Matrix metalloproteinases (MMPs) play a role in many physiological processes, have additional roles in reorganization of tissues during pathological conditions such as inflammation\[[@ref1][@ref2]\] and in immunosurveillance by leukocytes against infections.\[[@ref3]\] This classical paradigm was further developed by tumor biologists who discovered that an increased expression of proteinases, including MMPs, is a marker of invasion and metastasis of cancer cells. At that time, the hope was to use MMP inhibitors (MMPIs) to halt the spreading of cancer cells. However, during clinical trials of metastatic cancer, severe side effects were observed, thus leading researchers to reappraise the use of MMPIs for the therapy of invasive cancer.\[[@ref4][@ref5]\] The positive effect of these cancer researches and clinical studies is that the marker functions of MMPs have been refined in several ways. In this review, we discuss important biological aspects of MMPs in view of the uses of MMPIs in inflammatory periodontal disease.

MMPs - Redundancy, Expression Patterns, and Balances {#sec1-2}
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MMPs are multidomain enzymes containing a zinc ion, which are coordinated by three histidine residues in their active site. Although all MMPs possess different primary structures, they are composed of shared modules, known as protein domains. The hallmark of the MMP family is a catalytic domain that possesses a zinc-binding consensus sequence, a characteristic shared with other metalloproteinase families such as the ADAMs (a disintegrin and metalloproteinases) and the ADAMTSs (ADAMs with a thrombospondin motif). Another signature of the MMP is its activation by the so-called cysteine switch.\[[@ref6]\] When cells produce MMPs, most of the enzymes are secreted in a latent pro-form and removal of the pro-peptide (about 10 kDa) from the active site, for example, by proteolysis, leads to activation of the enzymes.\[[@ref7]\]

Expression levels of MMPs depend on the biological context, for example, some constitutive or homeostatic MMP genes possess simple promoter enhancer regions with cis-acting elements for basal transcription and are switched on in most cells under steady-state conditions. Other MMPs have complex promoter regions. The expression of these MMPs is regulated by various agonists. The biological milieu will determine the levels of expression of these inducible or inflammatory MMPs. Interesting examples of this dichotomy are the constitutive MMP2 and the inducible MMP9. Several other redundant enzymes exist with similar catalytic functions in extracellular matrix (ECM) biology. The three classical collagenases, interstitial collagenase (MMP1), neutrophil collagenase (MMP8), and collagenase 3 (MMP13), all cleave a specific scissile bond in the triple-helical collagens at one specific site. This redundancy, also observed for the stromelysins, ensures that the biological processes of ECM remodeling can take place under various conditions by different cell types, and so if one enzyme is inactivated then the host can still survive.

In addition to regulation by activation processes and gene expression, the activities of MMPs are also controlled by the four natural tissue inhibitors of metalloproteinases (TIMPs).\[[@ref8]\] This implies that the biological processes involving MMPs are always dependent on balances between proteinases and natural inhibitors. In conclusion, treatment with MMPIs will cause a distortion of these natural balances and so the targeting of MMPs is a challenging exercise in selectivity.

MMPs and Inflammation {#sec1-3}
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During an inflammatory response, leukocyte trafficking through tissue barriers, including basement membranes, is only possible if these cells are equipped with enzymes that can remodel the extracellular matrix.\[[@ref9]\] MMPs are therefore crucial effector molecules of inflammatory cells.\[[@ref10]\] However, MMPs can also modify cytokines and chemokines.\[[@ref11]\] They can act as switches or as delicate tuners in acute and chronic inflammation and in the regenerative phase after inflammation. Thus, MMP biology is important in the initiation, execution, and resolution phases of acute and chronic inflammatory and ischemic processes and consequently, MMPIs might interfere with these.

MMPs and Inflammatory Periodontal Disease {#sec1-4}
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Inflammatory destruction of periodontal attachment apparatus is the hallmark of periodontal disease. The inflammatory reaction associated with periodontitis may damage the surrounding cells and connective tissue structures, including alveolar bone, causing tooth loss.\[[@ref12]--[@ref14]\] During this event, the most important component of periodontium lost is the collagen type I that is found in the periodontal ligament and alveolar bone organic matrix. Four distinct pathways may be involved with this destruction: Plasminogen-dependent, phagocytic, osteoclastic, and MMP pathway.\[[@ref15]\] A wide range of evidences has indicated that the most important pathway is the one which involves MMPs.\[[@ref16]\]

Resident ligament cells such as fibroblasts, macrophages, osteoblasts, keratinocytes, and endothelial cells are activated in response to stimulus, contributing to the synthesis of cytokines and MMPs. MMPs are present in both active and latent forms in chronically inflamed gingival tissues and gingival crevicular fluid. Active collagenase and gelatinase are found in the crevicular gingival fluid of patients with periodontitis in much larger amounts than in control subjects.\[[@ref16]\] In contrast, high concentrations of the natural TIMPs are found in the gingival crevicular fluid of healthy gingiva.\[[@ref17]\] The area occupied by collagen fibers in gingival tissue specimens with periodontitis is significantly decreased, and the presence of MMP1, MMP2, and MMP9 is increased.\[[@ref18][@ref19]\]

