Characterization and targeting BRIT1 deficiency in breast cancer

乳腺癌中 BRIT1 缺陷的表征和靶向治疗

基本信息

批准号：
8470134
负责人：
KAIYI LI
金额：
$ 30.53万
依托单位：
BAYLOR COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-05-01 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8470134
关键词：
8p23.1 Antineoplastic Agents BRCA1 gene BRCA2 gene Binding Proteins Biological Assay Biometry Breast Cancer Cell Breast Cancer Model Breast Cancer Treatment Cancer Patient Carcinogens Cell Culture Techniques Cell Survival Cells Chromatin Chromosomal Instability Chromosome abnormality Chromosomes Clinic Clinical Code Cytogenetics DNA Damage DNA Repair DNA Sequence Deletion Mutation Development Diagnostic Neoplasm Staging Drug Targeting ERBB2 gene Exons Gene Dosage Gene Proteins Genetic Transcription Genome Stability Genotoxic Stress Human Human Genome In Vitro Incidence Introns Investigation Knockout Mice Lead Location Maintenance Malignant Neoplasms Mammary Neoplasms Mammary gland Mouse Mammary Tumor Virus Mus Mutagens Mutant Strains Mice Mutation NBS1 gene Oncogenic Pathway interactions Patients Pharmaceutical Preparations Prognostic Marker Proteins RNA Radiation Reverse Transcriptase Polymerase Chain Reaction Role Sampling Sequence Deletion Signal Transduction Site Specificity Specimen Spectral Karyotyping Staining method Stains Stress Testing Therapeutic Transgenic Mice Tumor Suppression Tumor Suppressor Proteins Tumor stage ataxia telangiectasia mutated protein cancer cell cancer therapy comparative genomic hybridization dimethylbenzanthracene effective therapy genome sequencing homologous recombination improved in vivo inhibitor/antagonist irradiation malignant breast neoplasm mouse model mutant novel protein expression public health relevance ras Oncogene recombinational repair response tumor tumor xenograft tumorigenesis

8p23.1 Antineoplastic Agents BRCA1 gene BRCA2 gene Binding Proteins Biological Assay Biometry Breast Cancer Cell Breast Cancer Model Breast Cancer Treatment Cancer Patient Carcinogens Cell Culture Techniques Cell Survival Cells Chromatin Chromosomal Instability Chromosome abnormality Chromosomes Clinic Clinical Code Cytogenetics DNA Damage DNA Repair DNA Sequence Deletion Mutation Development Diagnostic Neoplasm Staging Drug Targeting ERBB2 gene Exons Gene Dosage Gene Proteins Genetic Transcription Genome Stability Genotoxic Stress Human Human Genome In Vitro Incidence Introns Investigation Knockout Mice Lead Location Maintenance Malignant Neoplasms Mammary Neoplasms Mammary gland Mouse Mammary Tumor Virus Mus Mutagens Mutant Strains Mice Mutation NBS1 gene Oncogenic Pathway interactions Patients Pharmaceutical Preparations Prognostic Marker Proteins RNA Radiation Reverse Transcriptase Polymerase Chain Reaction Role Sampling Sequence Deletion Signal Transduction Site Specificity Specimen Spectral Karyotyping Staining method Stains Stress Testing Therapeutic Transgenic Mice Tumor Suppression Tumor Suppressor Proteins Tumor stage ataxia telangiectasia mutated protein cancer cell cancer therapy comparative genomic hybridization dimethylbenzanthracene effective therapy genome sequencing homologous recombination improved in vivo inhibitor/antagonist irradiation malignant breast neoplasm mouse model mutant novel protein expression public health relevance ras Oncogene recombinational repair response tumor tumor xenograft tumorigenesis

