TCR Engineering of Quiescent Primary Human T Cells

描述（由申请人提供）：对肿瘤细胞表达的抗原的免疫应答可用于治疗癌症患者。最近的临床试验表明，过继性T细胞转移（ACT）联合非清髓性化疗经常导致晚期黑色素瘤患者的客观临床反应。然而，肿瘤反应性T细胞难以从大多数恶性肿瘤中获得，因此大多数癌症患者不适合ACT。为了克服这一障碍，我们开发了一种TCR基因转移方法，该方法可以从任何患者提供自体肿瘤反应性T细胞的来源。使用TCR基因修饰的T细胞的早期临床试验结果相对令人失望。一个潜在的原因是，为了有效地转导，所使用的T细胞被完全激活。令人遗憾的是，这可能使效应细胞不能持续存在并分化成体内有效抗肿瘤活性所需的“正确”表型。我们最近开发了一种基于在我们的病毒载体中包含修饰的CD 34盒来选择转导细胞的新方法。这种经修饰的CD 34盒允许我们使用临床适用的方法快速富集用于重定向效应细胞的转导的T细胞培养物至高纯度。在该探索性应用中，我们提出将该CD 34选择系统与慢病毒载体联合收割机组合以确定是否可行以中等频率的重定向T细胞富集转导的静止人T细胞至足够的纯度用于治疗。将在体外和异种体内模型中评价静息重定向T细胞的功能，以确定静息TCR转导的T细胞是否上级其完全活化的对应物。这项研究代表了对静息TCR基因修饰的T细胞的首次评估，这项研究的结果可以迅速纳入未来的TCR基因转移临床试验，为患者提供一种具有更大治疗潜力的新型自体肿瘤反应性T细胞。 公共卫生相关性：该提议与公共卫生的相关性在于产生可用于过继性T细胞转移的新型效应T细胞群。使用我们开发的新技术的组合，我们将重新定向naove静息人T细胞的特异性以识别黑素瘤/黑素细胞相关抗原酪氨酸酶;我们希望在人黑素瘤的小鼠模型中证明这些TCR基因修饰的naove T细胞比它们完全活化的对应物更上级。如果这是真的，下一步将是在治疗晚期黑色素瘤患者的临床试验中评估这些新的效应物。

DESCRIPTION (provided by applicant): Immune responses to antigens expressed by tumor cells can be exploited for treating cancer patients. Recent clinical trials have shown that adoptive T cell transfer (ACT) in combination with non-myeloablative chemotherapy frequently cause objective clinical responses in patients with advanced melanoma. However, tumor reactive T cells are difficult to obtain from most malignancies and the majority of cancer patients are thus ineligible for ACT. To circumvent this hurdle, we have developed a TCR gene transfer approach which can provide a source of autologous tumor reactive T cell from any patient. Results from early clinical trials using TCR gene-modified T cells have been relatively disappointing. One potential reason is that the T cells used, in order to be efficiently transduced, were fully activated. Regrettably, this may render the effector cells incapable of persisting and differentiating into the "correct" phenotype needed for effective anti-tumor activity in vivo. We have recently developed a novel method for selecting transduced cells based on the inclusion of a modified CD34 cassette in our viral vectors. This modified CD34 cassette allows us to rapidly enrich transduced T cell cultures for redirected effector cells to high purity using clinically applicable methodologies. In this exploratory application, we proposed to combine this CD34 selection system with lentiviral vectors to determine if it is feasible to enrich transduced quiescent human T cells, with moderate frequency of redirected T cells, to sufficient purity for therapy. The function of the resting redirected T cells will be evaluated in vitro and in a xenogenic in vivo model to determine if quiescent TCR transduced T cells are superior to their fully activated counterparts. This study represents the first evaluation of resting TCR gene-modified T cells and the results from this study could be rapidly incorporated into future TCR gene transfer clinical trials that offer patients a novel type of autologous tumor reactive T cells with greater therapeutic potential. PUBLIC HEALTH RELEVANCE: The relevance of this proposal to public health is to generate a novel type of effector T cell population that can be used for adoptive T cell transfer. Using a combination of novel technologies we developed, we will redirect the specificity of naove resting human T cells to recognize the melanoma/melanocyte associated antigen tyrosinase; we hope to demonstrate in a mouse model for human melanoma that these TCR gene- modified naove T cells are superior to their fully activated counterparts. If true, the next step would be to evaluate these novel effectors in clinical trial treating advanced stage melanoma patients.