A Leukemia Cell-Specific Coiled-Coil Protein for Treatment of Chronic Myeloid Leukemia

用于治疗慢性粒细胞白血病的白血病细胞特异性卷曲螺旋蛋白

基本信息

批准号：
10319608
负责人：
Carol S. Lim
金额：
$ 34.19万
依托单位：
UNIVERSITY OF UTAH
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：
2021-01-01 至 2025-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10319608
关键词：
Acute Lymphocytic Leukemia Affinity Amino Acids Apoptosis Apoptotic Area Bcr-Abl tyrosine kinase Binding Biology Blood Circulation Bypass Cell Line Cell physiology Cells Charge Chemicals Chromosomal translocation Chromosome abnormality Chronic Myeloid Leukemia Clinical Clinical Trials Collaborations Combination Drug Therapy Combined Modality Therapy Computer Models Dimerization Disease Drug resistance Event Experimental Leukemia Genes Genetic Diseases Goals Growth Heterodimerization Homo Homodimerization Hydrophobicity Individual Inflammatory Intravenous Lead Length Leukemic Cell Ligation Location Malignant Neoplasms Mediating Modeling Oncogenic Pain Patients Peptide Synthesis Peptides Permeability Phase Phosphotransferases Physiological Processes Production Protein Tyrosine Kinase Proteins Proteolysis Research Personnel Resistance Sampling Serum Shapes Signal Transduction Solid Specificity Technology Tertiary Protein Structure Testing Therapeutic Therapeutic Intervention Translations Tyrosine Kinase Inhibitor Variant Viral cell killing chronic myeloid leukemia cell clinically relevant design dimer drug discovery improved in vitro testing in vivo inhibitor leukemia mouse model mutant mutational status new technology novel novel therapeutic intervention pre-clinical protein oligomer resistance mutation solid state standard of care stapled peptide success targeted agent targeted treatment therapeutic protein tumorigenesis

项目摘要

Project Summary/Abstract The long-term goals of this proposal are to explore the protein dimerization interface as an area for therapeutic intervention. Protein dimerization/oligomerization is a recurring theme in biology representing the mechanism by which hundreds of proteins regulate key cellular processes such as enzymatic activity and signal transduction. This non-covalent protein homo- or heterodimerization is mediated by hydrophobicity and both shape and charge complementarity. Once thought to be undruggable, dimer interfaces are emerging as an area for powerful therapeutic intervention for inflammatory diseases, pain, genetic diseases, cancer, and other diseases. The goal of this proposal is to develop a clinically relevant small protein dimerization inhibitor. As a model, we will use our dimerization coiled-coil (cc) inhibitor of Bcr-Abl. Bcr-Abl is an example of a protein that must dimerize to enable its oncogenic activity. Bcr-Abl, results from an abnormal chromosomal translocation, manifests as a constitutively active tyrosine kinase and causes of 95% of chronic myeloid leukemias (CML). We build on our novel, computationally designed Bcr coiled-coil mutant (ccmut) that selectively dimerizes with Bcr-Abl and inhibits its activity. When virally delivered as a gene, ccmut is effective against wild-type and mutant forms of Bcr-Abl. Our ccmut specifically favors heterodimerization with Bcr-Abl to disrupt Bcr-Abl dimerization, a necessary step for oncogenesis, and thus represents a novel therapeutic strategy. We have also fused ccmut to a non-toxic cell- penetrating peptide with known leukemia cell specificity and showed that it disrupts Bcr-Abl dimerization and enters and kills leukemia cells. For this proposal, we will explore peptide stapling technologies (to increase proteolytic stability) and native chemical ligation to synthesize 2 shorter stapled peptides into a longer therapeutic protein domain. We will first computationally model possible staple locations that maintain target affinity. These stapled versions (CPP-St-ccmut) are predicted to enter cells, resist serum proteolysis, bind to Bcr-Abl and inhibit its activity. We will then test the activity of our constructs with and without TKI ponatinib to test “multidomain targeting” of Bcr-Abl in CML cell lines, CML patient samples, and a CML animal model. Aims are as follows: Aim 1: Computationally design and synthesize with solid state peptide synthesis and native chemical ligation, a leukemia-specific, stapled cc inhibitor (CPP-St-ccmut) against Bc-Abl. Aim 2: Determine cell internalization, binding, and apoptotic ability of CPP-St-ccmut candidates in leukemic cell lines including those with clinically relevant mutations that are resistant to TKIs, and cells derived from patient samples, alone and in combination with ponatinib. Aim 3: Demonstrate efficacy of CPP-St-ccmut with and without ponatinib in a simple, pre-clinical mouse model of CML (syngeneic mouse model using intravenously injected BaF/3 cells expressing drug- resistant Bcr-Abl variants, including compound mutants). Our goal is to develop a stapled protein domain targeting the protein-protein dimerization interface Bcr-Abl kinase.

