lichen planus also has the potential of malignant transformation [7].

Patients with Fanconi anemia, ataxia telangiectasia, Bloom's syndrome, and Li-Fraumeni syndrome have the risk of developing head and neck cancer. In addition, the risk of developing malignancy increases in immunosuppressed patients. Although Kaposi's sarcoma and non-Hodgkin's lymphoma are the most prevalent lesions seen among HIV-positive patients, the risk of oropharyngeal squamous cell carcinoma is increased [7].

If you do not pay enough attention to the signs and symptoms of head and neck cancer and do not treat premalignant lesions and premalignant conditions, the requirement of a more aggressive treatment and even early death of the patient can occur [4]. Thus, be careful! It can be cancer.

#### **Overview of the chapters of this book**

**Second chapter:** "**Head and Neck Cancer: Epidemiology and Role of MicroRNAs" written by Muhammad Babar Khawar, Naz Fatima, Muddasir Hassan Abbasi, Rabia Mehmood, Saira Kainat, and Nadeem Sheikh**. This chapter starts with the epidemiology and then focuses on the role of microRNAs on head and neck cancer. The authors give information about the discovery and biology of microRNAs. Recently identified microRNAs that undergo deregulation in head and neck cancer and their role in nasopharyngeal carcinoma and oral carcinogenesis are presented in detail. A list of microRNAs, identified as potent biomarkers of head and neck cancer, is presented. This is particularly helpful to the readers. The authors emphasize that microRNAs may be used to improve treatment strategies of head and neck cancer.

**Third chapter: "Salivary Gland Cancers: A Survey Through History, Classifications and Managements" written by Mohammad Hossein Khosravi, Ali Bagherihagh, Masoumeh Saeedi, Payman Dabirmoghaddam, Ali Kouhi, and Mohammad Hosein Amirzade-Iranaq**. This chapter starts with the anatomy, embryology, and structure of salivary glands. Then, a detailed histological classification of epithelial tumors according to WHO and TNM classification and staging of major salivary gland tumors are given. Preoperative assessment and management of the tumors based on recently published data in the literature are presented. These topics are useful for the clinicians.

**Fourth chapter: "Thyroid Cancers: Considerations, Classifications and Managements" written by Mohammad Hossein Khosravi, Ali Kouhi, Masoumeh Saeedi, Ali Bagherihagh, and Mohammad Hosein Amirzade-Iranaq**. This chapter starts with a detailed epidemiology of thyroid cancer, following clinical features and categories of thyroid malignancies according to a new classification. Staging and management of these lesions are also presented. The authors' description of thyroid cancer categories and treatment options peculiar to each lesion is particularly informative.

**Fifth chapter: "Medullary Thyroid Carcinoma: Recent Updates on the Diagnosis and Management" written by Andrei Cismaru, Iulia Coroian, Gabriel Cismaru, and Adrian Udrea**. This chapter describes updates on the diagnosis and treatment of medullary thyroid carcinoma. Fine-needle aspiration, serum calcitonin, computed tomography, and fludeoxyglucose-positron emission tomography (FDG-PET) are summarized. Biomarkers with prognostic value, such as plasma calcitonin, carcinoembryonic antigen, germ-like RET mutation, and matrix metalloproteinase, are given. Updates on the management and treatment of the pathology including surgical treatment, radiation therapy, systemic therapy angiogenesis inhibitors as well as transcatheter arterial embolization, percutaneous ethanol injection and gene therapy are described. Finally, the authors present a case of medullary thyroid carcinoma.

**Sixth chapter: "Interventional Techniques for Head and Neck Cancer Pain" written by Victor M. Silva Ortíz**, **Guillermo E. Aréchiga Ornelas, José A. Flores Cantisani, J. Ignacio Reyes Torres, and Fernando Cantú Flores**. This chapter gives information about factors associated with pain resulting from head and neck cancer and the methods used for the management of pain.  In addition, vertebroplasty for cancerrelated cervical vertebral compression fractures, intrathecal drug delivery systems, peripheral nerve blocks, cervical epidural and medial branch block, and Botulinum neurotoxin are also included. This chapter provides valuable information about these techniques to the clinician.

**Seventh chapter: "Oral Side Effects of Head and Neck Irradiation" written by Vlaho Brailo, Vanja Vučićević Boras, Danica Vidović Juras, Ana Andabak Rogulj, Božana Lončar Brzak, and Ivan Alajbeg**. This chapter gives information about the acute and chronic side effects of head and neck irradiation. Besides, the dentist's role in head and neck cancer team, dental management before radiation therapy, and oral care of these patients during and after head and neck radiotherapy are given in detail. The role of dentists for the management of such patients' is stressed in the chapter. The readers can benefit from practical strategies which could be applied in clinical practice.

**Eighth chapter: "Health-Related Quality of Life in Maxillectomy Patients Rehabilitated with Obturator Prostheses: A Literature Review" written by Kadriye Peker**. This chapter gives information about the results of the studies including manufacturing individualized obturators, different retention mechanisms, obturator replacement with free flap, obturator functioning, health-related quality of life (HRQOL), and self-reported problems among patients wearing an obturator prosthesis according to a literature search performed in PubMed, EMBASE, and Google Scholar. The tables summarizing results of different studies are particularly informative to the readers.

## **Author details**

Premalignant lesions of the oral cavity are also suspected as risk factors. These lesions do not have a high-risk factor like the others addressed earlier but should be carefully examined and treated. Leukoplakia and erythroplakia are the common premalignant lesions. Oral lichen

Patients with Fanconi anemia, ataxia telangiectasia, Bloom's syndrome, and Li-Fraumeni syndrome have the risk of developing head and neck cancer. In addition, the risk of developing malignancy increases in immunosuppressed patients. Although Kaposi's sarcoma and non-Hodgkin's lymphoma are the most prevalent lesions seen among HIV-positive patients, the

If you do not pay enough attention to the signs and symptoms of head and neck cancer and do not treat premalignant lesions and premalignant conditions, the requirement of a more aggressive treatment and even early death of the patient can occur [4]. Thus, be careful! It can be cancer.

**Second chapter:** "**Head and Neck Cancer: Epidemiology and Role of MicroRNAs" written by Muhammad Babar Khawar, Naz Fatima, Muddasir Hassan Abbasi, Rabia Mehmood, Saira Kainat, and Nadeem Sheikh**. This chapter starts with the epidemiology and then focuses on the role of microRNAs on head and neck cancer. The authors give information about the discovery and biology of microRNAs. Recently identified microRNAs that undergo deregulation in head and neck cancer and their role in nasopharyngeal carcinoma and oral carcinogenesis are presented in detail. A list of microRNAs, identified as potent biomarkers of head and neck cancer, is presented. This is particularly helpful to the readers. The authors emphasize that

**Third chapter: "Salivary Gland Cancers: A Survey Through History, Classifications and Managements" written by Mohammad Hossein Khosravi, Ali Bagherihagh, Masoumeh Saeedi, Payman Dabirmoghaddam, Ali Kouhi, and Mohammad Hosein Amirzade-Iranaq**. This chapter starts with the anatomy, embryology, and structure of salivary glands. Then, a detailed histological classification of epithelial tumors according to WHO and TNM classification and staging of major salivary gland tumors are given. Preoperative assessment and management of the tumors based on recently published data in the literature are presented.

**Fourth chapter: "Thyroid Cancers: Considerations, Classifications and Managements" written by Mohammad Hossein Khosravi, Ali Kouhi, Masoumeh Saeedi, Ali Bagherihagh, and Mohammad Hosein Amirzade-Iranaq**. This chapter starts with a detailed epidemiology of thyroid cancer, following clinical features and categories of thyroid malignancies according to a new classification. Staging and management of these lesions are also presented. The authors' description of thyroid cancer categories and treatment options peculiar to each lesion

**Fifth chapter: "Medullary Thyroid Carcinoma: Recent Updates on the Diagnosis and Management" written by Andrei Cismaru, Iulia Coroian, Gabriel Cismaru, and Adrian Udrea**. This chapter describes updates on the diagnosis and treatment of medullary

microRNAs may be used to improve treatment strategies of head and neck cancer.

planus also has the potential of malignant transformation [7].

risk of oropharyngeal squamous cell carcinoma is increased [7].

**Overview of the chapters of this book**

2 Diagnosis and Management of Head and Neck Cancer

These topics are useful for the clinicians.

is particularly informative.

Zühre Akarslan

Address all correspondence to<EMAIL_ADDRESS>
Department of Oral and Maxillofacial Radiology, Faculty of Dentistry, Gazi University, Ankara, Turkey

## **References**

**Provisional chapter**

## **Head and Neck Cancer: Epidemiology and Role of MicroRNAs of MicroRNAs**

**Head and Neck Cancer: Epidemiology and Role**

DOI: 10.5772/intechopen.69418

Muhammad Babar Khawar, Naz Fatima, Muddasir Hassan Abbasi, Rabia Mehmood, Saira Kainat Suqaina and Nadeem Sheikh Muddasir Hassan Abbasi, Rabia Mehmood, Saira Kainat Suqaina and Nadeem Sheikh Additional information is available at the end of the chapter

Muhammad Babar Khawar, Naz Fatima,

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.69418

#### **Abstract**

**References**

4 Diagnosis and Management of Head and Neck Cancer

405-427

[1] Mehanna H, Paleri V, West CML, Nutting C. Head and neck cancer—Part 1: Epidemiology,

[2] Scully C, de Almeida OP, Bagan J, Diz Dios P, Taylor AM. Oral Medicine and Pathology

[3] National Institute for Health and Clinical Excellence. Improving Outcomes in Head and Neck Cancers—The Manual. 2004. Available from: http://guidance.nice.org.uk/csghn/

[4] Wood RE. Malignant diseases of the jaws. In: White SC, Pharoah MJ, editors. Oral Radiology. Principles and Interpretation. 6th ed. St. Louis Mosby: Elsevier; 2009. pp.

[5] Blot WJ, McLaughlin JK, Winn DM, Austin DF, Greenberg RS, Preston-Martin S, Bernstein L, Schoenberg JB, Stemhagen A, Fraumeni JF Jr. Smoking and drinking in rela-

[6] Leemans CR, Braakhuis BJM, Brakenhoff RH. The molecular biology of head and neck

[7] Shaw R, Beasley N. Aetiology and risk factors for head and neck cancer: United Kingdom National Multidisciplinary Guidelines. Journal of Laryngology & Otology. 2016;**130**:9-12

tion to oral and pharyngeal cancer. Cancer Research. 1988;**48**:3282-3287

presentation, and prevention. British Medical Journal. 2010;**341**:c4684

at a Glance. 1st ed. Wiley-Blackwell; England. 2010. p. 51

guidance/pdf/English [Accessed: June 2, 2017]

cancer. Nature Review Cancer. 2011;**11**:9-22

Head and neck cancer (HNC) is referred to the cancers of aerodigestive tract covering number of structures *viz*, oral and nasal cavity, paranasal sinuses, lips, salivary glands, oropharynx, hypopharynx, pharynx, larynx, and local lymph nodes. It is the sixth most common cancer in the world. MicroRNAs (miRNAs) are small single‐stranded noncoding RNAs (ncRNAs) of about 19–25 nucleotides. These miRNAs have been reported to influence number of biological activities, i.e., gene regulation, differentiation, organ formation, cell death, cell proliferation, and stress responses. The first ever study involving miRNAs in HNC was published in 2005. Since then, association between dysregulation of miRNAs and head and neck tumorigenesis has been documented by a number of researchers. This chapter has covered a comprehensive state of the art literature review of the recent studies about the role of miRNAs in HNC includ‐ ing oral squamous cell carcinoma (OSCC) and human nasopharyngeal carcinoma. Despite significant improvement in multimodal treatment, the prognosis of advanced HNC is quite poor. Recent studies are promising regarding the potential role of miRNAs as prognostic indi‐ cators. Recently, some miRNAs have been discovered as important diagnostic biomarkers. In fact, miRNAs are found circulated stably in different body fluids, i.e., urine, blood, saliva, as well as in breath. Hence, these miRNAs can be assessed easily with noninvasive methods. miRNAs are the key therapeutic targets in addition to their prognostic and diagnostic value. Use of synthetically designed "miRNAs sponges," miR mimics (agomiRs), miR antagonists ("antagomiRs"), and miR inhibitors (antimiRNAs oligonucleotides) is an innovative strategy to modulate oncogenic and tumor‐suppressive pathways. Our understanding of miRNAs involvement in HNC is in its infancy. The discovery of miRNAs heralds a complete new para‐ digm in the understanding of exact molecular pathways involved in HNC development. More detailed studies are required for better understanding and therapeutic targets to treat HNC.

**Keywords:** biomarker, gene regulation, head and neck cancer, microRNAs, nasopharyngeal carcinoma, noncoding RNAs, oral squamous cell carcinoma (OSCC), therapeutic targets

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

## **1. Head and neck cancer epidemiology**

Head and neck cancer (HNC) is referred to the cancers of aerodigestive tract covering number of structures *viz*, oral and nasal cavity, paranasal sinuses, lips, salivary glands, oropharynx, hypopharynx, pharynx, larynx, and local lymph nodes [1]. It is the sixth most common cancer in the world with more than 500,000 cases reported annually [2, 3]. HNC has found to be more prevalent among males than females [3]. More than 90% of all the reported cases of HNC are squamous cell carcinomas (HNSCCs) which usually arise from mucosal lining in these sites [1]. Among these cases, 80–90% cases were found to be associated with prolonged usage of alcohol and tobacco [4].

