Supplementary Figure S9:
Cell proliferation assay (BrdU incorporation) demonstrates that combination of DNA-PK inhibition with
low-dose doxorubicin reduces LMS03 and LMS05 cell proliferation compared to either drug alone.
Peposertib and AZD7648 are structurally unrelated DNA-PK inhibitors. Cells were treated for 7 days
with each drug and the combinations at the indicated doses; BrdU incorporation over 24h was
measured at day 7 with a luminescence-based ELISA assay (Roche)
Funding
Leiomyosarcoma:360 Program
National Cancer Institute (NCI)
United States Department of Health and Human Services
Find out more...Sarcoma Alliance for Research through Collaboration (SARC)
Erica's Entourage
Liddy Shriver Sarcoma Initiative
LeioMyoSarcoma Direct Research Foundation
National LeioMyoSarcoma Foundation
Deutsche Forschungsgemeinschaft (DFG)
National Secretary for Higher Education, Science, Technology and Innovation of Ecuador
ARTICLE ABSTRACT
Leiomyosarcoma (LMS) is an aggressive sarcoma for which standard chemotherapies achieve response rates under 30%. There are no effective targeted therapies against LMS. Most LMS are characterized by chromosomal instability (CIN), resulting in part from TP53 and RB1 co-inactivation and DNA damage repair defects. We sought to identify therapeutic targets that could exacerbate intrinsic CIN and DNA damage in LMS, inducing lethal genotoxicity.
We performed clinical targeted sequencing in 287 LMS and genome-wide loss-of-function screens in 3 patient-derived LMS cell lines, to identify LMS-specific dependencies. We validated candidate targets by biochemical and cell-response assays in vitro and in seven mouse models.
Clinical targeted sequencing revealed a high burden of somatic copy-number alterations (median fraction of the genome altered =0.62) and demonstrated homologous recombination deficiency signatures in 35% of LMS. Genome-wide short hairpin RNA screens demonstrated PRKDC (DNA-PKcs) and RPA2 essentiality, consistent with compensatory nonhomologous end joining (NHEJ) hyper-dependence. DNA-PK inhibitor combinations with unconventionally low-dose doxorubicin had synergistic activity in LMS in vitro models. Combination therapy with peposertib and low-dose doxorubicin (standard or liposomal formulations) inhibited growth of 5 of 7 LMS mouse models without toxicity.
Combinations of DNA-PK inhibitors with unconventionally low, sensitizing, doxorubicin dosing showed synergistic effects in LMS in vitro and in vivo models, without discernable toxicity. These findings underscore the relevance of DNA damage repair alterations in LMS pathogenesis and identify dependence on NHEJ as a clinically actionable vulnerability in LMS.
