High Prevalence Of Mutations Research Articles

High prevalence of the MLH1 V384D germline mutation in patients with HER2-positive luminal B breast cancer

HER2-positive luminal B breast cancer (BC), a subset of the luminal B subtype, is ER-positive and HER2-positive BC which is approximately 10% of all BC. However, HER2-positive luminal B BC has received less attention and is less represented in previous molecular analyses than other subtypes. Hence, it is important to elucidate the molecular biology of HER2-positive luminal B BC to stratify patients in a way that allows them to receive their respective optimal treatment. We performed molecular profiling using targeted next-generation sequencing on 94 HER2-positive luminal B BC to identify its molecular characteristics. A total of 134 somatic nonsynonymous mutations, including 131 nonsynonymous single nucleotide variants and three coding insertions/deletions were identified in 30 genes of 75 samples. PIK3CA was most frequently mutated (38/94, 40.4%), followed by TP53 (31/94, 33.0%), and others were detected at lower frequencies. Recurrent germline mutations of MLH1 V384D were found in 13.8% (13/94), with a significantly high TP53 mutations rate. The frequency of MLH1 V384D germline mutation in individuals with HER2-positive luminal B BC was significantly higher than that observed in the controls. All 13 cases were classified as microsatellite stable tumors. Tumor mutation burdens (TMB) were not significantly different between MLH1 V384D carrier and wild type. The concordant results of microsatellite instability (MSI) and TMB suggest that the haploinsufficiency of MLH1 plays a role as a tumor predisposition factor rather than a direct oncogenic driver. Our study identified, for the first time, that MLH1 V384D germline variant is frequently detected in HER2-positive luminal B BC. MLH1 V384D germline variant may not only contribute to gastrointestinal cancer predisposition but may also contribute to BC in East Asians.

Systematic Review of Somatic Mutations in Splenic Marginal Zone Lymphoma

The aims of this systematic review are to refine the catalogue of somatic variants in splenic marginal zone lymphoma (SMZL) and to provide a well-annotated, manually curated database of high-confidence somatic mutations to facilitate variant interpretation for further biological studies and future clinical implementation. Two independent reviewers systematically searched PubMed and Ovid in January 2019 and included studies that sequenced SMZL cases with confirmed diagnosis. The database included fourteen studies, comprising 2817 variants in over 1000 genes from 475 cases. We confirmed the high prevalence of NOTCH2, KLF2 and TP53 mutations and analysis of targeted genes further implicated TNFAIP3, KMT2D, and TRAF3 as recurrent targets of somatic mutation based on their high incidence across studies. The major limitations we encountered were the low number of patients with whole-genome, unbiased analysis and the relative sensitivities of differing sequencing approaches. Overall, we showed that there is little concordance between whole exome sequencing studies of SMZL. We strongly support the continuing unbiased analysis of the SMZL genome for mutations in all protein-coding genes and provide a valuable database resource to facilitate this endeavour that will ultimately improve our understanding of SMZL pathobiology.

High mutation prevalence of precore and basal core promoter in pregnant women who underwent spontaneous HBeAg seroconversion within one year postpartum

BackgroundSeroconversion of hepatitis B e antigen (HBeAg) is a critical event in the natural course of hepatitis B virus (HBV) infection. AimWe herein characterize the virological factors associated with postpartum spontaneous HBeAg seroconversion. MethodsA total of 214 pregnant women positive for both hepatitis B surface antigen (HBsAg) and HBeAg were followed up at 7–12 months postpartum. ResultsOf the subjects, 26 (12.1%) achieved spontaneous HBeAg seroconversion. Receiver operating curve analysis indicated that HBV DNA level <1.0 × 107 IU/mL, HBsAg <1.0 × 104 IU/mL and HBeAg <7.36 × 102 S/CO each independently predicted HBeAg seroconversion within 12 months postpartum. At delivery, 73.1% (19/26) women with postpartum HBeAg seroconversion had precore (PC) and/or basal core promoter (BCP) mutations, higher than that (5/36, 13.9%) in the women without postpartum seroconversion. Binary logistic regression analysis indicated that the presence of mutations in PC, BCP, and both PC and BCP at delivery was associated with an increased likelihood (OR = 13.286, 16. 238, and 22.143 respectively, all P < 0.05) to undergo postpartum spontaneous HBeAg seroconversion. ConclusionThese results suggest that quantitative determination of virological markers and sequencing PC and BCP can predict spontaneous HBeAg seroconversion, which could be valuable in deciding antiviral therapy against HBV.