One of the frequently diagnosed malignancies, globally, is head and neck cancer. Frequent relapses and distant metastasis have been observed in patients with advanced disease stages, resulted in an endurance rate of 5 years in almost 60% patients despite of substantial advances in multimodality rehabilitation [5]. Mucosal malignancy of head and neck is a potent mela‐ noma with poor prognosis. The nasal pit, paranasal sinuses, and oral cavity are the most widely recognized areas. Survival rates of 1, 3, and 5 years were 63, 30, and 20%, respectively, around 2000–2007. All interpretations suggested that cigarette smoking is hazardous. Clinical marks and clues are typically nonspecific for it. Surgery is regarded as the backbone of treat‐ ment for most mucosal melanomas of the head and neck supplemented with radiotherapy [6]. The relationship amongst alcohol consumption and head and neck disease is quite clear and the outcomes reliably demonstrated an expanded head and neck tumor chance related with alcohol drinking. Bagnardi et al. reported a positive relationship in between alcohol drinking and head and neck cancer risk [7]. In Western nations, around 39% of head and neck malig‐ nancy can be ascribed to alcohol utilization (4% for only alcohol drinking and 35% for the joint impact of alcohol and tobacco) [8]. Head and neck squamous cell carcinomas (HNSCCs) cause more than 300,000 deaths globally every year. Locoregional and removed recurrences are more important prognostic indicators and acknowledged surrogate markers of patients' survival. No legitimate biomarker and rescue treatment exist to recognize and treat patients at high‐danger of recurrence [9]. Despite of reduction in smoking and alcohol utilization, the rate of oropharyngeal squamous cell carcinoma (OPSCC) is rising. It refers to human pap‐ illoma virus (HPV) infection contamination [10]. HPV is an entrenched prognostic marker for OPSCC [11]. Oropharyngeal tumors are firmly connected with HPV‐positivity [12]. Oral tumor constitutes the dominant part of head and neck diseases, which is the fifth most basic malignancy around the world, representing 984,430 cases in 2012. During 2000 and 2010, there were 1916 instances of OSCC in New Zealand with a male to female proportion of 1.85:1, and an age‐institutionalized rate of 42 for every 1,000,000 people [13]. Liquor utilization, trailed by tobacco, is considered the most common hazard in New Zealand. Given the high pervasive‐ ness of these two hazardous elements and their synergistic impact, it is vital for specialists to boost smoking cessation and limited liquor consumption. More research should be conducted to confirm use of tobacco and water‐pipe smoking in New Zealand, particularly because of changing demography and increments in transient populaces. UV radiation is an addition‐ ally imperative hazard element [13].  Rates of LSCC have been estimated to escalate recently [14]. Salivary gland pleomorphic adenoma (SGPA) is also one of the most widely recognized types of salivary organ tumor. In China, particularly in the South, nasopharyngeal cancer (NPC) is another widely recognized threatening tumors and hence remained unregistered even by National cancer registries, since little is known about its epidemiology [15]. The occurrence repre‐ sented around 40% of the world's new cases as indicated by the World Health Organization's GLOBOCAN revealed information of 2012 [16]. Individuals with a family history of NPC have a generously higher danger of NPC [17]. NPC's mortality indicates marked distinction between endemic (highly vulnerable territories), where nonkeratinizing carcinoma (NKC) is pervasive, and nonendemic (safe districts), where the keratinizing squamous cell carcinoma (KSCC) is more frequent. Fluctuations in smoking and alcohol consumption amongst genders and geographic regions may clarify the diverse rates and patterns fully observed for KSCC and partially for NKC. Dietary patterns and improvement in disease management can also be accountable for observed trends [18].

## **2. MicroRNAs discovery**

**1. Head and neck cancer epidemiology**

6 Diagnosis and Management of Head and Neck Cancer

alcohol and tobacco [4].

Head and neck cancer (HNC) is referred to the cancers of aerodigestive tract covering number of structures *viz*, oral and nasal cavity, paranasal sinuses, lips, salivary glands, oropharynx, hypopharynx, pharynx, larynx, and local lymph nodes [1]. It is the sixth most common cancer in the world with more than 500,000 cases reported annually [2, 3]. HNC has found to be more prevalent among males than females [3]. More than 90% of all the reported cases of HNC are squamous cell carcinomas (HNSCCs) which usually arise from mucosal lining in these sites [1]. Among these cases, 80–90% cases were found to be associated with prolonged usage of

One of the frequently diagnosed malignancies, globally, is head and neck cancer. Frequent relapses and distant metastasis have been observed in patients with advanced disease stages, resulted in an endurance rate of 5 years in almost 60% patients despite of substantial advances in multimodality rehabilitation [5]. Mucosal malignancy of head and neck is a potent mela‐ noma with poor prognosis. The nasal pit, paranasal sinuses, and oral cavity are the most widely recognized areas. Survival rates of 1, 3, and 5 years were 63, 30, and 20%, respectively, around 2000–2007. All interpretations suggested that cigarette smoking is hazardous. Clinical marks and clues are typically nonspecific for it. Surgery is regarded as the backbone of treat‐ ment for most mucosal melanomas of the head and neck supplemented with radiotherapy [6]. The relationship amongst alcohol consumption and head and neck disease is quite clear and the outcomes reliably demonstrated an expanded head and neck tumor chance related with alcohol drinking. Bagnardi et al. reported a positive relationship in between alcohol drinking and head and neck cancer risk [7]. In Western nations, around 39% of head and neck malig‐ nancy can be ascribed to alcohol utilization (4% for only alcohol drinking and 35% for the joint impact of alcohol and tobacco) [8]. Head and neck squamous cell carcinomas (HNSCCs) cause more than 300,000 deaths globally every year. Locoregional and removed recurrences are more important prognostic indicators and acknowledged surrogate markers of patients' survival. No legitimate biomarker and rescue treatment exist to recognize and treat patients at high‐danger of recurrence [9]. Despite of reduction in smoking and alcohol utilization, the rate of oropharyngeal squamous cell carcinoma (OPSCC) is rising. It refers to human pap‐ illoma virus (HPV) infection contamination [10]. HPV is an entrenched prognostic marker for OPSCC [11]. Oropharyngeal tumors are firmly connected with HPV‐positivity [12]. Oral tumor constitutes the dominant part of head and neck diseases, which is the fifth most basic malignancy around the world, representing 984,430 cases in 2012. During 2000 and 2010, there were 1916 instances of OSCC in New Zealand with a male to female proportion of 1.85:1, and an age‐institutionalized rate of 42 for every 1,000,000 people [13]. Liquor utilization, trailed by tobacco, is considered the most common hazard in New Zealand. Given the high pervasive‐ ness of these two hazardous elements and their synergistic impact, it is vital for specialists to boost smoking cessation and limited liquor consumption. More research should be conducted to confirm use of tobacco and water‐pipe smoking in New Zealand, particularly because of changing demography and increments in transient populaces. UV radiation is an addition‐ ally imperative hazard element [13]. Laryngeal squamous cell carcinoma (LSCC), being a

MicroRNAs (miRNAs) are single‐stranded, small RNA molecules whose presence was reported for the first time in 1993 in nematode *Caenorhabditis elegans*. A number of biological activities like gene regulation, differentiation, organ formation, cell death, cell proliferation, and stress responses have been reported to be influenced by these miRNAs [19–22]. These miR‐ NAs were found to regulate translation in larval development through an antisense RNA‐RNA interaction [19, 23, 24]. 1600 miRNAs of *Homo sapiens* have been reported and recorded by miRBase database in June 2013 [25]. Since 1970s, studies to determine and understand the function and genetics of gene regulation were performed [26]. *C*. *elegans* was used as a genetic model and tool to facilitate these kinds of ventures. More than 300 mutants of *C*. *elegans* with many developmental/birth defects and certain behavioral changes were generated by series of groundbreaking experiments by Brenner and Sulston [26, 27]. One of them was character‐ ized by reverberation of specific cell lineages and showing a flaccid and extremely elongated body [27, 28]. The nature of the gene(s) and the exact mechanism underlying such morpho‐ logical and behavioral changes were unknown at that time. This mutant was considered as the founding member of the miRNA family and was named as lin‐4. Lin‐4 was reported to influence specific developmental responses in a variety of cell types of *C*. *elegans* larva [29, 30]. In 1993, cloning of lin‐4 locus took about 2 decades after its early description [23]. Lin‐4 locus was found unique having characteristics different from other coding genes. It was found employing site‐directed mutagenesis, that lin‐4 gene encodes a small RNA molecule instead of a protein. Furthermore, the transcripts of lin‐4 were of smaller size compared to other genes as the two transcripts identified were of 22 and 60 nucleotides only. To suffice, lin‐4 mRNA transcripts were found to be negatively regulating the lin‐14 expression, as it possesses anti‐ sense complementarity to a number of sites in 3′untranslated region (UTR) to lin‐14 gene [23]. Therefore, these discoveries lead to a novel class of small noncoding RNA molecules, which via antisense‐like interaction regulate a number of pathways [31–33]. After 7 years following lin‐4 cloning and characterization, a second gene was discovered with similar characters [34]. The study resulted in the isolation of let‐7 (21‐nt RNA molecule) that was characterized by acting as a heterochronic switch in *C*. *elegans* development. It was fascinating that the expres‐ sion of this miRNA was reported in a wide range of animals other than *C*. *elegans*, including arthropods, mollusks, and mammals [35]. Further studies reported more than 100 different noncoding RNAs (ncRNAs) in a variety of species across the animal phyla [36–38]. A number of important observations were made from these key studies. First, specific small temporal RNAs (stRNAs) express only in few stages of development and are about 22 nt long [38]. Second, cell‐type specificity of different stRNAs was determined [38]. For example, miR‐1 (a small RNA) exclusively expressed only in cardiac tissues [37]. Presence of homologs of these molecules across a wide range of species is an indication of evolutionary conservation [38]. Abundance and wide existence of these unique molecules represents a novel sequence‐specific posttranscriptional gene regulation. So, based on above given account, the small RNAs with similar functions were named as microRNAs (miRNAs) [36–38]. miRNA gene family is increasing day by day since its discovery [39]. Therefore, a number of databases have been generated to cope with the ever‐growing list of miRNA genes. The first ever miRNA database is known as "miRBase (http://microrna.sanger.ac.uk/sequences/)" and it has access to clone and register miRNA sequences [40]. The total number of miRNAs is exceeding ten thousands in number up to date.

#### **3. MicroRNAs biology**

miRNAs are small ncRNAs of about 19–25 nucleotides and have been discovered recently in all metazoans tested so far [41, 42]. They comprised of a small fraction of expressed genome; however, genes encoding them are present in either introns or exons of coding as well as noncoding genes throughout the genome. miRNAs are involved in regulating number of processes including differentiation, maintenance of homeostasis, migration, pro‐ grammed cell death, and morphogenesis [42–45]. miRNAs are transcribed with the aid of RNA polymerase II enzyme from the miRNAs genes, which in turn transcribed into pri‐ mary microRNA transcripts following transformation into pre‐miRNAs which are then ultimately results in miRNAs [32, 46–50]. In recent years, a novel class of molecules naming miRNAs has showed a boom. With advancements in genomic technologies and methodolo‐ gies, miRNAs gene family is rising as new members discovered day by day. In 2005, it was speculated that there are about 1000 miRNAs genes in human genome [51]. However, 2042 mature miRNAs have been reported in miRBase up to date (version 19). miRNAs are char‐ acterized by having 21–23 nucleotides in length and are exclusively found in eukaryotes. They show partial complementarity to specific regions in targeted mRNA and hence are involved in gene regulation of 50% of human genome at posttranscriptional level [52]. Each of these noncoding RNAs can have sequence complementary to a large number of tran‐ scripts and regulate expression of an enormous number of genes. This can be achieved by multiple mechanisms including an increased mRNA degradation, site‐specific cleavage, and translational inhibition [53]. miRNAs dysregulation has been associated with a num‐ ber of different types of cancer, their initiation, and proliferation, since the first ever study claiming the association between chronic lymphoid leukemia and miR‐15a and miR‐16‐1 in the year 2002 [54–56]. Therefore, they may prove as potent therapeutic targets for cancers including HNSCC as well as biomarkers for timely diagnosis, prognosis, and recurrence of cancers as well. Two different pathways have been reported for the biogenesis of miRNAs. Usually, a canonical pathway is used for their biogenesis as most of the miRNA genes are intergenic (**Figure 1**). These miRNAs when transcribed possess a local hairpin structure and are named as primary transcripts (pri‐miRNA) and have characteristics to that of mRNA, as there is a 5′cap and a 3′poly‐A tail. Drosha (a protein complex having nuclease activ‐ ity) and Pasha (an RNA‐binding protein) process these pri‐miRNAs to pre‐miRNAs hav‐ ing 70‐nucleotide stem‐loop structure. pre‐miRNAs are converted into a double‐stranded miRNA/miRNA\* duplex by an RNase III endonuclease (Dicer) followed by transportation in cytoplasm by Exportin‐5. Further, a helicase destabilizes the duplex miRNA/miRNA\* to a mature miRNA and miRNA\*. Finally, the mature miRNA then integrates into RNA‐ induced silencing complex (RISC) also called miRNA ribonucleoprotein complex (miRNP).

via antisense‐like interaction regulate a number of pathways [31–33]. After 7 years following lin‐4 cloning and characterization, a second gene was discovered with similar characters [34]. The study resulted in the isolation of let‐7 (21‐nt RNA molecule) that was characterized by acting as a heterochronic switch in *C*. *elegans* development. It was fascinating that the expres‐ sion of this miRNA was reported in a wide range of animals other than *C*. *elegans*, including arthropods, mollusks, and mammals [35]. Further studies reported more than 100 different noncoding RNAs (ncRNAs) in a variety of species across the animal phyla [36–38]. A number of important observations were made from these key studies. First, specific small temporal RNAs (stRNAs) express only in few stages of development and are about 22 nt long [38]. Second, cell‐type specificity of different stRNAs was determined [38]. For example, miR‐1 (a small RNA) exclusively expressed only in cardiac tissues [37]. Presence of homologs of these molecules across a wide range of species is an indication of evolutionary conservation [38]. Abundance and wide existence of these unique molecules represents a novel sequence‐specific posttranscriptional gene regulation. So, based on above given account, the small RNAs with similar functions were named as microRNAs (miRNAs) [36–38]. miRNA gene family is increasing day by day since its discovery [39]. Therefore, a number of databases have been generated to cope with the ever‐growing list of miRNA genes. The first ever miRNA database is known as "miRBase (http://microrna.sanger.ac.uk/sequences/)" and it has access to clone and register miRNA sequences [40]. The total number of miRNAs is exceeding ten thousands

miRNAs are small ncRNAs of about 19–25 nucleotides and have been discovered recently in all metazoans tested so far [41, 42]. They comprised of a small fraction of expressed genome; however, genes encoding them are present in either introns or exons of coding as well as noncoding genes throughout the genome. miRNAs are involved in regulating number of processes including differentiation, maintenance of homeostasis, migration, pro‐ grammed cell death, and morphogenesis [42–45]. miRNAs are transcribed with the aid of RNA polymerase II enzyme from the miRNAs genes, which in turn transcribed into pri‐ mary microRNA transcripts following transformation into pre‐miRNAs which are then ultimately results in miRNAs [32, 46–50]. In recent years, a novel class of molecules naming miRNAs has showed a boom. With advancements in genomic technologies and methodolo‐ gies, miRNAs gene family is rising as new members discovered day by day. In 2005, it was speculated that there are about 1000 miRNAs genes in human genome [51]. However, 2042 mature miRNAs have been reported in miRBase up to date (version 19). miRNAs are char‐ acterized by having 21–23 nucleotides in length and are exclusively found in eukaryotes. They show partial complementarity to specific regions in targeted mRNA and hence are involved in gene regulation of 50% of human genome at posttranscriptional level [52]. Each of these noncoding RNAs can have sequence complementary to a large number of tran‐ scripts and regulate expression of an enormous number of genes. This can be achieved by multiple mechanisms including an increased mRNA degradation, site‐specific cleavage,

in number up to date.

**3. MicroRNAs biology**

8 Diagnosis and Management of Head and Neck Cancer

The resultant miRNP is involved in RNA interference which is initiated by both siRNA and miRNA. In case of intronic stem‐loops‐derived miRNA, no Drosha activity is involved for the maturation of miRNAs [32, 57] (**Figure 2**).

