Can CD40 agonists open up a new pattern of immunotherapy

1.?Mitazalimab achieves positive clinical results in first-line pancreatic ductal carcinoma

On January 2, 2023, Alligator Bioscience released the interim results of Mitazalimab clinical phase II, OPTIMIZE-1. This open-label, multicenter clinical trial is evaluating the safety and efficacy of mitazalimab in combination with chemotherapeutics and mFOLFIRINOX in patients with untreated metastatic ductal pancreatic carcinoma.

The interim results were positive, with an ORR of 52.2% (n = 23), a DCR (disease control rate) of over 90%, and a significant increase in the ORR of mFOLFIRINOX monotherapy (31.6%). At the same time, the safety data proved that mitazalimab combined with mFOLFIRINOX was safe and tolerable at 900 ug/kg. Alligator Bioscience will further discuss with regulatory authorities how to accelerate the drug development and approval process.

Design of the OPTIMIZE-1 clinical trial

In terms of antibody design, Mitazalimab retains the traditional IgG1, has comparable FcγRI and FcγRIIb affinity compared with APX005M, and has no FcγR binding ability, further reducing potential ADCC and toxicity caused by CDC effects. Mitazalimab relies on FcγR cross-linking to exert the agonistic ability of antibodies and kills tumor cells through APC, B cells, macrophages, DCs, and other mechanisms.

2.?Sotigalimab achieves positive clinical results in first-line melanoma and liposarcoma

At the SITC Annual Conference in November 2022, Apexigen announced that its CD40 antibody agonist sotigalimab had achieved positive efficacy in two clinical trials.

In clinical phase I and II, sotigalimab combined with pembrolizumab achieved an ORR of 47% (n = 32) in the first-line metastatic melanoma patients, including the dose escalation group. According to RP2D, the ORR reached 50% (n = 24), which was significantly better than the 34% ORR of standard pembrolizumab monotherapy.

In the simultaneous phase II liposarcoma clinical trial, Sotigalimab combined with Doxorubicin (n = 10), mPFS reached 12.5 m (1.4–25.3 m), significantly improving the median survival time of historical Doxorubicin alone (less than 5 m).

3.?CD40 monoclonal antibody and bi-specific antibody pipelines are under development

As of 2022, there are more than 60 antibodies targeting CD40 in research and clinical stages, among which Apexigen's sotigalimab has the fastest clinical progress. In addition to CD40 monoclonal antibody agonists, it also includes multiple TSA/CD40 bi-specific antibodies, such as MSLN, FAP, 5T4, EpCAM, GPC3, CEA, and other targets.

Table: List of commercial-grade antibodies in advanced stages of clinical trials

Table: CD40 bispecific antibody under development

4.?Sanyou Bio provides solutions for CD40 efficacy and safety in vivo?

Dozens of off-the-shelf CD40 strong agonist antibodies and CD40 weak agonist antibodies are available.

Case 1: CD40 monoclonal antibody anti-tumor efficacy evaluation?

Experimental aim: To evaluate the inhibitory effect of selicrelumab antibody on MC38 (mouse colon cancer) tumors in hCD40 mice.

Experimental method: 2E+6?MC38 cells were subcutaneously injected into the right back of hCD40 mice (6–8 weeks old, female), and the tumor volume grew to ~100 mm^3?for random grouping (N = 5), tail vein injection twice a week. With different doses (1 mpk and 10 mpk) of selicrelumab, the changes in tumor volume, body weight, and physical signs of mice were recorded and analyzed at different time points.

Experimental conclusion: selicrelumab antibody showed significant anti-tumor effect, and it was dose-dependent. On Day 27, the tumor inhibition rate (TGI) was 99.98% at a dose of 20 mpk, and the tumors were completely regressed; there was no significant change in the body weight of the mice during the administration period.

Case 2: CD40 monoclonal antibody anti-tumor efficacy evaluation?

Experimental aim: To evaluate the inhibitory effect of APX005 molecule on Ramos (human B lymphocytoma cells) tumor in Balb/c nude mice.

Experimental method: 5E+6?Ramos cells were subcutaneously injected into the right back of Balb/c nude mice (6–8 weeks old, female), and the tumor volume grew to ~300 mm3?for random grouping (N = 12), three times a week. Different doses (0.2 mpk, 1 mpk, and 5 mpk) of APX005 molecules were injected intraperitoneally, and the changes in tumor volume, body weight, and physical signs of mice were recorded and analyzed at different time points.

Experimental conclusion: APX005 molecule shows significant anti-tumor effect, and it is dose-dependent. On D27, the tumor inhibition rate (TGI) at the dose of APX005-5mpk was 92.6%; there was no significant change in the body weight of the mice during the administration.

