Daniel D. Buchanan

Background: Prior studies have demonstrated that the overall density of T cells in colorectal tumors is favorably associated with colorectal cancer (CRC) survival; however, few studies have considered the potentially distinct roles of heterogeneous T cell subsets in different tissue regions in relation to CRC outcomes. Methods: Including 1,113 CRC tumors from three observational studies, we conducted in-situ T cell profiling using a customized 9-plex [CD3, CD4, CD8, CD45RA, CD45RO, FOXP3, KRT (keratin), MKI67 (Ki-67), and DAPI] multispectral immunofluorescence assay. Multivariable-adjusted Cox proportional hazards models were used to estimate hazard ratios (HRs) and 95% confidence intervals (CIs) for the associations of T cell subset densities in both epithelial and stromal tissue areas in CRC with disease-specific survival. Results: Higher CD3+CD4+ and CD3+CD8+ naive, memory, and regulatory T cell densities were significantly associated with better CRC-specific survival in both epithelial and stromal tissue areas (HRs highest quantile versus lowest quantile ranging 0.41-0.68). These associations persisted in models further adjusted for stage at diagnosis and were largely consistent when stratified by microsatellite instability (MSI) status. However, the further stratification into CD4+ or CD8+ T cell subsets beyond CD3+ subsets did not significantly improve how well our model explains CRC prognosis. Conclusions: The density of T cells in CRC tissue, both overall and for several T cell subset populations, is significantly associated with CRC-specific survival independent of MSI status and stage at diagnosis. Conclusions: Higher levels of T cell densities in different locations with different functions are associated with better CRC-specific survival.

Recent research has identified a potential protective effect of higher numbers of circulating lymphocytes on colorectal cancer (CRC) development. However, the importance of different lymphocyte subtypes and activation states in CRC development and the biological pathways driving this relationship remain poorly understood and warrant further investigation. Specifically, CD4+ T cells - a highly dynamic lymphocyte subtype - undergo remodelling upon activation to induce the expression of genes critical for their effector function. Previous studies investigating their role in CRC risk have used bulk tissue, limiting our current understanding of the role of these cells to static, non-dynamic relationships only. Here, we combined two genetic epidemiological methods - Mendelian randomisation (MR) and genetic colocalisation - to evaluate evidence for causal relationships of gene expression on CRC risk across multiple CD4+ T cell subtypes and activation stage. Genetic proxies were obtained from single-cell transcriptomic data, allowing us to investigate the causal effect of expression of 1,805 genes across five CD4+ T cell activation states on CRC risk (78,473 cases; 107,143 controls). We repeated analyses stratified by CRC anatomical subsites and sex, and performed a sensitivity analysis to evaluate whether the observed effect estimates were likely to be CD4+ T cell-specific. We identified six genes with evidence (FDR-P<0.05 in MR analyses and H4>0.8 in genetic colocalisation analyses) for a causal role of CD4+ T cell expression in CRC development - FADS2, FHL3, HLA-DRB1, HLA-DRB5, RPL28, and TMEM258. We observed differences in causal estimates of gene expression on CRC risk across different CD4+ T cell subtypes and activation timepoints, as well as CRC anatomical subsites and sex. However, our sensitivity analysis revealed that the genetic proxies used to instrument gene expression in CD4+ T cells also act as eQTLs in other tissues, highlighting the challenges of using genetic proxies to instrument tissue-specific expression changes. Our study demonstrates the importance of capturing the dynamic nature of CD4+ T cells in understanding disease risk, and prioritises genes for further investigation in cancer prevention research.

Early-onset bowel cancer incidence (age <50 years) has increased worldwide and is highest in Australia, but how this varies across histology and anatomical site remains unclear. We aimed to investigate appendiceal, proximal colon, distal colon, rectal, and anal cancer incidence trends by age and histology in Australia. Cancer incidence rate data were obtained from all Australian cancer registries (1990-2020 period). Birth cohort-specific incidence rate ratios (IRRs) and annual percentage change in rates were estimated using age-period-cohort modelling and joinpoint regression. After excluding neuroendocrine neoplasms, early-onset cancer incidence rose 5-9% annually, yielding 5,341 excess cases (2 per 100,000 person-years; 12% appendix, 45% colon, 36% rectum, 7% anus; 20-214% relative increase). Trends varied by site, period, and age: appendiceal cancer rose from 1990-2020 in 30-49-year-olds; colorectal cancers rose from around 1990-2010 in 20-29-year-olds and from 2010-2020 in 30-39-year-olds; anal cancer rose from 1990-2009 in 40-49-year-olds. Across all sites, IRRs increased with successive birth cohorts since 1960. Notably, adenocarcinoma incidence in the 1990s versus 1950s birth cohort was 2-3-fold for colorectum and 7-fold for appendix. The greatest subtype-specific increases occurred for appendiceal mucinous adenocarcinoma, colorectal non-mucinous adenocarcinoma, and anal squamous cell carcinoma. Only later-onset (age ≥50) colorectal and anal adenocarcinoma rates declined. Appendiceal tumours, neuroendocrine neoplasms (all sites), anorectal squamous cell carcinomas, and colon signet ring cell carcinomas rose across early-onset and later-onset strata. Appendiceal, colorectal, and anal cancer incidence is rising in Australia with variation across age and histology, underscoring the need to identify factors driving these trends. ALM is supported by an Australian Government Research Training Program Scholarship, Rowden White Scholarship, and WP Greene Scholarship. DDB is supported by a National Health and Medical Research Council of Australia (NHMRC) Investigator grant (GNT1194896), a University of Melbourne Dame Kate Campbell Fellowship, and by funding awarded to The Colon Cancer Family Registry (CCFR, www.coloncfr.org) from the National Cancer Institute (NCI), National Institutes of Health (NIH) [award U01 CA167551]. MAJ is supported by an NHMRC Investigator grant (GNT1195099), a University of Melbourne Dame Kate Campbell Fellowship, and by funding awarded to the CCFR from NCI, NIH [award U01 CA167551].

