Preclinical Toxicology: Safety Evaluation of pNGVL4a-CRT-E6E7L2 (detox) Recombinant DNA Vaccine in Mice

临床前毒理学：pNGVL4a-CRT-E6E7L2（排毒）重组 DNA 疫苗在小鼠中的安全性评价

• 批准号: 8944247
• 负责人: LALITHA IYER
• 金额: $ 74.05万
• 依托单位: SRI INTERNATIONAL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-19 至 2015-09-18
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8944247
• 关键词: Amino Acids; Antibodies; Biological Availability; CD4 Positive T Lymphocytes; CD8B1 gene; Cellular Immunity; Cervarix; Cervical Intraepithelial Neoplasia; Clinical; Clinical Trials; Codon Nucleotides; Contractor; Contracts; DNA; DNA Vaccines; Data; Detox; Development; Devices; Dose; Electroporation; Evaluation; Future; Gardasil; Genes; Genotype; Goals; HIV; Human; Human Papilloma Virus Vaccine; Human Papillomavirus; Human papilloma virus infection; Human papillomavirus 16; Immune; Immune response; Immunocompromised Host; Intramuscular; Intramuscular Injections; L2 viral capsid protein; Lesion; Licensing; Link; Malignant Neoplasms; Malignant neoplasm of cervix uteri; Manufacturer Name; Mediating; Medical; Methods; Mus; Mutation; Needles; Neoplasms; Oncogenic; Patients; Persons; Phase; Pre-Clinical Model; Prevention; Proteins; Safety; Splenocyte; System; T-Lymphocyte; Testing; Therapeutic Effect; Toxicology; Translating; Vaccinated; Vaccination; Vaccines; Virion; Virus-like particle; Work; base; calreticulin; cell bank; gene gun; genital infection; head-to-head comparison; neoplastic cell; particle; phase 1 study; pre-clinical; programs; therapeutic vaccine; tumor; vaccine delivery

pNGVL4a-hCRT/E6E7L2, is a naked DNA vaccine directed at HPV E6, E7, and L2 proteins, with the goal to develop a therapeutic vaccine capable of inducing cellular immunity to eliminate HPV infection and HPV-associated lesions. The newly licensed HPV vaccines, Gardasil® and Cervarix®, both based upon L1 virus-like particles (VLPs), are effective only for prevention of new genital infections by the HPV genotypes included in the vaccines. Unfortunately, the L1 VLP vaccines do not alter the course of established HPV infections, and thus will not reduce rates of HPV-related cancer for several decades; additionally, they are not effective against all oncogenic HPV types. There is a clear unmet need for a broadly protective HPV vaccine that is also able to trigger the clearance of established HPV infections and associated neoplasia, particularly in immunocompromised persons such as human immunodeficiency virus (HIV)+ patients who have higher rates of HPV-related cancers. pNGVL4a-hCRT/E6E7L2 was generated by fusing human calreticulin (CRT) to HPV16 E6 and E7 genes, each containing multiple inactivating mutations and codons optimized for high level expression, and capsid L2 protein amino acid residues 11−200. In proof-of-concept (PoC) studies in mice, vaccination with the pNGVL4a-hCRT/E6E7L2 DNA vaccine was shown to be capable of generating a strong E6/E7-specific CD8+ T cell immune response that resulted in a potent therapeutic effect against E6/E7-expressing TC-1 tumor cells. Furthermore, significant L2-specific antibodies were generated that were able to neutralize HPV 16 virions. In another PoC study in mice, vaccination with hCRTE6E7L2 DNA elicited a potent antitumor effect against E7-expressing tumors even in mice depleted for CD4+ T cells, suggesting that this vaccine may be active even in immune-compromised hosts. A head-to-head comparison of various administration methods for delivery of a DNA vaccine encoding CRT linked to HPV16 E7 (pNGVL4a-CRT/E7) was performed in a preclinical model, and tested with conventional intramuscular (IM) needle administration, electroporation (EP)-mediated IM delivery, and particle-mediated epidermal delivery via gene gun. Results showed that EP-mediated IM delivery generated higher levels of circulating protein and higher numbers of E7-specific CD8+ T cells among splenocytes compared to conventional IM injection. The enhanced CD8+ T cell immune response detected in EP-vaccinated mice translated into superior antitumor effects in E7-expressing tumors in tumor-bearing mice suggesting that EP-mediated delivery may provide an advantage compared to conventional IM delivery of the vaccine. Based on this promising preclinical data, a contract manufacturer has generated a GMP Master Cell Bank and manufactured clinical grade pNGVL4a-hCRT/E6E7L2 DNA. A toxicology study to assess the safety of pNGVL4a-hCRT/E6E7L2 DNA vaccine in mice when administered using the TriGrid Delivery System (TDS)-IM EP device (Ichor Medical Systems, Inc.) is required to support a planned clinical trial evaluating the safety and feasibility of DNA vaccination of patients with HPV16-associated cervical intraepithelial neoplasia (CIN) 2/3, as part of the Cervical Cancer SPORE program.

