**7. Diagnosis of IDA**

Diagnosis of IDA involves reduced hemoglobin levels and disturbed iron parameters. The initial evaluation includes a complete blood count (CBC), reticulocyte count, peripheral blood smear, and serum iron parameters. The CBC is useful in determining the concentration of hemoglobin, the mean corpuscular volume, and the average size of RBC.-

The CBC highlights:-

RBC dimensions are variable, unequal in size and generally smaller, this situation being known- as anisocytosis. This reflects an increased red blood cell distribution width index (RDW). The number of white blood cells and their morphology is normal, whereas moderate thrombocytosis is frequently encountered, but it resolves after the correction of iron deficiency.-

After the diagnosis of anemia, one should proceed to explore the iron metabolism. Evaluation of serum ferritin is the most used diagnostic test, but its level increases in inflammatory- disease and in old persons. For the diagnosis of IDA, some parameters as transferrin, soluble transferrin receptor (sTfR), and the soluble transferrin receptor/ferritin index could be useful.- The accuracy of some of these markers is limited being influenced by many other conditions,- especially the level of serum ferritin. The most used parameter, serum ferritin level, can be increased in acute or chronic inflammation, old persons, chronic infection, or malignancy- status. One more sensitive parameter for IDA diagnosis is soluble transferrin receptor (sTfR)- that remains unaffected by inflammatory states being an indirect measure of erythropoiesis.-

Usually, in clinical practice, the diagnosis of IDA includes low hemoglobin level, low transferrin saturation (<15%), low levels of serum ferritin (<30 ug/dl), and high total iron binding capacity (>13.1 umol/l). Another method of exploring iron reserves consists in the appreciation of the medullary reticulocytes through a medullary smear (bone marrow cytology) and Perls staining. This highlights the absence of hemosiderin (iron reserves), the absence of sideroblasts (lack of iron in erythroblasts), and the absence of siderocytes (lack of iron in red blood cells). This exam is not routinely required.-

Finally, after establishing the diagnosis of IDA, an iron deficiency cause should be investigated, as the most common cause in men is digestive losses and in women genital losses. These investigations should include gynecological examination and the exploration of the digestive tract: gastroscopy, colonoscopy, and other investigations according to specific elements identified in the medical history and physical exam [33, 49–51].

### **8. Differential diagnosis-**

The differential diagnosis of IDA should be made with other forms of microcytic anemia (see **Table 4**):-

**(a) Anemia from chronic disease** where one of the main mechanisms is the blockage of iron in its reserves, as in macrophages and the inability to be used in erythropoiesis.

### *Common elements:*

#### *Elements of differential diagnosis:-*

### **(b) Thalassemia minor**

*Common elements:*

• Hypochromic microcytic anemia.

*Elements of differential diagnosis:-*

### **Anemia from chronic disease**

 Chronic inflammatory disorders Chronic infections Autoimmune diseases Kidney diseases Cancer- **Thalassemia minor**

**Sideroblastic anemia**

**Table 4.** Differential diagnosis of iron-deficiency anemia.-

Smear shows frequent red blood cells with basophile points and bull's-eye erythrocytes (codocytes):-

The diagnosis of certainty is determined by hemoglobin electrophoresis: increased amount of fetal hemoglobin (HbF) or hemoglobin A2.-

### **(c) Sideroblastic anemia**

*Common elements:*

• Hypochromic microcytic anemia.

*Elements of differential diagnosis:-*

#### *Difficulties in diagnosis:-*

### **9. Treatment**

### **9.1. General aspects**

Iron-deficiency anemia and/or iron deficiency should be treated, regardless of the presence or absence of symptoms. The correction of anemia is vital; depending on its severity, anemia could trigger cardiovascular complications, sometimes severe [55, 56].

Treatment of IDA has two major objectives:

### **9.2. Iron therapy**

The treatment of iron deficiency is either enteral or parenteral.-

Intramuscular administration of iron is rarely used due to local complications such as pain at administration site, uneven absorption, skin pigmentation, and even development of sarcomas at the injection site as some authors have reported [57, 58].

Transdermal administration has been experimented in animal models, but there is no evidence that it would be effective and safe in humans [59].

