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: 10617177
• 负责人: Steve M Patrick
• 金额: $ 2.14万
• 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 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10617177
• 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 Adduction; DNA Adducts; DNA Interstrand Cross-Link Repair; DNA Interstrand Crosslinking; DNA Repair; DNA Repair Inhibition; DNA Repair Pathway; DNA biosynthesis; 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; Mismatch Repair; Modeling; Mutation; Normal Cell; Nucleotide Excision Repair; Nucleotides; Pathway interactions; Patient-Focused Outcomes; Patients; Pharmaceutical Preparations; Phenotype; Platinum; Play; Polymerase; Process; Productivity; Protein Family; Protein Overexpression; Proteins; Resistance; Role; Sales; Sampling; Site; Structure; Testing; Therapeutic; Treatment Protocols; Uracil; Vertebral column; Work; analog; base; cancer drug resistance; cancer therapy; chemotherapy; cisplatin-DNA adduct; clinically relevant; crosslink; design; drug efficacy; drug resistance development; homologous recombination; malignant breast neoplasm; member; novel; patient response; patient stratification; phosphodiester; recruit; response; treatment 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.

顺铂已在临床上用于治疗各种癌症近四十年。沿着第二和 虽然它是第三代铂类似物，但它仍然是最广泛使用的化疗药物之一， 年销售额10亿美元。尽管其被广泛使用，但临床局限性包括对正常细胞的药物毒性， 癌症耐药性的发展限制了对癌症治疗的影响。了解如何 克服这些临床局限性对于实现更好的癌症反应和提高总体 患者生存率。为此，我们建立了一个新的机制模型，其中两个特定的DNA修复 碱基切除修复（BER）和错配修复（MMR）途径共同介导顺铂 功效该机理模型基于特异性顺式/卡铂DNA链间交联（ICL） 在这种结构中，ICL侧翼的螺旋外胞嘧啶是脱氨基作用的目标。在本提案中，我们 有新的初步数据支持一个蛋白质家族，APOBEC 3胞苷脱氨酶，在启动 与顺铂ICL相邻的螺旋外胞嘧啶脱氨基并激活BER途径。 胞嘧啶脱氨为尿嘧啶后，尿嘧啶DNA糖基化酶（UNG）启动BER途径。 切割以产生脱碱基位点，其随后被AP内切核酸酶（APE 1）加工以切割所述脱碱基位点。 磷酸二酯骨架与顺铂ICL相邻。聚合酶β（Polβ）被募集到3 '-OH位点 并在顺铂ICL下游合成DNA，但保真度差。我们是第一个展示 依赖Polβ核苷酸错误掺入激活MMR，最终抑制生产性ICL DNA修复和维持顺铂敏感性。在我们的初步数据中，我们证明了对 APOBEC 3表达介导顺式/卡铂敏感性并激活BER反应。这进一步 这得到了临床数据的支持，其中高APOBEC 3表达可以介导对顺铂的临床应答。我们 还表明影响聚合酶活性的Polβ突变导致对顺铂的超敏反应， ICL DNA修复抑制增强的结果。这表明，临床相关的突变， 在约30%的测试肿瘤中观察到Polβ，其改变聚合酶功能（例如，降低 催化活性和/或降低的保真度）可能有益于对顺式/卡铂的更好的临床反应 由于顺式/卡铂ICL的无效处理而进行治疗。在此，我们建议（i）澄清 APOBEC 3家族成员参与顺铂敏感性和ICL胞嘧啶脱氨，（ii）评估APOBEC 3家族成员对顺铂敏感性的影响。 顺铂ICL加工中APOBEC 3成员和BER/MMR蛋白之间的相互作用以及（iii）鉴定 介导顺铂疗效并决定对APOBEC 3活性依赖性的临床Polβ突变。 因此，该项目将为APOBEC 3蛋白如何激活提供一个全面的机制模型。 BER/MMR维持顺铂/卡铂疗效并帮助建立顺铂/卡铂的新范例 利用特异性Polβ突变和改变的APOBEC 3表达进行化疗以用于患者分层。