**Figure 1.** Canonical pathway of miRNAs biogenesis. miRNAs are transcribed into a primary transcript (pri‐miRNA) which is processed into pre‐miRNAs by Drosha/Pasha complex. After transportation to cytoplasm, these pre‐miRNAs are converted into a double‐stranded miRNA/miRNA\* duplex by Dicer. Further, a helicase destabilizes the duplex miRNA/miRNA\* to a mature miRNA and miRNA\*. Finally, the mature miRNA then integrates into RISC.

**Figure 2.** Noncanonical pathway of miRNAs biogenesis. In noncanonical pathway of miRNAs biogenesis, no Drosha activity is involved. Pre‐miRNAs are directly produced from the debranching of the introns; and after their transportation, they are processed similar to that produced by canonical pathway.

## **4. MicroRNAs deregulation in head and neck cancer**

The first ever study involving miRNAs in HNC was published in 2005 [58]. miRNAs showed greater cancer‐related potential than expected. miRNAs have been associated with all processes of physiological and pathological nature. Expression profiles of miRNAs have reported to be more specific to cancer tissue origin and have greater potential for early diagnosis and provided more information than mRNAs [59, 60]. Association between dysregulation of miRNAs and tumorigenesis in head and neck has been documented previously [61–63]. Many biological and molecular mechanisms cause resistance to the tumors radiotherapy. The principle mechanisms include changes in intracellular pathways that are required in damaging DNA as well as its repair, apoptosis, proliferation, and angiogenesis. The regulation of these perplexing proce‐ dures is regularly controlled by microRNAs. miRNAs are short endogenous RNA molecules that posttranscriptionally modulate gene expression. Their impaired expression has been seen in numerous tumors including head and neck cancer. Particular expression patterns of miRNAs have additionally been appeared to anticipate prognosis and therapeutic response in head and neck cancer [64]. Development and progression of different sorts of malignancies in human is attributed to the deregulation of miRNAs. Oncogenic part of miR‐214 is proposed in NPC. Bax inhibition initiated by siRNA weakens the advancing impact of miR‐214 deregulation on NPC cell apoptosis, recommending that Bax is a downstream effector in miR‐214 that is inter‐ vened NPC cell proliferation and death. Bax expression level is deregulated in NPC tissues [65]. miR‐145 expression in LSCC is downregulated, and its overexpression creates hindrance in multiplication and relocation of Hep‐2 cells through cell cycle arrest and apoptotic induction. SOX‐2 is overexpressed in tumor samples and exhibit restricted expression in miR‐145 overex‐ pressed Hep‐2 cells [14]. miRNAs deregulation plays a key role in HNSCC progression [66]. miR‐93‐5p (HN2092) and miR‐425‐5p (HN1957) are top candidates of downregulated miRNAs in primary HNSCC cell cultures and blood plasma of patients [67]. miRNAs deregulation plays a noteworthy part in head and neck/oral cancer [68]. miR‐10b and miR‐196a, not formerly linked with HNSCC, may show a role in oncogenesis through the deregulation of cell proliferation [69]. Deregulation of miRNA genes (such as miR‐138) plays a critical part in HNSCC. While down‐ regulation of miR‐138 has been observed in HNSCC and other cancer types often; however, the exact role of miR‐138 in tumorigenesis is unknown. Current bioinformatics analyses and *in vitro* and *in vivo* researches have recognized a number of functional targets for miR‐138. These include genes that take part in necessary biological processes that are exceptionally pertinent to the onset and proliferation of HNSCC, including cell migration, epithelial to mesenchymal transition, cell cycle progression, DNA damage and repair, senescence, and differentiation [70]. Mechanistic target of rapamycin (mTOR) and Insuline like growth factor 1 receptor (IGF1R) signaling pathways depict that the downregulation of miR‐99 family adds to HNSCC tumori‐ genesis [71]. Meta‐analysis of various Gene expression omnibus (GEO) datasets revealed that Uracil DNA glycosylation (UNG), Fucosidase alpha‐L‐2 plasma (FUCA2), Differentially expressed regulation analysis (DERA), Glia maturation factor beta (GMFB), Transferrin (TF) and Sorting nexin 2 (SNX2) were usually downregulated in HNSCC [72]. miRNAs including miR‐21 and miR‐31 are progressively deregulated in tongue epithelium [73]. Underexpression of miR‐375 might play oncogenic roles in HNSCC [74]. miR‐29c is considerably deregulated in

**4. MicroRNAs deregulation in head and neck cancer**

transportation, they are processed similar to that produced by canonical pathway.

10 Diagnosis and Management of Head and Neck Cancer

The first ever study involving miRNAs in HNC was published in 2005 [58]. miRNAs showed greater cancer‐related potential than expected. miRNAs have been associated with all processes of physiological and pathological nature. Expression profiles of miRNAs have reported to be more specific to cancer tissue origin and have greater potential for early diagnosis and provided more information than mRNAs [59, 60]. Association between dysregulation of miRNAs and tumorigenesis in head and neck has been documented previously [61–63]. Many biological and molecular mechanisms cause resistance to the tumors radiotherapy. The principle mechanisms include changes in intracellular pathways that are required in damaging DNA as well as its repair, apoptosis, proliferation, and angiogenesis. The regulation of these perplexing proce‐ dures is regularly controlled by microRNAs. miRNAs are short endogenous RNA molecules that posttranscriptionally modulate gene expression. Their impaired expression has been seen in numerous tumors including head and neck cancer. Particular expression patterns of miRNAs have additionally been appeared to anticipate prognosis and therapeutic response in head and neck cancer [64]. Development and progression of different sorts of malignancies in human is attributed to the deregulation of miRNAs. Oncogenic part of miR‐214 is proposed in NPC. Bax inhibition initiated by siRNA weakens the advancing impact of miR‐214 deregulation on NPC cell apoptosis, recommending that Bax is a downstream effector in miR‐214 that is inter‐ vened NPC cell proliferation and death. Bax expression level is deregulated in NPC tissues [65]. miR‐145 expression in LSCC is downregulated, and its overexpression creates hindrance

**Figure 2.** Noncanonical pathway of miRNAs biogenesis. In noncanonical pathway of miRNAs biogenesis, no Drosha activity is involved. Pre‐miRNAs are directly produced from the debranching of the introns; and after their

**Table 1.** Recently identified miRNAs that undergo deregulation in HNC.

NPC. However, there is dearth of knowledge regarding outcome and molecular mechanisms of action of miR‐29c downregulation in development and progression of NPC [75]. Some recent studies indicating miRNAs deregulation in HNC are given in **Table 1**.

## **5. miRNAs as oncogenes in oral squamous cell carcinoma (OSCC)**

A number of miRNAs have been reported to act as oncogenes and were found to be upregu‐ lated in OSCC. miR‐21 was found to be highly expressed and to regulate several biological activities in OSCC [68, 92–94]. Presence of an upregulated level of miR‐21 in oral premalig‐ nant lesions was an indication that variations in miR‐21 level may be prior event in OSCC development [95]. miR‐21 plays an oncogenic role in the progression of OSCC by promoting cell proliferation [96], antiapoptotic activity [92], invasion [93, 97], and chemoresistance [98] in both *in vitro* and *in vivo* studies.

Oral squamous cell carcinoma (OSCC) is a typical cause of cancer‐related death. A number of efforts have been made in investigating new medications and impressive progress in multimodality treatment; however, remedial tolls have not yet been achieved. The trouble of timely detection and more pervasiveness of metastasis associated with OSCC results in poor prognosis. In recent couple of decades, growing information from tumor biology and clinical trials prompted development of ncRNAs prognostic biomarkers that are believed to be promising biomarkers in this regard. miRNAs are one of the most studied ncRNAs in terms of their biogenesis, function, and significance in carcinogenesis [99]. An association between severity of pathogenesis and increment of miR‐31 and miR‐21 has been reported through staining in 4NQO‐induced injury in tongue epithelium. A dynamic rise in the level of *miR‐21*, *miR‐31*, and *miR‐146a* in saliva and plasma samples was noted. miR‐31 was the earliest miRNA to be released in the saliva. A rise in plasma level of miR‐146a, miR‐372, and miR‐184 was found and it was prominent at the most progressive lesion state [73]. miRNA deregulation assists in pathogenesis of various disorders, including human tongue squamous cell carcinoma (TSCC), where they act as powerful oncogenes or tumor suppres‐ sors. Widespread miRNA profiling in TSCC samples and further *in vitro* and *in vivo* func‐ tional studies unveiled their involvement to hidden molecular mechanisms in initiation, development, progression, metastasis, chemoradioresistance, and relapse of TSCC [100]. An upregulated expression of miR‐483‐5p in OSCC patient sera might be a novel indica‐ tive and prognostic biomarker for this ailment [101]. The upregulation of miR‐9 was also detected in primary esophageal squamous cell carcinoma (ESCC) tumor tissue. miR‐9 pro‐ motes cell migration and tumor metastasis, as these processes effectively retarded when expression of miR‐9 was deregulated. Furthermore, it was established that miR‐9 inter‐ acts with the 3′untranslated region of E‐cadherin and downregulates its expression, which leads to β‐catenin nuclear translocation and consequently upregulates c‐myc and CD‐44 expression. Moreover, an ESCC miR‐9 results in epithelial‐mesenchymal transition (EMT) in ESCC, a key occasion in tumor metastasis [102]. Significantly, low level of miR‐29b has been reported. However, overexpression of miR‐29b repressed the proliferation, migration, invasion, and progression of TSCC cells, and slowed down the cell death. In addition, miR‐29b brings about deregulation of Sp1 by targeting the 3′untranslated part that causes upregulation of phosphatase and tensin homolog (PTEN), and subsequently inhibit phosphorylation of Protein kinase B (AKT). Sp1 knockdown in TSCC cell lines reflected the effects of miR‐29b overexpression. miR‐29b expression is inversely related to Sp1 expres‐ sion and positively correlated with PTEN. Thus, miR‐29b works as a tumor suppressor, and miR‐29b/Sp1/PTEN/AKT alliance may speak of a possible therapeutic target for TSCC prevention [91]. MiR‐99a deregulation was confirmed in oral cancer cell lines and clinical specimen as well. Ectopic miR‐99a expression resulted in repression of oral cancer cell migration and invasion. Myotubularin‐related protein 3 (MTMR3) with one evolutionarily

NPC. However, there is dearth of knowledge regarding outcome and molecular mechanisms of action of miR‐29c downregulation in development and progression of NPC [75]. Some recent

**Sample used Assay Type of cancer Reference**

miR‐9 Cell lines, tissues qRT‐PCR NPC [76] miR‐378 Cell lines, tissues qRT‐PCR NPC [77] miR‐200 family Cell lines Microarray NPC [78] miR‐451 Tissue qRT‐PCR NPC [79] MiR‐99a Tissue, cell lines Luciferase assay Oral cancer [80] qRT‐PCR Western blot

miR‐24 Tissue, cell lines RT‐PCR Oral cancer [81]

miR‐16, miR‐125b Tissues Microarray OSCC [83]

miR‐138, miR‐222 Cell lines Microarray TSCC [85, 86] miR‐342, miR‐21 Cell line Microarray TSCC [87]

miR‐494 Tissue Microarray HNSCC [62] miR‐375 Tissue Microarray HNSCC [89] miR‐125a, miR‐125b Tissue Microarray HNSCC [90] miR‐133a, miR‐205 Tissue Microarray HNSCC [91] miR‐125b, miR‐375 Tissue qRT‐PCR HNSCC [75]

Tissues qRT‐PCR TSCC [82]

Cell lines qRT‐PCR OSCC [84]

Cell lines Microarray HNSCC [88]

Microarray, qRT‐PCR NPC [76]

A number of miRNAs have been reported to act as oncogenes and were found to be upregu‐ lated in OSCC. miR‐21 was found to be highly expressed and to regulate several biological

**5. miRNAs as oncogenes in oral squamous cell carcinoma (OSCC)**

studies indicating miRNAs deregulation in HNC are given in **Table 1**.

**Table 1.** Recently identified miRNAs that undergo deregulation in HNC.

**Downregulated miRNAs**

miR‐133a, miR‐133b, miR‐100, miR‐138

miR‐133b, miR‐138, miR‐137, miR‐184

miR‐342, miR‐346, miR‐373

miR‐29c Cell lines, clinical

12 Diagnosis and Management of Head and Neck Cancer

specimens

preserved seed locale in the 3′untranslated region was a novel miR‐99a target. Draining MTMR3 expression fundamentally diminishes cell proliferation, migration, and invasion. An inverse relation was found among miR‐99a and MTMR3 protein in oral cancer lines and clinical patients. miR‐99a has been found to suppress oral cancer cell migration and inva‐ sion partially through inhibiting MTMR3 expression [103]. Keratinization of tumors and overexpression of miR‐21 are major factors responsible for poor prognosis. Interestingly, most of the keratinized tumors were reported to express an elevated miR‐21 level [104]. Moreover, an upregulated level of miR‐127 and a reduced level of miR‐357 have been reported in OSCC [105]. In CD‐44 (high) oral CSCs, miR‐200s/miR‐205s were epigeneti‐ cally triggered in tumors and their expression was found to be suppressed in the absence of DNA hypermethylation [105]. miRNAs expressions and DNA methylation variations are typical occasion in OSCC, and miR‐375, miR‐127, miR‐137, miR‐205, and miR‐200 family are promising candidates for future investigations. miR‐200/miR‐205 downregulation in oral CSCs specify that cell‐specific silencing of these miRNAs may enhance tumor expansion and progression [105]. Deregulation of miR‐24 was found to be associated with high‐grade late stage tumor [92].

