Genetic mutations in HER2-positive breast cancer: possible association with response to trastuzumab therapy.

BACKGROUND: HER2-positive breast cancer occurs in 15-20% of breast cancer patients and is characterized by poor prognosis. Trastuzumab is considered the key drug for treatment of HER2-positive breast cancer patients. It improves patient survival; however, resistance to trastuzumab remains a challenge in HER2-positive breast cancer patients. Therefore, the prediction of response to trastuzumab is crucial to choose optimal treatment regimens. The aim of the study was to identify genetic variants that could predict response to anti-HER2-targeted therapy (trastuzumab) using next-generation sequencing. METHOD: Genetic variants in the hotspot regions of 17 genes were studied in 24 Formalin-Fixed Paraffin-Embedded (FFPE) samples using Ion S5 next-generation sequencing system. FFPE samples were collected from HER2­positive breast cancer patients previously treated with anti­HER2­targeted treatment (Trastuzumab). Patients were divided into two groups; trastuzumab-sensitive group and trastuzumab-resistant group based on their response to targeted therapy. RESULTS: We identified 29 genetic variants in nine genes that only occurred in trastuzumab-resistant patients and could be associated with resistance to targeted therapy including TP53, ATM, RB1, MLH1, SMARCB1, SMO, GNAS, CDH1, and VHL. Four variants out of these 29 variants were repeated in more than one patient; two variants in TP53, one variant in ATM gene, and the last variant in RB1 gene. In addition, three genes were found to be mutated only in resistant patients; MLH1, SMARCB1 and SMO genes. Moreover, one novel allele (c.407A > G, p. Gln136Arg) was detected within exon 4 of TP53 gene in one resistant patient. CONCLUSION: NGS sequencing is a useful tool to detect genetic variants that could predict response to trastuzumab therapy.

HOXA9 gene promotor methylation and copy number variation of SOX2 and HV2 genes in cell free DNA: A potential diagnostic panel for non-small cell lung cancer.

BACKGROUND: Most cases of lung cancer are diagnosed at advanced stage. Detection of genetic and epigenetic markers in cell-free DNA (cfDNA) is a promising tool for the diagnosis of lung cancer at an early stage. The aim of this study was to identify non-invasive diagnostic markers in cell free DNA (cfDNA) for non-small cell lung cancer (NSCLC) as it is the most common type of lung cancer. METHODS: We investigated the cfDNA HOXA9 gene promotor methylation by pyrosequencing. Copy number variation of SOX2 and HV2 genes were detected by real-time PCR in cfDNA extracted from plasma samples of 25 newly diagnosed NSCLC patients and 25 age and sex matched controls. RESULTS: Methylation level of HOXA9 was significantly higher in NSCLC patients than controls (p > 0.001). SOX2 showed significantly higher CNV and HV2 showed lower CNV in patients than controls (p > 0.001, p = 0.001 respectively). Receiver Operating Characteristic (ROC) curve analysis for HOXA9 methylation, SOX2 CNV and HV2 CNV showed a discrimination power of 79.4%, 80% and 77.5% respectively and the area under the curve for the combined analysis of the three genes was 0.958 with 88% sensitivity and 100% specificity. CONCLUSIONS: In this study, we suggest a potentially diagnostic panel that may help in detection of lung cancer with high sensitivity and specificity using cell free DNA. This Panel included HOXA9 gene methylation and the CNV of SOX2 and HV2 genes.

LncRNA HIF1A-AS2: a potential biomarker for early diagnosis of acute myocardial infarction and predictor of left ventricular dysfunction.

