Tumor-targeted disruption of mismatch repair in microsatellite stable colorectal cancer

微卫星稳定结直肠癌中错配修复的肿瘤靶向破坏

基本信息

批准号：
10578049
负责人：
Judy Lieberman
金额：
$ 24.82万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-12-01 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10578049
关键词：
Affinity CD8-Positive T-Lymphocytes CXCL13 gene Carcinoma Cells Chimera organism Clinical Colorectal Cancer Colorectal Neoplasms DNA DNA Adduction DNA Adducts DNA Damage Defect Dependence Disease Epithelium Fibroblasts Gene Expression Gene Expression Profile Genes Genetic Transcription Genetically Engineered Mouse Graft Enhancements Human Immune Immune checkpoint inhibitor Immune response Immunocompetent Immunologic Deficiency Syndromes Immunologic Surveillance Immunologics Immunotherapy Implant Infiltration Inflammation Inflammatory Interferons Intervention Intestines Killer Cells LGR5 gene Link Macrophage Malignant Neoplasms Mammary Neoplasms Metalloproteases Methods Microsatellite Instability Microsatellite Repeats Minority Mismatch Repair Mismatch Repair Deficiency Modeling Mus Mutate Mutation Myeloid Cells Neoplasm Metastasis Neutrophil Infiltration Operative Surgical Procedures Organoids Outcome Pharmaceutical Preparations Phenotype Prognosis Resistance Small Interfering RNA Somatic Mutation Subcutaneous Injections T-Lymphocyte TACSTD1 gene Testing Therapeutic Tissues Tumor Antigens Tumor Immunity Tumor Tissue Tumor-infiltrating immune cells aggressive breast cancer antitumor effect aptamer cancer cell cancer diagnosis cancer infiltrating T cells cancer stem cell checkpoint therapy chemokine chemotherapy colon cancer cell line colon cancer patients design immune checkpoint blockade improved knock-down metastatic colorectal monocyte mortality neoantigens neutrophil programs promoter recruit repair enzyme repaired resistance mechanism response small molecule inhibitor stem cells subcutaneous triple-negative invasive breast carcinoma tumor tumor growth virtual

项目摘要

Colorectal cancer (CRC) is the third most diagnosed cancer in the US with the second highest mortality rate. Locally advanced and metastatic disease have a poor prognosis. A small subset (5-15%) of metastatic CRC patients, who bear tumors deficient in DNA mismatch repair (MMR) that have high microsatellite instability (MSI- high), can respond to immunotherapy with checkpoint blockade (CPI), but most CRC do not. We hypothesize that to induce immune responses in microsatellite stable (MSS) CRC, new strategies that go beyond CPI are needed. In MSI-high CRC tumors, tumor-infiltrating myeloid cells, fibroblasts and the tumors themselves all have inflammatory gene expression signatures and are colocalized into inflammatory hubs with adjacent tumor- reactive CD8 T cells, suggesting that MSI-high CRCs selectively provoke a robust inflammatory multicellular network within the tumor that recruits anti-tumor killer cells. Disruption of MMR in CRC mouse tumor grafts enhances tumor neoantigen expression and recruitment of tumor-specific T cells to trigger immune surveillance. We hypothesize that interventions that convert MSS CRC to an MMR-deficient MSI phenotype could enhance antitumor immunity and make resistant MSS CRC sensitive to CPI. This proposal will investigate whether tumor-targeted gene knockdown can convert MSS CRC to immune responsive MSI-high tumors. There are no known small molecule inhibitors of MMR. We will exploit a method of epithelial cancer-targeted gene knockdown that uses subcutaneous injection of aptamer-small interfering RNAs (AsiCs), which link a high affinity EpCAM aptamer for epithelial tumor targeting to small interfering RNAs, for tumor-selective gene knockdown. EpCAM, the first described tumor antigen, is ~100-1000-fold more highly expressed in epithelial cancers (including virtually all CRC) than normal epithelia, making it attractive for selective CRC tumor targeting. We previously showed that EpCAM-AsiCs that knockdown tumor-dependency genes or genes whose knockdown promotes immune recognition can strongly suppress aggressive breast cancer in orthotopic, metastatic and genetically engineered mouse tumor models and induce an effective immune response in immunologically cold tumors. In this proposal we will evaluate EpCAM-AsiCs to knockdown Mlh1, encoding a key MMR enzyme, in MSS CRC subcutaneous and caecal tumor implants of 4 mouse CRC cell lines and 2 MMR-proficient, MSS organoid lines derived from an aggressive genetically engineered mouse model, which conditionally express mutations in intestinal stem cells of four key genes that are frequently mutated in human CRC – Apc, Kras, Tgfbr2 and Trp53. We will evaluate the effect of MMR disruption on tumor growth in immunodeficient and immunocompetent mice and dissect in detail how microsatellite stability, tumor mutational burden and gene expression in the tumor and the immune response to the tumor are altered. We will also investigate whether CPI enhances the antitumor effect of tumor-targeted Mlh1 knockdown.

