Case report: response to immunotherapy and association with the fh gene in hereditary leiomyomatosis and renal cell cancer-associated renal cell cancer.

Hereditary leiomyomatosis and renal cell cancer (HLRCC) is a rare autosomal dominant syndrome caused by a germline mutation in the fumarate hydratase (FH) gene that manifests with cutaneous leiomyomas, uterine fibroids, and renal cell cancer (RCC). Patients with HLRCC-associated RCC (HLRCC-RCC) have aggressive clinical courses, but there is no standardized therapy for advanced HLRCC-RCC. In this study, we described a case of aggressive HLRCC in a 33-year-old female who exhibited a novel heterozygous germline insertion mutation in exon 8 of the FH gene (c.1126 C > T; p.Q376*). The patient underwent laparoscopic resection of the right kidney, but metastases appeared within 3 months after surgery. Histological staining of the resected tumor revealed high expression levels of programmed cell death-ligand 1 (PD-L1). Therefore, the patient was treated with immunotherapy. The patient achieved a partial response to immunotherapy, and the treatment of metastatic lesions has continued to improve. A thorough literature review pinpointed 76 historical cases of HLRCC-RCC that had undergone immunotherapy. From this pool, 46 patients were selected for this study to scrutinize the association between mutations in the FH gene and the effectiveness of immunotherapy. Our results indicate that immunotherapy could significantly improve the overall survival (OS) of patients with HLRCC-RCC. However, no influence of different mutations in the FH germline gene on the therapeutic efficacy of immunotherapy was observed. Therefore, our study suggested that immunotherapy was an effective therapeutic option for patients with HLRCC regardless of the type of FH germline mutation.

Clinical and Molecular Characterization of SMAD4 Splicing Variants in Patients with Juvenile Polyposis Syndrome.

Juvenile polyposis syndrome (JPS) is an inherited autosomal dominant condition that predisposes to the development of juvenile polyps throughout the gastrointestinal (GI) tract, and it poses an increased risk of GI malignancy. Germline causative variants were identified in the SMAD4 gene in a subset (20%) of JPS cases. Most SMAD4 germline genetic variants published to date are missense, nonsense, and frameshift mutations. SMAD4 germline alterations predicted to result in aberrant splicing have rarely been reported. Here, we report two unrelated Italian families harboring two different SMAD4 intronic variants, c.424+5G>A and c.425-9A>G, which are clinically associated with colorectal cancer and/or juvenile GI polyps. In silico prediction analysis, in vitro minigene assays, and RT-PCR showed that the identified variants lead to aberrant SMAD4 splicing via the exonization of intronic nucleotides, resulting in a premature stop codon. This is expected to cause the production of a truncated protein. This study expands the landscape of SMAD4 germline genetic variants associated with GI polyposis and/or cancer. Moreover, it emphasizes the importance of the functional characterization of SMAD4 splicing variants through RNA analysis, which can provide new insights into genetic disease variant interpretation, enabling tailored genetic counseling, management, and surveillance of patients with GI polyposis and/or cancer.

Hereditary Cancer Screening and Outcomes at an Urban Safety-Net Hospital.

PURPOSE: Patients with hereditary cancer syndromes (HCS) have a high lifetime risk of developing cancer. Historically underserved populations have lower rates of genetic evaluation. We sought to characterize demographic factors that are associated with undergoing HCS evaluation in an urban safety-net patient population. METHODS: All patients who met inclusion criteria for this study from 2016 to 2021 at an urban safety-net hospital were included in this analysis. Inclusion criteria were pathologically confirmed breast, ovarian/fallopian tube, colon, pancreatic, and prostate cancers. Patients also qualified for hereditary breast and ovarian cancers or Lynch syndrome on the basis of National Comprehensive Cancer Network guidelines. Institutional review board approval was obtained. Demographic and oncologic data were collected through retrospective chart review. Univariable and multivariable logistic regression models were constructed. RESULTS: Of the 637 patients included, 40% underwent genetic testing. Variables associated with receiving genetic testing on univariable analysis included patients living at the time of data collection, female sex, Latinx ethnicity, Spanish language, family history of cancer, and referral for genetic testing. Patients identifying as Black, having Medicare, having pancreatic or prostate cancer, having stage IV disease, having Eastern Cooperative Oncology Group (ECOG) prognostic score ≥1, having medium or high Charlson comorbidity index, with current or previous cigarette use, and with previous alcohol use were negatively associated with testing. On multivariable modeling, family history of cancer was positively associated with testing. Patients identifying as Black, having colon or prostate cancer, and having ECOG score of 2 had significantly lower association with genetic testing. CONCLUSION: Uptake of HCS was lower in patients identifying as Black, those with colon or prostate cancer, and those with an ECOG score of 2. Efforts to increase HCS testing in these patients will be important to advance equitable cancer care.

