Claudin 6 vaccines effectively inhibit tumor growth in a syngeneic mouse colon cancer model

Human Claudin 6 (CLDN6) is a member of the Claudin family of proteins, which plays a crucial role in the structure and function of tight junctions. In humans, CLDN6 is primarily expressed in stem cells and embryonic tissues, with minimal mRNA detected in testis, pancreas, and placenta. Elevated expression of CLDN6 has been found in several types of cancers, suggesting that CLDN6 is an ideal tumor-associated antigen for targeted immunotherapy, albeit its precise role in cancer remains to be defined. Here, we generated recombinant proteins consisting of multiple human CLDN6 B- and T-cell epitopes (CLDN6-B+T) or T-cell epitopes only (CLDN6-T), which were selected based on computationally predicted B/T-cell epitope sequences and fused with the transmembrane domain of diphtheria toxin (DTT) as an immune enhancer. The immunogenicity and anti-tumor activity of these two vaccines were evaluated in mouse models. In both wildtype and HLA*02:01-humanized mice, CLDN6-B+T and CLDN6-T with adjuvant CpG induced robust cellular immune response as assessed by IFNgamma ELISpot assay using splenocytes collected from mice vaccinated weekly for 6 weeks. In a syngeneic mouse tumor model using CLDN6-humanized MC38 colon cancer cell line (MC38-CLDN6(K)), both vaccines significantly suppressed the growth of tumor cells inoculated one day after the 6th vaccine dose, compared to control mice that received adjuvant only (n=8 in each group). In this study, 7/8 mice vaccinated with CLDN6-B+T and 8/8 mice vaccinated with CLDN6-T were tumor-free at the endpoint. A replicate study using an independently-derived CLDN6-humanized MC38 cell line, MC38-hCLDN6(C), demonstrated 72.6% and 73.2% tumor growth inhibition by CLDN6-B+T and CLDN6-T vaccines, respectively, compared to adjuvant controls(n=10 in each group). In a therapeutic study in which MC38-hCLDN6(C) cells were inoculated subcutaneously one day before weekly vaccination for 5 times, tumor growth was inhibited by 67.0% and 81.0% in mice vaccinated with CLDN6-T (n=11) and CLDN6-B+T (n=12), respectively, compared to mice that received adjuvant only (n=10). Collectively, our findings demonstrate that CLDN6-targeted vaccines significantly suppressed the growth of CLDN6-positive tumors in mice. These results support CLDN6-targeted vaccines as a promising therapeutic strategy for CLDN6-positive cancers in humans, with their potential to be further validated in upcoming studies.