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项目摘要

DESCRIPTION (provided by applicant): The intact and effective DNA damage response (DDR) is essential for the maintenance of genomic stability and it acts as a critical barrier to suppress tumorigenesis. We have recently identified BRIT1/MCPH1 as a novel key regulator in DDR pathway. Importantly, my lab has recently generated the BRIT1 knockout mice and demonstrated the essential roles of BRIT1 in homologous recombination DNA repair and in maintaining genomic stability in vivo. We also found significant decreases of BRIT1 gene copy number and its RNA and protein expression in multiple breast cancer lines. Significantly, we identified a BRIT1 deletion mutation in one of the 10 breast cancer specimens and this deletion led to truncation of BRIT1 protein and impaired its function in DNA damage response. Therefore, BRIT1 may function as a novel tumor suppressor for breast cancer via preserving genome stability, and targeting BRIT1 deficiency may provide novel and effective treatment for the breast cancer patients. Three specific aims are proposed to test this hypothesis: (1) To determine the effects of BRIT1 deficiency on mammary tumor formation using the BRIT1 knockout mouse. By crossing our unique BRIT1-/- mice with the MMTV-Ras transgenic mice, our initial study shows that loss of BRIT1 potentiates the Ras-induced mammary tumor development. We will compare the mammary tumor incidence, tumor grade/type and metastatic potential between MMTV-Ras/BRIT1-/- and MMTV-Ras/BRIT1+/+ mice by histological analysis. The underlying mechanisms will be investigated by analyzing the DNA repair function of major BRIT1 targets. In addition, we will determine if BRIT1 deficiency accelerates irradiation or carcinogen-induced mammary tumor development using BRIT1-conditional knockout mice. (2) To identify BRIT1 aberrations in clinical breast cancer specimens. We will identify BRIT1 aberrations from180 breast cancer samples stratified by tumor grade and HER2/ER/PR status. BRIT1 mutations in the coding region and exon/intron junction will be determined by DNA sequencing. The protein expression and subcellular location of BRIT1 will be also assessed by immunohistochemical staining, and its RNA level will be assessed using quantitative RT-PCR. In addition, we will determine if BRIT1 deficiency is correlated with HER2/ER/PR status and patient survival. (3)To develop novel treatment for BRIT1-deficient breast cancers using synthetic lethality approach. Our initial study shows that BRIT1-deficient cells are very sensitive to PARP inhibitors. To test if PARP inhibitors can serve as a potent drug targeting BRIT1-deficient breast cancers, we will systematically assess the response of the breast cancer cells to several potent PARP inhibitors in cell culture. Two mouse models carrying BRIT1-deficient mammary tumors will be used to further evaluate the efficacy of PARP inhibitors in vivo. We will also combine these inhibitors with clinic anti-cancer agents to establish the optimal therapeutic remedies. In summary, our study will contribute to an improved understanding of the key pathological alterations in breast cancer development and will provide the immediate clinic impact on the treatment of BRIT1-deficient breast cancer.

描述（由申请人提供）：完整有效的DNA损伤反应（DDR）对于维持基因组稳定性至关重要，并且是抑制肿瘤发生的关键障碍。我们最近将BRIT1/MCPH1确定为DDR Pathway中的新型关键调节剂。重要的是，我的实验室最近产生了BRIT1敲除小鼠，并证明了BRIT1在同源重组DNA修复和维持体内基因组稳定性方面的重要作用。我们还发现，在多个乳腺癌系中，BRIT1基因拷贝数及其RNA和蛋白质表达的显着降低。值得注意的是，我们在10个乳腺癌标本之一中鉴定了BIT1缺失突变，并且这种缺失导致Brit1蛋白的截断，并损害了其在DNA损伤反应中的功能。因此，BRIT1可以通过保持基因组稳定性来充当乳腺癌的新型肿瘤抑制剂，而靶向BRIT1缺乏症可以为乳腺癌患者提供新颖有效的治疗方法。提出了三个特定的目的来检验这一假设：（1）使用BRIT1敲除小鼠确定Brit1缺乏症对乳腺肿瘤形成的影响。通过将我们独特的BRIT1 - / - 小鼠与MMTV-RAS转基因小鼠跨越，我们的初步研究表明，BRIT1的损失增强了RAS诱导的乳腺肿瘤的发展。我们将通过组织学分析比较MMTV-RAS/BIRT1 - / - 和MMTV-RAS/BIRT1+/+小鼠之间乳腺肿瘤的发病率，肿瘤等级/类型和转移潜力。基本机制将通过分析主要BRIT1目标的DNA修复功能来研究。此外，我们还将确定BIRT1缺乏症是否会加速使用BIRT1-核敲除小鼠的辐照或致癌物诱导的乳腺肿瘤的发展。（2）鉴定临床乳腺癌标本中的BRIT1畸变。我们将确定180个按肿瘤级和HER2/ER/PR状态分层的乳腺癌样品的Brit1畸变。编码区域和外显子/内含子连接中的BRIT1突变将通过DNA测序确定。 BRIT1的蛋白质表达和亚细胞位置也将通过免疫组织化学染色来评估，其RNA水平将使用定量RT-PCR进行评估。此外，我们将确定BRIT1缺乏症是否与HER2/ER/PR状态和患者生存相关。（3）使用合成的致死性方法为Brit1缺陷乳腺癌开发新的治疗方法。我们的初步研究表明，BRIT1缺陷细胞对PARP抑制剂非常敏感。为了测试PARP抑制剂是否可以用作靶向BIT1缺陷乳腺癌的有效药物，我们将系统地评估乳腺癌细胞对细胞培养中几种有效PARP抑制剂的反应。两种携带BIT1缺陷乳腺肿瘤的小鼠模型将用于进一步评估PARP抑制剂在体内的疗效。我们还将将这些抑制剂与临床抗癌药结合起来，以建立最佳的治疗疗法。总而言之，我们的研究将有助于改善对乳腺癌发育的关键病理改变的理解，并将为临床对BRIT1缺陷型乳腺癌的治疗产生直接的影响。