CARP-1: A Potential Therapeutic Agent for Breast Cancer

CARP-1：乳腺癌的潜在治疗剂

基本信息

批准号：
8141847
负责人：
Arun Kumar Rishi
金额：
--
依托单位：
JOHN D DINGELL VA MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-01-01 至 2015-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8141847
关键词：
Adriamycin PFS Adverse effects Antibodies Apoptosis Apoptotic Attenuated BRCA1 gene BRCA2 gene Binding Biological Assay Breast Breast Cancer Cell Breast Cancer Treatment Breast Carcinoma Cancer Cell Growth Caspase Cell Cycle Cell Death Cell Line Cell Survival Cells Development Disease Dose Drug Kinetics Drug resistance Effectiveness Epidermal Growth Factor Receptor Epitopes Estrogen Receptors Etoposide Family member Fluorescence Polarization Gefitinib Goals Growth Healthcare Human In Vitro Intravenous Investigation Knowledge Laboratories Lead MAPK14 gene MCF7 cell Malignant Neoplasms Maps Mission Mitotic Molecular Molecular Target Mus New Agents Nuclear Oncogenic Pathway interactions Pharmaceutical Preparations Phenotype Progesterone Receptors Protein p53 Proteins Proteome Reagent Resistance Resistance development Retinoic Acid Receptor Risk Factors SCID Mice Signal Transduction Tail Tamoxifen Testing Therapeutic Therapeutic Agents Transducers Tumor Suppressor Proteins Ubiquitin Veins Veterans Woman Work Xenograft Model Xenograft procedure Yeasts acronyms analog anaphase-promoting complex base cancer cell cancer therapy caspase-9 cell growth cellular transduction chemotherapy combat cyclin B1 design drug development expectation fighting genetic regulatory protein high throughput screening human MAPK14 protein in vivo inhibitor/antagonist innovation insight loss of function malignant breast neoplasm mimetics multicatalytic endopeptidase complex mutant novel outcome forecast prevent protein function response small molecule small molecule libraries stress activated protein kinase therapeutic target treatment strategy tumor ubiquitin-protein ligase upstream kinase yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): The molecular complexity of breast cancers and therapy-associated side effects often limit effectiveness of many therapies, warranting the development of new agents for specific molecular targets while minimizing the off-target effects. Our goal is to develop novel, safer, and effective human breast cancer (HBC) therapies by exploiting functions of an apoptosis regulatory protein CCAR1/CARP-1. CARP-1 functions to transduce cell growth as well as chemotherapy (adriamycin, etoposide, or Iressa)-dependent inhibitory signaling. CARP-1 expression correlates inversely with HBC tumor grades. A yeast-two-hybrid (Y2H) screen revealed that CARP- 1 binds with the E3 ubiquitin ligase subunit anaphase promoting complex (APC)-2 protein. Following mapping of the interacting epitopes of CARP-1 and APC-2, we developed a fluorescence polarization assay (FPA) and screened a chemical library to identify small molecule inhibitors (SMIs) of CARP-1:APC-2 binding. The lead SMI termed CARP-1 Functional Mimetic (CFM)-4, inhibits HBC growth in vitro and in vivo. CFM-4 also attenuates growth of tamoxifen (TAM) or adriamycin (ADR)-resistant HBC cells but does not suppress growth of the non-tumorigenic and immortalized MCF-10A cells. CFM-4 binds with CARP-1, leads to increased CARP- 1 levels, activates pro-apoptotic p38 mitogen-activated protein kinase (MAPK), caspases-9 and 3, and apoptosis. CFM-4 induces loss of mitotic cyclin B1 and Cdc20 proteins, possibly through caspase-dependent but ubiquitin proteasome pathway (UPP)-independent mechanism(s). Hypothesis: CARP-1, a peri-nuclear phospho-protein, functions to regulate HBC cell growth and apoptosis, and is a key regulator of ADR signaling. We further hypothesize that CFM-4 stimulation of CARP-1 levels will provide a novel mechanism for treating breast and other cancers. Objectives: Our long-term objective is to elucidate mechanisms of CARP-1-dependent HBC growth inhibition, and to exploit this knowledge to develop agents for targeting of HBC. The rationale for this objective is based on the facts that loss of tumor suppressor p53 promotes development of aggressive HBCs and that resistance to ADR continues to be a problem. Since CARP-1 regulates HBC growth inhibition by ADR and is a co- activator of p53, the CFMs that bind and elevate CARP-1, activate pro-apoptotic p38 MAPK and caspases, and suppress Cdc20 and cyclin B1 levels have potential to target HBCs including those that have mutant p53 as well as have resistance to ADR or TAM. We will test our hypothesis by pursuing the following objectives. (1) To investigate molecular mechanism(s) of p38 activation, and the extent to which CARP-1 binding with p38 regulates HBC growth in the presence of CFM-4 or ADR. (2) To elucidate mechanism(s) of CARP-1-dependent activation of caspases-9 and -3, and determine the extent to which activated caspases target Cyclin B1 in the presence of CFM-4. (3) To conduct pharmacokinetic (PK) profiling and to demonstrate the anti-tumor activities of CFM(s) in mouse xenograft models of HBC. Potential Impact on Veterans Health Care: This investigation will facilitate development of strategies to prevent/combat breast cancer that will benefit women in the VA workforce, contribute to Veterans Health Care and further the mission of the VA.

