DEVELOPING a Pan-Coronavirus Vaccine
Actively involved with the group led by Prof.Lbachir BenMohamed at the University of California, Irvine in developing a pan-corona virus vaccine meant for COVID-19, all emerging variants of concern of SARS-CoV-2, common cold coronaviruses, and MERS. The multi-epitope pan-corona virus vaccine is in advanced stages of pre-clinical trial now. Our group has been awarded with multiple fundings including a R01 grant by the NIH to work on this pan corona virus vaccine. (1R01AI158060-01).
Prakash S, Dhanushkodi NR, Zayou L, Ibraim IC, Quadiri A, Coulon PG, Tifrea DF, Suzler B, Amin M, Chilukuri A, Edwards RA, Vahed H, Nesburn AB, Kuppermann BD, Ulmer JB, Gil D, Jones TM, BenMohamed L. Cross-Protection Induced by Highly Conserved Human B, CD4 +, and CD8 + T Cell Epitopes-Based Coronavirus Vaccine Against Severe Infection, Disease, and Death Caused by Multiple SARS-CoV-2 Variants of Concern. bioRxiv [Preprint]. 2023 May 24:2023.05.24.541850. doi: 10.1101/2023.05.24.541850. PMID: 37292861; PMCID: PMC10245830.
Prakash S, Srivastava R, Coulon PG, Dhanushkodi NR, Chentoufi AA, Tifrea DF, Edwards RA, Figueroa CJ, Schubl SD, Hsieh L, Buchmeier MJ, Bouziane M, Nesburn AB, Kuppermann BD, BenMohamed L. Genome-Wide B Cell, CD4+, and CD8+ T Cell Epitopes That Are Highly Conserved between Human and Animal Coronaviruses, Identified from SARS-CoV-2 as Targets for Preemptive Pan-Coronavirus Vaccines. J Immunol. 2021 Jun 1;206(11):2566-2582. doi: 10.4049/jimmunol.2001438. Epub 2021 Apr 28. PMID: 33911008.
Pierre-Gregoire Coulon, Swayam Prakash, Nisha R. Dhanushkodi, Ruchi Srivastava, Latifa Zayou, Delia F. Tifrea, Robert A. Edwards, J. Figueroa Cesar, Sebastian D. Schubl, Lanny Hsieh, Anthony B. Nesburn, Baruch D. Kuppermann, Elmostafa Bahraoui, Hawa Vahed, Daniel Gil, Trevor M. Jones, Jeffrey B. Ulmer, Lbachir BenMohamed. High Frequencies of PD-1+TIM3+TIGIT+CTLA4+ Functionally Exhausted SARS-CoV-2-Specific CD4+ and CD8+ T Cells Associated with Severe Disease in Critically ill COVID-19 Patients. bioRxiv 2022.01.30.478343; doi: https://doi.org/10.1101/2022.01.30.478343
https://www.nbclosangeles.com/on-air/uci-scientists-developing-more-effective-vaccine/2510578/
Further, my research focused on accessing the molecular profiling of genes associated with herpes pathophysiology. Knowing herpes specific milieu of genes and major biological pathways and the existing knowledge of immunology will aid in the development and implementation of novel immunotherapeutic strategies. Working on various projects related to Vaccine Development against Herpes Simplex Virus (HSV) infection. Major research involvements are:
Ø Examined the efficacy of a therapeutic vaccine against ocular HSV-1 infection using HLA-A2 Transgenic rabbits.
Ø Studied the roles of PD-1 and LAG-3 blockade towards an increased frequency of functional HSV-specific memory CD8+ T cells among (i) symptomatic and asymptomatic humans, and (ii) HSV-1 infected “humanized” HLA- Transgenic rabbits. Identified how the transcription milieu changes in response to the T cell exhaustion molecules' blockade during herpes infection through high throughput single-cell RNA sequencing and bulk-RNA sequencing. Validated the functional relevance of prominent genes through Flow Cytometry, Immunohistochemistry, Western Blot, and Real-Time PCR. Accessed how the HSV-1 viral titer changes in response to PD-1 and LAG-3 specific blockade by performing ex-vivo culture on trigeminal ganglia explants.
Ø Evaluated the immune evasion mechanisms by which HSV-1 and HSV-2 employ their acute or latent genes to interfere with the host immune system during acute and latent viral infections. Further, examined the role of HSV- 1 latency-associated transcript (LAT) using the mouse model through UV-B induced reactivation. Profiled the differentially expressed genes and micro-RNAs using NanoString based gene and miRNA expression assays.
Ø Identified epitopes/peptides from the HSV seropositive individuals by the genome-based bioinformatics searches and the prediction algorithms. Immunization of these peptides induced CD8+ T cell-dependent protective immunity against ocular herpes infection and disease in “humanized” HLA-A*02:01 transgenic mice. These findings should guide the development of a safe and effective T-cell-based herpes vaccine.
These projects involved conducting experiments, method development, optimization, and troubleshooting using different molecular/immunology/cell biology techniques, data analysis and presenting data in conferences and group meetings. I have already published over 10 research articles being part of this research group since May 2018, with two lead authorship papers.
Prakash S, Roy S, Srivastava R, Coulon PG, Dhanushkodi NR, Vahed H, Jankeel A, Geertsema R, Amezquita C, Nguyen L, Messaoudi I, Burkhardt AM, BenMohamed L. Unique molecular signatures of antiviral memory CD8+ T cells associated with asymptomatic recurrent ocular herpes. Sci Rep. 2020 Aug 14;10(1):13843. doi: 10.1038/s41598-020-70673-z. PMCID: PMC7427992.
