Ax-101, a small-molecule immunomodulator with a yet-to-be elucidated MoA, enhances lymphocyte proliferation, induces a Th1 profile, increases T-cell cytotoxicity, and improves the M1/M2 ratio. Specific Ax-101 effects include upregulated cell-surface TNFalpha (sTNF) on human T cells and monocytes, increased (>7.5x) T-cell cytotoxicity, upregulation by human PBMC of antitumor cytokines (e.g., IL-2, IFN-gamma, TNFalpha) and downregulation of cytokines that mediate immune evasion (e.g., IL-10), and decreased immune-suppressive myeloid cells. Syngeneic mouse cancer models including bladder (MBT-2), colorectal (CT-26), melanoma (Cloudman), and lymphoma (A20) show the addition of Ax-101 to anti-PD1 to improve outcomes. For example, neither Ax-101 nor anti-PD1 monotherapy had significant effects on mean tumor volume (MTV) in MBT-2 mice, while an Ax-101/anti-PD1 (same doses/regimen) combo resulted in up to 44% reduction in MTV c/w anti-PD1 alone (p < 0.05). Further, 30% (3/10) of MBT-2 mice achieved CR with the combo, whereas no CRs were seen with anti-PD1 alone. In CT-26 mice, 25% (2/8) achieved CR, whereas no CRs occurred with anti-PD1 only; subset analysis of mice exhibiting at least some antitumor response to anti-PD1 alone (6/8) or the Ax-101/anti-PD1 combo (5/8) revealed a 40% CR rate and MTV reduction of up to 75% in combo-treated animals. In Cloudman melanomas, all (5/5) mice receiving Ax-101/anti-PD1 throughout their treatment achieved CR. Importantly, in mice with rapidly growing melanomas refractory to anti-PD1 for 12 d, Ax-101 addition reversed growth, with 4/5 mice achieving CR. Of note, 9 Cloudman mice achieving CR were re-injected with 3x the melanoma cells they initially received and followed with no therapy. Only 1/9 mice developed tumor over 73 d; thus, Ax-101/anti-PD1 may elicit a vaccine-like effect. Anti-PD1 alone and the Ax-101/antiPD1 combo in A20 mice resulted in 23% and 40% lower MTVs c/w control. Ax-101 was previously in Ph 1/2 monotherapy studies in follicular lymphoma (FL) and myeloma (M) ( IDs: NCT00006466, NCT00007839). Thirty-two patients received 2 µg SQ Ax-101 q2 wkly or wkly for up to 18 months. No Grade 3 or higher toxicities were observed. Five of 14 FL patients showed tumor reduction from 16-64%, while 1/17 M patients showed 50% reduced M-protein; maximum responses occurred between 43 days and over 1 year. Increased PBMC sTNF was seen in all patients tested (8 with FL, 6 with M) (p = 0.0006). Consistent with Ax-101 activity depending on a functional immune system, delayed type hypersensitivity (DTH) testing showed all responders to be DTH+ (5 tumor reduction, 4 SD); 5 anergic (DTH-) patients had SD at best (2 SD, 3 worse). Current Ax-101 clinical development will leverage its beneficial immunostimulatory properties to augment the efficacy of anti-PD1 inhibitors such as pembrolizumab, nivolumab and cemiplimab.

Citation Format: Stephan W. Morris, Betsy Fisher, Kent Murphy. Ax-101, an immunostimulatory small molecule, markedly enhances the antitumor activity of PD1 immune checkpoint blockade in multiple preclinical solid tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1720.