Coordinate immune attacks against cancer

Tumor associated antibodies in cancer patients have been found to provide therapeutic benefits, partly by engaging innate immune responses for the clearance of tumor cells. Specifically, A post-mortem analysis study from Harvard University revealed that cancer patients with high isotype-switched antibody responses within tumor tissues are highly correlated with survival benefits in patients with breast cancer or melanoma. The observed survival benefit was attributed to the elevated activities of natural killer cells, a type of innate immune cells relying on antibodies for target recognition to eliminate cancer cells. Other previous research also showed monoclonal antibodies against tumor antigens can dramatically improve the efficacy of cancer immunotherapies. A preclinical study from Dr. Irvine and Dr. Wittrup at MIT showed that the combination treatment strategy, including tumor vaccines, checkpoint blockade therapy, and tumor antigen-specific monoclonal antibodies, is highly effective in several murine tumor models, demonstrating tumor-specific antibodies amplifying effect on the treatment efficacies.

However, vaccine-induced tumor antigen-specific antibodies, as a standalone therapy, have largely been ineffective in tumor treatments, suffering from immune suppression within tumor microenvironment or antigenic escape through tumor mutations. Considering vaccine therapy is potentially a safer and cheaper way to induce host antibody responses against cancer, as compared to monoclonal antibodies, we sought to develop a strategy to stimulate host responses against multiple tumor antigens, to generate both antibodies and T cells for cancer treatments.

In Dr. Collier lab at Duke, I have previously developed a self-assembled peptide nanofiber vaccine platform, namely Coil29, that can generate both T cell and antibodies responses against different peptide antigens. More importantly, Coil29 nanofiber vaccine is superior in inducing antigen-specific antibody responses, relative to several commercial adjuvants. In a follow-up study, we demonstrated the potential reason behind this outstanding ability is that Coil29 peptide has inherent T-cell epitopes that enhance the generation of Tfh cells, a specific cell type critical for antibody generation. Therefore, we think Coil29 platform is a great candidate for this cancer vaccine study. We hypothesize that the combination between antibody and T cell responses elicited by vaccine will synergize and improve the therapeutic effect, as compared to just antibodies or T cells alone.

Because of the self-assembling nature, Coil29 peptide appended with antigens can spontaneously form nanofibers carrying multiple different peptide antigens. As expected, this vaccine can stimulate high antibody and T cell responses, with antibodies targeting tumor-specific receptor, EGFRvIII (Epidermal Growth Factor Receptor variant III), and T cells against murine melanoma-associated protein, Trp2 (Tyrosinase-Related Protein 2). The responses elicited by vaccines are stronger than peptide antigens delivered with adjuvant CFA (Complete Freund’s Adjuvant).

This antibody and T cell responses induced by nanofibers provided protection for mice against tumor challenges with EGFRvIII-expressing murine melanoma cells (B16vIII), outperforming both antibody responses alone and T cell responses alone. In a therapeutic model, where mice were inoculated with B16vIII cells one day prior to receiving vaccine treatments, neither antibody alone nor T cell alone can inhibit tumor growth, but mice that were treated with vaccines generating both arms of immunity showed slower tumor growth. The inhibition effect is statistically significant, but the effect size is admittedly small, leaving a lot of room for improvements.

Among most murine cancer models, B16 murine melanoma is relatively harder to treat, due to its rapid growth and immune suppressive microenvironment. To overcome this challenge, I initially had considered to add checkpoint blockade antibodies, such as anti-PD-L1/PD-1 and anti-CTLA-4, to the treatment regimen, but ultimately decided that this plan lacks the specificity for a strategy centered around improving vaccine-induced antibody performance in cancer treatment, because both anti-PD-L1/PD-1 and anti-CTLA-4 are designed to help T cells overcome suppressive tumor environment, but provides little benefit to antibody therapy.

While searching for ways to further improve vaccine performance, I took a break and attended a cancer nanotechnology GRC conference. During this conference, Dr. Dennis E. Discher from UPenn gave a wonderful talk about mentorship and scientific discovery. In it, he mentioned his own research about CD47, a cell surface receptor “mark for self”, which overexpressed on cancer cells to evade immunosurveillance, and studies showed that blocking CD47 can improve phagocytosis of cancer cells by macrophages. This talk sparked an idea in my mind that vaccine-induced antibody responses can be improved with anti-CD47 mAbs (monoclonal antibodies), because antibodies inhibit tumor cells partly by engaging macrophages for phagocytosis. I still remember the excitement and anxiety when I was frantically searching for literatures in the back row of the conference room, to see if there have already been studies carrying out this idea. What I found was that, at that moment, anti-CD47 treatment had shown clinical success in a clinical trial, but has not gained much attention as a suitable strategy to combine with cancer vaccines, possibly because most cancer vaccines solely focusing on T cell responses, but not antibody responses. Meanwhile, because Coil29 nanofiber vaccine induces anti-tumor antibodies, combination with anti-CD47 mAbs should improve the treatment effect.

To determine whether anti-CD47 mAbs can improve the therapeutic effect of the nanofiber vaccine, we carried out several cell culture assays to evaluate cytotoxic functions of NK cells and macrophages agaisnt tumors, in the presences of anti-CD47 mAbs and sera from immunized mice. After a lot of optimizations for experimental conditions, we found that anti-CD47 treatments significantly improved the functions of NK cells and macrophages, and the cytotoxic effects rely on both tumor-specific antibodies and anti-CD47 mAbs. Encouraged by these findings, we examined the therapeutic effect in mice when we combine anti-CD47 and anti-PD-L1 with the nanofiber vaccine. We found that mice experiment results corroborated with our cell assays, in that combining these checkpoint blockade antibodies with vaccines greatly inhibited tumor growth and improved the mice survival in this melanoma model.

After a long while, at the time of writing, we will finally see this study published in July 2022. During the review process, all four reviewers provided many thoughtful comments and questions, and helped us improve the manuscript in many ways. Some more critical comments are focused on our treatment schedules, and the relatively “not impressive” therapeutic effect by nanofiber vaccines. These are all very welcomed critiques. I am also very aware of these shortcomings of this study myself before submitting the manuscript. Part of our future effort here at Syracuse will be exploring strategies to overcome some of these flaws. While this study is merely a small step toward future clinical treatment, we hope that this study can highlight the powerful potential that vaccine-induced antibodies hold for cancer treatment, and demonstrate that the combination between anti-CD47 and antibody-inducing vaccines can potentially benefit other types of cancer treatment.

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