As we all know, T cells are one of the main forces in the fight against tumors.
At present, the mainstream tumor immunotherapy includes immune checkpoint inhibitors, CAR-T cells, etc., based on this, we have a good understanding of the contribution of CD8+ T cells in the fight against cancer [1-3], but we do not fully understand the function of CD4+ T cells in addition to the auxiliary role.
Recently, a research team led by Dr. Lily Pao, Dr. Zhi-Yong Yang, and Dr. Gary J. Nabel from the R&D Department of Sanofi and Mode X Therapeutics published important research results in Nature[4].
They found that HER2/CD3× CD28 trispecific antibodies targeting HER2 and T cells can not only promote the killing of breast cancer cells by CD8+ T cells, but also directly inhibit the division of tumor cells in a CD4+ T cell-dependent manner, thereby achieving the effect of inhibiting tumor growth. Their findings suggest that CD4+T has a direct anti-tumor function in addition to its adjuvant effects.
Screenshot of the first page of the paper
Multispecific antibodies are emerging tumor immunotherapy drugs that are artificially designed to target different antigens in both arms of a Y-type antibody, allowing one antibody to bind to different antigens on a single cell or multiple cells. Multispecific antibodies exert antitumor effects by mechanisms such as blocking immunosuppressive receptors, or pulling immune cells closer to tumor cells and activating immune cells [5].
Trispecific antibodies constructed in this study
HER2 is a member of the family of human epidermal growth factor receptor tyrosine kinases and is highly expressed in tumor cells of some breast and gastric cancers, allowing us to treat patients with monoclonal antibodies that target HER2 [6,7], while in triple-negative breast cancer, her2 expression is significantly downregulated, creating resistance to monoclonal therapies that target HER2 [8]. Therefore, we need more effective treatments to overcome the resistance of HER2-low-expression tumors to targeted HER2 therapies.
Nabel's team wanted to use the power of multi-specific antibodies to direct T cells to HER2+ tumor cells and activate T cells to promote its killing of tumor cells. Considering that T cells both expressed cd3 and CD28 molecules, they constructed a trispecific antibody for HER2/CD3× CD28.
In vitro tests, it was found that the antibody can stimulate T cells to produce IL-2, activate the NF-κB pathway, and can mediate T cells to kill HER2+ tumor cells in an antigen-specific manner. At the same time, they noted that the antibody-mediated ability of T cells to kill tumor cells was positively correlated with the expression level of HER2 on tumor cells.
Trispecific antibodies activate T cells in vitro, inducing T cells to kill HER2-positive tumor cells
To study the therapeutic effect of this trispecific antibody in vivo, Nabel's team transfused human primary T cells to heavily immunodeficient mice with tumors, followed by trispecific antibody treatment. The results showed that both HER2 high-expression tumors and HER2 low-expression tumors were effectively inhibited after using antibody therapy. These data suggest that the antibody has the potential to treat HER2-low-expression tumors.
So which group of cells plays a major role in the treatment process?
Nabel's team transferred human CD3+ mixed T cells, CD4+ T cells, or CD8+ T cells to tumor-bearing mice, respectively, and treated them with trispecific antibodies. In the end, it was found that after receiving trispecific antibody treatment, the tumors of mice transfused with CD3+ mixed T cells and CD4+ T cells disappeared almost completely, while the tumors of mice with only CD8+ T cells were almost unrestrained. This may be due to the lack of Th cell adjuvant cd8+ T cells in this experimental system, and the survival rate is reduced.
Trispecific antibodies show therapeutic effects in humanized mouse models
At this point, Nabel's team noticed an interesting phenomenon, with only CD4+T present, the growth of tumors was completely inhibited!
To further explore the reasons behind this, Nabel's team cultured CD4+ T cells with HER2+ tumor cells for 24 h. The results showed that in the presence of trispecific antibodies, CD4+ T cells effectively inhibited the division of tumor cells, so that the cell cycle of tumor cells stagnated in the G1/S stage. CD8+ T cells, on the other hand, do not have the ability to inhibit the cell cycle.
These data suggest that CD4+ T cells can inhibit tumor growth by stalling the cell cycle of tumor cells. It also shows that trispecific antibodies can stimulate CD4+ T and CD8+ T cells to inhibit tumor growth through different but complementary mechanisms.
In the presence of trispecific antibodies, CD4+ T cells stall the cell cycle of tumor cells in the G1/S phase
Why do trispecific antibodies cause the cell cycle of tumor cells to stagnate?
Nabel's team sequenced RNA from tumor cells co-incubated with T cells for 24 hours. The results of the data analysis showed that in the presence of trispecific antibodies, gene expression related to cell cycle and cell division was significantly downregulated in tumor cells co-incubated with CD4+ T cells, while genes in pro-inflammatory responses— TNF-α and IFN-γ pathways— were significantly upregulated. This is consistent with the phenomenon that CD4+ T cells inhibit tumor cell division.
Since the use of antibodies to stimulate CD4+ T cells will promote the secretion of large amounts of IFN-γ and TNF-α, what kind of cytokines mediate the inhibition of tumor cell division? Through in vitro blocking experiments, Nabel's team found that when the antibody was used to neutralize TNF-α, the ability of CD4+ T cells to inhibit tumor cell growth was greatly weakened (but CD8+ T cells were not weakened). This suggests that CD4+ T cells inhibit the cell cycle of tumor cells by secreting TNF-α.
CD4+ T cells inhibit tumor cell growth by secreting TNF-α
Overall, the study not only constructed a trispecific antibody that induces T cells to attack HER2 low-expression tumor cells, but also found for the first time that CD4+ T cells can exert anti-tumor effects by directly inhibiting the cell cycle of tumor cells, an anti-tumor immune mechanism that was previously recognized.
At the same time, it is also suggested that follow-up studies should pay attention to the role of CD4+ T cells, which will help us develop drugs that improve the anti-tumor activity of CD4+ T and CD8+ T cells at the same time to achieve better therapeutic effects.
bibliography:
2. Porter DL, Levine BL, Kalos M, Bagg A, June CH. Chimeric antigen receptor-modified T cells in chronic lymphoid leukemia [published correction appears in N Engl J Med. 2016 Mar 10;374(10):998]. N Engl J Med. 2011;365(8):725-733. doi:10.1056/NEJMoa1103849
3. Sharma P, Allison JP. The future of immune checkpoint therapy. Science. 2015;348(6230):56-61. doi:10.1126/science.aaa8172
6. Berchuck A, Kamel A, Whitaker R, et al. Overexpression of HER-2/neu is associated with poor survival in advanced epithelial ovarian cancer. Cancer Res. 1990;50(13):4087-4091
7. Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science. 1987;235(4785):177-182. doi:10.1126/science.3798106
8. Pohlmann PR, Mayer IA, Mernaugh R. Resistance to Trastuzumab in Breast Cancer. Clin Cancer Res. 2009;15(24):7479-7491. doi:10.1158/1078-0432.CCR-09-0636
Responsible editor 丨IoTalker