## **6. MicroRNAs in human nasopharyngeal carcinoma (NPC)**

Paul Ahlquist's group working at National Cancer Institute (NCI) published the first ever study of global profiling of miRNAs involved in NPC in 2008 [88]. They discovered a num‐ ber of deregulated miRNAs, using a micro‐array‐based approach, in laser‐capture micro dissected (LCM) 31 NPCs and 10 epithelial samples as control [88]. miRNA downregulation and change in pathways have been involved in NPC which is profoundly invasive and met‐ astatic widespread in Southern China. miR‐9 is commonly downregulated in NPC with sig‐ nificant functional consequences. Diminished expression of miR‐9 is contrary to clinical stages and denotes the movement from locoregional to metastatic tumors. CpG island hypermethylation adds to inhibition of miR‐9 in NPC cell lines and tissues. Ectopic miR‐9 expression significantly hinders proliferative, transient, and obtrusive limits of NPC cells, both *in vitro* and *in vivo*. miR‐9 strongly reduces expression of CXCR4 in NPC cells. miR‐9 works as a tumor‐suppressor in NPC, and is interfered by combating CXCR4 expression [106]. NPC is exceptional worldwide, yet profoundly intrusive in later stages. It cannot be identified in normal medical examination at initial stages. Advancement of particular bio‐ markers ought to spare lives against this sort of ailment. Among them, increased levels of miR‐16, miR‐21, miR‐24, and miR‐155 and decreased level of miR‐378 have been observed in NPC patients. Plasma miRNA expression and cancer progression are negatively correlated. Blend of miR‐16, miR‐21, miR‐24, miR‐378, and miR‐155 gives affectability of 87.7% and specificity of 82.0% for marking NPC. Except miR‐16, mixture of the rest of four miRNAs gives a similar affectability, however, a somewhat diminished specificity. After treatment, levels of the five miRNAs were reverted to normal levels [88]. Regardless of the fact that enormous miRNAs have been discovered in the previous decade, with the progressions in DNA sequencing, the discovery of some more NPC‐related miRNAs is expected. Circulating miRNAs related with NPC are quite compelling, as they present an exploitable instrumenta‐ tion for early detection, diagnosis, and staging. A few worldwide profiling depicted the altered miRNA expression in a range of head and neck malignancies. In NPC, downregula‐ tion of the tumor silencer miRNAs (miR‐29c, miR‐9, let‐7 family, miR‐200 family), and over‐ expression of oncogenic miRNAs, (miR‐18a/b, miR‐141, miR‐155, miR‐214) have been observed [107]. miR‐9 is the most concerned miRNA in NPC pathogenesis [108], which works in the direction of various crucial cellular processes, facilitating proliferation, apop‐ tosis, incursion, metastasis, angiogenesis, and EMT [109–111]. Restricted expression of miR‐ 9in NPC is related with phenotypes that are more destructive and lesser survival. miR‐9 has been appeared to work as a tumor silencer in NPC by focusing on chemokine (CXC theme) receptor 4 (CXCR4) to repress cell multiplication, relocation, and invasion [87]. Similarly, miR‐9 directs innate response in NPC through control of various genes that are activated by interferon, and in addition major histocompatibility complex (MHC) class I members [112]. Imperatively, the capacity of miR‐9 to work as an individual prognostic biomarker for NPC metastasis has additionally been illustrated, whereas miR‐9 expression is related with decreased expansion, movement, and intrusion in NPC cells, and low‐levels of miR‐9 expression are associated with last stage [106]. Correspondingly, miR‐200 family is deregu‐ lated in NPC, bringing about expanded cell development, relocation, and intrusion of NPC cells according to suppression of the putative miR‐200 targets zinc finger E‐box restricting homeobox 2 (ZEB2), beta 1 (CTNNB1), and catenin (cadherin‐related protein) bringing about expanded NPC cell development, movement, and invasion. Moreover, diminished miR‐200a expression is related with initiating epithelial‐mesenchymal transition [89]. Various miRNAs have been reported as tumor silencers/suppressors in NPC, including miR‐34c, miR‐451, miR‐98, miR‐216b, miR‐375, and miR‐26a. miR‐375 has been accounted for a potential tumor silencer in NPC, working through restraint of the oncogenic protein metadherin (MTDH). Strangely, NPC cases with MTDH overexpression showed an expanded danger of recurrence of disease [113]. Low miR‐451 expression was related with diminished survival in NPC patients, and has been appeared to work by expanding cell development and invasion by focusing on macrophage migration inhibitory factor (MIF) in NPC cells [114]. miR‐26a, miR‐98, and miR‐101 also have been accounted to work as tumor suppressors in NPC [98]. Underexpression of these miRNAs in NPC gives clues about dere‐ pression of Enhancer of zeste homolog 2 (EZH2), instigating loss of suppression of targets of EZH2 regulation, comprising cyclins D3, c‐Myc, and E2, and cyclin‐dependent kinase 6 (CDK6) and CDK4 [115, 116]. Finally, miR‐216b has been appeared to increase NPC growth and incursion by pursuing Kirstan rat sarcoma (K‐RAS) [117]; on the other hand, miR‐34c has been revealed to stifle growth and metastasis by focusing on the proto‐oncogene MET [118], both of which are ominously downregulated in primary NPC tissues. In NPC, miR‐ 18a is profoundly overexpressed [119], which results in direct hang‐up of the miRNA bio‐ genesis regulatory protein Dicer1, leading to the downregulation of miRNA in NPC [119]. Likewise, miR‐18b is overexpressed in NPC, accompanying malady advancement and poor outcome. miR‐18b functions to inhibit connective tissue growth factor (CTGF) and thereby augmenting cellular proliferation [120]. Over‐articulation of miR‐141 in NPC has been con‐ nected with more cell growth, relocation, and invasion, and also with loss of cell cycle

preserved seed locale in the 3′untranslated region was a novel miR‐99a target. Draining MTMR3 expression fundamentally diminishes cell proliferation, migration, and invasion. An inverse relation was found among miR‐99a and MTMR3 protein in oral cancer lines and clinical patients. miR‐99a has been found to suppress oral cancer cell migration and inva‐ sion partially through inhibiting MTMR3 expression [103]. Keratinization of tumors and overexpression of miR‐21 are major factors responsible for poor prognosis. Interestingly, most of the keratinized tumors were reported to express an elevated miR‐21 level [104]. Moreover, an upregulated level of miR‐127 and a reduced level of miR‐357 have been reported in OSCC [105]. In CD‐44 (high) oral CSCs, miR‐200s/miR‐205s were epigeneti‐ cally triggered in tumors and their expression was found to be suppressed in the absence of DNA hypermethylation [105]. miRNAs expressions and DNA methylation variations are typical occasion in OSCC, and miR‐375, miR‐127, miR‐137, miR‐205, and miR‐200 family are promising candidates for future investigations. miR‐200/miR‐205 downregulation in oral CSCs specify that cell‐specific silencing of these miRNAs may enhance tumor expansion and progression [105]. Deregulation of miR‐24 was found to be associated with high‐grade

**6. MicroRNAs in human nasopharyngeal carcinoma (NPC)**

Paul Ahlquist's group working at National Cancer Institute (NCI) published the first ever study of global profiling of miRNAs involved in NPC in 2008 [88]. They discovered a num‐ ber of deregulated miRNAs, using a micro‐array‐based approach, in laser‐capture micro dissected (LCM) 31 NPCs and 10 epithelial samples as control [88]. miRNA downregulation and change in pathways have been involved in NPC which is profoundly invasive and met‐ astatic widespread in Southern China. miR‐9 is commonly downregulated in NPC with sig‐ nificant functional consequences. Diminished expression of miR‐9 is contrary to clinical stages and denotes the movement from locoregional to metastatic tumors. CpG island hypermethylation adds to inhibition of miR‐9 in NPC cell lines and tissues. Ectopic miR‐9 expression significantly hinders proliferative, transient, and obtrusive limits of NPC cells, both *in vitro* and *in vivo*. miR‐9 strongly reduces expression of CXCR4 in NPC cells. miR‐9 works as a tumor‐suppressor in NPC, and is interfered by combating CXCR4 expression [106]. NPC is exceptional worldwide, yet profoundly intrusive in later stages. It cannot be identified in normal medical examination at initial stages. Advancement of particular bio‐ markers ought to spare lives against this sort of ailment. Among them, increased levels of miR‐16, miR‐21, miR‐24, and miR‐155 and decreased level of miR‐378 have been observed in NPC patients. Plasma miRNA expression and cancer progression are negatively correlated. Blend of miR‐16, miR‐21, miR‐24, miR‐378, and miR‐155 gives affectability of 87.7% and specificity of 82.0% for marking NPC. Except miR‐16, mixture of the rest of four miRNAs gives a similar affectability, however, a somewhat diminished specificity. After treatment, levels of the five miRNAs were reverted to normal levels [88]. Regardless of the fact that enormous miRNAs have been discovered in the previous decade, with the progressions in DNA sequencing, the discovery of some more NPC‐related miRNAs is expected. Circulating

late stage tumor [92].

14 Diagnosis and Management of Head and Neck Cancer

control and decreased apoptosis. This regulation is thought to happen through downregula‐ tion of the supposed target gene phosphatase and tensin homolog (PTEN), bromo‐domain 3 (BRD3), and ubiquitin‐associated protein 1 (UBAP1) in NPC. Moreover, expression of miR‐141 is structured by the oncogenes short palate, lung, nasal epithelium clone 1 (SPLUNC1), and c‐Myc [120]. Expression of miR‐155 fortifies proliferation, migration, and incursion by regulation of target genes Jumonji Domain 1A (JMJD1A) and BTB and CNC homology 1 (BACH1), and its expression has been strongly related with tumor stage and patient endurance. Interestingly, regulation of miR‐155 occurs through the Epstein–Barr virus (EBV)‐encoded Latent membrane protein (LMP1) and LMP2A proteins [121]. Further‐ more, miR‐144 is overexpressed in NPC and inhibits PTEN expression, and brings about expanded cell proliferation, invasion, and metastasis [122]. miR‐214 has been firmly con‐ nected with augmented metastasis in NPC, both in cell lines and primary human samples, functioning at least in part via inhibition of the tumor suppressor lactotransferrin (LTF) [123]. Besides, miR‐214 has been appeared to upgrade proliferation and promote an anti‐ apoptotic phenotype in NPC cells [124]. In addition to miR‐144, miR‐155, and miR‐214, vari‐ ous other miRNAs that are overexpressed in NPC also contribute considerably to enhance the metastatic phenotype of NPC. miR‐30a has been demonstrated both *in vitro* and *in vivo* to build metastasis and intrusion by hindering E‐cadherin activity [125]. Moreover, expres‐ sion of miR‐149 was lifted profoundly in metastatic NPC cells, adding increased migration, invasion, and epithelial‐mesenchymal phenotypes over E‐cadherin inhibition [126]. miR‐93 has been accounted to inhibit transforming growth factor‐β receptor II (TGFβRII) [127] and disabled homolog‐2 (DAB2) [128], in this way directing tumor cell growth incursion and metastasis. Finally, miR‐504 is found to be involved as an oncogenic miRNA in NPC, work‐ ing to specifically target nuclear respiratory factor 1 (NRF1), whereas expanded expression connected with poor reaction to radiation treatment [129].

## **7. miRNAs as prognostic indicators**

Despite significant improvement in multimodal treatment, the prognosis of advanced HNC is quite poor. Recent studies are promising regarding the potential role of miRNAs as prognostic indicators. Downregulation of Let‐7 (a family of tumor suppressing miR‐ NAs) has been reported in HNSCCs by many researchers [74, 92, 95, 103, 130–138]. Similarly, underexpression of miR‐146a, miR‐155, and Let‐7 has been correlated with the progression of cancer [138]. Moreover, a decreased level of Let‐7 miRNA in nasopharyn‐ geal carcinoma cells was also suggestive of regulating the proliferation of carcinoma cells via c‐MYC downregulation [139]. Furthermore, role of Let‐7 in the Kirsten rat sarcoma (KRAS) regulation has been demonstrated recently by some studies on nonsmall cell lung cancer [140]. A variant allele in the V‐Ki‐ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) 3′untranslated region (KRAS‐LCS6) has been demonstrated to be involved in high expression of KRAS and decreased levels of Let‐7. Presence of KRAS‐LCS6 variant has been reported in Squamous cell head and neck cancer (SCHNC) and was correlated with poor prognosis [130]. TP53, a tumor suppressor gene, encodes a protein product that is involved in cell cycle regulation. It is a major change involved in cancer induction with a remarkable frequency of 53% as reported by Poeta et al. in a hefty cohort of SCHNC. The ratio of TP53 mutations were 75 and 56.7% in patients suffering from hypopharynx and larynx tumor, respectively. Furthermore, a strong association was described between TP53 mutations in SCHNC and high risk of recurrence and poor survival [141]. Ganci et al. has previously reported a strong link among 49 miRNAs and TP53 status. Among them, a more specific correlation of a subset of 12 miRNAs was correlated with a brief recurrence free‐survival while four of these were correlated with comparatively lower cancer‐specific survival [142]. Expression of some particular miRNAs, such as miR‐375 and miR‐210, has been correlated with the outcome of SCHNC patients. A low expression of miR‐375 results in poor survival and distant metastases while high expression of miR‐210 results in locore‐ gional recurrence [143, 144].

## **8. miRNAs as biomarkers**

control and decreased apoptosis. This regulation is thought to happen through downregula‐ tion of the supposed target gene phosphatase and tensin homolog (PTEN), bromo‐domain 3 (BRD3), and ubiquitin‐associated protein 1 (UBAP1) in NPC. Moreover, expression of miR‐141 is structured by the oncogenes short palate, lung, nasal epithelium clone 1 (SPLUNC1), and c‐Myc [120]. Expression of miR‐155 fortifies proliferation, migration, and incursion by regulation of target genes Jumonji Domain 1A (JMJD1A) and BTB and CNC homology 1 (BACH1), and its expression has been strongly related with tumor stage and patient endurance. Interestingly, regulation of miR‐155 occurs through the Epstein–Barr virus (EBV)‐encoded Latent membrane protein (LMP1) and LMP2A proteins [121]. Further‐ more, miR‐144 is overexpressed in NPC and inhibits PTEN expression, and brings about expanded cell proliferation, invasion, and metastasis [122]. miR‐214 has been firmly con‐ nected with augmented metastasis in NPC, both in cell lines and primary human samples, functioning at least in part via inhibition of the tumor suppressor lactotransferrin (LTF) [123]. Besides, miR‐214 has been appeared to upgrade proliferation and promote an anti‐ apoptotic phenotype in NPC cells [124]. In addition to miR‐144, miR‐155, and miR‐214, vari‐ ous other miRNAs that are overexpressed in NPC also contribute considerably to enhance the metastatic phenotype of NPC. miR‐30a has been demonstrated both *in vitro* and *in vivo* to build metastasis and intrusion by hindering E‐cadherin activity [125]. Moreover, expres‐ sion of miR‐149 was lifted profoundly in metastatic NPC cells, adding increased migration, invasion, and epithelial‐mesenchymal phenotypes over E‐cadherin inhibition [126]. miR‐93 has been accounted to inhibit transforming growth factor‐β receptor II (TGFβRII) [127] and disabled homolog‐2 (DAB2) [128], in this way directing tumor cell growth incursion and metastasis. Finally, miR‐504 is found to be involved as an oncogenic miRNA in NPC, work‐ ing to specifically target nuclear respiratory factor 1 (NRF1), whereas expanded expression

Despite significant improvement in multimodal treatment, the prognosis of advanced HNC is quite poor. Recent studies are promising regarding the potential role of miRNAs as prognostic indicators. Downregulation of Let‐7 (a family of tumor suppressing miR‐ NAs) has been reported in HNSCCs by many researchers [74, 92, 95, 103, 130–138]. Similarly, underexpression of miR‐146a, miR‐155, and Let‐7 has been correlated with the progression of cancer [138]. Moreover, a decreased level of Let‐7 miRNA in nasopharyn‐ geal carcinoma cells was also suggestive of regulating the proliferation of carcinoma cells via c‐MYC downregulation [139]. Furthermore, role of Let‐7 in the Kirsten rat sarcoma (KRAS) regulation has been demonstrated recently by some studies on nonsmall cell lung cancer [140]. A variant allele in the V‐Ki‐ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) 3′untranslated region (KRAS‐LCS6) has been demonstrated to be involved in high expression of KRAS and decreased levels of Let‐7. Presence of KRAS‐LCS6 variant has been reported in Squamous cell head and neck cancer (SCHNC) and was correlated with poor prognosis [130]. TP53, a tumor suppressor gene, encodes a protein product that is

connected with poor reaction to radiation treatment [129].