### *9.2.1. Oral iron therapy*

### *<IP_ADDRESS>. Treatment with oral iron products*

Treatment with oral iron products is preferable due to the low cost and lack of anaphylactic side effect. Usually, it is used to treat iron-deficiency anemia in children and adolescents.-

### **Indication:**

The main indications of oral iron are treatment of IDA, treatment of iron deficiency, and prophylaxis of iron deficiency in subjects with high needs such as pregnant women, adolescents, and preterm children.

A wide range of iron products is available in forms of tablets or liquid. Liquid iron products have the advantage of allowing titration of the dose. The most commonly used oral iron products are polysaccharide iron complex, ferrous gluconate, ferrous sulfate, and ferrous fumarate.

### **Dosage:**

Regarding the dosage, various over-the-counter tablets exist depending on the products, but generally, there are no differences between these products in effectiveness or adverse events. The daily dose needed to correct iron deficiency depends on age, iron deficiency, and urgency. In general, daily dose for the correction of IDA with oral products is 150–200 mg elemental iron. Reports show that this dose taken every other day is most effective and has a better absorption and fewer side effects [60, 61].

Toxicity of iron products is more important in elderly patients, so doses are smaller. In a randomized study of 90 patients, of over 80 years old with iron deficiency, doses of 10, 15, or 150 mg for 2 months were equally effective [62].

The bioavailability and absorption of oral iron products may be influenced by food (e.g., phytates, phosphates, and tannins can bind iron and prevent absorption) or by drugs which diminish gastric secretion (antacids, histamine receptor inhibitors, proton pump inhibitors, etc.) reducing the absorption of iron. This is why iron products are administered 2 hours before and 4 hours after the administration of antacid absorption of iron as ferrous iron depending on acidic environment.

In general, iron product administration is associated with acidic foods (orange juice) or vitamin C to promote iron absorption, but there are no studies that show a positive effect of vitamin C in the absorption of iron [63].

There is no consensus on the duration of treatment with oral iron products. Some experts end treatment after correcting anemia to allow early detection of secondary anemia, due to recurrent blood loss. Others recommend continuing treatment for 6 months after correction of anemia in order to supplement iron reserves. Generally, 6 weeks of treatment are needed to correct anemia, followed by 6 months to replenish the reserves.

#### **Side effect of oral therapy:-**

Concerning the side effects of oral iron therapy, the most common side effects involving gastrointestinal tract are metallic taste, nausea, flatulence, constipation, diarrhea, epigastric pain, and/- or vomit. Black stools create anxiety as they may be misinterpreted as gastrointestinal bleeding.- Some measures can be taken in order to diminish iron side effects and increase tolerability as increasing the time between administrations (given every other day, not daily), changing diet (concomitant administration of food, even if absorption is reduced), reducing daily dose of elemental iron, switching from tablets to liquid preparations that allow titration of doses, or switching from tables to IV administration.

### *<IP_ADDRESS>. Treatment with IV iron products*

### **Indication:**

There are some specific indications for IV administered iron products when oral administration is inefficient or contraindicated (see **Table 5**):-

**Table 5.** Iron therapy: Indication and side effects of enteral versus parenteral administration.-

Administration of IV products leads to reduced transfusion requirements for certain patients reducing the risk of posttransfusion adverse reactions.-

Several iron products are available for IV administration; the most important are ferric carboxymaltose (FCM), ferric gluconate (FG), ferumoxytol, iron sucrose (IS), iron isomaltoside, and low molecular weight iron dextran (LMW ID).-

Differences between these products consist in cost and the number of visits required for the administration of the total dose, namely, the time of administration and the number of doses required.

The total amount of iron required depends on the intended purpose, correcting anemia or supplementing the iron reserves.

Dose calculation is based on body weight, patient hemoglobin levels, and elemental iron/ml concentration of the product. In clinical practice, a 1000 mg dose of iron is generally enough to treat anemia. Generally, no premedication prior to IV administration is needed, except for patients with a history of asthma or drug allergies or patients with inflammatory arthritis for which 125 mg of methylprednisolone before iron administration is recommended [64].

Generally, antihistamines are not used in preventing side effects [65–67].

