SPECTRUM OF BRAF MUTATIONS IN PAKISTANI COHORT

MOTIVATION BEHIND THE STUDY "BRAF mutations and the association of V600E with CD133 and CDX2 expression in a Pakistani colorectal carcinoma cohort " https://doi.org/10.1186/s12885-024-12925-z

Published in Cancer and Biomedical Research

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BioMed Central
BioMed Central BioMed Central

BRAF mutations and the association of V600E with CD133 and CDX2 expression in a Pakistani colorectal carcinoma cohort - BMC Cancer

Background Despite a high incidence of colorectal carcinoma, data regarding genetic aberrations in colorectal carcinoma (CRC) patients in Pakistan is scarce. This study aimed to determine the frequency of BRAFV600E mutations in colorectal carcinoma tissue in the Pakistani population and to associate BRAFV600E expression with CD133, a marker of colorectal stem cells, and CDX2 marker of differentiation. Methods Sanger Sequencing of exon 15 (426 bp) including the hotspot V600E was performed on formalin-fixed-paraffin-embedded (FFPE) CRC tissue samples of 115 patients. The samples were subjected to immunohistochemistry (IHC) to assess the expression of BRAFV600E, CDX2, and CD133. Additionally, homology modelling and docking were performed to investigate novel deletions revealed in sequencing. Results Twenty-four (20.8%) BRAF variants were identified in the coding region, with V600E mutations detected in 14 (12.2% )cases (GenBank: PP003258.1; Pop Set: 2678087296). Moreover, a wide spectrum of novel non-V600E mutations (8.6%) were identified, including deletions and missense variations. In-silico analysis revealed that due to large deletions in the coding region of three samples, the affinity of the anti-BRAF drugs (Encorafenib and Vemurafenib) for the active site decreased in comparison to the wild type. The IHC analysis showed that BRAFV600E expression was significantly associated with CD133 expression (χ2(1, n=115) = 26.351; p = < 0.001) and with CDX2 expression (χ2(1, n=115) = 14.88; p = 0.001). Multivariate analysis using binary logistic regression revealed association of BRAFV600E mutations with age (OR = 1.123; CI = 1.024–1.232; p = 0.014), gender (OR = 0.071; CI = 0.006–0.831; p = 0.035), grade (0.007; CI = 0-0.644) and CD133 expression (OR = 65.649; CI = 2.153-2001.556; p = 0.016). Conclusion The present study demonstrates a notably high V600E frequency (12.2%) in comparison to global reported data, which ranges from 0.4 to 18%. This finding reflects the importance of upfront BRAF testing of the genetically distinct population of Pakistan. Previously unreported mutations identified in the sample may be of clinical significance and warrant further investigation. The concomitant high expression and significant association between CD133 and BRAFV600E represent vital actionable genes that may be targeted together to improve CRC patient management.

Colorectal cancer remains a major cause of cancer-related morbidity and mortality worldwide, yet genomic profiling and personalized oncology are far from being a clinical reality in many underdeveloped countries. Our motivation stemmed from a deep concern over this disparity. Despite a growing burden of colorectal cancer in our region, there was a notable absence of local molecular data, particularly regarding actionable mutations such as KRAS and BRAF, which guide prognosis and therapy.

We aimed to generate baseline genomic data regarding BRAF mutations for our local population and identify associated molecular events. Genetic testing services in our country are accessible only to a privileged few, and targeted therapy options are virtually non-existent in public-sector hospitals. This reality adds urgency to our efforts, as the lack of infrastructure perpetuates a cycle of late diagnosis, suboptimal treatment, and poor outcomes.

 We hope our work not only contributes to colorectal cancer research but also highlights the need for greater investment in molecular oncology infrastructure in under-resourced settings.

 

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