**7. miRNAs as prognostic indicators**

16 Diagnosis and Management of Head and Neck Cancer

Diagnosis of HNSCC more frequently occurs in advanced stage when it metastases to regional lymph nodes. Moreover, there is a high risk of recurrence even in patients with a combination of different therapeutic approaches. Therefore, for an early diagnosis, the most important goal would be the detection of biomarkers. Recently, some miRNAs have been discovered to fulfill the goal of diagnosis. In fact, miRNAs are found circulated stably in different body flu‐ ids, i.e., urine, blood, saliva, as well as in breath. Hence, these miRNAs can be assessed easily with noninvasive methods.

miRNAs can be utilized as biomarkers or novel therapeutic targets. Further investigation is needed to test its utilization [66].  The prognostic worth of this exciting perception requires affirmation using an independent patient cohort that includes clinical follow‐up data. Changes of miRNAs succeeding radio‐ chemotherapy in the blood plasma are related with the tumor response to therapy, and they might signify novel biomarkers for therapy monitoring [67]. The investigations of microRNA modifications in HNSCC are a fundamental stride to the mechanistic comprehension of tumorigenesis and could prompt the disclosure of clinically pertinent biomarkers [69]. For NPC diagnosis, plasma miRNAs expression proves to be a helpful biomarker [88]. Pathway improvement analysis of these four miRNAs (miR‐34c, miR‐140, miR‐154, and miR‐449b) sign posted a role in cell cycle regulation, highlighting a possibly important role for markers of cell cycle enactment as prognostic indicators in NPC [145]. miR‐200b expression is mostly con‐ nected with distant metastasis, while miR‐155 associated with local recurrence. miR‐155 and miR‐146a were recognized as surrogate markers for tumor‐invading lymphocytes in HNSCC [146]. miR‐21 expression could be an imperative tool for treatment planning and prognostic predictor for HNSCC patients [147]. Genetic variants of miR‐146a and miR‐1269b are biomark‐ ers for improvement of oral premalignant lesions (OPLs) and oral squamous cell carcinoma (OPSCC) [148]. hsa‐miR‐375‐3p appears to be a comparatively promising diagnostic marker

**Table 2.** A list of miRNAs identified as potent biomarker of HNC.

in HNSCC but is not appropriate for prognosis of patients [149]. Overexpression of TP53 mutation‐associated miR‐182 may contribute to proliferation and migration of tumour cell in HNSCC, hence suggest a possible biomarker for prognosis of tumour recurrence [150]. miR‐30a is a possible biomarker for metastasis in NPC patients [151]. miR‐148a and miR‐375 are sig‐ nificantly upregulated during LSCC and are conceivable biomarkers for early diagnosis of LSCC [152]. A list of miRNAs is given below which has been identified as potential biomarker by a number of researchers in HNC (**Table 2**).

## **9. Novel therapeutic targets**

miRNAs are key therapeutic agents (depends upon the type of mRNA affected by them) in addition to their prognostic and diagnostic value [21]. miRNAs are efficient molecules to be targeted as they regulate a number of biological activities by interacting with numerous other molecules [53]. Use of synthetically designed "miRNAs sponges" [153], miR mimics (agomiRs) [154], miR antagonists ("antagomiRs") [155, 156], and miR inhibitors (antimiRNAs oligonucleotides) [92] is an innovative strategy to modulate oncogenic and tumor‐suppres‐ sive pathways. MicroRNAs regulate each step of cell cycle and unraveling their altered expression may prove fruitful in designing new drugs and can open treatment regimes. One piece of evidence is a previous study abduction of miR‐122, which has been observed in a novel therapy for HCV patients by Miravirsen (first microRNA targeted drug) in clinical phase2a. This therapy resulted in diminution of HCV RNA levels in a dose‐dependent man‐ ner [157]. This research has raised a lot of hope for employing miRNAs therapies in num‐ ber of malignancies despite the disparity of the diseases from cancer. Some other miRNAs therapies are in preclinical and clinical phase 1. These approaches are anxiously awaited to be reported beyond the mere documentations. Few miRNAs, though not verified clinically, are very good applicants for being named for these therapies. Apoptotic pathway is one of the most significant candidates for novel anticancer therapies, as the neoplastic cells usually lose the ability to undergo programmed cell death. Therefore, any powerful proapoptotic agent may directly or indirectly reduce cancer progression by enhancing the apoptosis. *miR‐99a* mimics have been shown to decrease cell proliferation by inducing apoptosis in tongue SCC cell line [158]. Moreover, miR‐31‐3p inhibitor in oral leukoplakia cell lines resulted in decreased cell death [159]. Furthermore, restoration of miR‐100 resulted in inhibition of cell migration and proliferation by increasing the apoptosis in HNSCC cell lines [71]. Similarly, cancer cells showed a pronounced cell cycle arrest and enhanced cell death upon transfec‐ tion with miR‐1e [160]. On the other hand, a repression of apoptosis was observed in classi‐ cal oncogenic *miR*‐*21* [161]. These are only a few well‐known examples. A large number of other functionally investigated miRNAs in HNSCCs are directly or indirectly associated with apoptotic pathways promise an era of new and more potent therapeutic factors. miRNAs are utilized in another way in treating the HNSCC patients, as they are capable of highlight‐ ing the resistance to either chemotherapy or radiotherapy in patients of HNSCC patients. In addition to it, recently, miRNAs have also been reported to modulate the radiosensitivity and chemoresistance. For instance, let‐7 results in inhibition of cancer progression by reduc‐ tion in cancer‐proneness cells and repressing chemoresistance [162]. Similarly, miR‐21 in HA/ CD44‐activated head and neck cancer cells may prove a significant drug target to overcome chemoresistance and apoptosis as it is involved in regulation of Nanog/Stat‐3 signaling path‐ way [111]. Furthermore, transfection of pre‐miR‐98 in HNSCC cell lines led to an enhanced resistance to doxorubicin and cisplatin by downregulating the *HMGA2* expression [163].

in HNSCC but is not appropriate for prognosis of patients [149]. Overexpression of TP53 mutation‐associated miR‐182 may contribute to proliferation and migration of tumour cell in HNSCC, hence suggest a possible biomarker for prognosis of tumour recurrence [150]. miR‐30a is a possible biomarker for metastasis in NPC patients [151]. miR‐148a and miR‐375 are sig‐ nificantly upregulated during LSCC and are conceivable biomarkers for early diagnosis of LSCC [152]. A list of miRNAs is given below which has been identified as potential biomarker

NPC [166]

[169]

and oral squamous cell carcinoma

**miRNA biomarker Type of cancer References**

miR‐155 and miR‐146a HNSCC [167]

miR‐21 HNSCC [168]

(OPSCC)

hsa‐miR‐375‐3p HNSCC [170] miR‐182 HNSCC [171] miR‐30a NPC [172] miR‐148a and miR‐375 LSCC [173]

miR‐146a and miR‐1269b Oral premalignant lesions (OPLs)

miRNAs are key therapeutic agents (depends upon the type of mRNA affected by them) in addition to their prognostic and diagnostic value [21]. miRNAs are efficient molecules to be targeted as they regulate a number of biological activities by interacting with numerous other molecules [53]. Use of synthetically designed "miRNAs sponges" [153], miR mimics (agomiRs) [154], miR antagonists ("antagomiRs") [155, 156], and miR inhibitors (antimiRNAs oligonucleotides) [92] is an innovative strategy to modulate oncogenic and tumor‐suppres‐ sive pathways. MicroRNAs regulate each step of cell cycle and unraveling their altered expression may prove fruitful in designing new drugs and can open treatment regimes. One piece of evidence is a previous study abduction of miR‐122, which has been observed in a novel therapy for HCV patients by Miravirsen (first microRNA targeted drug) in clinical phase2a. This therapy resulted in diminution of HCV RNA levels in a dose‐dependent man‐ ner [157]. This research has raised a lot of hope for employing miRNAs therapies in num‐ ber of malignancies despite the disparity of the diseases from cancer. Some other miRNAs

by a number of researchers in HNC (**Table 2**).

**Table 2.** A list of miRNAs identified as potent biomarker of HNC.

**9. Novel therapeutic targets**

miR‐34c, miR‐140, miR‐154, and

18 Diagnosis and Management of Head and Neck Cancer

miR‐449b

miR‐155

Nevertheless, for an efficient and target specific miRNAs‐based drug delivery system, there lies enormous challenges [22, 53]. To access the targeted sites, therapeutic RNA must travel across the plasma membrane to enter in cytoplasm by leaving the circulatory system and avoiding endosomal vesicles to entering in the cell [53]. Moreover, nonconjugated therapeutic RNA molecules are either cleared by the kidney (<50kDa) or by the immune cells (7‐20kDa) [53]. Levels of mature miRNAs may be modulated or reduced by delivery of synthetic double‐stranded hairpin exogenously by complexing with lipids or proteins. Delivery of miR‐34a may prove effective in HNSCC cells as suppression of cellular proliferation and apop‐ tosis was observed in two cancer cell lines (colon) and experimental lung metastasis of murine melanoma upon miR‐34a delivery [164, 165]. Use of unmodified dsRNAs in environments, where local administration is possible, is of limited value *in vivo* as they are likely to be degraded by nucleases [166]. A high expression of these miRNAs from well‐defined transcription start and termination sites can be achieved using a viral vector with Pol III promoters for stable miR reintroduction; however, they are not cell‐specific [53, 165, 167]. In contrast, for a tissue‐specific approach or for ectopic expression of miRNAs, RNA Pol II promoters may be used for pri‐miR‐ NAs expression [168]. For an effective reintroduction of dysregulated miRNAs, later techniques may be utilized in HNSCC, e.g., miR‐375 [132]. However, use of a viral system for reintroduc‐ tion of miRNAs possesses a number of risks [53]. One of the major risks is that the delivered molecules integrate to an unpredictable site of host DNA and may activate proto‐oncogene and results in insertional mutagenesis [53]. These methods also have certain limitations as inser‐ tion of retroviral vectors is limited only to actively dividing cells and use of other units, like adenoviral vectors, may activate a severe immune response in host body [53, 169]. AntagomiRs (recently identified chemically engineered oligonucleotides) are commonly utilized today for silencing of host miRNAs [53, 156]. These AntagomiRs reverted the effects of upregulation of miRNAs in HNSCC, i.e., decreased cell viability and enhanced programmed cell death was observed in KB cells overexpressing miRNA‐155 upon introduction of antagomir‐155 in nude mice [155]. Moreover, to boost the efficient delivery of these antagomiR, they are delivered intravenously by complexing with a recently identified molecule the interfering nanoparticle (iNOP) [170]. Systemic delivery of iNop complexed with antimir‐122 in mice did not initi‐ ate any immune response and successfully silenced the miRNA‐122 with long lasting effects [170]. Chemically modified antimiRNAs oligonucleotides (AMOs) that are highly specific and efficient in binding to targeted RNA are another choice for miRNAs silencing but they do not have any specific delivery system [92, 164]. An enhanced apoptosis, limited survival, and pro‐ liferation were observed in tongue SCC cell lines upon inhibition of miRNA‐21 with AMO [92]. Moreover, a pronounced apoptosis and decreased cell proliferation was observed in nude mice upon repeated injections of miR‐21‐AMO which ultimately resulting in tumor suppression [92]. In HNSCC, miRNAs sponges or miRNAs masks can also be used to inhibit oncomiRNAs. miRNAs sponges block specific miRNAs attachment to their targets by expressing an mRNA having a number of tandem‐binding sites for entire family of targeted miRNAs, thus blocking their effects [53, 153]. On the other hand, miRNAs‐masking antisense oligonucleotides tech‐ nology shows its effect by destabilizing the association between specific miR‐mRNA pairs, as it comprised of complementary antisense oligonucleotides to miRNAs interacting site in the 3'UTR of a targeted mRNA [53, 167]. This methodology has advantage over others as there are few off‐target/undesirable effects and may be helpful in cancer therapy as multiple path‐ ways are targeted there simultaneously [53]. Furthermore, miRNAs may enhance sensitivity to radiotherapy [168]. In addition to it, some epigenetic drugs may also revert irregular meth‐ ylation or acetylation of a tumoral phenotype by restoration of tumor‐suppressive miRNAs expression [53]. Most of above‐mentioned techniques are in experimental phases. Our future approach should aim to aberrant miRNAs networks, as multiple miRNAs along with multiple miRNAs‐transcriptome interactions are involved in cancer pathogenesis. To achieve this aim, miRNAs biogenesis machinery or regulatorypathways must be targeted [53, 169]. However, exploration of the full potential ofthese drugs is highly recommended.

#### **10. Future challenges**

For last 3 decades, dysregulation of tumor suppressor genes and protein‐coding oncogenes were thought to be involved in cancer. However, discovery of noncoding genes, i.e., miRNAs raised a question on the notion that this mechanism is solely responsible for cancer. To unveil the involvement of miRNAs in cancer, massive efforts have been made but still enormous challenges lie ahead. Detection of the exact pathways and genes regulated by these miRNAs will be of prime importance. A better understanding of the wide‐ranging effects of these novel molecules will equip the researchers in the selection of specific pathways for treat‐ ment of cancers. Our understanding of miRNAs involvement in HNC is in its infancy. However, studies have been confirmed the dysregulation of a number of miRNAs in both benign and malignant HNC. There are real prospects that miRNAs in near future may be used as prognostic and diagnostics of HNC to improve its treatment strategies. Growing evidences make these miRNAs viable targets for the development of new and better anti‐ cancer therapies. However, before translation to be carried out in clinical settings, a more clear insight of efficacy and "off‐target" effects of miRNAs is necessarily suggested. It is quite fascinating that recently a number of miRNAs including miR‐21, miR‐155, and let‐ 7b have been nominated as key players of human carcinogenesis including head and neck tumors. The discovery of miRNAs heralds a complete new paradigm in the understanding of exact molecular pathways involved in HNC development. More detailed studies are required for better understanding and therapeutic targets to treat HNC.

However, a number of efforts have been made but truly comprehensive profiling of miRNAs is still needed in HNSCC. Even, technology utilized for profiling also needs to be improved. Use of Polymerase chain reaction (PCR) and microarrays is a common practice for profiling but for a better picture and more deep insight next‐generation sequencing (NGS), cross‐platform analysis and new approaches like NanoString nCounter system must be applied. Functional studies are growing, highlighting the potentially targetable miRNAs and some related path‐ ways. Tumor cells survival and their development have been reported to be affected by the forced expression or even the inhibition of a number of miRNAs *in vitro*, representing a new class of novel therapeutic targets [158, 171, 172]. To deliver therapeutic miRNAs, delivery of pre‐miR‐107 and siRNA nanoparticles proves to be an effective strategy [173, 174]. Therefore, the future of tumor biology seems quite bright.

