A Potent Tumor-Reactive p53-Specific Single-Chain TCR without On- or Off-Target Autoimmunity In Vivo.

Genetic engineering of T cells with a T cell receptor (TCR) targeting tumor antigen is a promising strategy for cancer immunotherapy. Inefficient expression of the introduced TCR due to TCR mispairing may limit the efficacy and adversely affect the safety of TCR gene therapy. Here, we evaluated the safety and therapeutic efficiency of an optimized single-chain TCR (scTCR) specific for an HLA-A2.1-restricted (non-mutated) p53(264-272) peptide in adoptive T cell transfer (ACT) models using our unique transgenic mice expressing human p53 and HLA-A2.1 that closely mimic the human setting. Specifically, we showed that adoptive transfer of optimized scTCR-redirected T cells does not induce on-target and off-target autoimmunity. Furthermore, ACT resulted in full tumor protection and led to a long-lived effective, antigen-specific memory T cell response in syngeneic and xenograft models. Taken together, the study demonstrated that our scTCR specific for the broadly expressed tumor-associated antigen p53(264-272) can eradicate p53+A2.1+ tumor cells without inducing off-target or self-directed toxicities in mouse models of ACT. These data strongly support the improved safety and therapeutic efficacy of high-affinity p53scTCR for TCR-based immunotherapy of p53-associated malignancies.

A high-content EMT screen identifies multiple receptor tyrosine kinase inhibitors with activity on TGFß receptor.

An epithelial to mesenchymal transition (EMT) enables epithelial tumor cells to break out of the primary tumor mass and to metastasize. Understanding the molecular mechanisms driving EMT in more detail will provide important tools to interfere with the metastatic process. To identify pharmacological modulators and druggable targets of EMT, we have established a novel multi-parameter, high-content, microscopy-based assay and screened chemical compounds with activities against known targets. Out of 3423 compounds, we have identified 19 drugs that block transforming growth factor beta (TGFß)-induced EMT in normal murine mammary gland epithelial cells (NMuMG). The active compounds include inhibitors against TGFß receptors (TGFBR), Rho-associated protein kinases (ROCK), myosin II, SRC kinase and uridine analogues. Among the EMT-repressing compounds, we identified a group of inhibitors targeting multiple receptor tyrosine kinases, and biochemical profiling of these multi-kinase inhibitors reveals TGFBR as a thus far unknown target of their inhibitory spectrum. These findings demonstrate the feasibility of a multi-parameter, high-content microscopy screen to identify modulators and druggable targets of EMT. Moreover, the newly discovered "off-target" effects of several receptor tyrosine kinase inhibitors have important consequences for in vitro and in vivo studies and might beneficially contribute to the therapeutic effects observed in vivo.