MMPs and Alveolar Bone Resorption {#sec1-5}
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MMPs play a key role during this event. Osteoclasts are unable to attach to the bone surface if the mineralized bone matrix is covered by an osteoid layer.\[[@ref20]--[@ref22]\] Osteoblast-derived collagenase (MMP13) seems to be mainly responsible for degrading the nonmineralized osteoid layer covering bone surfaces, exposing the mineralized matrix to osteoclasts.\[[@ref23]\] MMP13 (collagenase-3) is expressed in osteoblasts, periosteal cells, and fibroblasts during bone remodeling.\[[@ref24]\] MMP13 is as efficient as MMP1 and MMP8 in the digestion of type I collagen.\[[@ref25]\] Cleavage of collagen I by MMP13 seems to be the initial step of the entire bone resorption process\[[@ref26]--[@ref28]\] and subsequently, denatured collagen fragments are also degraded by gelatinases, MMP2 and MMP9. Studies have demonstrated that MMP3, MMP9, and MMP13 mRNA levels are increased when osteoblast cultures are stimulated by resorptive factors such as cytokines interleukin (IL)-1b and tumor necrosis factor (TNF)-α, parathyroid hormone, and prostaglandin E~2~.\[[@ref29]\] Other direct evidence of the participation of MMPs in bone resorption is the fact that inhibition of MMPs by chemically modified tetracyclines can prevent bone loss.\[[@ref30]\]

MMPIs - Current Scenario and Their Drawbacks {#sec1-6}
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Both macromolecular inhibitors (natural TIMPs and monoclonal antibodies) and small molecules (synthetic and natural products) have been considered as potential therapies for diseases in which excess MMP activity has been implicated.\[[@ref31]\]

Monoclonal antibody derivatives are promising drugs to be used as therapeutics, especially if a high MMP specificity is required.\[[@ref32]\] TIMPs that have affinity for MMPs in the picomolar range seem to be ideal inhibitors, but they lack good selectivity and possess other biological functions, which could lead to side effects.\[[@ref33]\]

Hydroxamate-based MMPIs, succinyl hydroxamate MMPIs (batimastat, BB-1101, and marimastat), CGS27023A, an N-sulfonyl amino-acid hydroxamate, and new generation hydroxamate-based MMPIs, MMI-270 and Prinomastat, have been extensively studied. Carboxylate MMPIs, new generation thiol-based MMPIs (Rebimastat), broad-spectrum peptidomimetic inhibitors, pyrimidine-based inhibitors, hydroxypyrone-based MMPIs, and phosphorous-based MMPIs are the non-hydroxamate-based MMPIs that have been evaluated for therapeutic use in various fields of medicine.\[[@ref34]\]

Unfortunately, these MMP intervention strategies have met with limited clinical success and severe toxicities. Most of the MMPIs eventually demonstrated side effects after short-term dosing (e.g. Marimastat) or prolonged treatment (e.g. BMS-275291), related to musculoskeletal pain and inflammation. The mechanism of these toxicities is widely assumed to be due to the poor selectivity of these compounds, but this has not been confirmed. In addition, it is now recognized that among the MMPs, some possess cancer-promoting activities while others have tumor-inhibiting functions underlining the risk of using broad-spectrum MMPIs.\[[@ref35]\]

Bisphosphonates\[[@ref36]\] have been studied for their MMPI effects and their clinical performance has not met their anticipated utility.

Tetracyclines are antibiotics that also inhibit the breakdown of connective tissue. Chemically modified tetracyclines (CMTs) without antibiotic activities have several potential advantages over conventional tetracyclines due to the absence of gastrointestinal side effects or toxicity and higher plasma concentrations can be reached for prolonged periods of time.\[[@ref37]\] Minocycline is an inexpensive MMPI and is currently being evaluated as a leading compound for the treatment of multiple sclerosis and vascular neurological disorders, as it crosses the blood-brain barrier easily.\[[@ref38]\] This subantimicrobial-dose doxycycline (SDD) has become widely established as an effective MMPI.

Next Generation of MMPIs: Antibody-based Therapeutic Agents {#sec1-7}
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Successful therapeutic intervention may critically depend on potently inhibiting one or more MMPs that contribute to disease progression while not inhibiting related MMPs that may be beneficial to the host or if inhibited lead to clinical toxicities. Monoclonal-antibody derivatives are promising drugs to be used as therapeutics, especially if a high MMP specificity is required.\[[@ref35]\] The high binding affinity of a monoclonal antibody to its target confers the potential for high potency and selectivity coupled to a drug scaffold with excellent pharmacological properties.