Adenomatous polyposis syndromes are hereditary conditions characterised by the development of multiple adenomas in the gastrointestinal tract, particularly in the colon and rectum, significantly increasing the risk of colorectal cancer and, in some cases, extra-colonic malignancies. These syndromes are caused by germline pathogenic variants (PVs) in genes involved in Wnt signalling and DNA repair. The main autosomal dominant adenomatous polyposis syndromes include familial adenomatous polyposis (FAP) and polymerase proofreading-associated polyposis (PPAP), caused by germline PVs in APC and the POLE and POLD1 genes, respectively. Autosomal recessive syndromes include those caused by biallelic PVs in the DNA mismatch repair genes MLH1, MSH2, MSH6, PMS2, MSH3 and probably MLH3, and in the base excision repair genes MUTYH, NTHL1 and MBD4. This review provides an in-depth discussion of the genetic and molecular mechanisms underlying hereditary adenomatous polyposis syndromes, their clinical presentations, tumour mutational signatures, and emerging approaches for the treatment of the associated cancers. Considerations for genetic testing are described, including post-zygotic mosaicism, non-coding PVs, the interpretation of variants of unknown significance and cancer risks associated with monoallelic variants in the recessive genes. Despite advances in genetic testing and the recent identification of new adenomatous polyposis genes, many cases of multiple adenomas remain genetically unexplained. Non-genetic factors, including environmental risk factors, prior oncologic treatments, and bacterial genotoxins colonising the intestine - particularly colibactin-producing Escherichia coli - have emerged as alternative pathogenic mechanisms.

Approximately 30% of sebaceous skin lesions (or sebaceous neoplasia) demonstrate DNA mismatch repair (MMR)-deficiency. MMR-deficiency can be caused by Lynch syndrome, resulting from germline pathogenic variants in the DNA MMR genes MLH1, MSH2, MSH6 and PMS2, but other causes include somatic MLH1 gene promoter hypermethylation, constitutional MLH1 gene promoter hypermethylation (MLH1 epimutation), or biallelic somatic MMR gene mutations. In colorectal (CRCs) and endometrial cancers (ECs), tumour MMR-deficiency showing loss of MLH1 and PMS2 protein expression (MLH1/PMS2-deficiency) is predominantly caused by somatic MLH1 hypermethylation, however, it is not clear if somatic MLH1 hypermethylation is a cause of MLH1/PMS2-deficiency in sebaceous neoplasia. This study investigated the causes of MLH1/PMS2-deficiency in 28 cases with sebaceous neoplasia. Germline pathogenic variants in MLH1 were identified in 11 of 28 cases. Of the remaining 17 non-Lynch syndrome cases, two (11.8%) were positive for MLH1 hypermethylation in blood-derived DNA (constitutional MLH1 epimutations). The corresponding sebaceous tissue of these two cases also showed MLH1 hypermethylation. None of the other eight cases with sufficient sebaceous tissue-derived DNA for testing showed somatic MLH1 hypermethylation. Multi-gene panel testing of sebaceous tissue and matched blood-derived DNA identified four cases with biallelic somatic MLH1 mutations as the cause of MLH1/PMS2-deficiency. No cause of MLH1/PMS2-deficiency could be identified in one case. This study demonstrates that biallelic somatic MLH1 mutations and constitutional MLH1 epimutations underlie MLH1/PMS2-deficiency in sebaceous neoplasms after excluding Lynch syndrome. Unlike CRCs and ECs, somatic MLH1 hypermethylation was not identified suggesting it is not a common cause of MLH1/PMS2-deficiency in sebaceous neoplasia.