BACKGROUND: Rapid diagnosis of acute myocardial infarction (AMI) is the subject of many clinical studies as it enables an effective therapy, preventing adverse progression of AMI and increasing survival rates. Recent studies have revealed that specific blood-based long non-coding RNAs (lncRNAs) are deregulated in patients with AMI and serve as promising diagnostic and prognostic tools. The current study aimed to determine the potential role of a hypoxia-responsive lncRNA, hypoxia-inducible factor 1A antisense RNA 2 (HIF1A-AS2), as a biomarker for early diagnosis and predictor of left ventricular dysfunction (LVD). METHODS: This study was carried out on 48 patients with AMI and 50 age-and sex-matched controls. The relative quantification of HIF1A-AS2 expression was done using reverse transcription real-time polymerase chain reaction. RESULTS: Compared to the control group, HIF1A-AS2 were significantly higher in MI patients (P < 0.001). Interestingly, patients presenting within 3 h of chest pain onset had elevated levels of HIF1A-AS2 as compared to patients with late presentation. The ROC curve was constructed to assess HIF1A-AS2 as an early marker. It demonstrated higher sensitivity (94%) and specificity (86%). Moreover, the multivariate regression analysis revealed that HIF1A-AS2 was significantly associated with LVD in the patient group after 6 months follow up (p = 0.018). CONCLUSION: Our study suggests that HIF1A-AS2 may be a potential early diagnostic biomarker of AMI with high sensitivity. In addition, it might have a promising role as a predictor of left ventricular dysfunction.

Preliminary results of targeted sequencing of BRCA1 and BRCA2 in a cohort of breast cancer families: New insight into pathogenic variants in patients and at­risk relatives.

Breast cancer (BC) is the most commonly diagnosed cancer worldwide and a major health concern in Egypt. There is a known association between pathogenic variants identified in breast cancer susceptibility gene (BRCA)1 and 2 and the risk of developing BC. However, the number of studies investigating mutations in BRCA1 and BRCA2 in Egypt remains limited. Thus, the aim of the present study was to investigate the frequency of BRCA1 and BRCA2 variants in patients with BC and their relatives. For this purpose, 11 families (11 patients and 16 relatives) were included in the present study. BRCA1 and BRCA2 variants were investigated using the Ion S5 next­generation sequencer. It was found that pathogenic variants were more frequent in patients with familial BC (FBC) than in those with sporadic BC, with 71% of variants in BRCA2, including the first reported identification of c.9089del, c.5583_5584dup, c.8243G>A and c.7976G>A pathogenic variants in Egyptian patients with BC. Pathogenic variants in relatives were confined to FBC c.1278delA, c.1960_1961del, and c.1224delT, with a higher incidence of variants of uncertain significance (VUS), such as BRCA2 intron 2 c.68­16delT. Of note, two cold spot benign variants, c.3113A>G and c.4837A>G, were repeatedly found (67%) in patients and relatives. In conclusion, to the best of our knowledge, novel pathogenic variants and VUS in Egyptian patients with BC and their high­risk relatives were identified for the first time in the present study. These findings should be integrated with other genomic data concerning Egyptian families and carefully interpreted during genetic counseling.

Lesson to Learn From Cellular infiltrate in Scalp Biopsy of Alopecia Areata.

BACKGROUND: Alopecia areata (AA) is a common cause of hair loss. It is mediated by T lymphocytes. Scalp biopsy findings in AA differ according to the disease phase and activity. OBJECTIVES: To study the cellular infiltrate in the transverse section of scalp biopsy of AA at different disease stages and in relation to disease activity. METHODS: The study was performed on 40 subjects with AA. A 4-mm punch biopsy was obtained from an AA scalp lesion. Biopsies were sectioned horizontally; 2 anatomical levels were studied (mid dermal and deep dermal levels). RESULTS: Ninety-five percent of AA showed noncicatrical alopecia. A significant relation was found between the course of AA and the terminal:vellus ratio. Peribulbar lymphocytic infiltration was seen in 70% of cases. Mast cells were observed in 87.5% of cases, including fibrous tract and around the arrector pili muscles. Eosinophils were detected in the scalp biopsy of 22.5% of cases. Course and activity of AA were significantly related to the peribulbar lymphocytic cell infiltration but not to mast cells and eosinophils. CONCLUSION: Although a peribulbar lymphocytic infiltrate is the classical finding of AA, it is absent in the chronic phase. Mast cells are commonly found in the scalp biopsy of AA and could explain the potential therapeutic effect of antihistamines.

MicroRNA155 Expression in Different Phenotypes and Genotypes of Behçet's Disease in a Sample of Egyptian Patients.

