ABL kinase signaling networks regulate Transcription-Replication Collisions in Small Cell Lung Cancer

About This Grant

Maintenance of genomic stability requires coordinated spatial and temporal regulation of the replication and transcription machineries on DNA. Deregulated transcription in cancer cells can promote transcription-replication conflicts (TRCs) through increased collisions with the replication machinery, thereby generating breaks on the DNA. Unexpectedly, we discovered that inhibition of the ABL tyrosine kinase enhances accumulation of TRCs in small cell lung cancer (SCLC), thereby revealing a novel role for ABL in the regulation of collisions between the transcription and DNA replication machineries. Notably, we found that ABL inactivation markedly impairs outgrowth of SCLC tumors in mouse models, resulting in prolonged animal survival. Further, ABL inactivation impairs the expression and function of proteins implicated in transcription-coupled homologous recombination (TC-HR). SCLC is a highly aggressive neuroendocrine lung cancer with an overall 5-year survival rate of only ~7%, which has remained unchanged for over 30 years. SCLC tumors exhibit a high degree of genomic instability with upregulation of proteins in the TC-HR pathway. TC-HR promotes repair of DNA double strand breaks (DSBs) at sites of active transcription and is associated with formation of R-loops comprised of DNA-RNA hybrids and displaced single-stranded DNA. Abnormal accumulation of R-loops increases TRCs, leading to enhanced DNA damage and cell death. Because ABL inactivation markedly increased TRCs and DNA damage, we evaluated whether blocking the activity DNA damage-repair pathways by targeting ATR in the presence of ABL inhibitors might synergize to promote SCLC cell death. We found that co-inactivation of ABL and ATR kinases induces profound synergistic inhibition of SCLC cell survival and impairs SCLC dissemination and outgrowth in mouse models. Here we will evaluate the hypothesis that ABL inactivation enhances SCLC vulnerabilities by increasing TRCs, resulting in enhanced DNA damage, and rendering SCLC cells hypersensitive to inactivation of components of DNA damage-repair pathways. The specific aims are: 1) Define the mechanisms by which ABL inactivation increases TRCs and DNA damage in SCLC subtypes. 2) Identify novel combination therapies to treat SCLC revealed by unbiased ABL inhibitor-mediated sensitization screens with SCLC cell lines, clinicalbiopsy- derived micro-organospheres, and SCLC metastases from mouse models treated with single and combination therapies. 3) Define the transcriptional landscapes of patient-derived SCLC subtypes, and metastatic SCLC in situ in mouse models treated with single and combination therapies. Justification for use of animal models: The use of mouse models is required for lung cancer metastasis studies to assess the contribution of multiple cell types, including immune cells, implicated in the metastatic cascade. There are no in vitro co-culture systems that faithfully mimic the complex interactions of SCLC metastases and cell types in the tumor microenvironment required for the multi-step process of metastasis observed in vivo. Completion of the proposed research will reveal novel mechanistic insights into pathways that promote SCLC metastasis, uncover the mechanisms whereby ABL signaling networks regulate TRCs, and reveal the effectiveness of co-inactivation of ABL kinase and DNA damage-repair pathways to impair dissemination and outgrowth of deadly SCLC tumors.