Comparative analysis of mutational patterns in triple negative breast cancer before and after neoadjuvant chemotherapy in patients with residual disease.

Triple-negative breast cancer (TNBC) is a heterogeneous disease with poor survival compared to other subtypes. Patients with residual disease after neoadjuvant chemotherapy (NAC) face an increased risk of relapse and death. We aimed to characterize the mutational landscape of this subset to offer insights into relapse pathogenesis and potential therapeutic targets. We retrospectively analyzed archived paired (pre- and post-NAC) tumor samples from 25 patients with TNBC with residual disease using a targeted 72-gene next-generation sequencing panel. Our findings revealed a stable mutational burden in both pre- and post-NAC samples, with a median count of 12 variants (IQR 7-17.25) per sample. TP53, PMS2, PTEN, ERBB2, and NOTCH1 variants were observed in pre-NAC samples predominantly. Notably, post-NAC samples exhibited a significant increase in AR gene mutations, suggesting potential prognostic and predictive implications. No difference in mutational burden was found between patients who did and did not receive platinum (p = 0.94), or between those with and without recurrence (p = 0.49). We employed K-means clustering to categorize the patients based on their variant profiles, aiding in the prediction of possible patterns associated with recurrence. Our study was limited by its small sample size and retrospective design, suggesting the need for further validation in larger prospective cohorts.

Droplet digital PCR (ddPCR) using FFPE DNA to assess methylation status of MGMT gene among patients with IDH mutant astrocytoma and IDH wild-type glioblastoma.

MGMT promoter methylation analysis in formalin-fixed paraffin-embedded (FFPE) tissues can be challenging since the DNA obtained is often fragmented. Bisulfite conversion, which is essential to determine methylation status, further degrades DNA. While conventional methylation-specific PCR (MSP) and pyrosequencing assays have long been used to determine the methylation status of MGMT, this study was designed to determine the utility of one-tube DNA extraction method coupled with a droplet digital PCR (ddPCR) assay, to study the epigenetic changes in the promoter region of the MGMT gene using DNA obtained from FFPE.The FFPE blocks of 30 (n=30) patients with Central Nervous System (CNS) WHO grade 4 tumours, previously tested by MSP (2011-2021) were retrieved; DNA was extracted using one-tube extraction method and bisulfite converted. All converted samples were analyzed for methylation status of the MGMT promoter region with a laboratory designed Methylation-Specific ddPCR (MS ddPCR) using degenerate primers and probes that were labelled with FAM or HEX flurocein dye.Of the 30 cases, 20 cases were MGMT methylated and 10 cases were unmethylated by MS ddPCR. The results of MS ddPCR were then compared with those obtained by MSP and found to be concordant in 93.3% (28/30) of the cases and discordant in 2 cases. The Cohen's kappa coefficient (κ) was 0.84. The sensitivity, specificity, positive predictive value and negative predictive value of the assay in detecting the methylation status was found to be 95%, 90%, 95% and 90%.The results show that MS ddPCR is a valuable tool to detect the methylation status of MGMT in FFPE with high sensitivity. This method is cost-effective and easy to perform and could be an attractive alternative to the routine method of MSP.

Effect of Arg399Gln single-nucleotide polymorphism in XRCC1 gene on survival rate of Indian squamous cell head-and-neck cancer patients.

BACKGROUND: Head-and-neck squamous cell carcinoma (HNSCC) is one of the most common cancers that contribute to 20%-40% of all cancer incidences in India. Indian patients with HNSCC are mostly associated with tobacco usage and may have different genetic alterations compared with Western patients who are mostly associated with human papillomavirus infection. Polymorphisms in DNA repair genes are correlated to individuals' susceptibility and progression of cancer. XRCC1 is a DNA repair enzyme. MATERIALS AND METHODS: In the present prospective study, Indian population of HNSCC patients (n = 45) were screened for Arg399Gln variant of XRCC1 using polymerase chain reaction-restriction fragment length polymorphism technique, prospective evaluation of the patients was done after treatment, and the single-nucleotide polymorphism results were correlated to survival functions. RESULTS: Out of 45 patients, 28 patients were Arg/Arg, 12 patients were Arg/Gln, and 5 patients were Gln/Gln. Overall survival for the entire cohort and Arg/Arg, Arg/Gln, and Gln/Gln cohort was 36.3 (95% confidence interval [CI]: 33-39.5), 38.6 (95% CI: 35.3-41.9), 35.8 (95% CI: 28.6-42.9), and 26.4 (95% CI: 13.7-39.1) months (P = 0.097), respectively. Progression-free survival (PFS) of the entire patient cohort and Arg/Arg, Arg/Gln, and Gln/Gln cohort was 35.2 (95% CI: 31.4-39.1), 38.2 (95% CI: 34.3-42.1), 32.7 (95% CI: 26.2-39.1), and 22.3 (95% CI: 9.4-35.3) (P = 0.061), respectively. CONCLUSIONS: This study suggests that HNSCC patients with Gln substitution in place of Arg at position 399 (both homozygous and heterozygous) in XRCC1 protein have significantly inferior survival functions, higher recurrence rate, and events after radical treatment.

Photocatalytic degradation of synthetic food dye, sunset yellow FCF (FD&C yellow no. 6) by Ailanthus excelsa Roxb. possessing antioxidant and cytotoxic activity.

The purpose of our work is to identify the bioactive compounds of bark and leaves extract from Ailanthus excelsa Roxb. and to explore its effectiveness against synthetic food dye. The presence of primary and secondary metabolites was confirmed by carrying out phytochemicals analysis. With the prior knowledge accessible on the indispensable secondary metabolites holding antioxidant and cytotoxicity activity, the quantitative screening of total phenolic and flavonoid content in methanolic and aqueous extract of bark and leaves from Ailanthus excelsa were done. Comparatively, a higher value of flavonoid (161±0.3µg/mg) and phenolic acid content (152.4±0.14µg/mg) was found in bark extract. By FTIR analysis, the characteristic peak was obtained at 1581.63 and 1598.99cm-1 confirmed the presence of functional groups associated to flavonoids and other phenolic groups respectively. In bark extract, 81% of DPPH inhibition was observed when compared to ascorbic acid (standard) 92% of free radical scavenging activity. Bark extract from Ailanthus excelsa exhibited 71% cytotoxicity against HeLa cell line (cervical cancer). In examining the toxicity level of crude extracts with red blood cells (RBC), the bark extract was showed a very less (2.8%) haemolytic activity. They also showed maximum zone of inhibition in antibacterial activity i.e. 13±0.5mm against Escherichia coli culture. At a concentration of 10mg/mL of crude extract from A. excelsa, 55% degradation of sunset yellow dye was observed. It concludes that, the compounds present in the A. excelsa, especially the bark extract showed better photocatalytic, haemolytic, antioxidant, cytotoxicity and antibacterial activity when compared to leaves extract.