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Germline mutations in the mismatch repair genes lead to the Lynch syndrome/ HNPCC, and genetic testing is offered to identify individuals at risk of developing this disease. About a third of all genetic alterations detected in the mismatch repair gene MutL homolog 1 (MLH1) are missense variants. The majority of these variants are of equivocal clinical significance. In this report, we investigate the molecular basis of the pathogenicity of three missense variants localized to distinct functional domains. Our results demonstrate that the MLH1 variants p.Arg265Cys (c.793C>T) and p.Lys618Ala (c.1852_1853AA>GC) significantly decrease the stability of the protein, while another missense variant p.Leu749Gln (c.2246T>A) is able to compromise heterodimerization of the MLH1-PMS2 complex.

AimsEpidermal growth factor receptor (EGFR) T790M mutations can be detected in the circulating tumour DNA from plasma of patients with non-small cell lung cancer (NSCLC) to triage patients for osimertinib eligibility and monitor patients longitudinally for development of T790M-mediated resistance.MethodsUsing droplet digital PCR (ddPCR), we examined the EGFR T790M status of 343 sequential patients with NSCLC and correlated mutational status with demographic and clinical features. Where available, serial T790M blood test results were assessed to identify clinical triggers and timing of repeat testing.ResultsOf the 343 patients with liquid biopsy test results, 24% were T790M positive. No clear clinical correlation with a T790M positive test result was identified in this study, although the number of metastatic sites did correlate significantly with the presence of EGFR sensitising mutations (L858R or exon 19 deletion) in patient plasma, as a measure of tumour DNA shedding. Of the 59 serial blood tests from patients that initially tested negative, 14% were positive on sequential testing, at a time interval up to 6 months after an initially negative blood test.ConclusionsThe ddPCR test for EGFR T790M mutations effectively triaged 24% of patients for treatment with osimertinib, avoiding the need for invasive tissue biopsy in these patients. Our findings suggest that initial and repeat ctDNA testing can be used to monitor for acquired EGFR T790M resistance for NSCLC.

Abstract. Defective mismatch repair leads to the microsatellite instability (MSI) phenotype of colorectal cancer (CRC). We previously showed that the MLH1-93G>A promoter polymorphism is strongly associated with MSI tumours, suggesting a modifier role for this polymorphism in CRC. The MLH1 promoter is bi-directional with the EPM2AIP1 gene located on the antisense strand. In order to evaluate the functional effects of this polymorphism, we transfected a panel of CRC, endometrial cancer and nontumourigenic cell lines with MLH1 luciferase promoter constructs. We used constructs in reverse orientation to assess the effect of this polymorphism on EPM2AIP1. The luciferase activities were compared using a two-sided Student's t-test. Electrophoretic mobility shift assays (EMSAs) were used to evaluate whether differential protein binding was responsible for the differences in promoter activity. We observed a higher level of activity with the -93G allele in all the cell lines observed; including the CRC cell line, HCT116 (P=0.002), the endometrial cancer cell line, HEC-1-A (P<0.001) and the normal colonic cell line, CCD-841-CoTr (P=0.002). This polymorphism also affected EPM2AIP1 transcription with the -93A allele demonstrating higher promoter activity in the HCT116 (P=0.007) and HEC-1-A (P=0.004) cells. The EMSA results suggest that this polymorphism alters the affinity of nuclear factors that bind to this region. Our findings indicate that the -93G>A polymorphism modifies the efficiency of MLH1/EPM2AIP1 transcription.

The results of the sharp trial established sorafenib, a tyrosine kinase inhibitor (TKI), as the sole first-line treatment option in advanced hepatocellular carcinoma (HCC) for more than a decade. In 2020, there has been a surge in new therapies for hcc, including immunotherapeutic strategies and the approval of a number of novel tkis. In addition to sorafenib, lenvatinib and combination atezolizumab–bevacizumab now represent standard first-line treatment options. As those systemic options begin to be better utilized, assurance of adequate liver function and optimal timing are required to improve patient outcomes. Furthermore, sequencing of the agents will have to be carefully tailored, given the increasing armamentarium of choices. Here, we discuss the role of lenvatinib and sorafenib in the first-line management of HCC.

Germline mutations in mismatch repair genes predispose patients to Lynch Syndrome and the majority of these mutations have been detected in two key genes, MLH1 and MSH2. In particular, about a third of the missense variants identified in MLH1 are of unknown clinical significance. Using the PeakPicker software program, we have conducted a proof-of-principle study to investigate whether missense variants in MLH1 lead to allelic imbalances. Lymphocyte RNA extracted from patients harboring known MLH1 variants was used to quantify the ratio of variant to wild-type transcript, while patient lymphocyte DNA was used to establish baseline allelic expression levels. Our analysis indicated that the missense variants c.350C>T, c.793C>T, and c.1852_1853AA>GC, as well as the truncating variant c.1528C>T were all associated with significantly unbalanced allelic expression. However, the variants c.55A>T and c.2246T>C did not demonstrate an allelic imbalance. These results illustrate a novel and efficient method to investigate the pathogenicity of unclassified genetic variants discovered in mismatch repair genes, as well as genes implicated in other inherited diseases. In addition, the PeakPicker methodology has the potential to be applied in the diagnostic setting, which, in conjunction with results from other assays, will help increase both the accuracy and efficiency of genetic testing of colorectal cancer, as well as other inherited diseases.

Background Gender equity has historically been a challenge within gastroenterology. Aims The Canadian Association of Gastroenterology (CAG) developed a survey to identify issues pertaining to equity and gender faced by its membership and to determine areas of action. Methods In 2014, the survey was emailed to all 1155 CAG members, and the data were analyzed using statistical methods. Results One hundred eleven CAG members responded to the survey. Of those, 52% were male, 75% were between 26 and 45 years of age, and 55% were in their first decade of practice. More males held the status of full professor (21% versus 0%; P=0.022). Male CAG members reported working more hours per week than their female counterparts (58.3 ± 15.4 versus 52.3 ± 11.8, P=0.025). Regarding commitments outside the workplace, 81% of respondents had a spouse/partner, and 52% had children under 18 years of age, both of which did not significantly differ based on gender. Overall, 70% were satisfied or very satisfied with their career path. However, significantly more females felt their age/ethnicity/gender/marital status hindered career advancement (36% versus 14%; P=0.008). Furthermore, more females reported difficulties attaining work-life balance (45% versus 22%; P=0.015). Conclusions This survey highlights that gender and equity challenges continue to exist within gastroenterology. The needs assessment highlights that work-life balance, physician well-being, negotiation skills and mentorship are areas of importance to many CAG members.

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