Opportunities for Improving Detection of Cancer Predisposition Syndromes in Pediatric Solid Tumor Patients.

BACKGROUND: Detection of cancer predisposition syndromes (CPS) depends on identifying risk factors, including tumor type, family history, and physical findings, to prompt referral for genetic counseling/testing. Whether pediatric oncology providers (POPs) collect adequate family history information is unknown. METHODS: A single-institution retrospective chart review of solid tumor patients <18 years of age referred for a CPS evaluation between January 1, 2017 and January 31, 2019 was performed. POP adherence to American Society of Clinical Oncology (ASCO) family history collection recommendations was measured and compared with genetic counselor performance. Whether sufficient family history was documented to satisfy the criteria of three genetic counseling referral guidelines [American College of Medical Genetics (ACMG), updated Jongmans (UJ), and McGill Interactive Pediatric OncoGenetic Guidelines (MIPOGG)] was evaluated. RESULTS: POPs and genetic counselors achieved all 6 ASCO family history metrics in 3% and 99% of 129 eligible cases, respectively. POPs failed to document sufficient family history to satisfy genetic counseling referral criteria in most cases (74% ACMG, 73% UJ, 79% MIPOGG). CONCLUSIONS: POPs perform poorly in family history collection, raising concern that some patients at risk for a CPS based on their family history may not be referred for genetic counseling/testing. Interventions to improve family history collection are needed to enhance CPS detection.

Open-Source Bioinformatic Pipeline to Improve PMS2 Genetic Testing Using Short-Read NGS Data.

The molecular diagnosis of mismatch repair-deficient cancer syndromes is hampered by difficulties in sequencing the PMS2 gene, mainly owing to the PMS2CL pseudogene. Next-generation sequencing short reads cannot be mapped unambiguously by standard pipelines, compromising variant calling accuracy. This study aimed to provide a refined bioinformatic pipeline for PMS2 mutational analysis and explore PMS2 germline pathogenic variant prevalence in an unselected hereditary cancer (HC) cohort. PMS2 mutational analysis was optimized using two cohorts: 192 unselected HC patients for assessing the allelic ratio of paralogous sequence variants, and 13 samples enriched with PMS2 (likely) pathogenic variants screened previously by long-range genomic DNA PCR amplification. Reads were forced to align with the PMS2 reference sequence, except those corresponding to exon 11, where only those intersecting gene-specific invariant positions were considered. Afterward, the refined pipeline's accuracy was validated in a cohort of 40 patients and used to screen 5619 HC patients. Compared with our routine diagnostic pipeline, the PMS2_vaR pipeline showed increased technical sensitivity (0.853 to 0.956, respectively) in the validation cohort, identifying all previously PMS2 pathogenic variants found by long-range genomic DNA PCR amplification. Fifteen HC cohort samples carried a pathogenic PMS2 variant (15 of 5619; 0.285%), doubling the estimated prevalence in the general population. The refined open-source approach improved PMS2 mutational analysis accuracy, allowing its inclusion in the routine next-generation sequencing pipeline streamlining PMS2 screening.

Clinical Updates and Surveillance Recommendations for DNA Replication Repair Deficiency Syndromes in Children and Young Adults.

