Biology subjects

Murugesan, M.

Publications and source records attributed to Murugesan, M..

4 recordsLinked to original sources

bioRxiv2026

Serum free expansion and transduction of human Vγ9δ2 T cells for adoptive immunotherapy

Background and AimGamma delta T cells are multivalent immune cells with innate and adaptive features that sense a broad spectrum of tumor-associated stress patterns to lyse them without prior potentiation. They are therefore a good option for developing an off-the-shelf therapeutic immune cell product with antitumor functions. Therefore, we aimed to develop an optimized cGMP compatible protocol for expanding V gamma 9-V delta 2 T cells in a serum-free media for adoptive immunotherapy applications. Methods and ResultsPeripheral blood mononuclear cells (PBMC) from healthy donors were activated with Zoledronic Acid (ZOL) and IL-2 for 14 days. Cell proliferation, fold expansion, and phenotype were monitored. Among the regular donors, the baseline levels (day 0) of CD3+Vgamma9+delta2+ T cells were (05.10+/-0.74%). There was a robust expansion of V gamma 9-V delta 2 T cells in the serum-free media (110+/-29.89-fold). During processing, an abrupt reduction of {beta} T cells was observed as early as day +07. After two weeks, 87.82+/-5.11% (n=8) of T cells were CD3+Vgamma9+ in Optimizer with 97.15+/-0.7% of the CD3+Vgamma9+ T cells positive for delta2. The CD3+Vgamma9+ NKG2D+ increased during expansion, reaching 93.76+/-1.55% expression on day +14. V gamma 9-V delta 2 T cells expanded in the serum-free (Optimizer) media had relatively reduced variance and reduced over all yield and fold expansion but comparable percentage of Vgamma9+ and {beta} TCR+ T cells on day 14. A flow cytometry-based tumor-toxicity assay gauged the antitumor functions against K562 cell lines. Pretreatment of K562 cells with ZOL differentially enhanced (2.48+/-0.76 fold) the cytotoxic capacity of V gamma 9-V delta 2 T cells in a donor-dependent manner. The conditions for lentiviral transduction and transgene expressions of V gamma 9-V delta 2 T cells were improvised as gauged by GFP expression driven by CMV promoter (39.09+/- 8.94%) without compromising the viability. ConclusionWe have optimized a cGMP compatible protocol for expansion of human V gamma 9-V delta 2 T cells in a serum-free media with high purity and viability in as early as 07 days. Pretreatment of target tumor cells with ZOL enhanced the cytotoxicity, revealing the impact of V gamma 9-V delta 2 T cell intrinsic factors in tumor lysis. This protocol, may be scaled up for clinical translation for adoptive immunotherapy.

bioengineering↗

bioRxiv2025

Single Nuclei-Derived Molecular Subtypes of Gastrointestinal Stromal Tumors Correlate with Clinicopathologic Features and Predict Clinical Outcomes

Historically, gastrointestinal stromal tumor (GIST) has been subtyped by oncogenic driver mutations. However, tumors with the same mutational profile can have variable biology. To further explore the impact of molecular diversity on GIST biology, we performed single nucleus RNA sequencing on 16 primary GIST and utilized an integrated single cell atlas of the normal GI tract composed from multiple publicly available datasets to identify six distinct GIST cell states. We then statistically estimated the relative abundances of these profiles in bulk transcriptomic data. These were used to define six common GIST molecular subtypes based upon one or two predominant tumor cell states. We found that these molecular subtypes correlate with tumor locations, mutational profiles, and patient outcomes, and validated these subtypes in an independent international cohort. These molecular subtypes have the potential to be used for clinical prognostication for patients with GIST, identifying new therapeutic targets, and studying the cell of transformation of GIST.

cancer biology↗

bioRxiv2022

Correlating the differences in the receptor binding domain of SARS-CoV-2 spike variants on their interactions with human ACE2 receptor

Spike protein of SARS-CoV-2 variants play critical role in the infection and transmission through its interaction with hACE2 receptor. Prior findings using molecular docking and biomolecular studies reported varied findings on the difference in the interactions among the spike variants with hACE2 receptor. Hence, it is a prerequisite to understand these interactions in a more precise manner. To this end, firstly, we performed ELISA with trimeric spike proteins of Wild (Wuhan Hu-1), Delta, C.1.2 and Omicron variants. Further, to study the interactions in a more specific manner by mimicking the natural infection, we developed hACE2 receptor expressing HEK-293T cell line and evaluated binding efficiencies of the variants and competitive binding of spike variants with D614G spike pseudotyped virus. In lines with the existing findings, we observed that Omicron had higher binding efficiency compared to Delta in both ELISA and Cellular models. Intriguingly, we found that cellular models could differentiate the subtle differences between the closely related C.1.2 and Delta in their binding to hACE2. From the analysis in receptor binding domain (RBD) revealed that a single common modification, N501Y, present in both Omicron and C.1.2 is driving the enhanced spike binding to the receptor and showed two-fold superior competitive binding than Delta. Our study using cellular model provides a precise method to evaluate the binding interactions between spike sub-lineages to hACE2 receptors and signifies the role of single common modification N501Y in RBD towards imparting superior binding efficiencies. Our approach would be instrumental in understanding the disease progression and developing therapeutics. Author SummarySpike proteins of evolving SARS-CoV2 variants demonstrated their signature binding to hACE2 receptor, in turn contributed to driving the infection and transmission. Prior studies to scale the binding efficiencies between the spike variant and the receptor had consensus in distinct variants, but discrepancies in the closely related ones. To this end, we compared spike variants-receptor interactions with ELISA, from cells expressing hACE2 receptor. Intriguingly, we found that cellular models could differentiate the subtle differences between the closely related C.1.2 and Delta in their binding to hACE2. More importantly, competitive binding studies in presence of pseudovirus, demonstrated that a single common modification, N501Y, present in both Omicron and C.1.2 showed two fold superior competitive binding than Delta. Collectively, our study suggests a precise approach to evaluate the binding interactions between spike sub-lineages to hACE2 receptor. This would be instrumental in understanding the disease progression and developing therapeutics.

pharmacology and toxicology↗

bioRxiv2022

Re-sensitising XDR biofilms using a combination of bacteriophage cocktails and colistin: a natural approach

Persistent antibiotic use results in the rise of antimicrobial resistance with limited or no choice for multidrug resistant (MDR) and extensively drug resistant (XDR) bacteria. This necessitates a need for alternative therapy to effectively combat clinical pathogens that are resistant to last resort antibiotics. The study investigates hospital sewage as a potential source of bacteriophages to control MDR/XDR bacterial pathogens. 81 samples were screened for phages against selected clinical pathogens. 10 phages were isolated against A. baumannii, 5 phages against K. pneumoniae and 16 phages obtained against P. aeruginosa. The novel phages were observed to be strain-specific with a complete growth inhibition of up to 6 hrs. Phage plus colistin combinations further reduced the MBEC of colistin up to 16 folds. Notably, cocktail of phages exhibited supreme efficacy with complete killing at 0.5-1 {micro}g/ml colistin concentrations. Thus, phages specific to clinical strains has a higher edge in treating nosocomial pathogens with their proven anti-biofilm efficacy. In addition, analysis of phage genomes revealed close phylogenetic relations with phages reported from Europe, China and other neighbouring countries. This study serves as a reference and can be extended to other antibiotics and phage types to assess optimum synergistic combinations to combat various drug resistant pathogens in the ongoing AMR crisis.

microbiology↗