TCR Engineering of Quiescent Primary Human T Cells

描述（由申请人提供）：肿瘤细胞对抗原表达的免疫反应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引起晚期黑色素瘤患者的客观临床反应。然而，肿瘤反应性T细胞很难从大多数恶性肿瘤中获得，因此大多数癌症患者不符合ACT的条件。为了克服这一障碍，我们开发了一种TCR基因转移方法，可以提供来自任何患者的自体肿瘤反应性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。一个可能的原因是，为了有效地转导，使用的T细胞被完全激活。遗憾的是，这可能导致效应细胞无法持续和分化为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CD34盒来选择转导细胞的新方法。这种改良的CD34卡带使我们能够使用临床适用的方法快速丰富转导T细胞培养物，使重定向效应细胞达到高纯度。在这个探索性应用中，我们提出将这种CD34选择系统与慢病毒载体结合起来，以确定是否可行，以中等频率的重定向T细胞来富集转导的静止人T细胞，以获得足够的纯度用于治疗。静止重定向T细胞的功能将在体外和异种体内模型中进行评估，以确定静止TCR转导的T细胞是否优于完全激活的T细胞。这项研究代表了对静止TCR基因修饰T细胞的首次评估，该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。

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.