Replication repair deficiency (RRD) is a pan-cancer mechanism characterized by abnormalities in the DNA mismatch repair (MMR) system due to pathogenic variants in the PMS2, MSH6, MSH2, or MLH1 genes, and/or in the polymerase-proofreading genes POLE and POLD1. RRD predisposition syndromes (constitutional MMR deficiency, Lynch, and polymerase proofreading-associated polyposis) share overlapping phenotypic and biological characteristics. Moreover, cancers stemming from germline defects of one mechanism can acquire somatic defects in another, leading to complete RRD. Here we describe the recent advances in the diagnostics, surveillance, and clinical management for children with RRD syndromes. For patients with constitutional MMR deficiency, new data combining clinical insights and cancer genomics have revealed genotype-phenotype associations and helped in the development of novel functional assays, diagnostic guidelines, and surveillance recommendations. Recognition of non-gastrointestinal/genitourinary malignancies, particularly aggressive brain tumors, in select children with Lynch and polymerase proofreading-associated polyposis syndromes harboring an RRD biology have led to new management considerations. Additionally, universal hypermutation and microsatellite instability have allowed immunotherapy to be a paradigm shift in the treatment of RRD cancers independent of their germline etiology. These advances have also stimulated a need for expert recommendations about genetic counseling for these patients and their families. Future collaborative work will focus on newer technologies such as quantitative measurement of circulating tumor DNA and functional genomics to tailor surveillance and clinical care, improving immune surveillance; develop prevention strategies; and deliver these novel discoveries to resource-limited settings to maximize benefits for patients globally.

[Multiple primary tumors in children: a clinicopathological analysis of four cases].

Objective: To investigate the clinicopathological features of children with metachronous or synchronous primary tumors and to identify related genetic tumor syndromes. Methods: The clinicopathological data of 4 children with multiple primary tumors diagnosed in the Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China from 2011 to 2023 were collected. The histological, immunophenotypic and molecular characteristics were examined using H&E staining, immunohistochemical staining, PCR, Sanger sequencing and next-generation sequencing (NGS). The patients were followed up. Results: Case 1 was an 8-year-old boy with the adrenal cortical carcinoma, and 5 years later a poorly differentiated gastric adenocarcinoma was detected. Case 2 was a 2-year-old boy, presented with a left ventricular choroid plexus carcinoma, and a hepatoblastoma was detected 8 months later. Case 3 was a 9-month-old girl, diagnosed with renal rhabdoid tumor first and intracranial atypical teratoid/rhabdoid tumor (AT/RT) 3 months later. Case 4 was a 7-year-old boy and had a sigmoid colon adenocarcinoma 3 years after the diagnosis of a glioblastoma. The morphology and immunohistochemical features of the metachronous or synchronous primary tumors in the 4 cases were similar to the corresponding symptom-presenting/first-diagnosed tumors. No characteristic germ line mutations were detected in cases 1 and 2 by relevant molecular detection, and the rhabdoid tumor predisposition syndrome was confirmed in case 3 using NGS. Case 4 was clearly related to constitutional mismatch repair deficiency as shown by the molecular testing and clinical features. Conclusions: Childhood multiple primary tumors are a rare disease with histological morphology and immunophenotype similar to the symptom-presenting tumors. They are either sporadic or associated with a genetic (tumor) syndrome. The development of both tumors can occur simultaneously (synchronously) or at different times (metachronously). Early identification of the children associated with genetic tumor syndromes can facilitate routine tumor screening and early treatment.

Studying the Effect of the Host Genetic Background of Juvenile Polyposis Development Using Collaborative Cross and Smad4 Knock-Out Mouse Models.