P-020 - Surprisingly high prevalence of DPYD gene mutation in UK regional trust: implications for treatment

P-020 - Surprisingly high prevalence of DPYD gene mutation in UK regional trust: implications for treatment

Abstract 2513: Mutational and transcriptional landscape of nodal marginal zone lymphoma

Abstract Nodal marginal zone lymphoma (NMZL) is a rare B-cell neoplasm lacking disease-defining markers. In this study, we aimed to determine the mutational profile of Asian NMZLs and report novel genetic features of this disease. We performed WES on FFPE samples of eight Korean NMZLs and two matched non-neoplastic tissue samples. For unmatched cases, additional variant filtering was performed to minimize germline contamination. RNAseq was performed on six NMZL FFPE samples with acceptable samples quality. A total of 394 non-synonymous variants in 370 genes were found with the mean number of variants per case of 49.3 (range 14 – 98). We selected 93 candidate NMZL genes (CNGs) based on the following criteria: (i) genes with mutations in ≥ 2 patients; (ii) genes with mutations in either of two cases with matched control; (iii) genes with mutations in at least one case and reported to have biological roles in B-cell lymphomas. Mutations in genes involved in chromatin remodeling including KMT2D (25%), HIST1H1E (25%), and HIST1H1B (13%) were observed, which was consistent with previous studies. Genes encoding key components of B-cell receptor (BCR) signaling and related pathways were also affected, including CD79B (25%), CARD11 (13%) and BCL6 (13%). Of note, NFKBIE (c.759_762delTTAC) frameshift mutation was found in one patient; this mutation was proven to be loss-of-function in nature, resulting in deregulation of NF-κB signaling. Identical mutation was reported in Hodgkin lymphoma and primary mediastinal B-cell lymphoma, however, this is the first report of this mutation in NMZL. Genes involved in immune response including MYD88 (13%), PIM1 (13%), CD70 (13%) and TNFRSF14 (13%) were also mutated, suggesting the linkage between immune dysregulation and NMZL pathogenesis. At the transcriptome level, three samples were consistently clustered together. This group of patients was characterized by higher Ki-67 proliferation index compared to the others, and was associated with older age, higher serum LDH level and higher IPI score. Gene set analysis (GSA) based on lymphoma-specific gene sets from SignatureDB (https://lymphochip.nih.gov/signaturedb/) showed significant association between XBP1 target genes and these high Ki-67 group (P = 0.024). In summary, this study showed high prevalence of mutations in chromatin remodeling and BCR signaling pathway, readdressing the importance of these pathways in NMZL. Several novel genetic alterations in NMZL were found including NFKBIE c.759_762delTTAC, warranting further validation to clarify the significance of these alterations. We also found that NMZLs with high Ki-67 index were associated with specific gene expression profile, providing clue to the explanation for the heterogeneity of NMZLs. Citation Format: Jiwon Koh, Seongmin Choi, Insoon Jang, Sehui Kim, Cheol Lee, Jin Ho Paik, Kwangsoo Kim, Yoon Kyung Jeon. Mutational and transcriptional landscape of nodal marginal zone lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2513.

The first Japanese nationwide multicenter study of BRCA mutation testing in ovarian cancer: CHARacterizing the cross-sectionaL approach to Ovarian cancer geneTic TEsting of BRCA (CHARLOTTE)

IntroductionBRCA gene mutations are associated with hereditary ovarian cancer. BRCA plays a key role in genome integrity, and mutations result in an increased risk for ovarian cancer. Although various guidelines...

GPCR-mediated PI3K pathway mutations in pediatric and adult thyroid cancer.

