Understanding and targeting non-genetic mechanisms of drug resistance

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PROJECT SUMMARY/ABSTRACT Drug resistance is one of the key contributing factors to treatment failure in infectious disease. Thus, overcoming drug resistance would significantly improve patient outcome and reduce healthcare costs. Extensive research on genetic mechanisms of drug resistance have revealed how drugs are rendered ineffective via changes in the DNA sequence. However, emerging evidence suggest that non-genetic mechanisms such as chromatin- and protein-based epigenetic states may explain how drug resistance arises frequently and rapidly. Understanding the non-genetic mechanisms that drive the evolution of drug resistance could pave the way for the development of novel therapeutic strategies to combat drug resistance. The Research Training Plan will leverage systems biology approaches in a leading human fungal pathogen Candida albicans to dissect the molecular underpinning of pararesistance, a non-genetic mechanism of drug resistance (Aims 1 and 2) and examine the clinical relevance and therapeutic potential of targeting pararesistance (Aim 3). In Aim 1, the applicant Dr. Jing Lin (Lucy) Xie will train in single-cell RNA sequencing and ODE modeling in the K99 phase to identify the transcriptional response and estimate the switching frequency associated with the establishment of pararesistance, developing a quantitative framework to understand how pararesistance is established. In Aim 2, Dr. Xie will train in pooled screen with CRISPRi in the K99 phase and complete the analysis in R00 phase to identify regulators of pararesistance and determine how pararesistance is maintained. In Aim 3, Dr. Xie will establish the prevalence of pararesistance in a collection of ~1,000 wild C. albicans isolates and identify chemical modulators of pararesistance in a library of >1,000 bioactive small molecules to investigate the clinical and therapeutic implications of pararesistance in the R00 phase. The Career Development Plan is designed to provide Dr. Xie with the opportunity and support to acquire additional expertise in single-cell RNA-sequencing and CRISPRi technologies and statistical and modeling methodologies, as well as additional professional development training in teaching, writing, and lab management skills. Mentor Prof. Daniel Jarosz is a leading expert in yeast prions and non-genetic inheritance. Complementary expertise is offered by co-mentor Prof. James Ferrell (quantitative biology), and advisors Prof. Ami Bhatt (clinical microbiology) and Prof. Judith Berman (antifungal drug tolerance and resistance), and collaborators Prof. Michael Hallett (single-cell technologies) and Prof. Rebecca Shapiro (CRISPR technologies). The Stanford University School of Medicine fosters a highly collaborative and supportive research environment and provides excellent infrastructures within a vibrant scientific community. In summary, Dr. Xie will receive high-quality training in research and career development, and is poised to launch her own independent research program. The proposed studies will offer mechanistic insight into non-genetic mechanisms of drug resistance and establish a framework for developing novel therapeutics that would circumvent drug resistance, with important implications for microbial pathogens.