Project Summary/Abstract The treatment for children and young adults with T-cell lymphoblastic leukemia (T-ALL) and T-cell lymphoblastic lymphoma (T-LL) has harmonized over time as T-LL patients were shown to have superior outcomes with T-ALL therapy. The recently completed Children’s Oncology Group (COG) phase 3 clinical trial AALL1231, however, found that T-LL but not T-ALL patients had improved event-free survival (EFS) and overall survival (OS) when randomized to the proteasome inhibitor bortezomib plus cytotoxic chemotherapy vs chemotherapy alone. In addition, corticosteroids were intensified on AALL1231 to eliminate prophylactic cranial radiation in most children. T-ALL patients benefited from the corticosteroid intensification and T-LL patients did not. This trial changed the treatment paradigm for T-ALL and T-LL and it is critical we identify the mechanisms underlying the differential response to therapy. For T-ALL patients, we performed comprehensive -omic analyses under R01CA193776 and an associated Gabriella Miller Kids First X01 award (X01HD100702). These studies included proteomic profiling on over 250 children with T-ALL and whole genome, whole exome, and RNA sequencing (WGS, WES, RNAseq) on over 1250 children with T-ALL. We also performed single cell genomic profiling (scRNASeq) on bone marrow from 30 T-ALL cases to understand clonal architecture and microenvironment. We now have access to tissues from over 250 children with T-LL which we will use to test our central hypothesis that specific proteomic, transcriptomic, and genomic profiles can: (1) identify patients with T-ALL and T-LL that have a higher risk of relapsing, (2) define patients that have a higher likelihood to benefit from novel therapies, and (3) delineate intrinsic (tumor) and extrinsic (microenvironment) biologic differences that lead to the differential response to therapy. We will leverage and expand our existing T-ALL data and generate new data in T-LL to test our hypothesis with the following specific aims. We will perform bulk genomic, transcriptomic, and proteomic profiling on T-LL tissues, as well as single cell profiling on T-LL bone marrow and T-ALL and T-LL CSF and compare the tumor and microenvironment in T-ALL and T-LL to identify biologic differences between the two (Aim 1). We will perform transcriptome and proteome profiling to define mechanisms of sensitivity and resistance to corticosteroids (Aim 2) and proteasome inhibitors (Aim 3) in T-ALL and T-LL and target dysregulated proteins driving resistance with small molecule inhibitors and CRISPR-Cas9 to overcome drug resistance. Impact: We are uniquely positioned to perform highly innovative studies in samples collected from children and young adults with T-ALL and T-LL treated on multi-center phase 3 trials. We will improve understanding of T- ALL and T-LL biology and define intrinsic (cancer cell) and extrinsic (microenvironment) biologic factors that distinguish the two. We will identify which T-AL...