Abstract A029: c-MYC/BCL2 induced replication stress drives dependence on POLQ-mediated microhomology-mediated end joining in aggressive large B-cell lymphoma
Cecilia Ayala-Zambrano, Dongni Yi, Michelle Manske, Kimberly Gwin, Richar Kandasamy, Surendra Dasari, Anne Novak, Shuhei Asada, Xiaosheng Wu, Zhenkun Lou, Thomas Witzig, Jithma AbeykoonAbstract
Introduction:
Large B-cell lymphoma (LBCL) harboring chromosomal rearrangements (double-hit, DH) or coexpression of MYC and BCL2 (double-expressor, DE) represents biologically aggressive subgroups with inferior clinical outcomes. Oncogene-driven DNA replication stress (ODRS) is a hallmark of MYC and BCL2 dysregulation; however, the DNA damage response (DDR) pathways that sustain replication and cell survival under ODRS remain incompletely defined in DLBCL. Identifying DDR dependencies induced by ODRS may reveal therapeutically actionable vulnerabilities in DH/DE-LBCL.
Methods:
We studied DH/DE-LBCL cell lines (OCILY1, SUDHL2, DOHH2), non-DH/DE-LBCL cell lines (HT, KARPAS422, HBL1), and benign retinal pigment epithelial (RPE) cells engineered to overexpress MYC and/or BCL2. ODRS and DNA damage were assessed by phosphorylation of RPA (S4/S8, S33) and H2AX histone (γH2AX), respectively, using immunofluorescence, immunohistochemistry, and western blot (WB). Activity of homologous recombination (HR), nonhomologous end-joining (NHEJ), and microhomology-mediated end joining (MMEJ) pathways were quantified with established reporter assays. POLΘ (gene: POLQ) expression, an MMEJ marker, was evaluated by qPCR and WB. DDR activity was assessed using genomic signatures from whole-exome sequencing data of 404 LBCL patients. In vitro sensitivity of LBCL cells to POLΘ and PARP inhibitors was assessed with publicly available small-molecule inhibitors using CellTiterGlo and clonogenic assays, and therapeutic efficacy was validated in a DH-LBCL xenograft model in NRG mice.
Results:
DH/DE-LBCL cells and primary tumors exhibited increased ODRS and persistent DNA damage compared with non-DH/DE-LBCL models, including accumulation of γH2AX in mitosis. Functional profiling revealed selective upregulation of MMEJ in MYC/BCL2-overexpressing cells, with minimal changes in HR or NHEJ activity. POLQ expression was significantly increased in DH/DE-LBCL cell lines and primary samples, and the ID6 signature (associated with MMEJ activity) was significantly enriched in DH-LBCL tumors compared to non-DH/DE-LBCL tumors. POLΘ inhibition demonstrated greater toxicity in DH/DE-LBCL and in MYC-overexpressing RPE cells compared to non-DH/DE-LBCL and wild-type RPEs. Combined inhibition of POLΘ and PARP resulted in marked synergy in vitro. The therapeutic efficacy of POLΘ inhibition, with the synthetic lethality of concurrent PARP inhibition, was validated in the DH-LBCL xenograft model. Concurrent POLΘ and PARP inhibition with small-molecule inhibitors induced an antitumor effect in vivo, with increased overall survival in mice compared with vehicle- or olaparib-single-agent-treated groups.
Conclusions:
ODRS in DH/DE-LBCL creates a selective dependence on MMEJ mediated by POLΘ. Therapeutic targeting of POLΘ, alone or in combination with PARP inhibition, exploits this vulnerability and represents a promising strategy for this high-risk LBCL subset. These findings provide a biological rationale for clinical translation of POLΘ-directed therapies in DH/DE-LBCL.
Citation Format:
Cecilia Ayala-Zambrano, Dongni Yi, Michelle Manske, Kimberly Gwin, Richar Kandasamy, Surendra Dasari, Anne Novak, Shuhei Asada, Xiaosheng Wu, Zhenkun Lou, Thomas Witzig, Jithma Abeykoon. c-MYC/BCL2 induced replication stress drives dependence on POLQ-mediated microhomology-mediated end joining in aggressive large B-cell lymphoma [abstract]. In: Proceedings of the Fifth AACR International Meeting on Advances in Malignant Lymphoma: From Discovery to Clinical Impact; 2026 Jun 24-27; Philadelphia, PA. Philadelphia (PA): AACR; Blood Cancer Discov 2026;7(3_Suppl):Abstract nr A029.