Juvenile polyposis syndrome (JPS) is a rare autosomal dominant disorder characterized by multiple juvenile polyps in the gastrointestinal tract, often associated with mutations in genes such as Smad4 and BMPR1A. This study explores the impact of Smad4 knock-out on the development of intestinal polyps using collaborative cross (CC) mice, a genetically diverse model. Our results reveal a significant increase in intestinal polyps in Smad4 knock-out mice across the entire population, emphasizing the broad influence of Smad4 on polyposis. Sex-specific analyses demonstrate higher polyp counts in knock-out males and females compared to their WT counterparts, with distinct correlation patterns. Line-specific effects highlight the nuanced response to Smad4 knock-out, underscoring the importance of genetic variability. Multimorbidity heat maps offer insights into complex relationships between polyp counts, locations, and sizes. Heritability analysis reveals a significant genetic basis for polyp counts and sizes, while machine learning models, including k-nearest neighbors and linear regression, identify key predictors, enhancing our understanding of juvenile polyposis genetics. Overall, this study provides new information on understanding the intricate genetic interplay in the context of Smad4 knock-out, offering valuable insights that could inform the identification of potential therapeutic targets for juvenile polyposis and related diseases.

Novel Cancer Prevention Strategies in Individuals With Hereditary Cancer Syndromes: Focus on BRCA1, BRCA2, and Lynch Syndrome.

Germline pathogenic variants (PVs) in the BRCA1 and BRCA2 genes confer elevated risks of breast, ovarian, and other cancers. Lynch syndrome (LS) is associated with increased risks of multiple cancer types including colorectal and uterine cancers. Current cancer risk mitigation strategies have focused on pharmacologic risk reduction, enhanced surveillance, and preventive surgeries. While these approaches can be effective, they stand to be improved on because of either limited efficacy or undesirable impact on quality of life. The current review summarizes ongoing investigational efforts in cancer risk prevention strategies for patients with germline PVs in BRCA1, BRCA2, or LS-associated genes. These efforts span radiation, surgery, and pharmacology including vaccine strategies. Understanding the molecular events involved in the premalignant to malignant transformation in high-risk individuals may ultimately contribute significantly to novel prevention strategies.

Benign splenic lesions in BAP1-tumor predisposition syndrome: a case series.

BAP1-Tumor Predisposition Syndrome (TPDS) is caused by germline variants in BAP1 and predisposes to solid tumors. After observation of a radiologically malignant-appearing splenic mass with benign pathology in a patient with BAP1-TPDS, we sought to retrospectively characterize splenic lesions in individuals with BAP1-TPDS seen at a comprehensive cancer center. A dedicated radiology review for splenic abnormalities was performed. We identified 37 individuals with BAP1-TPDS, 81% with a history of cancer. Of 33 individuals with abdominal imaging, 10 (30%) had splenic lesions, and none were shown to be malignant on follow-up. Splenectomy in an individual with suspected splenic angiosarcoma showed a benign vascular neoplasm with loss of nuclear staining for BAP1 in a subset of cells. Benign splenic lesions appear to be common and potentially BAP1-driven in individuals with BAP1-TPDS; confirmation of these findings could lead to more conservative management and avoidance of splenectomy.

NLRC5 promotes endometrial carcinoma progression by regulating NF-κB pathway-mediated mismatch repair gene deficiency.

The innate immune molecule NLR family CARD domain-containing 5 (NLRC5) plays a significant role in endometrial carcinoma (EC) immunosurveillance. However, NLRC5 also plays a protumor role in EC cells. Mismatch repair gene deficiency (dMMR) can enable tumors to grow faster and also can exhibit high sensitivity to immune checkpoint inhibitors. In this study, we attempted to determine whether NLRC5-mediated protumor role in EC is via the regulation of dMMR. Our findings revealed that NLRC5 promoted the proliferation, migration, and invasion abilities of EC cells and induced the dMMR status of EC in vivo and in vitro. Furthermore, the mechanism underlying NLRC5 regulated dMMR was also verified. We first found NLRC5 could suppress nuclear factor-kappaB (NF-κB) pathway in EC cells. Then we validated that the positive effect of NLRC5 in dMMR was restricted when NF-κB was activated by lipopolysaccharides in NLRC5-overexpression EC cell lines. In conclusion, our present study confirmed the novel NLRC5/NF-κB/MMR regulatory mechanism of the protumor effect of NLRC5 on EC cells, thereby suggesting that the NLRC5-mediated protumor in EC was depend on the function of MMR.

