Development of sd-rxRNAs as Therapy for Retinoblastoma and Other Malignancies

描述（申请人提供）：视网膜母细胞瘤是一种儿童肿瘤，其研究为癌症生物学提供了许多见解。也许最重要的是，这项工作揭示了癌症可以在肿瘤抑制基因突变的反应中发展。肿瘤抑制突变占所有癌症突变的约80%，但与癌基因的激活突变不同，抑制突变基因产物的药物无法靶向。Cobrinik实验室最近表明，视网膜母细胞瘤的生长不仅受到视网膜母细胞瘤蛋白（RB）突变的驱动，还受到视网膜锥体前体“正常”信号通路的驱动。该正常回路包括小鼠双分钟癌基因（MDM2）、V-myc髓细胞瘤病毒相关癌基因（MYCN）、甲状腺激素受体¿2 （THRB2）和视黄酸受体γ (RXRG)核受体的极高表达。这些基因的表达对视网膜母细胞瘤细胞的生长和存活至关重要，这表明尽管它们在视网膜母细胞瘤中没有基因改变，但它们可能是有效的治疗靶点。MYCN是一个特别有吸引力的开发靶点，因为它驱动多种癌症的肿瘤发生，包括神经母细胞瘤、髓母细胞瘤和其他中枢神经系统恶性肿瘤。此外，短暂的MYC下调似乎在机体水平上具有良好的耐受性，同时显著抑制肿瘤发生。然而，目前MYCN蛋白被认为是“不可药物的”。基于sirna的药物为治疗包括癌症和视网膜疾病在内的多种疾病提供了一种新的、潜在的重要治疗范式。由于缺乏有效和无毒的体内给药系统，将这项技术应用于人类疾病的能力受到阻碍。我们已经开发了一类新的疏水修饰的RNAi化合物，“自我递送rxRNA”或sd-rxRNATM，不需要递送载体进入细胞，与传统的sirna相比，具有改进的药理学。sd-rxRNA在啮齿类动物眼睛的玻璃体内注射导致视网膜完全穿透和有效的肿瘤摄取。此外，有效的持久沉默(高达21

DESCRIPTION (provided by applicant): The study of retinoblastoma, a childhood tumor, has provided numerous insights into cancer biology. Perhaps most significantly, this work has revealed that cancers can develop in response to mutations in tumor suppressor genes. Tumor suppressor mutations account for ~80% of all cancer mutations, but unlike activating mutations in oncogenes, cannot be targeted by drugs that inhibit a mutant gene product. The Cobrinik lab has recently shown that retinoblastoma growth is driven not only by retinoblastoma protein (RB) mutations, but also by "normal" signaling circuitry of retinal cone precursors. This normal circuitry includes extremely high expression of murine double minute oncogene (MDM2), V-myc myelocytomatosis viral related oncogene (MYCN), thyroid hormone receptor ¿2 (THRB2) and retinoic acid receptor gamma (RXRG) nuclear receptor. The expression of these genes is critical to retinoblastoma cell growth and survival, suggesting that they may be effective therapeutic targets despite the fact that they are not genetically altered in retinoblastoma. MYCN is an especially attractive target for development because it drives tumorigenesis in diverse cancers, including neuroblastoma, medulloblastoma, and other CNS malignancies. Moreover, transient MYC down-regulation appears to be well tolerated at the organismal level, while dramatically suppressing tumorigenesis. However, MYCN proteins are considered to be 'undruggable' at the present time. siRNA-based drugs represent a new and potentially significant therapeutic paradigm for the treatment of multiple diseases including cancer and retinal disorders. The ability to apply this technology to human disease has been impeded by the absence of efficient and non-toxic in vivo delivery systems. We have developed a novel class of hydrophobically modified RNAi compounds, "self-delivering rxRNA" or sd-rxRNATM, that do not require a delivery vehicle to enter cells and have improved pharmacology compared to traditional siRNAs. Intravitreal administration of sd-rxRNA in a rodent eye results in complete retina penetration and efficient tumor uptake. In addition, potent long lasting silencing (up to 21 days post single injection) was demonstrated. The objective of this proposal is to develop novel sd-rxRNA based therapeutics for treatment of retinoblastoma and other cancers. We believe that sd-rxRNA based treatment of retinoblastoma is feasible and will have minimal systemic toxicity which is specifically important in the context of pediatric patients. As part of this proposal, sd-rxRNA distribution via different routes of administration (intravitreal and intra-arterial injection) and target mRNA silencing in retinoblastoma cell tumors seeded into the eye will be evaluated in vivo. In parallel, sd-rxRNA compounds against MYCN and other retinoblastoma targets (as back up) will be identified and confirmed to be efficacious in vitro. A phase II application will focus on silencing of MYCN and these other targets in a retinoblastoma model. The impact of silencing on progression of a human tumor xenograft in mouse eye will be evaluated. The most potent compounds will be moved into preclinical development with a focus on developing a data package sufficient to support filing an IND. PUBLIC HEALTH RELEVANCE: Retinoblastoma is cancer of the eye that usually begins in the retina and primarily affects young children. Current treatments include chemotherapy, radiation therapy, laser therapy and possible surgery. We have recently developed a novel class of hydrophobically modified RNAi compounds and have demonstrated potent and long lasting silencing in a rodent eye. The focus of this grant application is to evaluate the applicability of d-rxRNA to silence genes in a retinoblastoma model in vivo.