MMPIs in Periodontal Diseases {#sec1-8}
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Currently, clinical therapy inhibiting the mediators of connective tissue breakdown is used for the adjunctive treatment of periodontitis.\[[@ref39]\] This is accomplished through the non-antimicrobial activities of low-dose tetracycline and tetracycline analogs via the inhibition of MMP8 and MMP13 protease mechanisms.\[[@ref40]\] The tetracycline analog, doxycycline hyclate, available for use specifically in periodontal disease, is the only collagenase inhibitor approved by the United States Food and Drug Administration (FDA) for any human disease.\[[@ref41]\] The therapeutic action witnessed is primarily due to the modulation of the host response because the low-dose formulations of these drugs have lost their antimicrobial activity.\[[@ref40]\]

This SDD approach has become widely established as an effective adjunctive systemic therapy in the management of periodontitis, along with the traditional mechanical therapies of scaling and root planing (SRP). For example, initial Phase III clinical trials of MMP inhibition by SDD in subjects with periodontal disease over a 6-month period led to maintained alveolar bone height compared to bone height loss with placebo.\[[@ref42]\] In chronic periodontitis, the more severe the periodontitis, the greater is the observed attenuation of disease activity by SDD therapy.\[[@ref43]\] In a recent systematic review, the effectiveness of SRP accompanied by MMP inhibition (by SDD), as an adjunctive treatment, showed improved outcomes that persisted for 9 months in adults with chronic periodontitis as observed in gains of clinical attachment level (CAL) and probing depth (PD) reduction.\[[@ref44]\] Most recently, in a double-masked, randomized, placebo-controlled, multicenter study of 266 subjects with periodontal disease, those individuals treated with a modified-release SDD formulation taken once daily as an adjunct to SRP displayed significantly greater clinical benefits (improved CAL gain and PD reduction) than individuals treated with SRP alone.\[[@ref45]\]

There have been other therapeutic approaches that involve SDD in the treatment of periodontitis. Notably, a recent proof-of-principle study was designed to examine aspects of the biologic response brought on by SDD combined with access flap surgery (AFS) on the modulation of periodontal wound repair in subjects with severe periodontitis who were not candidates for regenerative therapy, for the augmentation of periodontal wound healing (through improving CAL and PD), increased bone stabilization, and decreased MMP expression. The results of this investigation demonstrated that SDD, in combination with AFS, may improve the response to surgical therapy during drug dosing by reducing PD in cases of severe periodontitis compared to AFS alone.\[[@ref46]\] SDD tended to reduce post-surgical bleeding on probing (BOP), PD, and periodontal bone resorption during drug administration, yet it did not affect the periodontal microflora beyond the contribution of surgery alone. Other accumulating evidence demonstrated the ability of MMP inhibitors to be used in the management of periodontal disease in patients with decreased bone mass (as in the situation of postmenopausal osteoporosis). Recent studies\[[@ref47][@ref48]\] demonstrated the ability of SDD to be used to maintain bone mass while reducing periodontal disease progression. Furthermore, oral fluid-derived (i.e. gingival crevicular fluid) biomarkers, such as collagen telopeptide fragments, were reduced in subjects following SDD dosing.\[[@ref49][@ref50]\]

There is strong evidence to suggest that inhibition of MMPs in patients with periodontal disease clearly offers potential in disease management when coupled with mechanical therapy, such as SRP, and preliminary evidence available suggests the value of MMP inhibitory therapies for patients with peri-implant disease or in those conditions requiring surgical management.

MMPI in the Periodontal Clinic {#sec1-9}
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A tetracycline derivative, doxycycline, in subantimicrobial doses (Periostat; CollaGenex Pharmaceuticals Inc., Newtown, PA, USA) is currently the only MMPI approved by the US FDA and is used as an adjunct therapy in chronic periodontitis.\[[@ref39]\]

Challenges and Future Directions of MMPIs in Periodontitis {#sec1-10}
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A number of questions need to be answered in the management of periodontitis considering that these are chronic conditions with reference to the long-term consequences of extended MMP inhibition in patients and also the downstream-extended effects on controlling cytokine processing, especially concerning the role of decreasing MMP8 activity over time. Perhaps through the development of more selective MMPIs for periodontal treatment (to reduce potential side effects) and by examining combination therapy approaches considering antimicrobial and host response targets, we may make further inroads into answering these questions. Encouragingly, a survey of the recent trends in scientific publications suggest a continued interest in understanding the role of MMPs in normal bone homeostasis and periodontal diseases, which suggests that many of the unanswered questions are being addressed.

Conclusion {#sec1-11}
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Host response modulation has emerged as a valid treatment concept for the management of periodontal disease and represents a significant step forward for clinicians and patients. To date, only SDD has been approved specifically as a host response modulator. Further research is necessary to develop MMPIs for the clinical practitioner with a strong potential for the modulation of the host response in aiding periodontal disease management, when coupled with traditional mechanical therapy.

**Source of Support:** Nil

**Conflict of Interest:** None declared.