Whole exome sequencing (WES) recently identified frequent mutations in the genes of GPCR-mediated PI3K pathway (LPAR4, PIK3CA, and PTEN) in a Chinese population with papillary thyroid cancers (PTCs). The study found LPAR4 mutations as novel gene mutations in adult population with differentiated thyroid cancer (DTC). Here, we determine the prevalence of somatic mutations in this pathway (LPAR4 (exon 1), PIK3CA (exons 9 and 20) and PTEN (exons 5, 6, 7 and 8) in 323 thyroid samples consisting of 17 multinodular goiters (MNG), 89 pediatric DTCs, 204 adult DTCs, and 13 aggressive thyroid cancers including 10 poorly differentiated (PDTC) and 3 anaplastic thyroid cancer (ATC) from another ethnic population. We found 3.37% and 2.45% (includes Q214H, a novel PTEN mutation) in GPCR-mediated PI3K pathway of pediatric and adult DTCs, respectively. Analyses of 507 DTCs from thyroid Cancer Genome Atlas data (TCGA) revealed a low prevalence of mutations in this pathway (1.18%). In 13 cases with PDTC and ATC, we found no mutation in genes of this pathway. By contrast, analyses of 117 aggressive thyroid cancers (PDTC and ATC) from TCGA showed 13% of mutations in this pathway. Moreover, analyses of 1080 pan-cancer cell lines and 9020 solid tumors of TCGA data revealed high rates of mutations in this pathway (cell lines, 24.8%; tumors, 24.8%). In addition, PIK3CA + PTEN (p = <0.001) and LPAR4 + PIK3CA (p = 0.003) significantly co-occurred. Our study reveals a low prevalence of GPCR-mediated PI3K pathway mutations both in pediatric and adult DTCs corroborating the TCGA data and suggests a significant role of this pathway only in a small portion of DTCs. The high prevalence of mutations in this pathway in other solid malignancies suggests an important role in their pathogenesis making it an attractive target for therapeutic intervention both in a small subset of DTCs and other solid cancers.

High prevalence of BRCA deleterious mutations in African-American women with ovarian and/or breast cancer

High prevalence of BRCA deleterious mutations in African-American women with ovarian and/or breast cancer

High Prevalence of BRAF V600E Mutations in Langerhans Cell Histiocytosis of Head and Neck in Chinese Patients

Langerhans cell histiocytosis (LCH) is characterized by clonal proliferation of Langerhans cells and has been classified as a hematolymphoid tumor. BRAF V600E mutation was found to be frequent in LCH; however, it has also been reported that Asia patients with LCH tend to show a lower rate of BRAF V600E mutation. In this study, we found LCH from the head and neck region often involved bone especially the posterior of the mandible and presented a high prevalence of BRAF V600E mutation in Chinese patients. Our findings also showed immunohistochemical detection correlated very well to DNA sequencing of BRAF alterations, which may be useful in the diagnosis of LCH, especially in cases with a low proportion of Langerhans cells, and BRAF inhibitors might be a treatment option for patients with LCH harboring BRAF V600E mutation.

29-OR: High Prevalence of Pathogenic Mutations in Genes Causing Monogenic Diabetes among Patients with Common Type 2 Diabetes

In contrast to monogenic diabetes, the genetic investigations of common forms of type 2 diabetes (T2D) have led to modest advances in precision medicine. We aimed to assess the prevalence in common T2D of pathogenic mutations across 33 genes causing monogenic diabetes, and to compare it to the prevalence found in normoglycemic individuals. Through next-generation sequencing, genes were sequenced in a case-control study including more than 6,000 individuals. We then applied stringent quality control steps before the analyses. The pathogenicity of variants was assessed using the American College of Medical Genetics and Genomics criteria. Primary outcome was the prevalence of pathogenic or likely pathogenic mutations in the case-control study. Secondary outcomes included age of T2D diagnosis, body mass index, T2D treatment and T2D family history. We found a significant association between pathogenic or likely pathogenic variants and T2D risk (p&amp;lt;0.001; 6.5% of carriers among cases). This association was explained in part by pathogenic mutations of two actionable genes, GCK and HNF1A. Overall, among cases, carriers were leaner (β=-1.5±0.5 kg/m²; p&amp;lt;0.01) and tended to develop T2D earlier (β=-1.2±0.7 years; p=0.05) compared to non-carriers, but use of antidiabetic drugs and T2D family history were not different. We reported a high prevalence of pathogenic mutations in monogenic diabetes genes among common T2D patients who did not present with clinical characteristics of monogenic diabetes (i.e., MODY). These results could lead to precision diabetic medicine, and to significant savings in health expenditure via systematic molecular diagnosis of newly diagnosed patients. Disclosure A. Bonnefond: None. P. Froguel: None. Funding European Research Council; French National Research Agency

