Germline Biallelic Loss in MBD4 leading to Early Onset AML with Hyper-Mutator Genomic Signature

MBD4, a gene encoding a DNA glycosylase functioning in base excision repair, has recently been described in association with predisposition to early onset acute myeloid leukemia (AML), uveal melanoma and colorectal polyposis. MBD4 is essential for guarding against methylation damage due to spontaneous deamination of 5-methylcytosine and biallelic loss of MBD4 allows CG>TG mutations to accumulate in the genome predisposing to AML (Sanders et al. Blood 2018). Further understanding of the full phenotypic spectrum and mechanisms leading to cancer evolution for this predisposition syndrome is critical for informing early recognition, diagnosis, management, and treatment for these individuals and their at-risk family members with the ultimate goal of improving outcomes. Here we report novel germline mutations in MBD4, expand the spectrum of cancers and describe the mutational signatures associated with this cancer predisposition syndrome. Two siblings (brother and sister) from a non-consanguineous family with strong family history of cancers were diagnosed with early-onset AML and colorectal polyposis in their 30s. Prior to the diagnosis of AML, the brother was diagnosed with colorectal cancer and lymphoma at 24 and 28 years old respectively. He was diagnosed with AML at 30 years old and underwent allogenic hematopoietic stem cell transplant (HSCT) using her sister as donor but subsequently passed away following relapse of his AML. The sister was diagnosed with colorectal polyposis at 33 years old and underwent total colectomy due to numerous (>70) hyperplastic and adenomatous colorectal polyps. At the age of 35 years old she was diagnosed with AML after presenting with progressive peripheral cytopenias, lymphadenopathy and splenomegaly. Her AML was characterized by normal karyotype and absent of NPM1, CEBPA and FLT3 aberrations. Further molecular profiling using targeted myeloid NGS panel identified multiple C>T missense mutations including hotspot DNMT3A mutation c.2644C>T, p.R882C (Table 1). SNP microarray revealed balanced genome but identified copy loss of heterozygosity in chromosome 1p and 1q. She received two cycles of remission induction chemotherapy (7+3 regimen) and achieved remission prior to allogenic HSCT using matched unrelated donor. Her post-transplant course was complicated by chronic graft versus host disease treated with tacrolimus. She currently remains in remission now six years post-transplant. She later developed papillary thyroid carcinoma at the age of 44 for which she underwent total thyroidectomy. Shortly after, she was evaluated with brain MRI due to sensorineural hearing loss, which identified bilateral vestibular schwannoma. Her right schwannoma was resected and the smaller, left schwannoma is being monitored. Tumor profiling on tissue from the right schwannoma detected two C>T truncating mutations in NF2 as well as multiple other C>T missense variants resulting in high tumor mutational burden calculated at 16.8 mutations/MB (Table 1). Tumor profiling performed on tissue from her thyroid carcinoma is pending. Due to concern for germline cancer predisposition syndrome, whole exome sequencing was performed and identified two germline nonsense variants in MBD4 c.1291 C>T, p.Arg431* and c.1688 T>A, p.Leu563* in trans configuration. These truncating variants are located in the glycosylase domain of MBD4 and predicted to abolish the catalytic function of the gene (Figure 1). Both truncating variants (Arg431* and Leu563*) are present in gnomAD with very low allele frequency of 3.18 X 10 -5 and 7.04 X 10 -5 respectively. No additional germline mutations were identified. This case further validates the role of MBD4 as germline predisposition to myeloid malignancies characterized by hyper-mutated genomic signatures. Additionally we expanded the spectrum of cancers associated with this novel cancer predisposition syndrome to include papillary thyroid carcinoma and vestibular schwannomas. Genomic profiling of the normal and affected tissue identified germline bi-allelic loss of function mutation in MBD4 as initiator of methylation defect in key driver genes in tissue specific manner leading to carcinogenesis. This conserved path to mutagenesis is unique to this cancer predisposition syndrome and further biological studies are needed to fully understand the spectrum of cancers associated with this syndrome. Figure 1 Figure 1. Disclosures Majhail: Anthem, Inc: Consultancy; Incyte Corporation: Consultancy.

