Novel role of APOBEC3 enzymes as key mediators of cisplatin sensitivity through aberrant processing of interstrand crosslinks

APOBEC3 酶通过链间交联的异常加工作为顺铂敏感性关键介质的新作用

• 批准号: 10368997
• 负责人: Steve M Patrick
• 金额: $ 36.67万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10368997
• 关键词: Affect; Antineoplastic Agents; Apoptosis; Base Excision Repairs; Cancer Patient; Carboplatin; Cell Line; Chemotherapy-Oncologic Procedure; Cisplatin; Clinical; Clinical Data; Cytidine Deaminase; Cytosine; DNA; DNA Adducts; DNA Interstrand Crosslinking; DNA Repair; DNA Repair Inhibition; DNA Repair Pathway; DNA-(apurinic or apyrimidinic site) lyase; Data; Deaminase; Deamination; Dependence; Development; Drug resistance; Drug toxicity; Enzymes; Excision; Family; Family member; Generations; Goals; Health; Hypersensitivity; Malignant Neoplasms; Malignant neoplasm of ovary; Mediating; Mediator of activation protein; Mismatch Repair; Modeling; Mutation; Normal Cell; Nucleotide Excision Repair; Nucleotides; Pathway interactions; Patient-Focused Outcomes; Patients; Pharmaceutical Preparations; Phenotype; Platinum; Play; Polymerase; Process; Protein Family; Protein Overexpression; Proteins; Resistance; Role; Sales; Sampling; Site; Structure; Testing; Treatment Protocols; Uracil; Vertebral column; Work; analog; base; cancer drug resistance; cancer therapy; chemotherapy; cisplatin-DNA adduct; clinically relevant; crosslink; design; drug development; drug efficacy; homologous recombination; malignant breast neoplasm; member; novel; patient response; patient stratification; phosphodiester; recruit; response; tumor; uracil-DNA glycosylase

Cisplatin has been used clinically to treat a variety of cancers for nearly four decades. Along with second and third generation platinum analogs, it is still one of the most widely used chemotherapeutic drugs with over four billion dollars in annual sales. Despite its wide use, clinical limitations including drug toxicity to normal cells and the development of drug resistance in cancers has limited the impact on cancer treatment. Understanding how to overcome these clinical limitations is critical for achieving better cancer responses and increasing overall patient survival. To this end, we have established a novel mechanistic model in which two specific DNA repair pathways, base excision repair (BER) and mismatch repair (MMR), work in conjunction to mediate cisplatin efficacy. This mechanistic model is based on the specific cis/carboplatin DNA interstrand crosslink (ICL) structure in which extrahelical cytosines that flank the ICL are targets for deamination. In this proposal, we have novel preliminary data that supports a family of proteins, APOBEC3 cytidine deaminases, in initiating the deamination of the extrahelical cytosines adjacent to the cisplatin ICLs and activating the BER pathway. Following deamination of the cytosines to uracil, the BER pathway is initiated by uracil DNA glycosylase (UNG) cleavage to produce an abasic site that is subsequently processed by AP endonuclease (APE1) to cleave the phosphodiester backbone adjacent to the cisplatin ICL. Polymerase beta (Polβ) is recruited to the 3'-OH site and synthesizes DNA downstream of the cisplatin ICL, but with poor fidelity. We were the first to demonstrate a dependence on Polβ nucleotide misincorporation to activate MMR, which ultimately inhibits productive ICL DNA repair and maintains cisplatin sensitivity. In our preliminary data, we demonstrate a dependence on APOBEC3 expression to mediate cis/carboplatin sensitivity and activate the BER response. This is further supported by clinical data in which high APOBEC3 expression can mediate a clinical response to cisplatin. We also demonstrate that mutations in Polβ that affect polymerase activity result in hypersensitivity to cisplatin as a consequence of enhanced inhibition of ICL DNA repair. This suggests that the clinically relevant mutations in Polβ, which have been observed in ~30% of tumors tested, that alter polymerase function (e.g., decreased catalytic activity and/or decreased fidelity) may be beneficial for better clinical response to cis/carboplatin treatment as a result of the futile processing of cis/carboplatin ICLs. Here, we propose to (i) elucidate the APOBEC3 family members involved in cisplatin sensitivity and ICL cytosine deamination, (ii) assess the interplay between APOBEC3 members and BER/MMR proteins in cisplatin ICL processing and (iii) identify clinical Polβ mutations that mediate cisplatin efficacy and determine the dependence on APOBEC3 activity. Therefore, this project will provide a comprehensive mechanistic model for how APOBEC3 proteins activate BER/MMR to maintain cis/carboplatin efficacy and help establish a new paradigm in cis/carboplatin chemotherapy utilizing specific Polβ mutations and altered APOBEC3 expression for patient stratification.