Juvenile polyposis syndrome in children: The impact of SMAD4 and BMPR1A mutations on clinical phenotype and polyp burden.

OBJECTIVE: A constitutional disease-causing variant (DCV) in the SMAD4 or BMPR1A genes is present in 40%-60% of patients with juvenile polyposis syndrome (JPS). The aim of this study was to characterize the clinical course and polyp burden in children with DCV-positive JPS compared to DCV-negative JPS. METHODS: Demographic, clinical, genetic, and endoscopic data of children with JPS were compiled from eight international centers in the ESPHGAN/NASPGHAN polyposis working group. RESULTS: A total of 124 children with JPS were included: 69 (56%) DCV-negative and 55 (44%) DCV-positive (53% SMAD4 and 47% BMPR1A) with a median (interquartile range) follow-up of 4 (2.8-6.4) years. DCV-positive children were diagnosed at an older age compared to DCV-negative children [12 (8-15.7) years vs. 5 (4-7) years, respectively, p < 0.001], had a higher frequency of family history of polyposis syndromes (50.9% vs. 1.4%, p < 0.001), experienced a greater frequency of extraintestinal manifestations (27.3% vs. 5.8%, p < 0.001), and underwent more gastrointestinal surgeries (16.4% vs. 1.4%, p = 0.002). The incidence rate ratio for the development of new colonic polyps was 6.15 (95% confidence interval 3.93-9.63, p < 0.001) in the DCV-positive group compared to the DCV-negative group, with an average of 12.2 versus 2 new polyps for every year of follow-up. There was no difference in the burden of polyps between patients with SMAD4 and BMPR1A mutations. CONCLUSIONS: This largest international cohort of pediatric JPS revealed that DCV-positive and DCV-negative children exhibit distinct clinical phenotype. These findings suggest a potential need of differentiated surveillance strategies based upon mutation status.

Fumarate hydratase-deficient renal cell carcinoma: an oncology care institutional experience.

Renal cell carcinoma (RCC) accounts for 2% of all cancer cases worldwide, and majority are sporadic. The latest World Health Organization (WHO) classification of renal cell tumors (fifth edition, 2022) has molecularly defined renal tumor entities, which includes fumarate hydratase (FH)-deficient RCC. FH-deficient RCC is an aggressive carcinoma caused by pathogenic alterations in FH gene, seen in 15% of patients with hereditary leiomyomatosis and renal cell cancer syndrome (HLRCC) syndrome. These tumors occur more frequently at a younger age and present at an advanced stage, carrying a dismal prognosis. We report a series of 10 cases of FH-deficient RCC. The mean age was 49.8 years, and all cases presented in advanced stages (III and IV). Morphologically, the cases had varied architectural patterns with characteristic eosinophilic macronucleoli and perinucleolar halo. On immunohistochemistry (IHC), all showed diffuse nucleo-cytoplasmic expression of S-(2-succino)-cysteine (2-SC), with loss of FH in seven cases. FH-deficient RCCs are aggressive neoplasms and can be diagnosed using specific IHC markers (FH and 2-SC). These patients should undergo germline testing for FH gene mutation, genetic counseling, and surveillance of family members.

Gastrointestinal Cancer Precursor Conditions and Their Detection.

Gastrointestinal cancers are a leading cause of cancer morbidity and mortality. Many gastrointestinal cancers develop from cancer precursor lesions, which are commonly found in individuals with hereditary cancer syndromes. Hereditary cancer syndromes have advanced our understanding of cancer development and progression and have facilitated the evaluation of cancer prevention and interception efforts. Common gastrointestinal hereditary cancer syndromes, including their organ-specific cancer risk and surveillance recommendations, are reviewed in this article. The management of common gastroesophageal, pancreatic, and colonic precursor lesions is also discussed, regardless of their genetic background. Further research is needed to advance chemoprevention and immunoprevention strategies.

Updated European guidelines for clinical management of familial adenomatous polyposis (FAP), MUTYH-associated polyposis (MAP), gastric adenocarcinoma, proximal polyposis of the stomach (GAPPS) and other rare adenomatous polyposis syndromes: a joint EHTG-ESCP revision.

