Integrated Multimodal Genomic Analyses Reveal Novel Mechanisms of Glucocorticoid Resistance in Acute Lymphoblastic Leukemia

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer.Much has been discovered in recent decades regarding ALL biology, and the outcome of patients with ALL has vastly improved, especially in pediatric ALL patients.Despite very promising overall cure rates, patients who relapse have a greatly decreased prognosis with survival rates ranging from 30-60%.These numbers stand to improve even further with new targeted therapies that seek to improve or maintain cure rates while reducing treatment related toxicities which affect patients both acutely and chronically.Glucocorticoids (GCs) are essential components of modern chemotherapeutic intervention for ALL.Resistance to glucocorticoids is an important factor in determining early treatment response and overall patient survival.Reduction of glucocorticoid induced toxicities, such as osteonecrosis, can significantly affect patient quality of life and are associated with high dose glucocorticoid treatment in pediatric patients.Both endogenous and exogenous glucocorticoids exert their mechanism of action through various pleiotropic effects that regulate numerous cellular functions and can cause selective cytotoxicity in lymphoid malignancies.The complex mechanism of action of glucocorticoids is evident in the number of diverse clinically relevant molecular pathways that have been previously associated with resistance to glucocorticoids in ALL.The identification of genomic and epigenomic mechanisms of glucocorticoid resistance are important for improving ALL treatment outcomes.We used an agnostic genome-wide method to interrogate multiple types of genomic information (mRNA and miRNA expression, DNA methylation, SNPs, CNAs and SNVs/ ABSTRACT Acute lymphoblastic leukemia (ALL) is the most common childhood cancer.Much has been discovered in recent decades regarding ALL biology, and the outcome of patients with ALL has vastly improved, especially in pediatric ALL patients.Despite very promising overall cure rates, patients who relapse have a greatly decreased prognosis with survival rates ranging from 30-60%.These numbers stand to improve even further with new targeted therapies that seek to improve or maintain cure rates while reducing treatment related toxicities which affect patients both acutely and chronically.Glucocorticoids (GCs) are essential components of modern chemotherapeutic intervention for ALL.Resistance to glucocorticoids is an important factor in determining early treatment response and overall patient survival.Reduction of glucocorticoid induced toxicities, such as osteonecrosis, can significantly affect patient quality of life and are associated with high dose glucocorticoid treatment in pediatric patients.Both endogenous and exogenous glucocorticoids exert their mechanism of action through various pleiotropic effects that regulate numerous cellular functions and can cause selective cytotoxicity in lymphoid malignancies.The complex mechanism of action of glucocorticoids is evident in the number of diverse clinically relevant molecular pathways that have been previously associated with resistance to glucocorticoids in ALL.The identification of genomic and epigenomic mechanisms of glucocorticoid resistance are important for improving ALL treatment outcomes.We used an agnostic genome-wide method to interrogate multiple types of genomic information (mRNA and miRNA expression, DNA methylation, SNPs, CNAs and SNVs/Indels) in primary human acute lymphoblastic leukemia cells.We identified 463 genomic features associated with glucocorticoid resistance.Gene-level aggregation by a novel statistical method (TAP) identified 118 overlapping genes, 15 of which were confirmed by genome-wide CRISPR screening.Upon review of known glucocorticoid resistance mechanisms, we directly identified 30 of 38 (79%) genes/miRNAs and all 38 known resistance pathways, revealing 14 of 15 of our top candidate genes were not previously associated with glucocorticoid-resistance. CELSR2, the top novel gene downregulated in glucocorticoid resistant ALL was corroborated by single cell RNAseq and network-based transcriptomic modeling (NetBID).shRNA knockdown of CELSR2 recapitulated glucocorticoid resistance in human leukemia cell lines and revealed a synergistic drug combination (prednisolone and venetoclax), based on high BCL-2 expression, that was able to mitigate glucocorticoid resistance in mouse xenografts.In summation, we illustrated the power of a multi-dimensional integrative genomic strategy for elucidating genes and pathways conferring glucocorticoid resistance in patients with ALL.These findings will provide important new targets for treating glucocorticoid resistant ALL.vii