Chaluvally Lab

Copy number variations (CNVs) such as deletions or amplifications of chromosomal loci are common events during the initiation and progression of cancer. While the role of coding genes and proteins are well characterized in the CNVs, non-coding RNAs such as microRNAs and long noncoding RNAs (lncRNAs) part of the CNVs are understudied in the context of tumor initiation and progression. Recently we have characterized the role of two microRNAs miR569 and miR551b in the progression of cancer. Our studies identified that miR569 downregulates the expression of tumor suppressor gene TP53INP1 (Chaluvally-Raghavan et al., Cancer Cell, 2014), and miR551b activates the transcription of master regulator STAT3 in ovarian cancer (Chaluvally-Raghavan et al., Cell Reports, 2014). While CNVs altered expression of microRNAs (miRs) in tumors, the mechanism of action of miRNAs deregulated by CNVs and how they intersect with neoplastic progression remains unclear.

The long-term goal of our research is to characterize the critical factors that regulate the expression of non- coding genes, novel actions of non-coding RNAs and the sequential changes in downstream signaling associated with non-coding RNA expression in women’s cancer. Projects in the lab cover each of these avenues of research. We are expecting that our studies will lead to a greater understanding of the versatile role of both coding and noncoding RNAs, their mechanism of action and novel therapeutic targets.

My research program provided a solid foundation for trainee research in non-coding RNAs, RNA Binding Proteins (RBPs), and transcription factors (TFs) in gynecological cancers and breast cancer. Success on my projects can be traced to the recruitment and training postdoctoral fellows, clinical oncology fellows, and research associates who were succeeded with several peer-reviewed publications.

We have five major focus areas in our research.

1. RNA binding proteins in cancer

In this project, we will characterize the mechanism of how RBP identifies and interacts with its targets and affect the stability of target mRNAs, then promote translation by cooperating with post-transcriptional modifications such as capping, splicing and polyadenylation. We will apply enhanced cross-linked immunoprecipitation followed by RNA sequencing called eCLIP, which allows efficient determination of all major RNA species bound by various RBPs. Using proximity ligation assays (PLA), RNA electrophoretic mobility shift assay and polysome profiling we are characterizing the targets of RBPs and their mechanism of action in cancer cells, tumor-infiltrated immune cells and stromal cells.
Aberrant expression or functions of RBPs are associated with many diseases including cancer; however, targeting RBPs with conventional drugs has proven difficult. In an independent approach, we are collaborating with the members of the Program in Chemical Biology at MCW to develop small molecule inhibitors or proteolysis targeting chimeric (PROTAC) agents to inhibit RBPs.

2. RBP-induced membraneless organelles formation through phase separation

Membraneless organelles are biomolecular condensates which are micron-scale cellular compartments that lack membranous enclosures where a specific set of proteins and RNAs are concentrated. Membraneless organelles are increasingly recognized as regions of liquid–liquid phase separation (LLPS) that organize cellular activities in both normal and pathological condition. Our research identified that an RNA Binding Protein called Fragile-X Mental Retardation Syndrome Related Protein-1 (FXR1) promotes membraneless organelles (MLOs) formation. While the basic rules driving MLO formation are being explored, their functions in pathological contexts like cancer are still unclear. Therefore, the goal of this project is to characterize the mechanism operated through RBPs on MLO formation and the contents of MLOs and their contributions on post-transcriptional of genes and their effects on protein synthesis.

3. Oncostatin signaling in cancer

Using single-cell RNA sequencing datasets of ovarian cancer, we identified that Oncostatin M Receptor (OSMR) is distinctly expressed in ovarian cancer cells compared to the immune cells or stromal cells. OSMR is one of the receptor proteins for oncostatin M (OSM), that in humans is encoded by the OSMR gene. OSMR is a member of the type I cytokine receptor family. This protein heterodimerizes with interleukin 6 signal transducer (IL6ST; a.k.a. GP130) when it binds with OSM or heterodimerizes with interleukin 31 receptor A (IL31RA) when it binds with interleukin 31 (IL-31) to transduce oncogenic signaling. OSMR-GP130 complex along the JAK1 pathway leads to IL-6 signaling, which is linked with the activation of MAPK cascade, PI3K-AKT cascade and STAT3-induced oncogenic transcription. Compared to other cytokines and interleukins, oncostatin family proteins are understudied as a potential therapeutic target for cancer therapy. Towards the goal of targeting OSMR, we developed a monoclonal antibody that can block the interaction of OSMR with its ligand and promote the internalization and degradation of OSMR through endocytosis and patented. We are now looking for industrial partnerships for developing our patented anti-OSMR antibodies as an investigational drug for clinical trials.
Please tell which color depicts what. In bracket internalization color can be written.