While working at the Sanjay Gandhi Postgraduate Institute of Medical Sceinces, Lucknow; I was part of two different projects which (i) Evaluated the impact of miRNAs on ESRD and renal allograft dysfunction and (ii) Evaluated the clinical and environmental factors associated with Chronic Kidney Disease of Unknown Etiology (CKDu) by the Indian Council of Medical Research and Indian Society of Nephrology. Major research outcomes were:
Ø Putative association of microRNAs with corresponding target genes and SNPs in altered renal pathophysiology: Post-transcriptional events account for nearly 97% of kidney function involving the activities of miRNA like ncRNAs. miRNA being a negative regulator may suppress or up-regulate the target gene expression. Our findings observed this, in turn, affects the vascularization process resulting in end-stage renal disease (ESRD). After exhaustive in-silico analysis, I figured out the association of 20 most significant miRNAs and their corresponding target genes with ESRD. The Real-Time PCR based genotyping revealed synergistic relation between mutant genotypes of miRNA198, let7e, miRNA296, and their corresponding target genes FGFR1, STAT3, and TGFβ1 towards susceptibility to ESRD. Further, through ELISA and western blotting, the functional relevance of the target genes were evaluated on serum and renal biopsy samples. [Funding: Indian Council of Medical Research]
DECIPHERING THE GENETICS AND PATHOLOGY ASSOCIATED WITH CHRONIC KIDNEY DISEASE OF UNKNOWN ETIOLOGY (CKDu)
Dr.Swayam Prakash while working among CKDu patients in tribal belts of India
Ø Deciphering the clinical and environmental causes behind Chronic Kidney Disease of Unknown etiology (CKDu): Performed genome-wide association study (GWAS) to decipher the potential biomarkers associated with a rather uncommon pattern of kidney disease called chronic kidney disease of unknown etiology (CKDu). The exact causes of CKDu are not known yet. However, the mortality rate associated with CKDu in different regions of the world is alarming. The GWAS involved a total of 96 participants; 56 cases and 40 controls and two covariates, i.e., eGFR and Urine analysis protein. The GWAS was performed on the AXIOM PMRA (Affymetrix) platform. Out of the 2,83,232 variants identified as polymorphic, a significantly high-risk association was found with NLRP4, PRKN, and KAZN. More-so pathway analysis revealed genes associated with insulin receptor signaling, PDGF signaling, chemokine, and cytokine signaling, interferon signaling, the B-cell receptor pathway, NOTCH signaling pathways with CKDu. Subsequently, Immunohistochemistry and ELISA performed on renal biopsy samples revealed higher expression of NLRP4, KAZN, and PRKN among CKDu patients. Also, studied the trace element analysis using MASS-Spectrometry and performed heat stress analyses to evaluate the environmental causes. This is the first study in the Indian Sub-continent looking at the effect of genes associated with CKDu which may help in shaping the future course of study. [Funding: LaRenon and Indian Society of Nephrology]
DOCTORAL RESEARCH
The research outcomes from my PhD (Thesis title: Molecular profiling of the killer immunoglobulin-like receptors (KIRs) and their HLA Class-I ligands and their clinical relevance to the end stage renal disease) suggest that an element of genetic risk exists which determines the pathogenesis of the end-stage renal disease (ESRD) and involves deregulation of NK cells, their activity being inclined towards more of an activating state. The high degree of polymorphism and the inter-individual variation in both the number and types of KIR genes can account for varied NK cell responses among individuals, which can influence the pathogenesis of the ESRD. Further, it was observed that the KIR gene and HLA ligand incompatibility might impact graft survival in solid organ transplantation. The effect of matches between KIR genes and known HLA ligands was evaluated. The presence of certain KIRs in a kidney recipient, where the donor lacked the corresponding HLA ligand was considered a mismatch. The allograft was considered matched when both KIR receptor and HLA alloantigen revealed compatibility among recipient and donor. The data revealed better survival among individuals with matched inhibitory KIR receptors and their corresponding HLA ligands. The findings concluded that the presence of an inhibitory KIR gene lead to better renal allograft survival, whereas less prolonged survival was observed with activating KIRs. Particularly susceptible, as well as protective combinations obtained were further studied for the allelic variants of KIR genes through direct sequence-based typing. This revealed prolonged renal allograft survival for subjects positive for 3DL1*0010101 and reduced survival for 3DL1*00401, and 3DS1*01301 alleles. The results suggest position 31 in the D0 domain and position 283 near the C-terminal end of the D2 domain in KIR3DL1/3DS1 molecule to be mostly affected by recombination. Interestingly bioinformatics analysis revealed D1, and D2 domains to form the ligand-binding site, and D0 domain to act as an enhancer that modulates the strength of receptor-ligand interaction. Thus, individuals possessing 3DL1*00401 allele with R31H and W283L point mutations revealed susceptible association with renal allograft survival. Identification of risk factors that influence the incidence and severity of acute rejection remains the priority of transplant biologists and the outcomes of my research contribute as an additional tool of donor-recipient matching before going for transplantation. The KIR genotyping data obtained for the northern Indian ESRD patients has been submitted to the international KIR database (www.allelefrequencies.net). I have published 7 research articles in lead authorship from my doctoral research.