BACKGROUND: Hereditary adenomatous polyposis syndromes, including familial adenomatous polyposis and other rare adenomatous polyposis syndromes, increase the lifetime risk of colorectal and other cancers. METHODS: A team of 38 experts convened to update the 2008 European recommendations for the clinical management of patients with adenomatous polyposis syndromes. Additionally, other rare monogenic adenomatous polyposis syndromes were reviewed and added. Eighty-nine clinically relevant questions were answered after a systematic review of the existing literature with grading of the evidence according to Grading of Recommendations, Assessment, Development, and Evaluation methodology. Two levels of consensus were identified: consensus threshold (≥67% of voting guideline committee members voting either 'Strongly agree' or 'Agree' during the Delphi rounds) and high threshold (consensus ≥ 80%). RESULTS: One hundred and forty statements reached a high level of consensus concerning the management of hereditary adenomatous polyposis syndromes. CONCLUSION: These updated guidelines provide current, comprehensive, and evidence-based practical recommendations for the management of surveillance and treatment of familial adenomatous polyposis patients, encompassing additionally MUTYH-associated polyposis, gastric adenocarcinoma and proximal polyposis of the stomach and other recently identified polyposis syndromes based on pathogenic variants in other genes than APC or MUTYH. Due to the rarity of these diseases, patients should be managed at specialized centres.

Multiple Onychopapillomas and BAP1 Tumor Predisposition Syndrome.

Importance: BRCA1-associated protein (BAP1) tumor predisposition syndrome (TPDS) is a cancer genodermatosis associated with high risk of uveal and cutaneous melanoma, basal cell carcinoma, and multiple internal malignant neoplasms, including mesothelioma and renal cell carcinoma. Early detection of the syndrome is important for cancer surveillance and genetic counseling of family members who are at risk. Objective: To determine the prevalence of nail abnormalities in individuals with pathogenic germline variants in BAP1. Design, Setting, and Participants: In this prospective cohort study, individuals who were known carriers of pathogenic BAP1 germline variants were consecutively enrolled between October 10, 2023, and March 15, 2024. Dermatologic evaluation for nail abnormalities was performed, including a history of nail abnormalities and associated symptoms, physical examination, medical photography, and nail biopsy for histopathology. This was a single-center study conducted at the National Institutes of Health Clinical Center. Main Outcomes and Measures: Primary outcomes were the prevalence and spectrum of nail changes and histopathologic characterization. Results: Among 47 participants (30 female [63.8%]; mean [SD] age, 46.4 [15.1] years) ranging in age from 13 to 72 years from 35 families, nail abnormalities were detected in 41 patients (87.2%) and included leukonychia, splinter hemorrhage, onychoschizia, and distal nail hyperkeratosis. Clinical findings consistent with onychopapilloma were detected in 39 patients (83.0%), including 35 of 40 individuals aged 30 years or older (87.5%). Nail bed biopsy was performed in 5 patients and was consistent with onychopapilloma. Polydactylous involvement with onychopapillomas was detected in nearly all patients who had nail involvement (38 of 39 patients [97.4%]). Conclusions and Relevance: This study found that BAP1 TPDS was associated with a high rate of nail abnormalities consistent with onychopapillomas in adult carriers of the disease. Findings suggest that this novel cutaneous sign may facilitate detection of the syndrome in family members who are at risk and patients with cancers associated with BAP1 given that multiple onychopapillomas are uncommon in the general population and may be a distinct clue to the presence of a pathogenic germline variant in the BAP1 gene.

SMAD4 mosaicism in juvenile polyposis: Essential contribution of somatic analysis in diagnosis.