PB1800 PROGNOSTIC RELEVANCE OF PROTEIN EXPRESSION, CLINICAL FACTORS, AND MYD88 MUTATION IN PRIMARY TESTICULAR DIFFUSE LARGE B‐CELL LYMPHOMA

Background:Primary testicular diffuse large B‐cell lymphoma (tDLBCL) is a rare, clinically aggressive form of extranodal lymphoma confined to the testis. It is known to have a poor prognosis, with an outcome inferior to nodal DLBCL. Only a few studies of tDLBCL have included immunohistochemical and molecular investigations, and little is known about the prognostic significance of the in situ tissue antigen expression profiles in tDLBCL.Aims:We performed a mutation analysis of the MYD88 L265P mutation in 17 tDLBCL patients and investigated the immunophenotypic biomarkers and clinicopathological factors associated with tDLBCL.Methods:Clinical date and paraffin‐embedded material from 17 patients presenting with tDLBCL were retrieved from the archives of the pathology department at the Nanjing Drum Tower Hospital.Immunohistochemistry was used to examine the expression of CD10, Bcl2, Bcl6, MUM1, C‐myc, P50, P65, P53, PSTAT3, OCT4, PIM1, CXCR4 and JAK2. MYD88 gene mutation analysis and complete clinical data evaluation were carried out and linked to overall survival time.Results:High expression of B‐cell lymphoma 2 was detected in 7/17 tDLBCL patients and was associated with a poor prognosis in a univariate model. Most of these patients (6/7) also expressed c‐myc. P50, JAK2, PSTAT3, PIM1 and CXCR4 were also upregulated in tDLBCLs. There were seventeen cases of tDLBCL; in total, 82.4% (14/17) of patients exhibited an activated B‐cell‐like phenotype (ABC), 56.3% (9/16) of patients harbored the MYD88 L265P mutation and showed a correlation between the MYD88 L265P mutation and the ABC phenotype (p = 0.009). However, there was no association between the MYD88 mutation status and overall survival.Summary/Conclusion:We demonstrated that clinical factors such as age, IPI score, stage, lack of B symptoms and the over‐expression of Bcl‐2 are independent prognostic factors determining overall survival and that the over‐expression of P50, JAK2, PSTAT3, PIM1 and CXCR4 may promote the pathogenesis of tDLBCL. Moreover, we found a high prevalence of the MYD88 L265P mutation in tDLBCL, which may reflect a fairly consistent genetic feature in tDLBCL.

PF247 CONSEQUENCES OF NRAS MUTATIONS IN PATIENTS WITH ACUTE MYELOID LEUKEMIA

Background:Acute myeloid leukemia (AML) is a malignancy with a high prevalence of NRAS mutations. Transforming point mutations, located dominantly at hotspot codons 12, 13 and 61 of NRAS gene, occur in 11 to 22% of patients. Mutations in NRAS gene stimulate signalling activity of RAS pathway, resulting in an increased cellular proliferation and reduced apoptosis. Prognostic significance of NRAS gene mutations in AML patients remains controversial.Aims:The aim of our work was to analyze the association of NRAS gene mutations in AML with patient characteristics and outcomes.Methods:A total of 297 consecutive AML (non‐acute promyelocyte leukemia) patients (median age 53 years; range 19–70) with curative therapy were analyzed by next generation sequencing (NGS) using ClearSeq AML panel (Agilent Technologies) on MiSeq and NextSeq (Illumina). The NRAS mutations detected by NGS with variant allele frequency (VAF) ≥1.0% were verified by NRAS StripAssay (ViennaLab). Only mutations scored as positive by both methods were included in further analyses. Overall survival (OS) was analyzed by log‐rank (Mantel‐Cox) test. The comparative statistical analyses were evaluated by t‐test and Fisher's exact test.Results:In total, 84 mutations were identified in 66/297 (22.2%) analyzed AML patients. Fifteen patients (22.7%) were carriers of multiple gene mutations (range 1–3 mutations per patient). The mutations were most frequently found in the codon G12 (44/84 mutations, 52.4%), the most frequent gene mutations were: G12D (26/84; 31.0% mutations), G13D (15/84; 17.9%) and G12S (14/84; 16.7%). Furthermore, rare mutation G60E was identified in one patient. The median VAF of NRAS mutations was 7.0% (range 1.0–53.4%). Frequency of co‐mutation with FLT3‐TKD reached statistical significance (p = 0.025) and a trend for co‐mutation with NPM1 (p = 0.082) and DNMT3A (p = 0.091). A trend for mutual exclusivity was observed for FLT3‐ITD mutations with allelic ratio above 0.5 (p = 0.063). Occurrence of NRAS mutations was associated with younger age (median 48 vs. 54 years, respectively, p = 0.012), and a trend towards higher white blood cell count (median 41.0 vs. 14.2, respectively, p = 0.069). There was no association with OS, blast percentage in BM or sex. The presence of multiple NRAS mutations was not associated with analyzed characteristics nor patient outcomes.Summary/Conclusion:Our study showed NRAS gene mutations detected by two independent methods in more than one fifth of AML patients with curative therapy. Most of them have low VAF, therefore suggesting their later occurrence during leukemogenesis and limited proliferative potential. Although approximately 5% of AML patients harbor multiple NRAS mutations, we were not able to identify the difference between patients with one or more NRAS mutations.Supported by Ministry of Health of the Czech Republic, grant nr. 15–25809A, project FNBr. 65269705 and project MUNI/A/1105/2018. All rights reserved.