Diagnostic challenges in a CMMRD patient with a novel mutation in the PMS2 gene: a case report

Background Constitutional mismatch repair deficiency (CMMRD) is a rare autosomal recessive condition, which is caused by biallelic mutations in mismatch repair genes: MSH2, MLH1, MSH6, and PMS2. Case presentation We reported a unique case of an 11-year-old Chinese girl with colorectal polyposis and café-au-lait macules who had no obvious family history of Lynch syndrome-associated tumors, followed by brain gliomas and colorectal carcinoma five years later. The diagnosis of CMMRD was based on gene sequencing analysis showing a homozygous deletion NM_00535.5:c.1577delA (p.Asp526fs) in exon 11 of the PMS2 gene. Although the patient underwent surgery and radiation therapy, and close surveillances including radiological, endoscopic and hematological screening have been recommended, she died of the exacerbation of neurological symptoms at the age of 18. Conclusions We identified a novel homozygous deletion in the PMS2 gene in a CMMRD patient with complex clinical features.

Germline loss-of-function variants in the base-excision repair gene MBD4 cause a Mendelian recessive syndrome of adenomatous colorectal polyposis and acute myeloid leukaemia

ABSTRACTInherited defects in base-excision repair (BER) predispose to adenomatous polyposis and colorectal cancer (CRC), yet our understanding of this important DNA repair pathway remains incomplete. By combining detailed clinical, histological and molecular profiling, we reveal biallelic germline loss-of-function (LOF) variants in the BER gene MBD4 to predispose to adenomatous polyposis and –uniquely amongst CRC predisposition syndromes– to myeloid neoplasms. Neoplasms from MBD4-deficient patients almost exclusively accumulate somatic CpG>TpG mutations, resembling mutational signature SBS1. MBD4-deficient adenomas harbour mutations in known CRC driver genes, although AMER1 mutations were more common and KRAS mutations less frequent. We did not find an increased risk for colorectal tumours in individuals with a monoallelic MBD4 LOF variant. We suggest that this condition should be termed MBD4-associated neoplasia syndrome (MANS) and that MBD4 is included in testing for the genetic diagnosis of polyposis and/or early-onset AML.

Use of sanger and next-generation sequencing to screen for mosaic and intronic APC variants in unexplained colorectal polyposis patients

In addition to classic germline APC gene variants, APC mosaicism and deep intronic germline APC variants have also been reported to be causes of adenomatous polyposis. In this study, we investigated 80 unexplained colorectal polyposis patients without germline pathogenic variants in known polyposis predisposing genes to detect mosaic and deep intronic APC variants. All patients developed more than 50 colorectal polyps, with adenomas being predominantly observed. To detect APC mosaicism, we performed next-generation sequencing (NGS) in leukocyte DNA. Furthermore, using Sanger sequencing, the cohort was screened for the following previously reported deep intronic pathogenic germline APC variants: c.1408 + 731C > T, p.(Gly471Serfs*55), c.1408 + 735A > T, p.(Gly471Serfs*55), c.1408 + 729A > G, p.(Gly471Serfs*55) and c.532-941G > A, p.(Phe178Argfs*22). We did not detect mosaic or intronic APC variants in the screened unexplained colorectal polyposis patients. The results of this study indicate that the deep intronic APC variants investigated in this study are not a cause of colorectal polyposis in this Dutch population. In addition, NGS did not detect any further mosaic variants in our cohort.

Germline and Somatic Whole-Exome Sequencing Identifies New Candidate Genes Involved in Familial Predisposition to Serrated Polyposis Syndrome

The serrated polyposis syndrome (SPS) is the most common and yet underdiagnosed colorectal polyposis syndrome. It is characterized by multiple and/or large colonic serrated polyps and a higher associated risk for colorectal cancer (CRC). The main objective of this study was to identify new candidate genes involved in the germline predisposition to SPS/CRC. Thirty-nine SPS patients from 16 families (≥2 patients per family) were recruited without alterations in well-known hereditary CRC genes, and germline and somatic whole-exome sequencing were performed. Germline rare variants with plausible pathogenicity, located in genes involved in cancer development, senescence and epigenetic regulation were selected. Somatic mutational profiling and signature analysis was pursued in one sample per family, when possible. After data filtering, ANXA10, ASXL1, CFTR, DOT1L, HIC1, INO80, KLF3, MCM3AP, MCM8, PDLIM2, POLD1, TP53BP1, WNK2 and WRN were highlighted as the more promising candidate genes for SPS germline predisposition with potentially pathogenic variants shared within families. Somatic analysis characterized mutational profiles in advanced serrated polyps/tumors, revealing a high proportion of hypermutated samples, with a prevalence of clock-like mutational signatures in most samples and the presence of DNA mismatch repair-defective signatures in some cases. In conclusion, we identified new candidate genes to be involved in familial SPS. Further functional studies and replication in additional cohorts are required to confirm the selected candidates.

