In the world of biotherapeutic development, one of the first keys to success remains the identification of new targets for innovative drugs, and a detailed understanding of their mechanism of action in the targeted pathology. Within the family of metalloproteinases, matrix metalloproteinase-9 (MMP-9) is one such immunotherapy target that has been the focus of particular attention for several years. MMP-9 is an enzyme that plays an essential role in various physiological and pathological processes, making it an interesting candidate for therapeutic intervention. So let's take a closer look at the importance of MMP-9, its role in health and disease, and together explore the promising route of SYnAbs monoclonal antibodies as a means of exploiting its therapeutic potential.

Matrix Metalloproteinases (MMPs) are a family of enzymes responsible for the degradation and remodeling of the extracellular matrix (ECM) in tissues. MMP-9, also known as Gelatinase B, is one of the key members of this family. It is a zinc-dependent endopeptidase that primarily targets gelatin, collagen, and elastin, which are crucial components of the ECM.

MMP-9 is synthesized as an inactive zymogen (proMMP-9) and requires activation to exhibit its enzymatic activity. Once activated, MMP-9 can cleave various ECM proteins, facilitating tissue remodeling during processes like wound healing, tissue repair, and embryogenesis. Its controlled activity is essential for maintaining tissue homeostasis.

The activity of MMP-9 is tightly regulated at multiple levels to prevent excessive tissue degradation. Endogenous tissue inhibitors of metalloproteinases (TIMPs) bind to activated MMP-9, inhibiting its proteolytic activity. Dysregulation of this balance can lead to pathological conditions, making MMP-9 an intriguing therapeutic target.

In normal physiological conditions, MMP-9 plays several essential roles in the body:

• Wound Healing: MMP-9 assists in tissue repair by breaking down ECM components to allow for cell migration and tissue remodeling at the site of injury. During wound healing, MMP-9 is suggested to be involved in keratinocyte migration and granulation tissue remodeling. MMP-9 production is enhanced by interleukin-1 beta (1).

• Embryonic Development: During embryogenesis, MMP-9 contributes to processes such as tissue morphogenesis and organogenesis. MMPs expressed in precursor stromal cells promote their differentiation and expansion (2), and MMP-9 play an important in trophoblast invasion during pregnancy (3).

• Immune Response: MMP-9 is involved in immune cell migration and infiltration during inflammatory responses, aiding in the resolution of infections and tissue recovery since MMP-9 is produced by neutrophils or macrophages (4).

While MMP-9 is indispensable in normal physiological processes, its dysregulation is implicated in various pathological conditions:

• Cancer Metastasis: The over-expression of MMP-9 facilitates metastasis and tumor progression by degrading the extracellular matrix (ECM), which allows tumor cells to migrate and colonize host tissues (5).

• Inflammatory Diseases: Chronic inflammatory conditions such as arthritis and inflammatory bowel disease exhibit increased MMP-9 activity, contributing to tissue damage. The abundance and activation of matrix metalloproteinases significantly increases in ulcerative colitis and Crohn's mucosa (6) and MMP9 is overexpressed in in latent and active form of rheumatoid arthritis and psoriatic arthritis (7).

• Neurological Disorders: In neurodegenerative diseases like Alzheimer's and multiple sclerosis, MMP-9 is involved in blood-brain barrier disruption and neuroinflammation (8)

• Cardiovascular Diseases: MMP-9 is associated with the degradation of blood vessel walls, contributing to atherosclerosis and aneurysm formation. MMP-9 levels and activity are higher in unstable plaques than in stable plaques (9).

Given the significant role of MMP-9 in various disease processes, targeting its activity has become a focus of therapeutic research. Various drugs have emerged as a promising avenue for inhibiting MMP-9 effectively.

MMPs are highly expressed in human cancer and are linked to every stage of cancer development. Indeed, the role of MMP-9 in angiogenesis, metastasis, invasion and survival of cancer cells has been widely described in several publications (10, 11), and it is therefore quite naturally that in the 1990s, the first inhibitors of matrix metalloproteinase 9 were designated for the treatment of cancer.

The very first generation of these inhibitors were primarily soluble peptide compounds designed to mimic the amino acids of the endogenous MMP9 ligand.

