Arming adoptively transferred T-cells with drug-loaded nanoparticles for cancer immunotherapy

Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biological Engineering, February 2016.Cataloged from PDF version of thesis. "November 2015."Includes bibliographical references (pages 95-103).In adoptive cell therapy (ACT), autologous tumor-specific T-cells isolated from cancer patients or genetically engineered lymphocytes are activated and expanded ex vivo, then infused back into the individual to eliminate metastatic tumors. A major limitation of this promising approach is the loss of ACT T-cell effector functions in vivo due to the highly immunosuppressive environment in solid tumors. Protection of T-cells from immunosuppressive signals can be achieved by systemic administration of supporting adjuvant drugs such as interleukins, chemotherapy, and other immunomodulators, but these adjuvant treatments are often accompanied by serious toxicities and may still fail to optimally stimulate lymphocytes in all tumor and lymphoid compartments. Here we propose a two-pronged approach to address this problem, namely 1) repeatedly reloading supporting drugs to T-cells and 2) extending the initial functional lifetime of drug carriers conjugated to cell surfaces before transfer. To achieve this, we developed a novel strategy to repeatedly stimulate or track ACT T-cells, using cytokines or ACT-cell-specific antibodies as ligands to target PEGylated liposomes to transferred T-cells in vivo. Using F(ab')2 fragments against a unique cell surface antigen on ACT cells (Thyl.1) or an engineered interleukin-2 (IL-2) molecule on an Fc framework as targeting ligands, we demonstrate that >95% of ACT cells can be conjugated with liposomes following a single injection in vivo. Further, we show that IL-2-conjugated liposomes both target ACT cells and are capable of inducing repeated waves of ACT T-cell proliferation in tumor-bearing mice. These results demonstrate the feasibility of repeated functional targeting of T-cells in vivo, which will enable delivery of imaging contrast agents, immunomodulators, or chemotherapy agents in adoptive cell therapy regimens. On the other hand, we identified CD45 as a non-internalizing receptor on T-cells that could be used as an anchor to block internalization of cell surface-conjugated nanoparticles. Anti-CD45 decorated nanogels consisting of IL-15 superagonists remained on T-cells surfaces for over 12 days and induced 15-fold T-cell expansion in tumors in vivo and significant tumor regression without toxicity, while equivalent doses of free IL-15Sa were lethal. These results show that anti-CD45 can be generally employed to decorate a broad array of nanoparticles to endow them enhanced stability on cell surfaces for extracelluar drug delivery, tracking, or diagnostic purposes. We also compared the efficacy of anti-Thyl.1 liposomes and anti-CD45 liposomes in delivering SB525334, an immunosuppression-reverting drug for inhibiting TGF-[beta] signaling pathway, to ACT T-cells. In the setting of pre-loading Tcells with liposomes in vitro, binding to T-cells through the non-internalizing receptor CD45 elicited greater granzyme expression in ACT T-cells systemically and particularly led to greater donor T-cell infiltration of tumors, which correlated with greater therapeutic efficacy. Nevertheless, as a proof of concept, anti-Thyl.1 liposomes allowed specific re-arming of ACT T-cells with SB525334 by in vivo targeting and slowed down tumor growth significantly compared to equivalent dose of free drug and anti-CD45 liposomes. These might provide us with insights into designing and selecting the right targeting nanoparticles or combination of them depending on the nature of drugs. All together, these results demonstrate the efficacy and specificity of surface-ligand decorated nanoparticles in enhancing in vivo persistence of transferred T-cells. These nanoparticles may be applied to significantly improve the therapeutic index of drugs in cancer immunotherapy.by Yiran Zheng.Ph. D