Abstract PR13: Inactivation of DNA repair triggers dynamic neoantigen evolution and impairs cancer growth

Colorectal, ovarian, endometrial and other tumors carrying defects in DNA mismatch repair often show favorable prognosis and indolent progression. The genomes of these tumors -also known as microsatellite unstable (MSI) cancers- bear hundreds of thousands of somatic mutations, a feature which fosters cancer progression and rapid evolution of resistance to targeted therapies. The molecular bases for the favorable outcome of MMR deficient cancers have long remained a mystery. Recent evidences that a subset of MSI tumors respond prominently to immune checkpoint blockade led to the seminal hypothesis that the presence of high number of somatic mutations may be responsible for effective immune-surveillance. However, several reports indicate that a relevant fraction of hyper-mutated tumors have unfavorable prognosis and do not respond to immune-modulators. Intrigued by these contradictory findings, we used the CRISPR system to genetically inactivated MutL homolog 1 (MLH1) in colorectal, breast and pancreatic mouse cancer cells. The growth of MMR deficient cells was comparable to their proficient counterparts in vitro and upon transplantation in immune-compromised mice. Strikingly however, isogenic MMR deficient colorectal, breast and pancreatic cancer cells were largely unable to form tumors when injected subcutaneously or orthotopically in syngeneic mouse models. MMR deficient tumors initially established in immune-deficient mice continued to grow exponentially when transplanted in syngeneic animals but regressed promptly when immune checkpoint inhibitors (anti PD-1 and anti CTLA-4) were administered. Exome sequencing of MMR proficient cells revealed mutational loads and neo-antigen profiles that were stable over time. MMR inactivation further increased the mutation burden and led to highly dynamic mutational profiles, resulting in persistent renewal of neoantigens. These results led us hypothesize that enforced increase of the number of mutations in cancer cells could be paradoxically- beneficial for therapeutic purposes. We therefore performed a pharmacological screen to identify agents capable of permanent inactivation of MMR in colorectal, breast and PDAC cancer cells. We found that temozolomide triggers MLH1 inactivation and leads to rapid clonal evolution and dynamic neoantigen profiles. Temozolomide-treated cells were unable to form tumors in syngeneic animals, while cells treated with other alkylating agents did. Genomic analysis of these tumor models revealed that fluctuating levels of neoantigens, rather than the absolute number of mutations is critical to provoke immune surveillance. These results provide the rationale for developing innovative anticancer therapies that target DNA repair proteins. This abstract is also being presented as Poster B11. Citation Format: Giovanni Germano, Simona Lamba, Giuseppe Rospo, Alessandro Magri, Federica Maione, Mariangela Russo, Nabil Amirouchene Angelozzi, Ludovuc Barault, Monica Montone, Federica Di Nicolantonio, Enrico Giraudo, Alberto Bardelli. Inactivation of DNA repair triggers dynamic neoantigen evolution and impairs cancer growth. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr PR13.