Batimastat/BB-94 (developed by British Biotech) and Marimastat/BB-2516 (also developed by British Biotech) are examples belonging to this class of therapeutic active ingredients. Batimastat was the first to undergo clinical trials in 1994 but as it could not be administered orally it was discontinued to be replaced by Marimastat. Despite this second generation of compounds, British Biotech decided to stop the development of these anti-MMP9 in Phase 3, the results not being to the satisfaction of the company's shareholders.

Several chemical compounds were subsequently developed like CGS-27023A, GM6001, CGS-25966, but all failed in clinical trials. Their low solubility, their low oral bioavailability and their serious side effects have in fact not spoken in their favor. Let us add that these compounds have the unfortunate tendency to inhibit other zinc-dependent metalloenzymes, in particular ADAMs, due to their too poor specificity.

To address these issues, Tanomastat/ BAY12-9566 (developed by Bayer),  Prinomastat/AG-3380 and Rebimastat were developed but did not experience any more success than their predecessors during their clinical trials.

We can therefore conclude at this stage that most MMP-9 inhibitors were not selective enough (these chemical and peptide compounds can potentially simultaneously inhibit other MMPs with a strong structural resemblance to MMP-9), presented a low efficiency and off-target risks.

Consequently, none of these inhibitors has obtained FDA approval for the treatment of cancer and have all been stopped in Phase 3. But if the approach of MMP inhibition does not seem devoid of interest , it is therefore necessary to proceed to the generation of compounds with higher specificity for the target and presenting less toxicity.

REFERENCES ABOUT MMP9

1. Salo T, Mäkelä M, Kylmäniemi M, Autio-Harmainen H, Larjava H. Expression of matrix metalloproteinase-2 and -9 during early human wound healing. Lab Invest. 1994 Feb;70(2):176-82. PMID: 8139259.
2. Alexander CM, Hansell EJ, Behrendtsen O, Flannery ML, Kishnani NS, Hawkes SP, Werb Z. Expression and function of matrix metalloproteinases and their inhibitors at the maternal-embryonic boundary during mouse embryo implantation. Development. 1996 Jun;122(6):1723-36. doi: 10.1242/dev.122.6.1723. PMID: 8674412.
3. Bischof P, Martelli M, Campana A, Itoh Y, Ogata Y, Nagase H. Importance of matrix metalloproteinases in human trophoblast invasion. Early Pregnancy 1995;1:263e9.
4. Luchian, I.; Goriuc, A.; Sandu, D.; Covasa, M. The Role of Matrix Metalloproteinases (MMP-8, MMP-9, MMP-13) in Periodontal and Peri-Implant Pathological Processes. Int. J. Mol.Sci. 2022, 23, 1806
5. Klein G, Vellenga E, Fraaije MW, et al. The possible role of matrix metalloproteinase (MMP)-2 and MMP-9 in cancer, e.g. acute leukemia. Crit Rev Oncol Hematol 2004;50:87-100.
6. Baugh M, Perry M, Hollander A, Rhodridavies D, Cross S, Lobo A, Taylor C, Evans G. Matrix Metalloproteinase levels are elevated in inflammatory bowel disease. Gastroenterology 1999;117:814±822
7. Giannelli G, Erriquez R, Iannone F, Marinosci F, Lapadula G, Antonaci S. MMP-2, MMP-9, TIMP-1 and TIMP-2 levels in patients with rheumatoid arthritis and psoriatic arthritis. Clin Exp Rheumatol. 2004 May-Jun;22(3):335-8. PMID: 15144129.
8. Singh D., Srivastava S.K., Chaudhuri T.K., Upadhyay G. Multifaceted role of matrix metalloproteinases (MMPs) Front. Mol. Biosci. 2015;2:19. doi: 10.3389/fmolb.2015.00019.
9. Galis Z. S., Sukhova G. K., Lark M. W., Libby P. Increased expression of matrix metalloproteinases and matrix degrading activity in vulnerable regions of human atherosclerotic plaques. The Journal of Clinical Investigation. 1994;94(6):2493–2503. doi: 10.1172/JCI117619.v
10. N. A. Bhowmick, E. G. Neilson and H. L. Moses: Stromal fibroblasts in cancer initiation and progression. Nature, 432(7015), 332-337 (2004) DOI: 10.1038/nature03096
11. R. Kalluri and M. Zeisberg: Fibroblasts in cancer. Nat Rev Cancer.