项目摘要/摘要该提案的长期目标是探索蛋白质二聚化界面作为治疗领域干涉。蛋白质二聚/寡聚是代表机制的生物学的反复主题数百种蛋白质调节关键细胞过程，例如酶活性和信号转导。这种非共价蛋白质均质或异二聚化是由疏水性和形状和电荷介导的完整性。一旦被认为是不可能的，二聚体界面就会成为功能强大的领域炎症性疾病，疼痛，遗传疾病，癌症和其他疾病的治疗干预。目标该建议是开发临床上相关的小蛋白二聚化抑制剂。作为模型，我们将使用我们的二聚化盘圈（CC）抑制剂的BCR-ABL抑制剂。 bcr-abl是必须二聚到的蛋白质的一个例子启用其致癌活性。 BCR-ABL是由异常染色体易位产生的组成性活性酪氨酸激酶和95％的慢性髓样白血病（CML）的原因。我们以我们的新颖的计算设计的BCR盘绕螺旋突变体（CCMUT），可选择性地用BCR-ABL二聚体并抑制它的活动。当病毒作为基因传递时，CCMUT对BCR-ABL的野生型和突变形式有效。我们的CCMUT专门利用BCR-ABL的异二聚化来破坏BCR-ABL二聚体，这是一个必要的步骤肿瘤发生，因此代表了一种新型的治疗策略。我们还将CCMUT融合到无毒细胞 - 具有已知白血病细胞特异性的穿透胡椒，并表明它破坏了BCR-ABL二聚体和输入并杀死白血病细胞。对于此建议，我们将探索肽钉技术（以增加蛋白水解稳定性）和天然化学连接，以合成2个较短的辣椒成较长的治疗蛋白质结构域。我们将首先在计算上建模可能维持目标亲和力的主食位置。这些预测上演版本（CPP-ST-CCUMUT）将进入细胞，抵抗血清蛋白水解，与BCR-ABL结合并抑制它的活动。然后，我们将在有或没有TKI PONATINIB的情况下测试我们的构建体的活性以测试“多域在CML细胞系，CML患者样品和CML动物模型中的BCR-ABL的靶向”。目的如下：AIM 1：计算设计和与固态胡椒合成和天然化学结扎的合成针对BC-ABL的白血病特异性，分期的CC抑制剂（CPP-ST-CCUMUT）。目标2：确定细胞内在化，在白血病细胞系中CPP-ST-CCUMUT候选物的结合和凋亡能力，包括临床上对TKI具有抗性的相关突变，单独和组合源自患者样品的细胞与Ponatinib。 AIM 3：在一个简单的临床前，证明具有和不带有Ponatinib的CPP-ST-CCUMUT效率 CML的小鼠模型（使用静脉注射的BAF/3细胞表达药物 - 抗性BCR-ABL变体，包括化合物突变体）。我们的目标是开发一个分阶段的蛋白质领域靶向蛋白质蛋白二聚化界面BCR-ABL激酶。