Title: Transcriptional responses to anticancer drugs in iPSC-derived heterogeneous cardiac cell cultures
Abstract: Doxorubicin (DOX) and 5-fluorouracil (5-FU) are commonly prescribed to treat a variety of cancers. Though efficacious, up to 48% of patients treated with DOX and 30% of patients treated with 5-FU experience adverse cardiac side effects, suggesting that risk for drug-induced cardiotoxicity (CT) may be genetically regulated. Indeed, we previously found that the genetic variants that regulate transcriptional responses (i.e., expression quantitative loci, or eQTLs) of cardiomyocytes to DOX in vitro can be used to predict individual risk for DOX-induced CT in cancer patients. We reasoned that by characterizing transcriptional responses to DOX in other cell types relevant for CT, we could identify additional drug response eQTLs to improve risk prediction. We included 5-FU in our study as it has been less explored in a gene regulatory context compared to DOX, despite also being widely prescribed and associated with cardiotoxic effects. By comparing the transcriptional effects of these two distinct compounds (an anthracycline and an antimetabolite, both of which are associated with diverse CT effects), we sought to identify general and drug-specific mechanisms of CT.
In principle, induced pluripotent stem cells (iPSCs) can differentiate into any cell type in the body, and when combined with single-cell RNA sequencing, iPSCs enable large-scale mapping of eQTLs across different contexts. The challenge is to find a way to rapidly expand the dimensionality of cell types and cell states we can characterize. To address this, we developed a guided iPSC differentiation protocol that generates a temporally and functionally diverse range of cardiac-relevant cell types in 16 days. To efficiently assay multiple CT-relevant cell types across many individuals, we modified our cardiomyocyte differentiation protocol to produce heterogeneous differentiating cultures of cardiac cell types (cardiac HDCs). Cardiac HDCs are three-dimensional, rhythmically contracting cell aggregates that include cardiomyocytes, epicardial cells, cardiac fibroblasts, endothelial cells, and hematopoietic cells, along with both ectodermal and endodermal derivatives. We independently treated cardiac HDCs from 46 individuals with 1 µM DOX and 5 µM 5-FU for 24 hours, followed by single-cell RNA sequencing to capture transcriptional responses. We targeted 12,000 cells per individual (4,000 cells per condition, plus a vehicle control) for over half a million cells (more than 20 billion reads in total). Our preliminary analyses reveal thousands of drug-responsive genes that are differentially expressed between treatment conditions, many of which show cell-type-specific expression patterns. These data allow us to identify and characterize drug response eQTLs, which facilitate the exploration of mechanisms of DOX- and 5-FU-induced CT and the development of a retrospective genetic predictor of CT in cancer patients. Ultimately, our work may enable clinicians to provide personalized cancer treatments based on patient genotype.
Our Seminar Lab Series will be offered in hybrid format:
The in-person seminar is each Tuesday during the semester from noon-1 p.m. at the Discovery Building, DeLuca Forum.
To join online, please click the zoom link below:
https://uwmadison.zoom.us/j/96958883460
*if you are asked for a passcode: 970506