Circulating cell-free DNAas a biomarker in the serum of colorectal cancer patients

Abstract

Background: The PI3K/AKT/mTOR pathway is frequently activaed across many cancers. AKT1 is a subfamily of serine/threonine protein kinases, affecting cell survival, proliferation and invasion. The prior reports from Western country have shown AKT1 mutation in 3-6% of breast cancer and in 2-3% of endometrial cancer. However, the clinico-pathological features of AKT1 E17K mutation in Asian female cancers is unclear. In this study, we examined breast cancer samples and endometrial cancer samples for mutation in AKT1 hotspot at our institution. Methods: For patients with breast cancer and endometrial cancer, we retrospectively analyzed tissue samples preserved between January 2008 and June 2015 in a biobank at national cancer center hospital. We extracted DNA and determined AKT1 mutation with PNA-LNA clamp methods. Kaplan-Meier analysis was performed to analyze relapse-free survival (RFS) after surgery and overall survival (OS) for breast cancer patient. Results: A total of 469 patients were analyzed. Three hundred and twenty-nine patients were breast and 140 were endometrial cancer. The median age was 52 (range 22-90). In breast cancer patients, 305 patients were initially stage I to III, and thirty-two patients were initially Stage IV. The number of tumor subtype was hormone receptor (HR) positive and HER2 negative tumor; 248 (74%), HR positive and HER2 positive tumor; 44 (13%), HR negative and HER2 positive tumor; 28 (8%), and Triple negative tumor; 17 (5%), respectively. We detected AKT1 mutation in 24 patients (7.3%) of breast-cancer patients and 7 patients (5.0%) of endometrial-carcinoma patients. In breast cancer patients, there were 176 relapses and 32 deaths. Median RFS was 79 months in patients with AKT1 wild type and 75 months in patients with AKT1 mutation (p-value:0.77). Conclusions: We detected AKT1 mutation in 31 patients (6.6%) of 469 breast and endometrial cancer. RFS in breast cancer patients is not significantly different by AKT1 mutation status. Background: Head and neck squamous cell carcinoma (HNSCC) in India is strongly associated with use of tobacco and areca nut products. Most patients present in their fifth and sixth decades at advanced stages, compared with age at onset in the North American population. These differences in etiology and onset suggest potential differences in pathogenesis and immuno-biology. During immune evasion by tumor, receptors such as CTLA-4, LAG-3, TIM-3, and programmed cell death protein 1 (PD-1), have been identified on exhausted and dysfunctional lymphocytes and shown to be upregulated in tumor cell lines and tissues by transcriptional methods. However, there is limited information currently available in the Indian population in this regard. In our study we intended to define the immune contexture in HNSCC cases by characterizing the tumor infiltrating lymphocytes (TILs) and the intra-tumoral heterogeneity of other relevant markers by immunohistochemistry (IHC) methods. Determining the prognostic and predictive value of these markers may guide the development of drugs for this cancer type, as it has for other carcinomas. It could help clinicians better predict clinical outcomes and thereby make more informed therapeutic decisions. Methods: 50 cases of HNSCC were studied for expression of immune cell markers such as PDL1, IDO, CD8, FOXP3, LAG-3 by IHC methods on formalin fixed paraffin embedded (FFPE) sections. The whole slide images of the stained slides were analyzed manually and by digital algorithms. Frequency and intensity of PDL1 and IDO were evaluated in both immune cells (IC) and tumor cells (TC). Proximity analyses between PDL1+ IC /IDO+IC with FOXP3+ IC /CD8+ IC/ CD3+ IC were performed using standard morphometric approaches and the pattern of immune cell infiltration documented. Results: Preliminary analysis reveals high concordance between pathologists for all markers and between manual and digital scoring methods. Significant positive correlation between FOXP3+ IC and PDL1+ IC and IDO+ TC with FOXP3 IC is seen with overall percentage of PDL1 and FOXP3 positive immune cells being low as compared to IDO and CD8. Conclusions: As these changes, at least in part, mirror the immune contexture of HNSCC studied in the North American population, it is surmised that treatment approaches based on PD1 blockade are likely to also provide significant benefit to Indian patients. More detailed analysis is being done. Data will be correlated with pathological stage, tumor and serum kynurenine levels, and two year follow up information , wherever available. Background: Hepatocellular carcinoma (HCC) is one of the most frequent cancers in Egypt in which there is high prevalence of hepatitis C virus (HCV) infection. Matrix metalloproteinases (MMPs) are multifunctional proteins that assume an urgent part in cell development, differentiation, inflammation and angiogenesis. MMPs gene poly-morphism might be included in development of HCV-related HCC. The study expected to explore relationship of MMP-1,3 & 9 genes polymorphisms with liver cir-rhosis and HCC patients. Methods: This study included 128 individuals who were enrolled from Menoufia University Hospital in the period between October 2015 and August 2016 classified into the following; Group I: included 48 patients with HCC on top of liver cirrhosis, they were 26 males and 22 females with mean age (58.6065.29); Group II: 50 patients with liver cirrhosis, they were 26 males and 24 females with mean age (56.7465.21); and Group III: 30 healthy subjects as controls, they were 15 males and 15 females with mean age (56.30 67.30). Diagnosis of HCC was done by their characteristic features in two imaging methods (abdominal US & triphasic CT). All subjects except controls were positive for serum HCV RNA. Liver functions tests, AFP & CHILD score were assessed. Genes polymorphisms were made by PCR-RFLP. Results: HCC patients had higher mutant G2G2 (35.4%) and G2 allele (62.5%) of MMP-1 gene than both cirrhotic (P < 0.05) and control groups (P < 0.001). For MMP-3 gene, additionally, HCC patients had the most noteworthy predominance of mutant 5A/5A (22.9%) and 5A allele (52.1%) than both cirrhotic (P < 0.05) and control groups (P < 0.001). The results of MMP-9 gene uncovered higher frequency of mutant TT genotype and T allele in both HCC (56.3% and 74%, respectively) and cirrhotic groups (10% and 35%, respectively) than in the control group. In HCC patients, we detected a significant correlation between heterozygote G1/G2 and G2/G2 of MMP-1 gene and homozygote TT of MMP-9 with higher CHILD score, tumor size and stage (P < 0.05). Moreover, MMP-3 5A/6A had higher CHILD score, portal vein thrombosis and advanced stage (P < 0.05) compared with other genotypes. Conclusions: MMP-1, 3, 9 gene mutation may be embroiled in progression of liver cir-rhosis and hazard relationship for HCC development. Background: Cell-free DNA is extracellular nucleic acids found in cell-free plasma/ serum of humans. Circulating extracellular DNA can be found in healthy persons, persons with nonmalignant diseases, as well as persons with various malignancies. Given the recent approval of a liquid biopsy, the use of circulating tumor DNA as a novel and