权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

FEN1-Nuclease-Targeted Therapy for Ewing Sarcoma

FEN1-核酸酶靶向治疗尤文肉瘤

基本信息

批准号：
10600872
负责人：
Sun Ha Choo
金额：
$ 59.16万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10600872
关键词：
Ablation Address Adolescent Affect Animals BRCA1 gene Bar Codes Biological Assay Biological Availability Biopsy Bone Tissue Bone neoplasms CRISPR library CRISPR/Cas technology Cell Death Cell Line Cells Cessation of life Child Chromosome Breakage Clinic Clustered Regularly Interspaced Short Palindromic Repeats Collection Coupled DNA DNA Binding Domain DNA Damage DNA Repair DNA biosynthesis Dependence Drug Kinetics Drug resistance EWS-FLI1 fusion protein EWSR1 gene Enzymes Ewings sarcoma FLI1 gene Family Future Genes Genetic Genetic Transcription Goals Growth Guide RNA Human In Vitro Institutional Review Boards Knock-out Legal patent Letters Libraries Malignant Childhood Neoplasm Malignant Neoplasms Measures Mitotic Molecular Molecular Medicine Mus Nude Mice Okazaki fragments Oncoproteins Operative Surgical Procedures Patients Pediatric Oncologist Pediatrics Pharmaceutical Preparations Pharmacotherapy Phenotype Primitive Neuroectodermal Tumor Principal Investigator Process Protocols documentation Radiation Reporting Research Research Project Grants Resected Resistance Sampling Signal Transduction Small Interfering RNA Specificity Statistical Methods Survival Rate Testing Therapeutic Transactivation Transplantation Tumor Cell Line Tumor Volume Xenograft procedure advanced disease aggressive therapy brca gene cancer cell cell transformation chemotherapy cytotoxic data mining endonuclease genome-wide improved in vivo in vivo evaluation inhibitor mouse model neoplastic cell novel novel therapeutics nuclease patient derived xenograft model patient prognosis preclinical evaluation preclinical study response small molecule soft tissue synergism targeted nucleases targeted treatment three dimensional cell culture tumor tumor growth tumorigenesis young adult

项目摘要

PROJECT SUMMARY Ewing Sarcoma Family of Tumors (ESFT) are bone and soft tissue cancers that affect children and adolescents. Surgery, radiation and multi-agent chemotherapy have improved patient prognosis but a therapeutic plateau has been reached for both localized and metastatic cases. Hence, there is an urgent need for novel and targeted therapeutic strategies. Towards that goal, this Multi-PI collaborative research project will investigate the hypothesis that ESFT cells are dependent on FEN1, an endonuclease that processes the 5’ flaps of Okazaki fragments during lagging strand DNA synthesis, for viability. This hypothesis is supported by: (a) Four genome- scale CRISPR library screens have independently found that ESFT cell lines are highly sensitive to FEN1- CRISPRs; (b) ESFT cells were reported to be phenotypically BRCA1-deficient, and we have shown that BRCA1/2-deficient cells are hypersensitive to FEN1 inhibition; and (c) we have found several ESFT cell lines to be sensitive to FEN1-CRISPRs, FEN1-siRNAs and two small molecular FEN1-inhibitors. A team of three principal investigators will jointly direct this project: RD Kolodner is a geneticist and an inventor of FEN1-inhibitors in documented patent applications, JYJ Wang is a cancer biologist with expertise in DNA damage response, and SH Choo is a pediatric oncologist with an ongoing IRB to conduct research with ESFT patient samples. Together, we will pursue four specific aims to investigate the ESFT-dependency on FEN1. AIM-1: To demonstrate and to quantify FEN1-essentiality in a panel of 10 ESFT cell lines by genetic ablation of FEN1 with validated siRNAs and CRISPR/CAS9, respectively. AIM-2: To determine the cytotoxic effects of small molecule FEN1 inhibitors on 10 ESFT cell lines by short-term growth and death assays and longer-term colony formation assays in 2D and 3D cultures. We will edit FEN1 in ESFT cells to express a drug-resistant FEN1R enzyme so as to demonstrate on-target effects. We will evaluate the potential synergistic interactions between FEN1-inhibitors and the chemotherapeutic drugs currently used in the clinic to treat ESFT patients. AIM-3: To investigate the mechanisms underlying ESFT dependency on FEN1 by addressing three mechanistic questions on whether (a) the EWS-FLI1 oncoprotein of ESFT induces FEN1-dependency, (b) FEN1-inhibitors cause irreversible blockade of DNA replication in ESFT cells, and (c) FEN1-inhibitors cause DNA breakage and mitotic catastrophe in ESFT cells. AIM-4: To determine the effects of FEN1 inhibitors on ESFT growth in mice. We have found that a FEN1- inhibitor (SMD154) reduced the growth of orthotopic xenografts from an ESFT cell line in athymic nude mice. The pharmacokinetics (PK) of SMD154 support its testing on orthotopic xenografts from two ESFT cell lines that show sufficiently low in vitro IC50 for this compound. We will construct 4-6 ESFT patient-derived xenografts (PDX) from biopsies and resected tumors. We will test SMD154 and future FEN1-inhibitors with optimized PKs on cell line- and patient-derived xenografts. This comprehensive preclinical study will produce a detailed understanding of the ESFT-dependency on FEN1 and pave the way towards developing FEN1-inhibitors to treat ESFT.

项目总结