Epidemiology and survival of lung cancer in a Latin American cohort.

e13101 Background: Lung cancer epidemiology and survival data is lacking in LA. This information is relevant for understanding regional cancer burden and for designing targeted interventions for cancer control. Methods: Retrospective analysis of lung cancer cases diagnosed at Instituto Nacional de Enfermedades Neoplasicas (INEN) from 2010 to 2014. Data was manually curated from clinical files. Results: 1384 patients were included. Median age at diagnosis was 64 years old (range 20-92) and 51.6% were females. The histologic type was adenocarcinoma, NOS carcinoma, squamous cell carcinoma and small-cell carcinoma in 72.3%, 12.8%, 9.4% and 3.5% of the cases, respectively. The adenocarcinoma/squamous cell carcinoma ratio was 7.7. Among NSCLC cases, most patients presented with advanced disease. Stage III (9.2%) and IV (85.5%) were the most frequent. With a median follow-up of 71.5 months, median overall survival (OS) was 9.2, 7.7 and 8.1 months for adenocarcinoma, NOS carcinoma and squamous carcinoma, respectively. Five-year OS for stage III was 6% and for stage 4, 4%. Detailed results are shown in the table. Conclusions: The epidemiological profile of lung cancer in Peruvian patients is different from that of other countries with a younger age at diagnosis, a preponderance of females over males and an unexpectedly high frequency of adenocarcinomas. This may be in concordance with the low prevalence of tobacco smoking and the high prevalence of EGFR mutations reported for our population as well as with particular exogenous exposures such as biomass fumes from inhouse cooking. The molecular characterization of this cohort of patients is ongoing. [Table: see text]

Detection of circulating tumor DNA in patients with metastatic clear cell renal cell carcinoma.

e16105 Background: Clear cell renal cell carcinoma (ccRCC) represents the most common presentation of kidney cancer. Patient outcomes have significantly improved with the approval of several targeted agents over the past decade. However, the lack of predictive biomarkers has hampered treatment optimization. Circulating tumor DNA (ctDNA) has emerged as an effective, minimally-invasive alternative to tissue for profiling the tumor genome in other tumors. Methods: Blood samples were collected from patients (pts) with metastatic ccRCC for next-generation sequencing of cell free DNA and germline DNA. Targeted sequencing was performed using the Roche Human Oncology Design panel of 981 genes to a median depth of 937x unique reads. Matched FFPE tissue was available from 14 pts and profiled using the same assay. Results: Samples from 52 metastatic, treatment-naïve pts were analyzed. Germline mutations were detected in 6/52 (11%) of the pts with the most frequent abnormalities affecting ATM, PALB2, RAD51D, CHEK2, BRCA1. Median ctDNA fraction was 3.9% (2-40%) with a median of 2 mutations/pt. Somatic mutations were detected in 29/52 (56%) of the samples; of those, 15/29 (51%) of pts harbored RCC-related genes mutations, 3/29 (10%) non-coding mutations and 10/29 (34%) alterations in non RCC-associated genes. Median variant allele frequency was 1.9% (1-22%). Consistent with tissue-based reports, VHL, BAP1, PBRM1, and TP53 were the most frequently altered genes. Lastly, 50% of the patient-matched FFPE tissue samples shared a fully concordant mutation profile. Conclusions: We confirmed a high prevalence of germline mutations in pts with ccRCC. The rate of ctDNA detection in metastatic ccRCC appears to be lower than in other metastatic solid tumors. Furthermore, it is not yet clear whether all the detected somatic alterations are strictly ccRCC-ctDNA dependent. Nevertheless, a quarter of pts exhibited clinically-informative ccRCC associated alterations in their liquid biopsy. These findings suggest that ctDNA is a promising tool for genomic profiling in a subset of patients with metastatic ccRCC.