描述（由申请人提供）：乳腺癌的分子复杂性和治疗相关的副作用常常限制许多疗法的有效性，因此需要开发针对特定分子靶点的新药物，同时最大限度地减少脱靶效应。我们的目标是通过利用细胞凋亡调节蛋白 CCAR1/CARP-1 的功能来开发新型、更安全且有效的人类乳腺癌 (HBC) 疗法。 CARP-1 的功能是转导细胞生长以及化疗（阿霉素、依托泊苷或易瑞莎）依赖性抑制信号传导。 CARP-1 表达与 HBC 肿瘤分级呈负相关。酵母双杂交 (Y2H) 筛选显示 CARP-1 与 E3 泛素连接酶亚基后期促进复合物 (APC)-2 蛋白结合。在绘制了 CARP-1 和 APC-2 相互作用表位的图谱后，我们开发了荧光偏振测定 (FPA) 并筛选了化学库来鉴定 CARP-1:APC-2 结合的小分子抑制剂 (SMI)。主要的 SMI 称为 CARP-1 功能模拟物 (CFM)-4，可在体外和体内抑制 HBC 生长。 CFM-4 还可减弱他莫昔芬 (TAM) 或阿霉素 (ADR) 耐药性 HBC 细胞的生长，但不会抑制非致瘤性和永生化 MCF-10A 细胞的生长。 CFM-4 与 CARP-1 结合，导致 CARP-1 水平升高，激活促凋亡 p38 丝裂原激活蛋白激酶 (MAPK)、caspases-9 和 3，以及细胞凋亡。 CFM-4 可能通过半胱天冬酶依赖性但不依赖于泛素蛋白酶体途径 (UPP) 的机制诱导有丝分裂细胞周期蛋白 B1 和 Cdc20 蛋白的丢失。假设：CARP-1 是一种核周磷酸蛋白，具有调节 HBC 细胞生长和凋亡的功能，并且是 ADR 信号传导的关键调节因子。我们进一步假设 CFM-4 刺激 CARP-1 水平将为治疗乳腺癌和其他癌症提供一种新机制。目标：我们的长期目标是阐明 CARP-1 依赖性 HBC 生长抑制机制，并利用这些知识开发靶向 HBC 的药物。这一目标的基本原理是基于以下事实：肿瘤抑制因子 p53 的缺失会促进侵袭性 HBC 的发展，并且对 ADR 的耐药性仍然是一个问题。由于 CARP-1 通过 ADR 调节 HBC 生长抑制，并且是 p53 的共激活剂，因此结合并升高 CARP-1、激活促凋亡 p38 MAPK 和半胱天冬酶以及抑制 Cdc20 和细胞周期蛋白 B1 水平的 CFM 具有靶向 HBC 的潜力包括那些具有突变 p53 以及对 ADR 或 TAM 具有抗性的细胞。我们将通过追求以下目标来检验我们的假设。 (1) 研究 p38 激活的分子机制，以及在 CFM-4 或 ADR 存在的情况下 CARP-1 与 p38 结合调节 HBC 生长的程度。 (2) 阐明 caspase-9 和 -3 的 CARP-1 依赖性激活机制，并确定在 CFM-4 存在的情况下激活的 caspase 靶向 Cyclin B1 的程度。 (3) 进行药代动力学 (PK) 分析并证明 CFM 在 HBC 小鼠异种移植模型中的抗肿瘤活性。对退伍军人医疗保健的潜在影响：这项调查将促进制定预防/抗击乳腺癌的战略，这将使退伍军人事务部的女性受益，为退伍军人医疗保健做出贡献，并进一步履行退伍军人事务部的使命。