## **Author details**

results in insertional mutagenesis [53]. These methods also have certain limitations as inser‐ tion of retroviral vectors is limited only to actively dividing cells and use of other units, like adenoviral vectors, may activate a severe immune response in host body [53, 169]. AntagomiRs (recently identified chemically engineered oligonucleotides) are commonly utilized today for silencing of host miRNAs [53, 156]. These AntagomiRs reverted the effects of upregulation of miRNAs in HNSCC, i.e., decreased cell viability and enhanced programmed cell death was observed in KB cells overexpressing miRNA‐155 upon introduction of antagomir‐155 in nude mice [155]. Moreover, to boost the efficient delivery of these antagomiR, they are delivered intravenously by complexing with a recently identified molecule the interfering nanoparticle (iNOP) [170]. Systemic delivery of iNop complexed with antimir‐122 in mice did not initi‐ ate any immune response and successfully silenced the miRNA‐122 with long lasting effects [170]. Chemically modified antimiRNAs oligonucleotides (AMOs) that are highly specific and efficient in binding to targeted RNA are another choice for miRNAs silencing but they do not have any specific delivery system [92, 164]. An enhanced apoptosis, limited survival, and pro‐ liferation were observed in tongue SCC cell lines upon inhibition of miRNA‐21 with AMO [92]. Moreover, a pronounced apoptosis and decreased cell proliferation was observed in nude mice upon repeated injections of miR‐21‐AMO which ultimately resulting in tumor suppression [92]. In HNSCC, miRNAs sponges or miRNAs masks can also be used to inhibit oncomiRNAs. miRNAs sponges block specific miRNAs attachment to their targets by expressing an mRNA having a number of tandem‐binding sites for entire family of targeted miRNAs, thus blocking their effects [53, 153]. On the other hand, miRNAs‐masking antisense oligonucleotides tech‐ nology shows its effect by destabilizing the association between specific miR‐mRNA pairs, as it comprised of complementary antisense oligonucleotides to miRNAs interacting site in the 3'UTR of a targeted mRNA [53, 167]. This methodology has advantage over others as there are few off‐target/undesirable effects and may be helpful in cancer therapy as multiple path‐ ways are targeted there simultaneously [53]. Furthermore, miRNAs may enhance sensitivity to radiotherapy [168]. In addition to it, some epigenetic drugs may also revert irregular meth‐ ylation or acetylation of a tumoral phenotype by restoration of tumor‐suppressive miRNAs expression [53]. Most of above‐mentioned techniques are in experimental phases. Our future approach should aim to aberrant miRNAs networks, as multiple miRNAs along with multiple miRNAs‐transcriptome interactions are involved in cancer pathogenesis. To achieve this aim, miRNAs biogenesis machinery or regulatorypathways must be targeted [53, 169]. However,

20 Diagnosis and Management of Head and Neck Cancer

exploration of the full potential ofthese drugs is highly recommended.

For last 3 decades, dysregulation of tumor suppressor genes and protein‐coding oncogenes were thought to be involved in cancer. However, discovery of noncoding genes, i.e., miRNAs raised a question on the notion that this mechanism is solely responsible for cancer. To unveil the involvement of miRNAs in cancer, massive efforts have been made but still enormous challenges lie ahead. Detection of the exact pathways and genes regulated by these miRNAs will be of prime importance. A better understanding of the wide‐ranging effects of these

**10. Future challenges**

Muhammad Babar Khawar1 , Naz Fatima1 , Muddasir Hassan Abbasi1,3, Rabia Mehmood1 , Saira Kainat Suqaina1 and Nadeem Sheikh1,2\*

\*Address all correspondence to<EMAIL_ADDRESS>
1 Cell & Molecular Biology Lab, Department of Zoology, University of the Punjab, Lahore, Pakistan

## **References**

[1] Alibek K, Kakpenova A, Baiken Y. Role of infectious agents in the carcinogenesis of brain and head and neck cancers. Infectious Agents and Cancer. 2013;**8**:1

[13] Yakin M, Gavidi RO, Cox B, Rich A. Oral cancer risk factors in New Zealand. The New Zealand Medical Journal. 2017;**130**:30‐38

[2] Warnakulasuriya S. Global epidemiology of oral and oropharyngeal cancer. Oral

[3] Vokes EE, Weichselbaum RR, Lippman SM, Hong WK. Head and neck cancer. New

[4] Nagadia R, Pandit P, Coman WB, Cooper‐White J, Punyadeera C. miRNAs in head and

[5] Courthod G, Franco P, Palermo L, Pisconti S, Numico G. The role of microRNA in head and neck cancer: Current knowledge and perspectives. Molecules. 2014;**19**:5704‐5716 [6] Ascierto PA, Accorona R, Botti G, Farina D, Fossati P, Gatta G, Gogas H, Lombardi D, Maroldi R, Nicolai P, Ravanelli M, Vanella V. Mucosal melanoma of the head and neck.

[7] Bagnardi V, Rota M, Botteri E, Tramacere I, Islami F, Fedirko V, Scotti L, Jenab M, Turati F, Pasquali E, Pelucchi C, Galeone C, Bellocco R, Negri E, Corrao G, Boffetta P, La VC. Alcohol consumption and site‐specific cancer risk: A comprehensive dose‐response

[8] Hashibe M, Brennan P, Chuang SC, Boccia S, Castellsague X, Chen C, Curado MP, Dal ML, Daudt AW, Fabianova E, Fernandez L, Wunsch‐Filho V, Franceschi S, Hayes RB, Herrero R, Kelsey K, Koifman S, La VC, Lazarus P, Levi F, Lence JJ, Mates D, Matos E, Menezes A, McClean MD, Muscat J, Eluf‐Neto J, Olshan AF, Purdue M, Rudnai P, Schwartz SM, Smith E, Sturgis EM, Szeszenia‐Dabrowska N, Talamini R, Wei Q, Winn DM, Shangina O, Pilarska A, Zhang ZF, Ferro G, Berthiller J, Boffetta P. Interaction between tobacco and alcohol use and the risk of head and neck cancer: Pooled anal‐ ysis in the International Head and Neck Cancer Epidemiology Consortium. Cancer

[9] Citron F, Armenia J, Franchin G, Polesel J, Talamini R, D'Andrea S, Sulfaro S, Croce CM, Klement W, Otasek D, Pastrello C, Tokar T, Jurisica I, French D, Bomben R, Vaccher E, Serraino D, Belletti B, Vecchione A, Barzan L, Baldassarre G. An integrated approach identifies mediators of local recurrence in Head & Neck Squamous Carcinoma. Clinical

Cancer Research. 2017: clincancers‐2814. DOI: 10.1158/1078‐0432.CCR‐16‐2814

noma. European Archives of Oto‐Rhino‐Laryngology. 2017; Mar **11**:1‐9.

[10] Qureishi A, Mawby T, Fraser L, Shah KA, Moller H, Winter S. Current and future tech‐ niques for human papilloma virus (HPV) testing in oropharyngeal squamous cell carci‐

[11] Fakhry C, Westra WH, Wang SJ, van ZA, Zhang Y, Rettig E, Yin LX, Ryan WR, Ha PK, Wentz A, Koch W, Richmon JD, Eisele DW, D'Souza G. The prognostic role of sex, race, and human papillomavirus in oropharyngeal and nonoropharyngeal head and neck

[12] Peterson CE, Khosla S, Jefferson GD, Davis FG, Fitzgibbon ML, Freels S, Johnson TP, Hoskins K, Joslin CE. Measures of economic advantage associated with HPV‐positive head and neck cancers among non‐Hispanic black and white males identified through

the National Cancer Database. Journal of Cancer Epidemiology. 2017;**48**:1‐7

Oncology. 2009;**45**:309‐316

22 Diagnosis and Management of Head and Neck Cancer

England Journal of Medicine. 1993;**328**:184‐194

neck cancer revisited. Cellular Oncology. 2013;**36**:1‐7

Critical Reviews in Oncology/Hematology. 2017;**112**:136‐152

meta‐analysis. British Journal of Cancer. 2015;**112**:580‐593

Epidemiology, Biomarkers & Prevention. 2009;**18**:541‐550

squamous cell cancer. Cancer. 2017;**123**(9):1566‐1575

[43] Williams AH, Liu N, Van Rooij E, Olson EN. MicroRNA control of muscle development and disease. Current Opinion in Cell Biology. 2009;**21**:461‐469

[25] John K, Wu J, Lee BW, Farah CS. MicroRNAs in head and neck cancer. International

[28] Horvitz HR, Sulston JE. Isolation and genetic characterization of cell‐lineage mutants of

[29] Chalfie M, Horvitz HR, Sulston JE. Mutations that lead to reiterations in the cell lineages

[30] Ambros V, Horvitz HR. The lin‐14 locus of *Caenorhabditis elegans* controls the time of expression of specific postembryonic developmental events. Genes & Development.

[31] Stefani G, Slack FJ. Small non‐coding RNAs in animal development. Nature Reviews

[32] Carthew RW, Sontheimer EJ. Origins and mechanisms of miRNAs and siRNAs. Cell.

[33] Ghildiyal M, Zamore PD. Small silencing RNAs: An expanding universe. Nature

[34] Reinhart BJ, Slack FJ, Basson M, Pasquinelli AE, Bettinger JC, Rougvie AE, Horvitz HR, Ruvkun G. The 21‐nucleotide let‐7 RNA regulates developmental timing in *Caenorhabditis*

[35] Pasquinelli AE, Reinhart BJ, Slack F, Martindale MQ, Kuroda MI, Maller B, Hayward DC, Ball EE, Degnan B, Muller P. Conservation of the sequence and temporal expression

[36] Lau NC, Lim LP, Weinstein EG, Bartel DP, An abundant class of tiny RNAs with prob‐

[37] Lee RC, Ambros V. An extensive class of small RNAs in *Caenorhabditis elegans*. Science.

[38] Lagos‐Quintana M, Rauhut R, Lendeckel W, Tuschl T. Identification of novel genes cod‐

[39] Griffiths‐Jones S. The microRNA registry. Nucleic Acids Research. 2004;**32**:D109‐D111 [40] Griffiths‐Jones S, Saini HK, van Dongen S, Enright AJ. miRBase: Tools for microRNA

[41] Zeng Y, Cullen BR. The biogenesis and function of MicroRNAs. Gene Expression and

[42] Hwang HW, Mendell JT. MicroRNAs in cell proliferation, cell death, and tumorigenesis.

of let‐7 heterochronic regulatory RNA. Nature. 2000;**408**:86‐89

ing for small expressed RNAs. Science. 2001;**294**:853‐858

genomics. Nucleic Acids Research. 2008;**36**:D154‐D158

British Journal of Cancer. 2006;**94**:776‐780

able regulatory roles in *Caenorhabditis elegans*. Science. 2001;**294**:858‐862

[26] Brenner S. The genetics of Caenorhabditis elegans. Genetics. 1974;**77**:71‐94

the nematode *Caenorhabditis elegans*. Genetics. 1980;**96**:435‐454

[27] Sulston JE, Brenner S. The DNA of *Caenorhabditis elegans*. Genetics. 1974;**77**:95‐104

Journal of Dentistry. 2013;**2013**

24 Diagnosis and Management of Head and Neck Cancer

of *C. elegans*. Cell. 1981;**24**:59‐69

Molecular Cell Biology. 2008;**9**:219‐230

Reviews Genetics. 2009;**10**:94‐108

*elegans*. Nature. 2000;**403**:901‐906

1987;**1**:398‐414

2009;**136**:642‐655

2001;**294**:862‐864

Regulation. 2006;481‐492

[72] Pashaei E, Guzel E, Ozgurses ME, Demirel G, Aydin N, Ozen M. A Meta‐Analysis: Identification of common Mir‐145 target genes that have similar behavior in different GEO datasets. PLoS One. 2016;**11**:e0161491

[58] Jiang J, Lee EJ, Gusev Y, Schmittgen TD. Real‐time expression profiling of microRNA precursors in human cancer cell lines. Nucleic Acids Research. 2005;**33**:5394‐5403

[60] MacFarlane LA, Murphy R. MicroRNA: Biogenesis, function and role in cancer. Current

[61] Chang SS, Jiang WW, Smith I, Poeta LM, Begum S, Glazer C, Shan S, Westra W, Sidransky D, Califano JA. MicroRNA alterations in head and neck squamous cell carcinoma.

[62] Chen HC, Chen GH, Chen YH, Liao WL, Liu CY, Chang KP, Chang YS, Chen SJ. MicroRNA deregulation and pathway alterations in nasopharyngeal carcinoma. British

[63] Jakymiw A, Patel RS, Deming N, Bhattacharyya I, Shah P, Lamont RJ, Stewart CM, Cohen DM, Chan EK. Overexpression of dicer as a result of reduced let−7 MicroRNA levels contributes to increased cell proliferation of oral cancer cells. Genes, Chromosomes and

[64] Ahmad P, Sana J, Slavik M, Slampa P, Smilek P, Slaby O. MicroRNAs involvement in radioresistance of head and neck cancer. Disease Markers. 2017;**2017**:8245345

[65] He J, Tang Y, Tian Y. MicroRNA214 promotes proliferation and inhibits apoptosis via targeting Bax in nasopharyngeal carcinoma cells. Journal of Molecular Medicine.

[66] Masood Y, Kqueen CY, Rajadurai P. Role of miRNA in head and neck squamous cell

[67] Summerer I, Niyazi M, Unger K, Pitea A, Zangen V, Hess J, Atkinson MJ, Belka C, Moertl S, Zitzelsberger H. Changes in circulating microRNAs after radiochemotherapy in head

[68] Chen D, Cabay RJ, Jin Y, Wang A, Lu Y, Shah‐Khan M, Zhou X. MicroRNA deregulations in head and neck squamous cell carcinomas. Journal of Oral & Maxillofacial Research.

[69] Severino P, Bruggemann H, Andreghetto FM, Camps C, Klingbeil MF, de Pereira WO, Soares RM, Moyses R, Wunsch‐Filho V, Mathor MB, Nunes FD, Ragoussis J, Tajara EH. MicroRNA expression profile in head and neck cancer: HOX‐cluster embedded microRNA‐196a and microRNA‐10b dysregulation implicated in cell proliferation. BMC

[70] Jin Y, Chen D, Cabay RJ, Wang A, Crowe DL, Zhou X. Role of microRNA‐138 as a poten‐ tial tumor suppressor in head and neck squamous cell carcinoma. International Review

[71] Chen Z, Jin Y, Yu D, Wang A, Mahjabeen I, Wang C, Liu X, Zhou X. Down‐regulation of the microRNA‐99 family members in head and neck squamous cell carcinoma. Oral

carcinoma. Expert Review of Anticancer Therapy. 2015;**15**:183‐197

and neck cancer patients. Radiation Oncology. 2013;**8**:296

of Cell and Molecular Biology. 2013;**303**:357‐385

[59] Lujambio A, Lowe SW. The microcosmos of cancer. Nature. 2012;**482**:347‐355

International Journal of Cancer. 2008;**123**:2791‐2797

Journal of Cancer. 2009;**100**:1002‐1011

Genomics. 2010;**11**:537‐561

26 Diagnosis and Management of Head and Neck Cancer

Cancer. 2010;**49**:549‐559

2015;**12**:6286‐6292

2013;**4**:e2

Cancer. 2013;**13**:533

Oncology. 2012;**48**:686‐691

[97] Reis PP, Tomenson M, Cervigne NK, Machado J, Jurisica I, Pintilie M, Sukhai MA, Perez‐Ordonez B, Grenman R, Gilbert RW. Programmed cell death 4 loss increases tumor cell invasion and is regulated by miR‐21 in oral squamous cell carcinoma. Molecular Cancer. 2010;**9**:1

[84] Kozaki K, Imoto I, Mogi S, Omura K, Inazawa J. Exploration of tumor‐suppressive microRNAs silenced by DNA hypermethylation in oral cancer. Cancer Research.