K-RasG12D Has a Potential Allosteric Small Molecule Binding Site.

KRAS is the most commonly mutated oncogene in human cancer, with particularly high mutation frequencies in pancreatic cancers, colorectal cancers, and lung cancers [Ostrem, J. M., and Shokat, K. M. (2016) Nat. Rev. Drug Discovery 15, 771-785]. The high prevalence of KRAS mutations and its essential role in many cancers make it a potentially attractive drug target; however, it has been difficult to create small molecule inhibitors of mutant K-Ras proteins. Here, we identified a putative small molecule binding site on K-RasG12D using computational analyses of the protein structure and then used a combination of computational and biochemical approaches to discover small molecules that may bind to this pocket, which we have termed the P110 site, due to its adjacency to proline 110. We confirmed that one compound, named K-Ras allosteric ligand KAL-21404358, bound to K-RasG12D, as measured by microscale thermophoresis, a thermal shift assay, and nuclear magnetic resonance spectroscopy. KAL-21404358 did not bind to four mutants in the P110 site, supporting our hypothesis that KAL-21404358 binds to the P110 site of K-RasG12D. This compound impaired the interaction of K-RasG12D with B-Raf and disrupted the RAF-MEK-ERK and PI3K-AKT signaling pathways. We synthesized additional compounds, based on the KAL-21404358 scaffold with more potent binding and greater aqueous solubility. In summary, these findings suggest that the P110 site is a potential site for binding of small molecule allosteric inhibitors of K-RasG12D.

Genomic Characterization of Prostatic Ductal Adenocarcinoma Identifies a High Prevalence of DNA Repair Gene Mutations.

Ductal prostate cancer (dPC) is a rare variant of prostatic adenocarcinoma associated with poor outcomes. Although its histopathologic features are well characterized, the underlying molecular hallmarks of this aggressive subtype are not well described. We sought to provide a comprehensive overview of the spectrum of mutations associated with dPC. Three case series across multiple institutions were assembled. All patients had a diagnosis of dPC, and histopathologic classification was confirmed by an expert genitourinary pathologist. Case series 1 included men who were prospectively enrolled in a tumor sequencing study at the University of Washington (n = 22). Case series 2 and 3 included archival samples from men treated at Johns Hopkins Hospital (n = 21) and University of Calgary (n = 8), respectively. Tumor tissue was sequenced on a targeted next-generation sequencing assay, UW-OncoPlex, according to previously published methods. The frequency of pathogenic/likely pathogenic mutations are reported. Overall, 25 patients (49%) had at least one DNA damage repair gene alteration, including seven (14%) with a mismatch repair gene mutation and 16 (31%) with a homologous repair mutation. Germline autosomal dominant mutations were confirmed or suspected in 10 patients (20%). Activating mutations in the PI3K pathway (n = 19; 37%), WNT pathway (n = 16; 31%), and MAPK pathway (n = 8; 16%) were common. This study strongly suggests that dPCs are enriched for actionable mutations, with approximately 50% of patients demonstrating DNA damage repair pathway alteration(s). Patients with dPC should be offered next-generation sequencing to guide standard-of-care treatment (eg, immune checkpoint inhibitors) or triaged toward an appropriate clinical trial (eg, poly [ADP-ribose] polymerase inhibitors).