Case 3: CD40/TSA bi-specific antibody anti-tumor efficacy evaluation

Experimental aim: To evaluate the inhibitory effect of CD40/TSA bispecific antibody on TSA-MC38 (mouse colon cancer) tumor in hCD40 mice.

Experimental method: 2E+6?TSA-MC38 cells were subcutaneously injected into the right back of hCD40 mice (6–8 weeks old, female), and the tumor volume grew to ~100 mm^3 for random grouping (N = 5), twice a week. Different doses (1 mpk and 10 mpk) of CD40/TSA double antibody were injected intravenously, and the changes in tumor volume, body weight, and physical signs of mice were recorded and analyzed at different time points.

Experimental conclusion: CD40/TSA bi-specific antibody molecule showed significant anti-tumor effect, and it was dose-dependent. On D25, the tumor inhibition rate (TGI) at the dose of CD40/TSA-20 mpk was 99.98%, and the tumors were completely regressed; there was no significant change in the body weight of the mice during the administration.

Case 4: CD40 monoclonal antibody spleen toxicity evaluation?

Experimental aim: to evaluate the spleen toxicity of selicrelumab in hCD40 mice in the MC38 (mouse colon cancer) tumor model.

Experimental method: 2E+6?MC38 cells were subcutaneously injected into the right back of hCD40 mice (6–8 weeks old, female), and the tumor volume grew to ~100 mm^3?for random grouping (N = 5), tail vein injection twice a week with Selicrelumab at different doses (1 mpk, 10 mpk). After the administration period, the mice were euthanized with CO2 and then dissected; the weight of the organs was measured, and the organ-to-body ratio was calculated.

Experimental results: Compared with the PBS control group, the visceral ratio of selicrelumab at a dose of 20 mpk was significantly increased, and there was no significant difference at a dose of 1 mpk, indicating that the toxicity of selicrelumab to the spleen was significantly dose-dependent.

Case 5: CD40/TSA bi-specific antibody spleen toxicity evaluation

Experimental aim: to evaluate the spleen toxicity of CD40/TSA bi-specific antibodies on hCD40 mice in the MC38 (mouse colon cancer) tumor model.

Experimental method: 2E+6?TSA-MC38 cells were subcutaneously injected into the right back of hCD40 mice (6–8 weeks old, female), and the tumor volume grew to ~100 mm^3?for random grouping (N = 5), twice a week. Different doses (1 mpk and 10 mpk) of CD40/TSA double antibodies were injected intravenously. After the administration period, the mice were euthanized with CO2 and then dissected, and the weight of the organs was measured and the organ-to- body ratio was calculated.

?Experimental results: Compared with the PBS control group, the CD40/TSA bi-specific antibody had no significant spleen toxicity at the doses of 20 mpk and 1 mpk.

Case 6: Selicrelumab Hepatotoxicity Evaluation?

Experimental Aim: To evaluate the hepatotoxicity of selicrelumab in hCD40 mice in the MC38 (mouse colon cancer) tumor model.

Experimental method: 2E+6?MC38 cells were subcutaneously injected into the right back of hCD40 mice (6–8 weeks old, female), and the tumor volume grew to ~100 mm^3?for random grouping (N = 5), tail vein injection twice a week with Selicrelumab at different doses (1 mpk, 10 mpk). Before the fifth administration, blood was collected from the orbital vein, and the ALT and AST test kit (Nanjing Jiancheng) was used to detect ALT and AST and conduct comparative analysis.

ALT and AST levels in the serum of selicrelumab-treated mice, a CD40 monoclonal antibody

Experimental results: Compared with the PBS control group, the serum ALT content of selicrelumab-treated group increased significantly at a dose of 20 mpk, and the serum ALT level tended to increase at a dose of 1 mpk, but there was no statistical difference. The AST indicators all had a rising trend in a dose-dependent manner, but there was no statistical difference.

Case 7: CD40/TSA bi-specific antibody hepatotoxicity evaluation

Experimental aim: to evaluate the hepatotoxicity of CD40/TSA bi-specific antibodies in hCD40 mice of MC38 (mouse colon cancer) tumor model.

Experimental method: 2E+6?MC38 cells were subcutaneously injected into the right back of hCD40 mice (6–8 weeks old, female), and the tumor volume grew to ~100 mm^3?for random grouping (N = 5), tail vein injection twice a week with different doses (1 mpk, 10 mpk) of CD40/TSA bi-specific antibodies. Before the fifth administration, blood was collected from the orbital vein, and the ALT and AST test kit (Nanjing Jiancheng) was used to detect ALT and AST and conduct comparative analysis.

?Levels of ALT and AST in the serum of mice treated with the CD40/TSA bispecific antibody

Experimental results: Compared with the PBS control group, CD40/TSA had no significant difference in serum ALT and AST at doses of 20 mpk and 1 mpk, indicating to some extent that it has no significant hepatotoxicity.