{"title":"A Precision Oncology Platform to Target CDK4/6 Inhibitor Resistance","authors":"","doi":"10.1016/j.ijrobp.2024.07.069","DOIUrl":null,"url":null,"abstract":"<div><h3>Purpose/Objective(s)</h3><div>Cyclin-dependent kinase (CDK)4/6 inhibitors such as palbociclib, in combination with endocrine therapies, are commonly used as first line treatment in patients with metastatic Estrogen Receptor-positive (ER+) breast cancer (BC). While CDK4/6 inhibitors significantly improve survival, development of acquired resistance to these agents is nearly universal, commonly through loss of <em>RB1.</em> Since new drug development is a time consuming and expensive endeavor, we hypothesized that high throughput screens (HTS) employing existing drugs and drug candidates is a rapid and cost-effective process for identifying novel targeted therapies that are effective against breast tumors that have developed resistance to current standard-of-care treatments such as CDK4/6 inhibitors.</div></div><div><h3>Materials/Methods</h3><div>We used the CRISPR-Cas9 system to knockout <em>RB1</em> in a panel of ER+ cell lines, including MCF-7, to generate palbociclib-resistant models of ER+ BC. For HTS, MCF-7 <em>RB1</em><sup>-/-</sup> cells were plated to a final density of 600 cells per well in 60 μL of medium in 384-well microtiter plates and incubated with the drug library (one drug/well). Cell Tier Glo assay was used to measure cell viability after 48-hour incubation with the drug library. Top 100 hits from the primary screen were validated in a confirmatory screen using a multidose format. Xenograft studies in athymic nude mice were employed to study the <em>in vivo</em> efficacy of top hits identified in HTS.</div></div><div><h3>Results</h3><div>We carried out a HTS using Selleck and Prestwick chemical libraries comprised of over 3000 compounds, many of which are FDA approved drugs and identified several “hits” that exhibited potent <em>in vitro</em> activity in inhibiting the growth of palbociclib-resistant MCF-7 <em>RB1</em><sup>-/-</sup> cells. One of the top hits, JIB-04 (a small molecule inhibitor of Jumonji histone demethylase) was further validated in <em>in vivo</em> xenograft models. Thus, JIB-04 (50 mg/kg by oral gavage, 3 days/week) completely inhibited tumor growth (<em>P</em> < 0.05 for JIB-04 vs vehicle and <em>P</em> < 0.05 for JIB-04 vs palbociclib by unpaired, 2-tailed t-test), although palbociclib (100 mg/kg by oral gavage, daily) was completely ineffective in inhibiting growth of these tumors (<em>P</em> > 0.5 for palbociclib vs vehicle by unpaired, 2-tailed t-test). Mice did not show any evidence of toxicity or weight loss</div></div><div><h3>Conclusion</h3><div>We used a HTS to rapidly identify JIB-04, a small molecule inhibitor of Jumonji histone demethylase, as a highly effective targeted therapy for treatment of ER+ breast cancers that have developed resistance to CDK4/6 inhibitors due to loss of <em>RB1</em>. Since treatment options for patients with CDK4/6 inhibitor-resistant ER+ breast cancer are severely limited, our findings provide therapeutic rationale for developing new clinical trials for evaluating the efficacy of Jumonji histone demethylase inhibitors in patients who developed resistance to CDK4/6 inhibitors through loss of <em>RB1.</em></div></div>","PeriodicalId":14215,"journal":{"name":"International Journal of Radiation Oncology Biology Physics","volume":null,"pages":null},"PeriodicalIF":6.4000,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"International Journal of Radiation Oncology Biology Physics","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0360301624008319","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"ONCOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Purpose/Objective(s)
Cyclin-dependent kinase (CDK)4/6 inhibitors such as palbociclib, in combination with endocrine therapies, are commonly used as first line treatment in patients with metastatic Estrogen Receptor-positive (ER+) breast cancer (BC). While CDK4/6 inhibitors significantly improve survival, development of acquired resistance to these agents is nearly universal, commonly through loss of RB1. Since new drug development is a time consuming and expensive endeavor, we hypothesized that high throughput screens (HTS) employing existing drugs and drug candidates is a rapid and cost-effective process for identifying novel targeted therapies that are effective against breast tumors that have developed resistance to current standard-of-care treatments such as CDK4/6 inhibitors.
Materials/Methods
We used the CRISPR-Cas9 system to knockout RB1 in a panel of ER+ cell lines, including MCF-7, to generate palbociclib-resistant models of ER+ BC. For HTS, MCF-7 RB1-/- cells were plated to a final density of 600 cells per well in 60 μL of medium in 384-well microtiter plates and incubated with the drug library (one drug/well). Cell Tier Glo assay was used to measure cell viability after 48-hour incubation with the drug library. Top 100 hits from the primary screen were validated in a confirmatory screen using a multidose format. Xenograft studies in athymic nude mice were employed to study the in vivo efficacy of top hits identified in HTS.
Results
We carried out a HTS using Selleck and Prestwick chemical libraries comprised of over 3000 compounds, many of which are FDA approved drugs and identified several “hits” that exhibited potent in vitro activity in inhibiting the growth of palbociclib-resistant MCF-7 RB1-/- cells. One of the top hits, JIB-04 (a small molecule inhibitor of Jumonji histone demethylase) was further validated in in vivo xenograft models. Thus, JIB-04 (50 mg/kg by oral gavage, 3 days/week) completely inhibited tumor growth (P < 0.05 for JIB-04 vs vehicle and P < 0.05 for JIB-04 vs palbociclib by unpaired, 2-tailed t-test), although palbociclib (100 mg/kg by oral gavage, daily) was completely ineffective in inhibiting growth of these tumors (P > 0.5 for palbociclib vs vehicle by unpaired, 2-tailed t-test). Mice did not show any evidence of toxicity or weight loss
Conclusion
We used a HTS to rapidly identify JIB-04, a small molecule inhibitor of Jumonji histone demethylase, as a highly effective targeted therapy for treatment of ER+ breast cancers that have developed resistance to CDK4/6 inhibitors due to loss of RB1. Since treatment options for patients with CDK4/6 inhibitor-resistant ER+ breast cancer are severely limited, our findings provide therapeutic rationale for developing new clinical trials for evaluating the efficacy of Jumonji histone demethylase inhibitors in patients who developed resistance to CDK4/6 inhibitors through loss of RB1.
期刊介绍:
International Journal of Radiation Oncology • Biology • Physics (IJROBP), known in the field as the Red Journal, publishes original laboratory and clinical investigations related to radiation oncology, radiation biology, medical physics, and both education and health policy as it relates to the field.
This journal has a particular interest in original contributions of the following types: prospective clinical trials, outcomes research, and large database interrogation. In addition, it seeks reports of high-impact innovations in single or combined modality treatment, tumor sensitization, normal tissue protection (including both precision avoidance and pharmacologic means), brachytherapy, particle irradiation, and cancer imaging. Technical advances related to dosimetry and conformal radiation treatment planning are of interest, as are basic science studies investigating tumor physiology and the molecular biology underlying cancer and normal tissue radiation response.