AIM: To display microRNA155 (miRNA155) expression in different entities of Behçet's disease (BD), and to find out if expression is affected in more than one of disease status than another, either phenotypically, according to HLA B51 expression, presence of family history, or patients' age. METHODS: Thirty BD patients (13 of which were HLAB51 positive) and 15 healthy subjects' samples were obtained. White blood cell miRNA155 expression in both types of samples was estimated. RESULTS: Results showed that there is a degree of relation between decrease of miRNA155 expression and different disease aspects, and also, that miRNA155 has an inverse relation with the patients' ages. CONCLUSION: MiRNA155 might be used as a measure of disease of different phenotypes, and that any manifestation of the disease can happen when the expression level decreases.

Serum expression of microRNA-16 in a cohort of Egyptian patients with ulcerative colitis and its correlation with disease extent and severity / Expressão sérica do microRNA-16 em uma coorte de pacientes egípcios com colite ulcerosa e sua correlação com a extensão e gravidade da doença

Abstract Ulcerative colitis is one of the IBDs. Its etiology and pathogenesis remain undefined with an interaction between environmental, genetic and immunological factors is the most accepted explanation. Several recent studies have examined microRNA expression in the peripheral blood and tissues from IBD patients. The study aims at assessing the expression of serum miR-16 in ulcerative colitis patients and its correlation with disease extent, activity and severity. It included 30 treatment naïve ulcerative colitis patients of different presentations. Serum miR-16 expression was assessed using reverse transcriptase quantitative real time PCR (RT-qPCR), and then correlated with that of a group of 20 healthy subjects to assess its role in diagnosis of ulcerative colitis. Also, it was correlated with disease extent (proctitis, left sided colitis, extensive colitis) and disease activity and severity indices (Truelove and Witts criteria, fecal calprotectin and UCEIS). Thirty ulcerative colitis patients were enrolled, 53% had mild, 37% had moderate, while 10% had severe disease. Concerning endoscopic extent, 8 had proctitis, 14 had left sided colitis and 8 had extensive colitis. Serum expression of miR-16 in the 30 patients were compared to that of the healthy control subjects. The patients' group showed median serum miR-16 expression of 1.91, 1.13 for the control group with a significant difference between both groups. Correlation between serum miR-16 expression with disease extent, activity and severity showed no significant relation. From the current study we can conclude that increased serum expression of miR-16 is associated with ulcerative colitis despite no significant relation to disease activity extent or severity.

Resumo A colite ulcerativa é uma das DII. Sua etiologia e patogênese permanecem indefinidas; a interação entre fatores ambientais, genéticos e imunológicos é a explicação mais aceita. Vários estudos recentes avaliaram a expressão de microRNA no sangue e tecidos periféricos em pacientes com DII. O presente estudo teve como objetivo avaliar a expressão do miR-16 sérico em pacientes com colite ulcerativa e sua correlação com a extensão, atividade e gravidade da doença. Foram incluídos 30 pacientes de colite ulcerativa, com diferentes apresentações, que ainda não haviam sido submetidos a nenhum tipo de tratamento. A expressão sérica de miR-16 foi avaliada usando transcrição reversa seguida de reação em cadeia da polimerase quantitativa (RT-qPCR) e, em seguida, correlacionada com a de um grupo de 20 indivíduos saudáveis para avaliar seu papel no diagnóstico de colite ulcerativa. Além disso, foi feita uma correlação com a extensão da doença (proctite, colite do lado esquerdo, colite extensa) e com os índices de atividade e gravidade da doença (critérios de Truelove e Witts, calprotectina fecal e UCEIS). Trinta pacientes com colite ulcerativa foram incluídos no estudo, classificada como leve em 53%, moderada em 37% e grave em 10%. Quanto à extensão endoscópica, oito apresentavam proctite, 14 apresentavam colite do lado esquerdo e oito apresentavam colite extensa. A expressão sérica de miR-16 nos 30 pacientes foi comparada à dos indivíduos controle saudáveis. No, grupo de pacientes, a expressão sérica de miR-16 foi de 1,91 (grupo controle: 1,13), uma diferença estatisticamente significativa entre os dois grupos. Não foi observada relação significativa entre a expressão sérica de miR-16 e a extensão, atividade e gravidade da doença. A partir do presente estudo, pode-se concluir que o aumento da expressão sérica do miR-16 está associado à colite ulcerativa, apesar de não haver relação significativa com a extensão ou gravidade da atividade da doença.