PNGVL4A-HCRT/E6E7L2是一种针对HPV E6，E7和L2蛋白的裸DNA疫苗，其目标是开发能够诱导细胞免疫学以消除HPV感染并与HPV相关的病态的治疗疫苗。新许可的HPV疫苗Gardasil®和Cervarix®均基于L1病毒样颗粒（VLP），仅对预防疫苗中包含的HPV基因型的新生殖器感染有效。不幸的是，L1 VLP疫苗不会改变已建立的HPV感染的过程，因此几十年来不会降低与HPV相关的癌症的发生率。此外，它们对所有致癌HPV类型都无效。对于广泛保护的HPV疫苗的需求明显未满足，该疫苗也能够触发已建立的HPV感染和相关肿瘤的清除，尤其是在人类免疫缺陷病毒（HIV）+患者HPV相关癌症患者等免疫功能低下的人中。 PNGVL4A-HCRT/E6E7L2是通过将人钙网蛋白（CRT）与HPV16 E6和E7基因融合而产生的，每个基因都包含用于高水平表达的多重失活突变和代码子，以及Capsid L2蛋白氨基酸氨基酸残基11-200。在对小鼠的概念验证（POC）研究中，用PNGVL4A-HCRT/E6E7L2 DNA疫苗接种疫苗被证明能够产生强大的E6/E7特异性CD8+ T细胞免疫响应，从而导致对E6/E7表达表达TC-1 tumor细胞的潜在治疗作用。此外，产生了能够中和HPV 16病毒的显着L2特异性抗体。在另一项对小鼠的POC研究中，HCRTE6E7L2 DNA的疫苗即使在耗尽CD4+ T细胞的小鼠中也会引起对E7表达肿瘤的潜在抗肿瘤作用，这表明该疫苗即使在免疫敏捷的宿主中也可以活跃。在临床前模型中进行了与HPV16 E7（PNGVL4A-CRT/E7）相关的DNA疫苗的各种给药方法的正面比较。结果表明，与常规IM注射相比，EP介导的IM递送产生了更高水平的循环蛋白和较高数量的E7特异性CD8+ T细胞。在EP-VACCCCAC的小鼠中检测到的增强的CD8+ T细胞免疫增强响应在表达E7的肿瘤肿瘤中转化为超级抗肿瘤作用，这表明与传统的IM递送相比，EP介导的递送可能具有优势。基于这些承诺的临床前数据，合同制造商生成了GMP大型细胞库，并制造了临床级PNGVL4A-HCRT/E6E7L2 DNA。 A toxicology study to assess the safety of pNGVL4a-hCRT/E6E7L2 DNA vaccine in mice when administered using the TriGrid Delivery System (TDS)-IM EP device (Ichor Medical Systems, Inc.) is required to support a planned clinical trial evaluating the safety and feasibility of DNA vaccination of patients with HPV16-associated cervical intraepithelial neoplasia (CIN) 2/3，作为宫颈癌孢子计划的一部分。