Effect of andecaliximab (anti-MMP9) on proteolysis of IL-7 in vitro, TCR diversity in mice, and serum IL-7 in gastric cancer patients in combination with chemotherapy.

Abstract

101 Background: Andecaliximab (andeca) is a monoclonal antibody that selectively inhibits matrix metalloproteinase 9 (MMP9). IL-7 selectively enhances the proliferation and survival of naïve, memory, and effector T-cells (but not regulatory T-cells) in the periphery. Previous clinical trials with systemic recombinant IL-7 therapy increased TCR diversity. In the disease setting, elevated circulating IL-7 may be due to a compensatory increase in IL-7 production as demonstrated in mice upon inhibition of IL-7 signaling. Methods: An in vitro screen, in which recombinant active human MMP9 was incubated with > 140 human recombinant folded proteins, identified IL-7 as the most efficient substrate of MMP9. Results: IL-7 proteolysis occurred between A128:L129. IL-7 proteolysis altered the global protein structure, evidenced by a loss of cooperative unfolding observed with intact IL-7 (intrinsic fluorescence, Tm = 59.3o C). Mouse MMP9 proteolyzed mouse IL-7 in vitro. In the orthotopic syngeneic NeuT mouse model, MMP9 inhibition reduced tumor growth (p = 0.0005). In a 7-day NeuT study, anti-MMP9 alone improved TCR diversity (decreased clonality) within tumor-infiltrating T-cells (Dunnett’s p = 0.0083). Further, anti-MMP9 and anti-PDL1 co-treatment promoted an increase in CD3+ cells (p = 0.01), CD4+ T cells (p = 0.006), and CD8+ T cells (p = 0.013) concomitant with a decrease in tumor-associated CD25+ FoxP3+ regulatory T cells (p = 0.04) in the tumor. In gastric cancer patient serum, pro-MMP9 (p < 0.0001), active MMP9 (p < 0.0001), and IL-7 (p < 0.0001) were higher than healthy controls. Serum IL-7 levels were normalized upon treatment with andeca plus mFOLFOX6 (N = 40; FDR-corrected p < 0.001) in a Phase I gastric cancer study. Conclusions: MMP9 proteolyzed IL-7 in vitro. Specific MMP9 inhibition in a mouse tumor model improved TCR diversity. Andeca +mFOLFOX6 therapy normalized serum IL-7 levels, which could be due to andeca, chemotherapy, or disease resolution. The functional implications of IL-7 proteolysis by MMP9 in gastric cancer are currently under investigation.

Local and Systemic IL-7 Concentration in Gastrointestinal-Tract Cancers

“…Similarly, systemic levels of IL-7 in adenocarcinomas of the gastric cardia were at the levels observed in controls. While our observations relating to local IL-7 expression corroborate those of Greenstein [22], Quan et al [36] listed IL-7 among 105 molecules, out of 507 examined, differently expressed in gastric tumors as compared to normal tissue. However, in their study, tumor IL-7 expression constituted 9% of that found in normal tissue.…”

“…Accordingly, the antitumor activity of IL-7 has been shown in vitro and in vivo in animal models of prostate cancer, melanoma, and some neurological and blood cancers (reviewed in Reference [23]). However, some doubts have arisen about its positive role in cancer resulting from observations on upregulated expression [9,10,11,12,13,14] and/or secretion [15,16,17,18,19,20,21,22] in several cancers. Moreover, IL-7/IL-7R expression [10,12,28], as well as IL-7 secretion [15,19,20,21], have been correlated with adverse clinical outcomes in most, but not all [13,24], studies.…”

“…However, the highest difference in IL-7 concentration between tumor and patient-matched normal tissue was in gastric cancer. Only recently, Greenstein et al [22] published an abstract reporting elevated systemic IL-7 in gastric cancer. In our group, systemic IL-7 in GC was the lowest in the examined cancers and did not significantly differ as compared to the controls.…”

“…Moreover, in line with the beneficial role proposed for IL-7, it has been suggested that IL-7 overexpression might be a compensatory mechanism to overcome hampered IL-7/IL-7R signaling [22]. Indeed, while Greenstein et al [22] showed that IL-7 is a substrate for the MMP-9 protease, an enzyme known to be overexpressed in cancer [39], others demonstrated that T-cell responsiveness to IL-7 was obstructed by cell exposure to inflammatory cytokines [40]. Moreover, the upregulation of MMP-9 expression and activity is associated with tumor angiogenesis, invasion, and metastasis [39] when IL-7 is overexpressed as well.…”

“…However, for such a renowned cytokine, surprisingly little is known on IL-7 in solid tumors. In fact, contradicting a beneficial role attributed to the cytokine, several reports have demonstrated IL-7 overexpression [9,10,11,12,13,14] and oversecretion [15,16,17,18,19,20,21,22] in cancer, and have linked it with unfavorable characteristics [9,12]. In light of the existing controversies, a need for further research on IL-7 has recently been stressed [23].…”