Juvenile polyposis syndrome (JPS) is a rare disease characterized by multiple hamartomatous polyps in the gastrointestinal tract, associated with pathogenic variants of BMPR1A and SMAD4. We present the description of SMAD4 mosaicism in a 30-year-old man who had caecum adenocarcinoma, 11 juvenile colon polyps and epistaxis since childhood. We conducted NGS polyposis and CRC panel analysis on DNA extracted from two polyps, revealing a likely pathogenic SMAD4 variant: NM_005359.5:c. 1600C>T, p.(Gln534*). This variant was then identified at a very low frequency on blood and normal colonic tissue, by targeted visualization of previously obtained NGS data. These findings support the presence of a likely pathogenic mosaic SMAD4 variant that aligns with the patient's phenotype. Given the relatively frequent occurrence of de novo SMAD4 mutations, somatic mosaicism could account for a significant proportion of sporadic JPS patients with unidentified pathogenic variants. This case underscores the diagnosis challenge of detecting mosaicism and emphasizes the importance of somatic analyses.

Variant classification changes over time in the clinical molecular diagnostic laboratory setting.

BACKGROUND: Variant classification in the setting of germline genetic testing is necessary for patients and their families to receive proper care. Variants are classified as pathogenic (P), likely pathogenic (LP), uncertain significance (VUS), likely benign (LB) and benign (B) using the standards and guidelines recommended by the American College of Medical Genetics and the Association for Molecular Pathology, with modifications for specific genes. As the literature continues to rapidly expand, and evidence continues to accumulate, prior classifications can be updated accordingly. In this study, we aim to characterise variant reclassifications in Ontario. METHODS: DNA samples from patients seen at hereditary cancer clinics in Ontario from January 2012 to April 2022 were submitted for testing. Patients met provincial eligibility criteria for testing for hereditary cancer syndromes or polycystic kidney disease. Reclassification events were determined to be within their broader category of significance (B to LB or vice versa, or P to LP or vice versa) or outside of their broader category as significance (ie, significant reclassifications from B/LB or VUS or P/LP, from P/LP to VUS or B/LB, or from VUS to any other category). RESULTS: Of the 8075 unique variants included in this study, 23.7% (1912) of variants were reassessed, and 7.2% (578) of variants were reclassified. Of these, 351 (60.7%) variants were reclassified outside of their broader category of significance. Overall, the final classification was significantly different for 336 (58.1%) variants. Importantly, most reclassified variants were downgraded to a more benign classification (n=245; 72.9%). Of note, most reclassified VUS was downgraded to B/LB (n=233; 84.7%). CONCLUSIONS: The likelihood for reclassification of variants on reassessment is high. Most reclassified variants were downgraded to a more benign classification. Our findings highlight the importance of periodic variant reassessment to ensure timely and appropriate care for patients and their families.

Death receptors 4/5 mediate tumour sensitivity to natural killer cell-mediated cytotoxicity in mismatch repair deficient colorectal cancer.

BACKGROUND: Identifying the target of natural killer (NK) cells in colorectal cancer (CRC) is critical for optimising the clinical use of NK cell-mediated immunotherapy. Mismatch repair deficiency (dMMR) is associated with high immune cell infiltration and MHC Class I defects. Whether dMMR CRC responses to NK cell therapy remains unclear. METHODS: MLH1, DR4, and DR5 knockout cell lines were established using CRISPR-Cas9 system. NK92-MI or NK cell isolated from BABL/C mice were used as effector cells against tumour cells. Inflammatory cytokines secretion by CRC cells was assessed via cytokine analysis. NK-cell-deficient/proficient animal models were used to validate the NK cell sensitivity. RESULTS: We observed that dMMR CRC cells were more sensitive to NK cell-mediated cytotoxicity than were mismatch-repair-proficient (pMMR) CRC cells. In dMMR CRC, Death receptor (DR)4/5 was upregulated and mediated sensitivity to NK cell-mediated cytotoxicity. DR4/5-mediated secretion of interleukin -12 sustained NK cell viability in dMMR CRC. NK cell depletion induced dMMR CRC tumour growth, and NK cell transfer inhibited lung metastasis of dMMR CRC with DR4/5 expression in vivo. TP53 upregulated DR4/DR5 expression in dMMR CRC. CONCLUSIONS: dMMR associated with increased sensitivity to NK cell-mediated cytotoxicity in CRC. DR4/DR5 sensitise dMMR CRC to NK cell-mediated cytotoxicity.