SAT-314 Malignant Pheochromocytoma: A Rare Neoplasm Easily Confounded with Metastasis and Associated with Lynch Syndrome

Introduction: Pheochromocytomas are a rare neoplasm of chromaffin cells of the adrenal medulla with an annual incidence in the United States between 500 to 1600 cases, of which about 10% are malignant. Due to this rarity, adrenal masses, especially in the context of other concurrent primary tumors, can be easily confounded with metastasis. Case Report: A 56-year-old male with diabetes mellitus and hypertension presents with left flank pain associated with hematochezia, constipation, and an 8.2kg weight loss over 2.5 months. Physical examination was unremarkable. Abdominal CT scan showed a 4x5cm left adrenal gland mass with infiltration of peri-adrenal tissues and hemorrhage. Dexamethasone suppression test was normal. Plasma metanephrines and aldosterone were not elevated. Repeat CT a month later showed marked increase of the mass to 6.2x5.8cm and an ascending colon mass. Colonoscopy with biopsy revealed a moderately differentiated adenocarcinoma with immunohistochemistry (IHC) staining negative for MSH2 and MSH6 mismatch repair proteins, typical of Lynch syndrome. The patient had a strong family history of malignancy, positive for endometrial cancer (mother) and colon cancer (mother/brothers). A right segmental hemicolectomy and left adrenalectomy-nephrectomy was performed with a presumption that the adrenal mass was a metastatic site. On pathology, the colon specimen confirmed a 5x4x2cm adenocarcinoma. Surprisingly, the adrenal specimen revealed a 7x6x6cm malignant pheochromocytoma with a Pheochromocytoma of the Adrenal Gland Scaled Score (PASS) of 18 with IHC also negative for MSH2 and MSH6. Repeat CT 4 months post-resection showed new lymphadenopathy followed by PET/CT revealing a 2.7x3.4cm FDG avid left para-aortic lymph node consistent with metastasis and a 2.5cm non-FDG avid cystic mass in the pancreatic tail. Discussion: There is a high prevalence of germline mutations associated with pheochromocytomas, commonly including succinate dehydrogenase (SDH) subunit genes, SDHAF2, TMEM127, MAX-gene, and three main associated syndromes including von Hippel Lindau, multiple endocrine neoplasia type 2, and neurofibromatosis type 1. Lynch Syndrome is diagnosed by a mutation in at least one of these mismatch repair genes: MLH1, MSH2, MSH6, and PMS2. It is often associated with extra-colonic cancers including the endometrium, gastric, biliary, urinary tract, pancreas, small intestine, brain, and skin. There is a growing number of cases reporting neuroendocrine tumors in the setting of Lynch Syndrome. To date, there are only two cases reporting an association between pheochromocytoma and Lynch Syndrome, neither of which were described as malignant pheochromocytomas. We present a case of MSH2 and MSH6 mismatch repair protein deficiency in both colon and adrenal masses, suggesting that Lynch Syndrome may be an additional rare risk factor for malignant pheochromocytoma.

High prevalence of Pfdhfr\u2013Pfdhps quadruple mutations associated with sulfadoxine\u2013pyrimethamine resistance in Plasmodium falciparum isolates from Bioko Island, Equatorial Guinea

BackgroundSulfadoxine–pyrimethamine (SP) is recommended for intermittent preventive treatment of malaria in Africa. However, increasing SP resistance (SPR) affects the therapeutic efficacy of the SP. As molecular markers, Pfdhfr (dihydrofolate reductase) and Pfdhps (dihydropteroate synthase) genes are widely used for SPR surveillance. This study aimed to assess the prevalence of Pfdhfr and Pfdhps genes mutations and haplotypes in Plasmodium falciparum isolates collected from Bioko Island, Equatorial Guinea (EG).MethodsIn total, 180 samples were collected in 2013–2014. The single nucleotide polymorphisms (SNPs) of the Pfdhfr and Pfdhps genes were identified with nested PCR and Sanger sequencing. The genotypes and linkage disequilibrium (LD) tests were also analysed.ResultsSequences of Pfdhfr and Pfdhps genes were obtained from 92.78% (167/180) and 87.78% (158/180) of the samples, respectively. For Pfdhfr, 97.60% (163/167), 87.43% (146/167) and 97.01% (162/167) of the samples carried N51I, C59R and S108N mutant alleles, respectively. The prevalence of the Pfdhps S436A, A437G, K540E, A581G, and A613S mutations were observed in 20.25% (32/158), 90.51% (143/158), 5.06% (8/158), 0.63% (1/158), and 3.16% (5/158) of the samples, respectively. In total, 3 unique haplotypes at the Pfdhfr locus and 8 haplotypes at the Pfdhps locus were identified. A triple mutation (CIRNI) in Pfdhfr was the most prevalent haplotype (86.83%), and a single mutant haplotype (SGKAA; 62.66%) was predominant in Pfdhps. A total of 130 isolates with 12 unique haplotypes were found in the Pfdhfr and Pfdhps combined haplotypes, 65.38% (85/130) of them carried quadruple allele combinations (CIRNI-SGKAA), whereas only one isolate (0.77%, 1/130) was found to carry the wild-type (CNCSI-SAKAA). For LD analysis, the Pfdhfr N51I was significantly associated with the Pfdhps A437G (P < 0.05).ConclusionBioko Island possesses a high prevalence of the Pfdhfr triple mutation (CIRNI) and Pfdhps single mutation (SGKAA), which will undermine the pharmaceutical effect of SP for malaria treatment strategies. To avoid an increase in SPR, continuous molecular monitoring and additional control efforts are urgently needed in Bioko Island, Equatorial Guinea.