[85] Liu X, Yu J, Jiang L, Wang A, Shi F, Ye H, Zhou X. MicroRNA‐222 regulates cell invasion by targeting matrix metalloproteinase 1 (MMP1) and manganese superoxide dismutase 2 (SOD2) in tongue squamous cell carcinoma cell lines. Cancer Genomics Proteomics.

[86] Liu X, Jiang L, Wang A, Yu J, Shi F, Zhou X. MicroRNA‐138 suppresses invasion and promotes apoptosis in head and neck squamous cell carcinoma cell lines. Cancer Letters.

[87] Yu ZW, Zhong LP, Ji T, Zhang P, Chen WT, Zhang CP. MicroRNAs contribute to the chemoresistance of cisplatin in tongue squamous cell carcinoma lines. Oral Oncology.

 Biochemical and Biophysical

[89] Avissar M, Christensen BC, Kelsey KT, Marsit CJ. MicroRNA expression ratio is predictive of head and neck squamous cell carcinoma. Clinical Cancer Research. 2009;**15**:2850‐2855

[90] Ramdas L, Giri U, Ashorn CL, Coombes KR, El‐Naggar A, Ang KK, Story MD. miRNA expression profiles in head and neck squamous cell carcinoma and adjacent normal tis‐

[91] Childs G, Fazzari M, Kung G, Kawachi N, Brandwein‐Gensler M, McLemore M, Chen Q, Burk RD, Smith RV, Prystowsky MB, Belbin TJ, Schlecht NF. Low‐level expression of microRNAs let‐7d and miR‐205 are prognostic markers of head and neck squamous cell

[92] Li J, Huang H, Sun L, Yang M, Pan C, Chen W, Wu D, Lin Z, Zeng C, Yao Y. MiR‐21 indicates poor prognosis in tongue squamous cell carcinomas as an apoptosis inhibitor.

[93] Kawakita A, Yanamoto S, Yamada Si, Naruse T, Takahashi H, Kawasaki G, Umeda M. MicroRNA‐21 promotes oral cancer invasion via the Wnt/beta‐catenin pathway by tar‐

[94] Hedback N, Jensen DH, Specht L, Fiehn AM, Therkildsen MH, Friis‐Hansen L, Dabelsteen E, von Buchwald C. MiR‐21 expression in the tumor stroma of oral squamous cell carci‐ noma: An independent biomarker of disease free survival. PloS One. 2014;**9**:e95193 [95] Brito JA, Gomes CC, Guimaraes AL, Campos K, Gomez RS. Relationship between microRNA expression levels and histopathological features of dysplasia in oral leuko‐

[96] Mydlarz W, Uemura M, Ahn S, Hennessey P, Chang S, Demokan S, Sun W, Shao C, Bishop J, Krosting J. Clusterin is a gene‐specific target of microRNA‐21 in head and neck

carcinoma. American Journal of Pathology. 2009;**174**:736‐745

geting DKK2. Pathology & Oncology Research. 2014;**20**:253‐261

plakia. Journal of Oral Pathology & Medicine. 2014;**43**:211‐216

squamous cell carcinoma. Clinical Cancer Research. 2014;**20**:868‐877

2008;**68**:2094‐2105

28 Diagnosis and Management of Head and Neck Cancer

2009;**6**:131‐139

2009;**286**:217‐222

2010;**46**:317‐322

Research Communications. 2007;**358**:12‐17

Clinical Cancer Research. 2009;**15**:3998‐4008

sue. Head Neck. 2009;**31**:642‐654

[120] Yu X, Zhen Y, Yang H, Wang H, Zhou Y, Wang E, Marincola FM, Mai C, Chen Y, Wei H, Song Y, Lyu X, Ye Y, Cai L, Wu Q, Zhao M, Hua S, Fu Q, Zhang Y, Yao K, Liu Z, Li X, Fang W. Loss of connective tissue growth factor as an unfavorable prognosis factor activates miR‐18b by PI3K/AKT/C‐Jun and C‐Myc and promotes cell growth in naso‐ pharyngeal carcinoma. Cell Death and Disease. 2013;**4**:e634

[109] Gwak JM, Kim HJ, Kim EJ, Chung YR, Yun S, Seo AN, Lee HJ, Park SY.  Breast Cancer Research and Treatment.

[110] Ma L, Young J, Prabhala H, Pan E, Mestdagh P, Muth D, Teruya‐Feldstein J, Reinhardt F, Onder TT, Valastyan S, Westermann F, Speleman F, Vandesompele J, Weinberg RA. miR‐9, a MYC/MYCN‐activated microRNA, regulates E‐cadherin and cancer metastasis.

[111] Yu T, Liu K, Wu Y, Fan J, Chen J, Li C, Yang Q, Wang Z. MicroRNA‐9 inhibits the prolif‐ eration of oral squamous cell carcinoma cells by suppressing expression of CXCR4 via

[112] Gao F, Zhao ZL, Zhao WT, Fan QR, Wang SC, Li J, Zhang YQ, Shi JW, Lin XL, Yang S, Xie RY, Liu W, Zhang TT, Sun YL, Xu K, Yao KT, Xiao D. miR‐9 modulates the expression of interferon‐regulated genes and MHC class I molecules in human nasopharyngeal carci‐ noma cells. Biochemical and Biophysical Research Communications. 2013;**431**:610‐616

[113] Hui AB, Bruce JP, Alajez NM, Shi W, Yue S, Perez‐Ordonez B, Xu W, O'Sullivan B, Waldron J, Cummings B, Gullane P, Siu L, Liu FF. Significance of dysregulated metadherin and microRNA‐375 in head and neck cancer. Clinical Cancer Research.

[114] Liu N, Jiang N, Guo R, Jiang W, He QM, Xu YF, Li YQ, Tang LL, Mao YP, Sun Y, Ma J. MiR‐451 inhibits cell growth and invasion by targeting MIF and is associated with

[115] Lu J, He ML, Wang L, Chen Y, Liu X, Dong Q, Chen YC, Peng Y, Yao KT, Kung HF, Li XP. MiR‐26a inhibits cell growth and tumorigenesis of nasopharyngeal carcinoma

[116] Yu L, Lu J, Zhang B, Liu X, Wang L, Li SY, Peng XH, Xu X, Tian WD, Li XP. miR‐ 26a inhibits invasion and metastasis of nasopharyngeal cancer by targeting EZH2.

[117] Deng M, Tang H, Zhou Y, Zhou M, Xiong W, Zheng Y, Ye Q, Zeng X, Liao Q, Guo X, Li X, Ma J, Li G. miR‐216b suppresses tumor growth and invasion by targeting KRAS in

[118] Li YQ, Ren XY, He QM, Xu YF, Tang XR, Sun Y, Zeng MS, Kang TB, Liu N, Ma J. MiR‐34c suppresses tumor growth and metastasis in nasopharyngeal carcinoma by tar‐

[119] Luo Z, Dai Y, Zhang L, Jiang C, Li Z, Yang J, McCarthy JB, She X, Zhang W, Ma J, Xiong W, Wu M, Lu J, Li X, Li X, Xiang J, Li G. miR‐18a promotes malignant progres‐ sion by impairing microRNA biogenesis in nasopharyngeal carcinoma. Carcinogenesis.

nasopharyngeal carcinoma. Journal of Cell Science. 2011;**124**:2997‐3005

survival in nasopharyngeal carcinoma. Molecular Cancer. 2013;**12**:123

through repression of EZH2. Cancer Research. 2011;**71**:225‐233

the Wnt/beta‐catenin signaling pathway. Oncogene. 2014;**33**:5017‐5027

2014;**147**:39‐49

30 Diagnosis and Management of Head and Neck Cancer

2011;**17**:7539‐7550

2013;**34**:415‐425

Nature Cell Biology. 2010;**12**:247‐256

Oncology Letters. 2013;**5**:1223‐1228

geting MET. Cell Death ad Disease. 2015;**6**:e1618

[142] Ganci F, Sacconi A, Ben‐Moshe NB, Manciocco V, Sperduti I, Strigari L, Covello R, Benevolo M, Pescarmona E, Domany E. Expression of TP53 mutation‐associated microRNAs predicts clinical outcome in head and neck squamous cell carcinoma patients. Annals of Oncology. 2013;**24**:3082‐3088

[131] Fletcher AM, Heaford AC, Trask DK. Detection of metastatic head and neck squamous cell carcinoma using the relative expression of tissue‐specific mir‐205. Translational

[132] Lajer CB, Nielsen FC, Friis‐Hansen L, Norrild B, Borup R, Garnaes E, Rossing M, Specht L, Therkildsen MH, Nauntofte B. Different miRNA signatures of oral and pharyngeal squamous cell carcinomas: A prospective translational study. British Journal of Cancer.

[133] Liu CJ, Tsai MM, Hung PS, Kao SY, Liu TY, Wu KJ, Chiou SH, Lin SC, Chang KW. miR‐31 ablates expression of the HIF regulatory factor FIH to activate the HIF pathway

[134] Kikkawa N, Hanazawa T, Fujimura L, Nohata N, Suzuki H, Chazono H, Sakurai D, Horiguchi S, Okamoto Y, Seki N. miR‐489 is a tumour‐suppressive miRNA target PTPN11 in hypopharyngeal squamous cell carcinoma (HSCC). British Journal of

[135] Nohata N, Hanazawa T, Kikkawa N, Sakurai D, Fujimura L, Chiyomaru T, Kawakami K, Yoshino H, Enokida H, Nakagawa M. Tumour suppressive microRNA‐874 regulates novel cancer networks in maxillary sinus squamous cell carcinoma. British Journal of

[136] Ramdas L, Giri U, Ashorn CL, Coombes KR, ElΓÇÉNaggar A, Ang KK, Story MD. miRNA expression profiles in head and neck squamous cell carcinoma and adjacent

[137] Rentoft M, Fahlen J, Coates PJ, Laurell G, Sjostrom B, Ryden P, Nylander K. miRNA analysis of formalin‐fixed squamous cell carcinomas of the tongue is affected by age of

[138] Scapoli L, Palmieri A, Muzio LL, Pezzetti F, Rubini C, Girardi A, Farinella F, Mazzotta M, Carinci F. MicroRNA expression profiling of oral carcinoma identifies new markers of tumor progression. International Journal of Immunopathology and Pharmacology.

[139] Wong TS, Man O‐Y, Tsang CM, Tsao SW, Tsang RK‐Y, Chan JY‐W, Ho WK, Wei WI, To VS‐H. MicroRNA let‐7 suppresses nasopharyngeal carcinoma cells proliferation through downregulating c‐Myc expression. Journal of Cancer Research and Clinical

[140] Chin LJ, Ratner E, Leng S, Zhai R, Nallur S, Babar I, Muller RU, Straka E, Su L, Burki EA. A SNP in a let‐7 microRNA complementary site in the KRAS 3 untranslated region

[141] Poeta ML, Manola J, Goldwasser MA, Forastiere A, Benoit N, Califano JA, Ridge JA, Goodwin J, Kenady D, Saunders J. TP53 mutations and survival in squamous‐cell car‐ cinoma of the head and neck. New England Journal of Medicine. 2007;**357**:2552‐2561

increases non‐small cell lung cancer risk. Cancer Research. 2008;**68**:8535‐8540

in head and neck carcinoma. Cancer Research. 2010;**70**:1635‐1644

Oncology. 2008;**1**:202‐208

32 Diagnosis and Management of Head and Neck Cancer

Cancer. 2010;**103**:877‐884

Cancer. 2011;**105**:833‐841

2010;**23**:1229‐1234

Oncology. 2011;**137**:415‐422

normal tissue. Head & Neck. 2009;**31**:642‐654

the samples. International Journal of Oncology. 2011;**38**:61

2011;**104**:830‐840

[166] Esquela‐Kerscher A, Trang P, Wiggins JF, Patrawala L, Cheng A, Ford L, Weidhaas JB, Brown D, Bader AG, Slack FJ. The let‐7 microRNA reduces tumor growth in mouse models of lung cancer. Cell Cycle. 2008;**7**:759‐764

[153] Ebert MS, Neilson JR, Sharp PA. MicroRNA sponges: Competitive inhibitors of small

[154] Bader AG, Brown D, Winkler M. The promise of microRNA replacement therapy.

[155] Rather MI, Nagashri MN, Swamy SS, Gopinath KS, Kumar A. Oncogenic MicroRNA‐155 Down‐regulates tumor suppressor CDC73 and promotes oral squamous cell carcinoma cell proliferation Implications for Cancer Therapeutics. Journal of Biological Chemistry.

[156] Krutzfeldt J, Rajewsky N, Braich R, Rajeev KG, Tuschl T, Manoharan M, Stoffel M. Silencing of microRNAs in vivo with antagomirs. Nature. 2005;**438**:685‐689

[157] Janssen HL, Reesink HW, Lawitz EJ, Zeuzem S, Rodriguez‐Torres M, Patel K, van der Meer AJ, Patick AK, Chen A, Zhou Y. Treatment of HCV infection by targeting

[158] Yan B, Fu Q, Lai L, Tao X, Fei Y, Shen J, Chen Z, Wang Q. Downregulation of microRNA 99a in oral squamous cell carcinomas contributes to the growth and survival of oral

[159] Xiao W, Bao ZX, Zhang CY, Zhang XY, Shi LJ, Zhou ZT, Jiang WW. Upregulation of miR‐31\* is negatively associated with recurrent/newly formed oral leukoplakia. PloS

[160] Nohata N, Sone Y, Hanazawa T, Fuse M, Kikkawa N, Yoshino H, Chiyomaru T, Kawakami K, Enokida H, Nakagawa M. miR‐1 as a tumor suppressive microRNA tar‐ geting TAGLN2 in head and neck squamous cell carcinoma. Oncotarget. 2011;**2**:29‐42

[161] Lu Z, Liu M, Stribinskis V, Klinge CM, Ramos KS, Colburn NH, Li Y. MicroRNA‐21 pro‐ motes cell transformation by targeting the programmed cell death 4 gene. Oncogene.

[162] Yu CC, Chen YW, Chiou GY, Tsai LL, Huang PI, Chang CY, Tseng LM, Chiou SH, Yen SH, Chou MY. MicroRNA let‐7a represses chemoresistance and tumourigenicity in head and neck cancer via stem‐like properties ablation. Oral Oncology. 2011;**47**:202‐210

[163] Hebert C, Norris K, Scheper MA, Nikitakis N, Sauk JJ. High mobility group A2 is a target for miRNA‐98 in head and neck squamous cell carcinoma. Molecular Cancer.

