A polyherbal and probiotic-based immunostimulatory formulation for managing gut dysbiosis in colorectal cancer.

INTRODUCTION: The prevention of colorectal cancer (CRC) and its possible relapse remains a daunting challenge due to the unclear mechanisms governing gut dysbiosis and its interaction with the gut-immune axis. The present study aimed to investigate the preventive role of a polyherbal immunostimulatory formulation to restoring microbial diversity in gut and preventive management of colon cancer. METHODS: We developed a novel polyherbal and probiotic-based immunostimulatory formulation with four variants (F3A, F3B, F3C, and F3D), incorporating botanical extracts and probiotics. Initially, plant-derived biological lead compounds were identified using LC/MS/MS. Next, the identified biological leads were subjected to in-silico molecular docking and molecular dynamics simulations with apoptotic biomarker, oncogenic marker, and immunomodulatory biomarkers (i.e., BCL2/BAX, KRAS, BRAF, IL-6, and IL-10), respectively. Followed by, the cellular cytotoxicity of the formulations were analyzed in murine colon cancer cell line (CT26), human colorectal adenocarcinoma cell line (HT29), and normal human embryonic kidney cells (HEK-293). All four formulations were tested for their immunomodulatory potential by western blot analysis in in CT26 cell line. Furthermore, E. coli growth inhibition, and T-cell and B-cell proliferation were examined by ex-vivo conditions. RESULTS: Formulation F3B showed maximal cytotoxicity in CT26 cell line however it considered for animal experiments. Thereafter, in-vivo experiments were conducted using adult female BALB/c mice to investigate the modulation of gut dysbiosis. In the present study, animals were divided in three groups (normal control, diseases control, treatment control) while each group consisted of n = 6 animals. DISCUSSION: Our findings demonstrate that the proposed formulations exhibit anti-proliferative, mild pro-apoptotic, immunomodulatory, and antimicrobial effect. Based on 16S rRNA sequencing results, F3B demonstrated better outcomes toward managing gut dysbiosis in female BALB/C mice. However, we can conclude that formula 3FB can be used as preventive approach to control gut dysbiosis and possible colon cancer relapse.