结直肠癌（CRC）是美国第三大诊断癌症，死亡率第二高。一小部分（5-15%）转移性 CRC 的局部晚期和转移性疾病预后较差。患有 DNA 错配修复 (MMR) 缺陷且具有高度微卫星不稳定性 (MSI- 高），可以对检查点阻断（CPI）的免疫疗法产生反应，但大多数结直肠癌没有反应。为了在微卫星稳定 (MSS) CRC 中诱导免疫反应，超越 CPI 的新策略是在高 MSI 的 CRC 肿瘤中，肿瘤浸润性骨髓细胞、成纤维细胞和肿瘤本身都具有这种能力。炎症基因表达特征与邻近肿瘤共定位于炎症中心反应性 CD8 T 细胞，表明 MSI 高的 CRC 选择性地激发强大的炎症多细胞肿瘤内招募抗肿瘤杀伤细胞的网络，破坏 CRC 小鼠肿瘤移植物中的 MMR。增强肿瘤新抗原的表达和肿瘤特异性 T 细胞的募集以触发免疫监视。我们努力将 MSS CRC 转化为 MMR 缺陷的 MSI 表型的干预措施可以增强抗肿瘤免疫并使耐药性 MSS CRC 对 CPI 敏感该提案将调查是否存在这种情况。肿瘤靶向基因敲除可以将 MSS CRC 转化为免疫反应性 MSI 高肿瘤。尚无已知的 MMR 小分子抑制剂，我们将开发一种针对上皮癌靶向基因的方法。使用皮下注射适配体小干扰 RNA (AsiC) 的敲低，该适配体小干扰 RNA (AsiC) 连接高亲和力用于上皮肿瘤的 EpCAM 适体靶向小干扰 RNA，用于肿瘤选择性基因敲除。 EpCAM 是第一个被描述的肿瘤抗原，在上皮癌中的表达水平高出约 100-1000 倍（包括几乎所有的结直肠癌）比正常上皮细胞，使其对选择性结直肠癌肿瘤靶向具有吸引力。先前表明 EpCAM-AsiCs 可以敲低肿瘤依赖性基因或敲低的基因促进免疫识别可以强烈抑制原位、转移性和转移性乳腺癌基因工程小鼠肿瘤模型并在免疫冷中诱导有效的免疫反应在本提案中，我们将评估 EpCAM-AsiCs 在肿瘤中敲除编码关键 MMR 酶的 Mlh1。 MSS CRC 皮下和盲肠肿瘤植入物，由 4 种小鼠 CRC 细胞系和 2 种 MMR 熟练的 MSS 组成源自攻击性基因工程小鼠模型的类器官系，其有条件地表达肠道干细胞中四个关键基因的突变，这些基因在人类结直肠癌中经常发生突变——Apc、Kras、我们将评估 MMR 破坏对免疫缺陷和肿瘤生长的影响。免疫活性小鼠并详细剖析微卫星稳定性、肿瘤突变负荷和基因如何我们还将研究 CPI 是否改变了肿瘤中的表达和对肿瘤的免疫反应。增强肿瘤靶向 Mlh1 敲低的抗肿瘤作用。