Therapeutic Vaccine for HIV/HPV-associated Oropharyngeal and Tonsillar Malignanci

描述（由申请人提供）：本项目的目的是进一步开发基于HPV在发病机制中的作用的HIV相关恶性肿瘤的免疫策略，并使用新药进行临床前实验，使我们能够进入临床试验以测试其安全性和有效性。HPV相关的癌症表达HPV的E6/E7癌蛋白，其是免疫诱导疫苗的理想靶标。我们开发了一种基于新型腺病毒血清型-5载体（Ad 5）平台的疫苗，该平台具有病毒DNA聚合酶和早期基因2b（E2 b）区域中的前末端蛋白的独特和额外缺失（Ad 5 [E1-，E2 b-]）。在使用传染病和癌症抗原的研究中，我们报道了当用于在多重同源免疫方案中诱导CMI应答时，新的Ad 5 [E1-，E2 b-]载体疫苗上级于当前的Ad 5 [E1-]载体疫苗（在早期基因1（E1）区域中含有缺失）。我们已经证明，尽管存在预先存在的Ad 5免疫，但在小鼠和猴中诱导了显著的抗原特异性CMI应答。我们的数据表明，新的Ad 5 [E1-，E2 b-]载体诱导针对肿瘤相关抗原（TAA）的稳健CMI应答，从而产生抗肿瘤活性，即使在预先存在的Ad 5免疫的情况下也是如此。我们已经构建并产生了经修饰的HPV-E6/E7基因，其表达非致癌早期基因6（E6）和早期基因7（E7）HPV蛋白。我们已经将这种修饰的HPV-E6/E7基因整合到新的重组Ad 5 [E1-，E2 b-]载体平台（Ad 5 [E1-，E2 b-]-HPV-E6/E7）中，以用作治疗患有HPV-E6/E7表达癌症的患者的免疫剂。该重组平台通过注射直接转染抗原呈递细胞后表达修饰的HPV-E6/E7抗原来诱导免疫应答。我们评估了该药物与toll样受体激动剂（TLRa）平台的组合，该平台被设计用于增强由载体诱导的免疫应答。在采用修饰的HPV-E6/E7基因插入物的Ad 5免疫鼠癌症模型中，比较了在添加或不添加Ad 5 [El-，E2 b-]-TLRa的情况下用Ad 5 [El-，E2 b-]-HPV-E6/E7重复免疫的免疫原性和体内抗肿瘤效果。与单独用Ad 5 [E1-，E2 b-]-HPV-E6/E7免疫的小鼠相比，在用Ad 5 [E1-，E2 b-]-HPV-E6/E7加Ad 5 [E1-，E2 b-]-TLRa免疫的小鼠中诱导了显著更高水平的HPV-E6/E7指导的CMI活性。重要的是，与注射Ad 5 [El-，E2 b-]-null（空载体）的对照小鼠相比，通过添加或不添加Ad 5 [El-，E2 b-]-TLRa的Ad 5 [El-，E2 b-]-HPV-E6/E7免疫疗法治疗的小鼠经历了肿瘤生长的大幅降低。这些结果表明，使用新的Ad 5 [El-，E2 b-]-HPV-E6/E7对表达HPV-E6/E7的肿瘤的免疫治疗诱导了有效的抗肿瘤活性。我们以前的研究数据也表明，这种新的免疫抑制药物可能与化疗/放疗联合使用，以诱导有效的抗肿瘤活性。本临床前项目中的研究将使我们能够推进生产方法并获得数据，从而使我们能够推进FDA批准的临床试验

DESCRIPTION (provided by applicant): The objective of this project is to further develop an immunotherapeutic strategy for HIV-associated malignancy based on the role of HPV in the pathogenesis and perform the pre-clinical experiments with a new drug which allow us to enter clinical trials to test its safety and effectiveness. HPV related cancers express the E6/E7 oncoproteins of HPV that are ideal targets for immune inducing vaccines. We developed a vaccine based upon a novel adenovirus serotype-5 vector (Ad5) platform with unique and additional deletions of the viral DNA polymerase and the pre-terminal protein in the early gene 2b (E2b) region (Ad5 [E1-, E2b-]). In studies employing infectious disease and cancer antigens, we reported that new Ad5 [E1-, E2b-] vector vaccines are superior to current Ad5 [E1-] vector vaccines (containing deletion in the early gene 1 (E1) region) when used to induce CMI responses in a multiple homologous immunization regimen. We have demonstrated that significant antigen specific CMI responses are induced in mice and monkeys despite the presence of pre- existing Ad5 immunity. Our data indicates that the new Ad5 [E1-, E2b-] vectors induces robust CMI responses against tumor associated antigens (TAA) resulting in anti-tumor activity, even in the presence of pre-existing Ad5 immunity. We have constructed and produced a modified HPV-E6/E7 gene that expresses non-oncogenic early gene 6 (E6) and early gene 7 (E7) HPV proteins. We have incorporated this modified HPV-E6/E7 gene into the new recombinant Ad5 [E1-, E2b-] vector platform (Ad5 [E1-, E2b-]-HPV-E6/E7) to be used as an immunotherapeutic for the treatment of patients with HPV-E6/E7 expressing cancers. This recombinant platform induces immune responses by expressing the modified HPV-E6/E7 antigens after direct transfection of antigen presenting cells via injection. We evaluated this dru combination with a toll-like receptor agonist (TLRa) platform designed to enhance immune responses induced by the vector. In an Ad5 immune murine cancer model employing the modified HPV-E6/E7 gene insert, the immunogenicity and in vivo anti-tumor effects of repeated immunizations with the Ad5 [E1-, E2b-]-HPV-E6/E7 with or without the addition of Ad5 [E1-, E2b-]-TLRa were compared. Significantly higher levels of HPV-E6/E7 directed CMI activity was induced in mice immunized with Ad5 [E1-, E2b-]-HPV-E6/E7 plus Ad5 [E1-, E2b-]-TLRa as compared with mice immunized with Ad5 [E1-, E2b-]-HPV-E6/E7 alone. Importantly, mice treated by immunotherapy with Ad5 [E1-, E2b-]-HPV-E6/E7 with or without the addition of Ad5 [E1-, E2b-]-TLRa experienced large reductions in tumor growth as compared with control mice injected with Ad5 [E1-, E2b-]-null (empty vector). These results indicate that immunotherapy of HPV-E6/E7 expressing tumors using the new Ad5 [E1-, E2b-]-HPV-E6/E7 induces potent anti-tumor activity. Our previous research data also indicates that this new immunotherapeutic drug may be used in conjunction with chemotherapy/irradiation treatment to induce effective anti-tumor activity. The studies in the present pre-clinical program will allow us to advance manufacturing approaches and acquire data that will allow us to advance to FDA approved clinical trials