New biallelic mutations in WEE2: expanding the spectrum of mutations that cause fertilization failure or poor fertilization

To investigate the genetic cause of fertilization failure or poor fertilization. Genetic analysis. University-affiliated center. Twenty-four Chinese women who underwent assisted reproductive technology (ART) and had repeated fertilization failure or poor fertilization. None. Twenty-four affected patients were subjected to whole-exome sequencing and candidate mutations were validated by Sanger sequencing. Single-cell reverse transcription was used to analyze the functional characterization of the splice-site mutation invivo. Evolutionary conservation and molecular modeling analyses were used to predict the impact of missense mutations on secondary protein structure. Immunofluorescence was used to analyze the protein levels of WEE2 and phosphorylated CDC2. Biallelic mutations in WEE2 were identified in 5 of 24 (20.8%) Chinese patients with fertilization failure or poor fertilization. Among these individuals we found a novel splice-site mutation, two novel missense mutations, and a previously reported frame-shift mutation. Splicing mutation c.1136-2A>G of WEE2 caused an alteration of the reading frame and introduced a premature stop codon (p.Gly379Glufs*6/p.Asp380Leufs*39). The missense mutations c.585G>C (p.Lys195Asn) and c.1228C>T (p.Arg410Trp) produced obvious changes in secondary protein structures. Immunostaining indicated that mutated WEE2 resulted in the loss of phosphorylated CDC2. The phenotypes of women carrying WEE2 mutations exhibited slight variability, from total fertilization failure to poor fertilization. Novel mutations in the known causative gene WEE2 were identified in 5 of 24 women with fertilization failure or poor fertilization, indicating a high prevalence of WEE2 mutations in Chinese women experiencing fertilization failure or poor fertilization.

BRAF gene mutations in ameloblastic fibromas

To investigate the BRAF gene mutations in ameloblastic fibroma (AF), and to further analyze the relationship between the BRAF mutation and clinical characteristics so as to provide new reference to the study of AF's molecular pathology. Sixteen cases diagnosed as AF at the Department of Oral Pathology, Peking University School of Stomatology between January 1990 and December 2017 were collected. Genomic DNA was extracted from formalin-fixed, paraffin embedded tissues using the QIAamp DNA Mini Kit (Qiagen, Germany) according to the manufacturer's instructions. Polymerase chain reaction (PCR) and direct sequencings were used to detect the BRAF gene mutations. The clinicopathological data, such as the age, location of the lesion, symptoms and treatments were retrospectively analyzed. The sixteen cases of AF involved nine women and seven men aged 2-67 years. Three lesions occurred in the maxilla and thirteen in the mandible. The most common presenting symptom of AF was a painless slowly enlarging mass with swelling. Ten patients received conservative treatment and the other six patients received radical surgery. Three cases relapsed during the study period. BRAF gene mutation was found in sixteen of all the sixteen samples analyzed (100%). The BRAF mutation was a point mutation with a thymine-adenine transversion at nucleotide 1 799 of 15 exons, resulting in a change at residue 600 that substituted glutamine for valine. This mutation was the strongest activator of the downstream RAS/RAF/MEK/ERK-MAPK signaling pathway. This helped to bring about a gain-of-function mutation due to a V600E substitution. Many studies identified that BRAF regulated survival, apoptosis, and proliferation of cells by inducing MAPK pathways activation. For the existing cases, none of the age, sex, location, recurrence and treatments had a statistically significant correlation with BRAF mutation. Our findings demonstrated high prevalence of BRAF V600E mutation in AF. The pathogenic role remains to be clarified．.