顺铂在临床上用于治疗多种癌症已有近四十年的历史。与第二个和 第三代铂类似物，仍然是使用最广泛的化疗药物之一，有超过四种 年销售额十亿美元。尽管其用途广泛，但临床局限性包括对正常细胞的药物毒性和 癌症耐药性的发展限制了对癌症治疗的影响。了解如何 克服这些临床限制对于实现更好的癌症反应和提高总体水平至关重要 患者生存。为此，我们建立了一种新颖的机制模型，其中两种特定的DNA修复 通路、碱基切除修复 (BER) 和错配修复 (MMR) 协同作用来介导顺铂 功效。该机制模型基于特定的顺式/卡铂 DNA 链间交联 (ICL) ICL 侧翼的螺旋外胞嘧啶是脱氨基作用的目标结构。在这个提案中，我们 拥有新的初步数据支持 APOBEC3 胞苷脱氨酶蛋白家族在启动 与顺铂 ICL 相邻的螺旋外胞嘧啶脱氨并激活 BER 通路。 胞嘧啶脱氨基为尿嘧啶后，尿嘧啶 DNA 糖基化酶 (UNG) 启动 BER 途径 裂解产生脱碱基位点，随后由 AP 核酸内切酶 (APE1) 加工以裂解 与顺铂 ICL 相邻的磷酸二酯主链。聚合酶 β (Polβ) 被招募到 3'-OH 位点 并在顺铂 ICL 下游合成 DNA，但保真度较差。我们是第一个展示 依赖 Polβ 核苷酸错误掺入来激活 MMR，最终抑制生产性 ICL DNA 修复并维持顺铂敏感性。在我们的初步数据中，我们证明了对 APOBEC3 表达可介导顺式/卡铂敏感性并激活 BER 响应。这进一步 临床数据支持 APOBEC3 高表达可以介导对顺铂的临床反应。我们 还证明影响聚合酶活性的 Polβ 突变会导致对顺铂过敏 ICL DNA 修复抑制增强的结果。这表明临床相关突变 Polβ，已在约 30% 的测试肿瘤中观察到，它改变聚合酶功能（例如，减少 催化活性和/或保真度降低）可能有利于顺式/卡铂更好的临床反应 由于顺式/卡铂 ICL 无效加工而导致的治疗。在此，我们建议 (i) 阐明 APOBEC3 家族成员参与顺铂敏感性和 ICL 胞嘧啶脱氨作用，(ii) 评估 APOBEC3 成员与 BER/MMR 蛋白在顺铂 ICL 处理中的相互作用，以及 (iii) 识别 临床 Polβ 突变介导顺铂疗效并决定对 APOBEC3 活性的依赖性。 因此，该项目将为 APOBEC3 蛋白如何激活提供全面的机制模型 BER/MMR 可维持顺式/卡铂疗效并帮助建立顺式/卡铂的新范例 利用特定 Polβ 突变和改变 APOBEC3 表达进行患者分层的化疗。