### **Administration and dosing of various iron products:**

*LMW Fe Dextran*: It can be given in a single dose of 1000 mg in 250 ml of saline, 1 hour infusion, or multiple doses of 2 ml, i.e., 100 mg elemental Fe (2 ml ampoule, 50 mg Fe/ml).-

If single dose is preferred, a test dose of 0.5 ml is administeredin 30 seconds prior to the infusion.

*Ferric gluconate (Ferrlecit)*: It is administered in a 10–15 ml dose, the equivalent of 125–187.5 mg (1 ampoule contains 12.5 mg of elemental Fe/ml). In patients with drug allergies, it is recommended to administer a test dose prior to infusion. Each dose can be given by bolus or infusion of 20–30 minutes.-

*Fe sucrose (Venofer)*: Multiple infusions with a maximum dose of 10–15ml, equivalent to- 200–300mg of elemental Fe (ampoule 20mg/ml), are administered. It is recommended- to administer a 1.25ml test dose (25mg IV, administered slowly to patients with drug- allergies).-

*Ferumoxytol*: It consists of superparamagnetic iron oxide nanoparticles coated with a low molecular weight semisynthetic carbohydrate. It is administered at a dose of 17 ml, the equivalent of 510 mg elemental Fe (ampoule 30 mg/ml), with a 15 minute infusion.-

Ferumoxytol may interfere with magnetic resonance imagery (MRI), which can lead to errors in interpreting the results.

*Ferric carboxymaltose (Ferinject)*: It is used in 20 ml dose, equivalent to 1000 mg of elemental iron (the vial contains 50 mg/ml, the maximum dose being 20 mg Fe/kg body weight). Doses are given in a 15 minute infusion. It is efficient and safe [68–71]. It can sometimes cause hypophosphatemia.

*Iron isomaltoside (Monofer)*: It has a matrix structure where lean iron is slowly removed, allowing administration of 20 mg/kg infusion at 15 minutes; the ampoule is 100 mg/ml. It does not require a test dose. Efficacy has been demonstrated in inflammatory bowel disease, chronic renal disease, heart surgery, and chemotherapy-induced anemia [72–76].

### **Side effects of IV iron products:-**

The most common side effects consist in allergic reactions, sometimes anaphylactic shock. However, these reactions are extremely rare and often overestimated.

Patients with inflammatory arthritis (rheumatoid arthritis) may experience an exacerbation of symptoms during iron infusion, which can be prevented by administering 125 mg of methylprednisolone.

Reducing the risk of allergic reactions can be achieved through the following measures:

Nonallergic reactions may occur, which include located rash, heart palpitations, dizziness, muscle spasms of the neck, scapula-vertebral region, etc.

The risk of infection may be increased because iron acts as a growth factor for bacteria. This is supported by some authors but denied by others [77, 78].

### **9.3. Monitoring and evaluating response to treatment**

Monitoring the response to iron therapy depends on the severity of anemia and on the way of iron administration. For patients treated with oral products, reassessment is done after 2 weeks of treatment by checking Hb levels and reticulocytes. For parenteral treatment, reevaluation is done after 4–8 weeks. Iron treatment is continued until ferritin and transferrin return to normal. There are some either clinical or laboratory criteria used to monitor the efficacy of treatment with iron.-

Amelioration of asthenia and installation of a well-being state in the first days of treatment besides the increased level of hemoglobin within 1–2 weeks of treatment seem to be the most important parameters related to iron-deficiency correction. In patients with severe anemia, a modest reticulocytic crisis occurs within 7–10 days after the initiation of treatment. In those with mild anemia, this reticulocytosis is not encountered.