Germline testing of patients with personal history of colorectal polyposis by cancer genetics counseling services.

47 Background: National Comprehensive Cancer Network (NCCN) guidelines recommend that individuals with >10 adenomatous polyps, ≥2 hamartomatous polyps, or ≥5 serrated polyps proximal to the sigmoid colon have detailed risk assessment and potential genetic testing to rule out polyposis syndrome. Here, we describe germline testing of patients with a personal history of colorectal polyposis by Cancer Genetics Counseling Services. Methods: This is an IRB-approved retrospective chart-review study. Between 2016 and 2020, 1011 unique genetic counseling visits were conducted. Germline testing was recommended by a certified genetic counselor if medically necessary. All patients with a personal history of colorectal polyposis were identified (N=20) and their germline testing results were summarized. Results: The reasons for referral to the Cancer Genetics Counseling Services were personal history of >10 adenomatous polyps (N=13), personal and family history of colorectal polyposis (N=3), personal history of juvenile colorectal polyps (N=3) or personal history of ≥2 hamartomatous polyps (N=1). The median age is 58 years-old (1-84). Ten (50%) patients were females. Caucasians, African Americans and other ethnic backgrounds represented 80%, 10% and 10% respectively. In our cohort, 6 out of 20 (30%) patients had a pathogenic germline mutation, 5 (25%) patients had variant of unknown significance (VUS) and 9 (45%) patients had negative testing. Among patients with pathogenic germline mutations, 3 patients had a pathogenic APC mutation (APC c.1659G>A, APC c.2802C>A and APC c.1643dupT) and were diagnosed with Familial Adenomatous Polyposis (FAP). One patient had 2 pathogenic MUTYH mutations (MUTYH c.536A>G and MUTYH c.1187G>A) and was diagnosed with One patient had a pathogenic PTEN c.634+5G>A mutation and was diagnosed with PTEN Hamartoma Tumor Syndrome. Among the 3 patients with a personal history of juvenile colorectal polyps, one patient had a CHEK2 c.190G>A mutation while the other two had negative genetic test results. The VUS mutations in our cohort were MRE11A c.826C>T, BLM c.3478T>C, BRCA2 c.2519T>C, CHEK2 p.V395L and CTNNA1 c.392dupT. Conclusions: In our cohort of patients with personal history of colorectal polyposis, the majority of patients (45%) had negative germline testing. An underlying pathogenic germline mutation and VUS were identified in 30% and 25% of the patients, respectively. FAP Syndrome was the most commonly diagnosed hereditary polyposis syndrome with 3 patients found to have APC germline mutations. Other pathogenic mutations were identified in the MUTYH, PTEN and CHEK2 genes. Patients with MUTYH and PTEN mutations were diagnosed with MAP and PTEN Hamartoma Tumor Syndromes respectively.

Familial adenomatous colorectal polyposis complicated by colonic obturation: a clinical case

Background. Colorectal obturation is a fairly rare complication in patients with colorectal polyposis. Case descriptions of colonic obturation with underlying familial adenomatous colorectal polyposis have not been reported to date in national and foreign literature.Clinical Case Description. Patient G., female, 31 yo, was emergently admitted to a surgical unit with a preliminary diagnosis: acute intestinal obstruction, complaints of abdominal pain, nausea, vomiting, stool and gas outlet blockage, marked general weakness. Clinical and biochemical blood tests without peculiarities. Signs of intestinal obstruction in abdominal ultrasonic and X-ray examination. Obstructive right hemicolectomy performed as emergent surgery. Diagnosis: transverse colonic C-r T3NoMo, stage II, clinical group 2. Patient had routine fibrocolonoscopy in six months; polyps were revealed in all operated colon portions. APC genetic test was positive, total colectomy was decided with single-barrel ileostomy excretion on anterior abdominal wall. Definitive diagnosis: transverse colonic C-r T3NoMo, stage II, developed with underlying familial adenomatous colorectal polyposis, clinical group 2.Conclusion. Diagnosis of familial adenomatous colorectal polyposis with acute intestinal obturation is challenging due to forced urgent surgical intervention and lack of time for a detailed deeper examination in avoidance of baleful consequences. The case reported demonstrates that clinical manifestations of familial adenomatous colorectal polyposis extend beyond the routine complaints of abdominal bloating, stool blockage and rectal bleeding towards a formidable complication of acute colonic obturation of polypoid genesis.