Strategies to Counteract Resistance Mechanisms in CAR+ T-Cell-Based Immunotherapy for Triple-Negative Breast Cancer
Technical Report,01 Sep 2017,31 Aug 2018
The University of North Carolina at Chapel Hill Chapel Hill United States
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The immunohistochemical assay IHC with monoclonal antibodies to detect CSPG4 in TNBC tumors was optimized. Formalin fixed, paraffin embedded TNBC tumor sections incubation of slides in EDTA buffer, pH8.0, was found to be the most sensitive substrate in IHC reactions. The combination of the mouse mAbs D2.8.5 3ug mL, 763.74 5ugmL and TP41.2 5ugmL yields the best results. Utilizing these conditions, a TMA containing 63 TNBC tumors was tested for expression of CSPG4. Fifty-five of the tumors expressed high levels of CSPG4. The remaining 8 tumors expressed low or barely detectable level of CSPG4. The latter tumors were tested for the expression of B7-H3, a distinct tumor antigen which is also an attractive target of CAR T cell-based immunotherapy. Two of the tumors expressed high levels of B7- H3. Utilizing anti-idiotypic monoclonal antibodies, an assay has been developed to monitor the level of CSPG4 Cars on transduced T cells. CSPG4 CARs were detected on about 50 percent of the transduced T cells. The level of CARs was upregulated on T cells by treatment with the HDAC inhibitor vorinostat. The upregulation of CARs on T cells was associated with an increased anti-tumor activity.
- Medicine and Medical Research