Immunological Profiles in Lynch Syndrome Colorectal Cancers Are Not Specific to Mismatch Repair Gene Defects

Background and aims : Colorectal carcinomas (CRCs) in patients with Lynch syndrome (LS) exhibit heightened immunogenicity due to mismatch repair deficiency (MMR-d), often resulting in favorable responses to T cell immune checkpoint therapies. Recent studies indicate that the phenotype and genotype of LS-associated CRCs vary depending on the specific MMR gene mutated. Here, we investigated whether the immune profiles of LS-associated CRCs differ based on the MMR gene defects. Methods : Tissue material from 18 MLH1 -, 16 MSH2 -, 40 MSH6 -, and 23 PMS2 -mutated CRCs and 35 sporadic MMR-d CRCs were included in the study. Imaging mass cytometry (IMC) analysis, along with targeted multiplex immunofluorescence imaging (mIF) and immunohistochemistry, were applied to examine the tumor immune microenvironment, including Human Leukocyte Antigen (HLA) class I and programmed death-ligand 1 (PD-L1) expression. Results : Unsupervised hierarchical clustering of cell phenotypes identified by IMC, followed by mIF validation, revealed comparable lymphoid and myeloid cell infiltration levels across CRCs from all MMR groups. Infiltrating T cell levels negatively correlated with the number of mutations at coding microsatellite sequences, particularly in MLH1 -mutated CRCs. HLA class I defects were observed in 76% of all CRCs. These defects were more frequently accompanied by β2M defects in hereditary MMR-d CRCs (67%) compared to sporadic MMR-d CRCs (37%), and did not associate with the number of γδ T cells, which were present in CRCs from all MMR groups. PD-L1 expression in tumor cells was only detected in 8% of all CRCs. Conclusion : Our findings illustrate that, from an immunological perspective, there is no evidence of differing immunogenic features across MMR defects. This is important to consider when developing preventive vaccine strategies and evaluating immunotherapy for LS patients and those with MMR-d CRCs. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was funded by the MLDS (Dutch Digestive Disease foundation, grant number FP16-06). The funder had no role in the study design, data acquisition and analysis, decision to publish, or manuscript preparation. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The Medical Ethical Committee of Leiden, The Hague, Delft (protocol P17.098) approved this study. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The datasets analyzed during this study are available from the corresponding author upon reasonable request.