4. microRNA-induced transcriptional activation

miRNAs were thought to primarily downregulate the gene expression by binding 3’UTR of target genes. Currently we are extending our research to study novel actions of microRNAs as critical modulators that can directly activate gene expression. Our recent results demonstrate that microRNAs also bind to the promoter, which in turn facilitates the recruitment of transcription factors and activates gene expression through the process called RNA activation (RNAa). We are employing biochemical and molecular biology approaches integrated with computational biology and bioinformatics tools to understand the process of RNA activation.

5. Develop RNA interference (RNAi) approaches for cancer therapy

RNA interference (RNAi) involves introducing miRNAs or siRNAs to a cell where they target, bind to, and destroy specific mRNAs—effectively shutting down the production of proteins. Over the last six years, we have made major scientific contributions in the RNAi field using anti-microRNAs and siRNAs to abrogate oncogenic mechanisms in cancer cells. We have adopted this approach pioneered by the Nobel prize-winning discovery of Drs. Craig Mello and Andrew Fire. FDA has approved the first-ever RNA interference (RNAi)-based drug (Patisiran) to treat hereditary transthyretin amyloidosis in 2018, then Givosiran and Lumasiran for treating hepatic porphyria and hyperoxaluria in the subsequent years 2019 and 2020. Excitingly, RNAi provides opportunities to target and inhibit any gene that was previously considered as undruggable. We have published several papers that RNAi approaches can be used to inhibit the growth and metastasis of breast and ovarian cancers (Cancer Cell 2014, Cell Reports 2016, Cell Reports 2019 and Cell Reports 2021).
The major hurdle of the potential use of siRNA-based therapies in the clinic is mainly due to the challenges associated with the delivery of the siRNAs. Therefore, translating this potential into a broad new family of therapeutics, it is necessary to optimize the efficacy of the RNA-based drugs. Several advances have been made in the RNAi research in the past for the improvement of oligonucleotide stability and pharmacokinetics by introducing nucleotide analogues over natural nucleotides such as phosphorothioates, 2′-O-methylation, 2′-O-allyl or 2′-deoxy-fluorouridine modification for siRNA therapy. Based on such advances in RNAi research, we are now modifying oligonucleotides in siRNAs for improved stability, target binding and target inhibition.

Chaluvally, Pradeep 05042016obgyn-3-featured
Chaluvally-Raghavan, Pradeep, PhD

Associate Professor; Linda G. and Herbert J. Buchsbaum, MD, Chair in Gynecologic Oncology
Research & Advanced Education

Geethadevi, Anjali-12212017OB-004
Geethadevi, Anjali, PhD

Postdoc Fellow - Dr. Chaluvally-Raghavan's Lab

George, Jasmine-08062018OBGYN_004
George, Jasmine, PhD

Postdoc Fellow - Dr. Chaluvally-Raghavan's Lab

12202022OBGYN-14-Anupama Nair
Nair, Anupama, PhD

Postdoc Fellow - Dr. Pradeep Chaluvally-Raghavan's Lab

Pulikkal, Ishaque 01102020OB-008-Ishaque-feature
Pulikkal Kadamberi, Ishaque

Research Associate - Dr. Chaluvally-Raghavan's Lab

Principal Investigator Title of Research Study
Awards / Publications / Presentations
Chaluvally-Raghavan, Pradeep, PhD Translational Regulation of Oncoproteins by FXR1 CNV in Ovarian Cancer
*Grant funded by Department of Defense (DoD)
Chaluvally-Raghavan, Pradeep, PhD Role of RNA activation in Tumor Progression and Metastasis
*Grant funded by NIH Cancer Institute
Chaluvally-Raghavan, Pradeep, PhD Targeting miR511b to Prevent Tumor Formation and Metastasis of Triple Negative Breast Cancer
*Grant funded by Department of Defense (DoD)