[164] Chen Y, Zhu X, Zhang X, Liu B, Huang L. Nanoparticles modified with tumor‐tar‐ geting scFv deliver siRNA and miRNA for cancer therapy. Molecular Therapy.

[165] Tazawa H, Tsuchiya N, Izumiya M, Nakagama H. Tumor‐suppressive miR‐34a induces senescence‐like growth arrest through modulation of the E2F pathway in human colon cancer cells. Proceedings of the National Academy of Sciences. 2007;**104**:15472‐15477

microRNA. New England Journal of Medicine. 2013;**368**:1685‐1694

cancer cells. Molecular Medicine Reports. 2012;**6**:675‐681

RNAs in mammalian cells. Nature Methods. 2007;**4**:721‐726

Cancer Research. 2010;**70**:7027‐7030

34 Diagnosis and Management of Head and Neck Cancer

2013;**288**:608‐618

One. 2012;**7**:e38648

2008;**27**:4373‐4379

2010;**18**:1650‐1656

2007;**6**:5

**Provisional chapter**

## **Salivary Gland Cancers: A Survey through History, Classifications and Managements Classifications and Managements**

**Salivary Gland Cancers: A Survey through History,**

DOI: 10.5772/intechopen.70127

Mohammad Hossein Khosravi, Ali Bagherihagh, Masoumeh Saeedi, Payman Dabirmoghaddam, Ali Kouhi and Mohammad Hosein Amirzade-Iranaq Ali Bagherihagh, Masoumeh Saeedi, Payman Dabirmoghaddam, Ali Kouhi and Mohammad Hosein Amirzade-Iranaq

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.70127

Mohammad Hossein Khosravi,

#### **Abstract**

In this chapter, we are going to discuss about salivary glands cancers, their clinical manifestations and categories, pathogenesis, diagnosis and treatment. We will go through details in each part in both clinical and surgical aspects based on recently prominent published studies and research in prestigious journals. After a short review on clinical features, epidemiology, pathogenesis, diagnosis and treatment, we will show staging and tumor node metastasis (TNM) classification of major salivary gland tumors and also basic principles of approach to salivary gland cancers.

**Keywords:** cancer, salivary gland, classification, benign tumors

## **1. Embryology, anatomy and structure of salivary glands**

There are three pairs of salivary glands in human: parotid (PG), submandibular (SMG) and sublingual gland (SLG). They all have the same architecture; an arborized duct which opens into the oral cavity with secretary end pieces called acini which produce the saliva [1]. A mixed structure of extracellular matrix, myoepithelial cells, myofibroblasts, immune cells, endothelial cells, stromal cells, and nerve fibers surround the acinar cells. The main excretory duct of PG is Stensen's duct which opens in buccal mucosa near the second maxillary molar after crossing the masseter muscle and penetrating buccinators muscle. Submandibular glands secrete the saliva through their main excretory duct, called Wharton's, which opens

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

into the oral cavity under the tongue by the lingual frenum at a structure called the sublingual caruncule. In contrast, sublingual gland has small ducts called ducts of Rivinus and a common duct, Bartholin's duct, which connects with Wharton's duct at the sublingual caruncle [1].

Blood supply of parotid gland is mainly provided by the transverse facial artery and SMG by the facial artery. Postganglionic nerves from the otic and submandibular ganglia provide parasympathetic innervation. Superior cervical ganglion makes sympathetic postganglionic nerves which innervate the glands along blood vessels [1, 2]. Facial nerve is closely associated with PG capsule and facial nerve injury resulting in hemifacial paralysis is a major complication of parotid gland surgery. Lingual nerve accompanies Wharton's duct and then lingual nerve injury may be a complication of surgical exploration of floor of mouth for salivary stones removal.

Major salivary glands produce over 90% of saliva in a healthy adult. Saliva excretion is stimulated by both parasympathetic and sympathetic autonomic nervous system necessary for lubricating the oral cavity to enable eating, talking, swallowing, dental health and maintaining oral homeostasis, tasting, while also providing protective functions and aiding in digestion. Different types of saliva are produced by two types of acinar cells. Parotid gland secretes watery serous saliva produced by serous acini. SMGs and SLGs contain both serous and mucous acinar cells. Most of the acinar cells in SMGs are serous, while the majority of SLG is composed by mucous acinar cells. In addition to major salivary glands, there are several minor glands which are widely distributed across the oral mucosa [3].

Major salivary glands start to develop at 6–8 weeks of gestation. Oral ectoderm starts thickening to produce placodes of major glands. Growth factors and other necessary molecular cues for epithelial branching morphogenesis are provided by neural crest-derived mesenchyme. SMGs seem well differentiated with desmosomes and microvilli projected from cells along lumens by 13–16 weeks of gestation. At 16 weeks, the striated and intercalated ducts can be recognized and acinar cells begin to predominate tissue by 20–24 weeks [4]. Salivary glands continue to develop up to 28 weeks and glands can secret saliva at birth.

## **2. Salivary gland neoplasms**

Salivary gland tumors are a diverse group of neoplasms in terms of both morphology and clinical manifestations. These tumors have an incidence of approximately 2.5 cases to 3.0 cases per 100,000 per year [5]. Salivary gland malignancies consist more than 0.5% of all malignancies and approximately 3–5% of all head and neck cancers [6]. Salivary gland tumors usually involve people in their sixth or seventh decade of life [7, 8]. Recent studies have indicated that incidence of major salivary glands are increasing [9].

The exact etiology of salivary gland malignancies has not been determined; however ionizing radiation is indicated as a cause of salivary gland cancer [6, 7, 10, 11].

Tumors of salivary gland are related to both major and minor salivary glands. Minor salivary gland lesions are more frequently seen in palate; however, they can be found throughout oral cavity as oral mucosa, posterior tongue, larynx, pharynx, paranasal sinuses, retromolar and peritonsillar areas, uvula and floor of mouth [6, 12]. More than 50% of salivary gland tumors are benign and about 70–80% of all salivary gland neoplasms originate from parotid gland [13]. Different sites have different rate of malignancy so that 20–25% of parotid, 35–40% of submandibular, 50% of palate and more than 90% of sublingual gland tumors are malignant [14].

Comprising about 50% of all salivary gland tumors and 65% of parotid gland tumors, pleomorphic adenoma is the most common benign minor and major salivary gland tumor. On the other hand, mucoepidermoid carcinoma is the most prevalent major and minor salivary gland tumor comprising 10% of all salivary gland and about 35% of malignant salivary neoplasms [15]. This neoplasm mostly occurs in parotid gland [15, 16].

Most benign major and minor salivary gland tumors present with painless swelling of the parotid, submandibular or sublingual glands. Numbness or weakness which are signs of nerve involvement, typically indicate malignancy. Facial nerve weakness accompanied by parotid tumor is not a pleasant sign. A majority of benign and malignant tumors of parotid gland present as asymptomatic mass in the gland [13].

Tumors have more favorable prognosis when present in major salivary glands; parotid gland tumors have the best prognosis followed by submandibular gland. On the other hand, sublingual and minor salivary gland tumors have less favorable prognosis. Adequate surgical resection is enough for curing early-stage low-grade major and minor salivary gland tumors. In contrary, high-grade tumors have poorer prognosis and may need postoperative radiation therapy [17]. In addition, histology, grade, extent of primary tumor (stage), facial nerve involvement, fixation to skin or deep structures and lymph node or distant metastasis are among other factors involved in prognosis [18, 19].

## **3. Classification of tumors**

into the oral cavity under the tongue by the lingual frenum at a structure called the sublingual caruncule. In contrast, sublingual gland has small ducts called ducts of Rivinus and a common duct, Bartholin's duct, which connects with Wharton's duct at the sublingual caruncle [1].

Blood supply of parotid gland is mainly provided by the transverse facial artery and SMG by the facial artery. Postganglionic nerves from the otic and submandibular ganglia provide parasympathetic innervation. Superior cervical ganglion makes sympathetic postganglionic nerves which innervate the glands along blood vessels [1, 2]. Facial nerve is closely associated with PG capsule and facial nerve injury resulting in hemifacial paralysis is a major complication of parotid gland surgery. Lingual nerve accompanies Wharton's duct and then lingual nerve injury may be a complication of surgical exploration of floor of mouth for salivary

Major salivary glands produce over 90% of saliva in a healthy adult. Saliva excretion is stimulated by both parasympathetic and sympathetic autonomic nervous system necessary for lubricating the oral cavity to enable eating, talking, swallowing, dental health and maintaining oral homeostasis, tasting, while also providing protective functions and aiding in digestion. Different types of saliva are produced by two types of acinar cells. Parotid gland secretes watery serous saliva produced by serous acini. SMGs and SLGs contain both serous and mucous acinar cells. Most of the acinar cells in SMGs are serous, while the majority of SLG is composed by mucous acinar cells. In addition to major salivary glands, there are several

Major salivary glands start to develop at 6–8 weeks of gestation. Oral ectoderm starts thickening to produce placodes of major glands. Growth factors and other necessary molecular cues for epithelial branching morphogenesis are provided by neural crest-derived mesenchyme. SMGs seem well differentiated with desmosomes and microvilli projected from cells along lumens by 13–16 weeks of gestation. At 16 weeks, the striated and intercalated ducts can be recognized and acinar cells begin to predominate tissue by 20–24 weeks [4]. Salivary glands

Salivary gland tumors are a diverse group of neoplasms in terms of both morphology and clinical manifestations. These tumors have an incidence of approximately 2.5 cases to 3.0 cases per 100,000 per year [5]. Salivary gland malignancies consist more than 0.5% of all malignancies and approximately 3–5% of all head and neck cancers [6]. Salivary gland tumors usually involve people in their sixth or seventh decade of life [7, 8]. Recent studies have indicated that

The exact etiology of salivary gland malignancies has not been determined; however ionizing

Tumors of salivary gland are related to both major and minor salivary glands. Minor salivary gland lesions are more frequently seen in palate; however, they can be found throughout

minor glands which are widely distributed across the oral mucosa [3].

continue to develop up to 28 weeks and glands can secret saliva at birth.

radiation is indicated as a cause of salivary gland cancer [6, 7, 10, 11].

**2. Salivary gland neoplasms**

incidence of major salivary glands are increasing [9].

stones removal.

38 Diagnosis and Management of Head and Neck Cancer

Several pathological classifications have been proposed for salivary gland tumors since 50 years by several authorities such as Armed Forces Institute of Pathology and the World Health Organization (WHO). In 1954, there were only 16 salivary gland epithelial tumor entities in pathological classification which increased to 36 entities in 1996 showing an evident progress [20]. **Table 1** shows WHO histological classification of epithelial tumors of the salivary glands.

#### **3.1. Acinic cell carcinoma**

Acinic cell carcinoma (AcCC) comprises one of six parotid cancers which is supported by a nationwide study in the Netherlands where 15% of parotid malignancies were AcCC [21–23]. Based on 1973–2009 Surveillance, Epidemiology, and End Results (SEER) analysis AcCC comprises 11% of salivary malignancies [24]. AcCC has an average annual incidence of 0.13 cases per 100,000 patients per year. Also approved by other studies, SEER program indicated that AcCC has a higher average incidence for females than males (0.15 cases vs. 0.11 cases per

#### **Malignant epithelial (n = 24)**

Acinic cell carcinoma

Oncocytic carcinoma

Mucoepidermoid carcinoma

Salivary duct carcinoma

Adenoid cystic carcinoma

Adenocarcinoma (NOS)

Polymorphous (LG)

Adenocarcinoma

Myoepithelial carcinoma

Epithelial-myoepithelial carcinoma

Carcinoma ex pleomorphic adenoma

Clear cell carcinoma (NOS)

Carcinosarcoma

Basal cell adenocarcinoma

Metastasising pleomorphic adenoma

Sebaceous carcinoma

Squamous cell carcinoma

Sebaceous lymphadenocarcinoma

Small-cell carcinoma

Cystadenocarcinoma

Large cell carcinoma

LG cribriform cystadenocarcinoma

Lymphoepithelial carcinoma

Mucinous adenocarcinoma

Sialoblastoma

#### **Benign epithelial (n = 10)**

Pleomorphic adenoma

Lymphadenoma

Sebaceous

Nonsebaceous

Ductal papilloma

Inverted ductal papilloma

Intraductal papilloma

Sialadenoma papilliferum

**Table 1.** WHO (2005) histological classification of epithelial tumors of the salivary glands, squamous cell carcinoma (SCC) and tumors like "carcinoma in pleomorphic adenoma".

100,000 patients) [25, 26]. Patients with AcCC have a median age of 52 years at the time of diagnosis and it occurs more in younger ages than other salivary gland tumors [27]. One-third of patients are under 40, one-third between 40 and 59 and one-third above 60 years of age [24]. AcCC more frequently occurs in whites (85%) than in blacks (7%) or other races (8%) [24, 28]. There are no known risk factors for AcCC; however, family history and radiation exposure have been proposed as potential risk factors [29].

The most common clinical feature of AcCC is a slow-growing swelling and its association with pain or fixation to neighboring structures is an indicator of poor prognosis [25]. Nodal metastasis is really uncommon at the time of diagnosis [26]. AcCC is less common in minor salivary glands and comprises only 9% according to SEER database [30]. AcCC of minor salivary glands mostly occur in buccal mucosa and upper lip in contrast to other types of minor salivary glands which mostly occur in palate [31, 32].

#### *3.1.1. Diagnosis and preoperative assessment*

**Malignant epithelial (n = 24)** Acinic cell carcinoma Oncocytic carcinoma

40 Diagnosis and Management of Head and Neck Cancer

Mucoepidermoid carcinoma Salivary duct carcinoma Adenoid cystic carcinoma Adenocarcinoma (NOS) Polymorphous (LG) Adenocarcinoma

Myoepithelial carcinoma

Clear cell carcinoma (NOS)

Basal cell adenocarcinoma

Sebaceous carcinoma Squamous cell carcinoma

Small-cell carcinoma Cystadenocarcinoma Large cell carcinoma

Sialoblastoma

**Benign epithelial (n = 10)** Pleomorphic adenoma Lymphadenoma Sebaceous Nonsebaceous Ductal papilloma

 Inverted ductal papilloma Intraductal papilloma Sialadenoma papilliferum

Carcinosarcoma

Epithelial-myoepithelial carcinoma Carcinoma ex pleomorphic adenoma

Metastasising pleomorphic adenoma

Sebaceous lymphadenocarcinoma

LG cribriform cystadenocarcinoma Lymphoepithelial carcinoma Mucinous adenocarcinoma

Since AcCC often presents symptomless, preoperative assessment seems necessary to determine location of tumor, extent and malignancy indicators as in parotid surgery these factors show the risk of facial nerve injury [33]. Imaging is highly suggested when a glandular swelling is accompanied by impaired mobility or when involvement of deeper structures or nerves is suspected [34–37].