*Integrins in Ovarian Cancer: Survival Pathways, Malignant Ascites and Targeted Photochemistry DOI: http://dx.doi.org/10.5772/intechopen.106725*

Fluorescence imaging of cancer relies on the selective accumulation of fluorescent agents in cancer cells. αvβ3 integrins are the most common targets in integrin-targeted fluorescent imaging studies. For instance, the fluorescent probe squaraine was covalently attached to one (monovalent) and two (divalent) cyclic RGD peptides by Shaw and colleagues to target ovarian cancer cells that overexpress αvβ3 integrins [167]. Uptake of the divalent probe in OVCAR4 cells was 2.2-fold higher than the monovalent probe, based on fluorescence imaging. Consistently, tumors grown in nude mice and imaged with the divalent probe were almost three times more fluorescent compared to tumors given the monovalent probe, and six times more fluorescent than tumors that received non-conjugated squaraine. To explore the potential of integrin-targeted, fluorescence-guided resection in ovarian cancer, Alvero *et al.* created a PLGA-PEG nanoparticle to target αvβ3 integrins in OCSC1-F2 ovarian cancer cells using an RGD peptide and three different fluorescent dyes: DIR, C6, and ICG [168]. The resulting conjugates enabled the investigators to visualize both the tumorassociated vasculature and intraperitoneal ovarian cancer micrometastases as small as 100 μm in a xenograft model. These studies demonstrate the potential of using αvβ3-targeted agents for fluorescence guided resection in ovarian cancer. An additional important consideration for this approach is the accuracy of tumor detection. This concern was addressed in a study by Harlaar *et al.*, who found that the diagnostic accuracy of an αvβ3-targeted agent in combination with an NIR fluorescence intraoperative imaging system was 96.5%, with a sensitivity of 95% and a specificity of 88% [169].

In comparison to RGD peptides that have been relatively widely used to target integrins, less commonly used peptides, such as "OA02", have been synthesized to bind an α3 integrin subunit [170]. An *in vivo* study by Aina *et al.* used nude mice bearing ES-2 tumors to evaluate three different forms of this peptide: OA02-biotin-Cy5.5, OA02-Cy5.5, and OA02-AlexaFluo 680. Results showed that OA02-Cy5.5 and OA02- AlexaFluo 680 exhibited fast and specific tumor uptake that sustained a fluorescence signal for approximately 70 minutes. Although the cellular uptake of OA02 biotin-Cy5.5 was slower than other peptide variants, the duration of the fluorescence signal was 24 hours. To confirm that α3 integrins were mediating the binding of OA02 peptides to ES-2 tumors, mice were injected with an anti-α3 monoclonal antibody, which blocked binding of the peptides to the tumors.

The value of targeting integrins has also been explored in the context of PTT, which involves the interaction of electromagnetic radiation (typically NIR light) with a photothermal agent to generate heat, leading to tissue hyperthermia. In a study by Zhou *et al.*, the selectivity of silica-coated gold nanorods for ovarian cancer cells increased using hyaluronic acid and an RGD peptide that bind to CD44 and αvβ3 integrin, respectively [171]. The targeted nanoparticle showed high selectivity for SKOV3 cells but not for non-cancerous HOSEpiC cells. The nanorods were also loaded with doxorubicin (DOX) to increase cytotoxicity. PTT using the dual-targeted, DOX-loaded gold nanorods, and irradiation with an 808 nm laser at a high-power density (2 W/cm2 ), exhibited the highest cytotoxicity to SKOV3 cells among all experimental groups. Subsequent experiments have revealed that the release of DOX is pH-sensitive and triggered by NIR irradiation. DOX release may be influenced by hyaluronidase-mediated degradation of hyaluronic acid in low pH environments, and the disruption of the interaction between DOX and silica, respectively. An overview of photochemistry-based studies that focus on modulating or targeting integrins in the context of ovarian cancer is presented in **Figure 5**.

### **Figure 5.**

*Integrins as targets for fluorescence imaging and photochemical or photothermal treatment in ovarian cancer. Current research focuses on using RGD tripeptide-conjugated PS or PTT agents to target ovarian cancer cells that overexpress integrins (left), or to modulate integrin activity and inhibit cancer cell adhesion to secondary sites by low level cellular photodamage (right).*

In summary, targeting integrins is a promising strategy for both anti-cancer PDT and fluorescence imaging. Since most PSs also have fluorescent properties, novel nanocarriers with integrin-targeting molecules can be used in theranostic applications and in real-time image-guided PDT of ovarian cancer. The potential of integrintargeted PDT warrants further evaluation.


**Acknowledgement**

The author would like to thanks Instituto Federal de Educação, Ciência e Tecnologia de São Paulo for the MATLAB license and hardware resources used at the development of this work.