Harnessing endogenous miR-181a to segregate transgenic antigen receptor expression in developing versus post-thymic T cells in murine hematopoietic chimeras.
MicroRNAs (miRNAs) are small, noncoding RNAs that regulate gene expression by targeting complementary sequences, referred to as miRNA recognition elements (MREs), typically located in the 3' untranslated region of mRNAs. miR-181a is highly expressed in developing thymocytes and markedly downregulated in post-thymic T cells. We investigated whether endogenous miR-181a can be harnessed to segregate expression of chimeric antigen receptors (CARs) and TCRs between developing and mature T cells. Lentiviral-encoded antigen receptors were tagged with a miR-181a-specific MRE and transduced into mouse BM cells that were used to generate hematopoietic chimeras. Expression of a CAR specific for human CD19 (hCD19) was selectively suppressed in late double-negative and double-positive thymocytes, coinciding with the peak in endogenous miR-181a expression. Receptor expression was fully restored in post-thymic resting and activated T cells, affording protection against a subsequent challenge with hCD19+ tumors. Hematopoietic mouse chimeras engrafted with a conalbumin-specific TCR prone to thymic clonal deletion acquired peptide-specific T cell responsiveness only when the vector-encoded TCR transcript was similarly engineered to be subject to regulation by miR-181a. These results demonstrate the potential of miRNA-regulated transgene expression in stem cell-based therapies, including cancer immunotherapy.
Papapetrou
Eirini P
EP
Center for Cell Engineering, Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center (MSKCC), New York, NY 10065, USA.
Kovalovsky
Damian
D
Beloeil
Laurent
L
Sant'angelo
Derek
D
Sadelain
Michel
M
D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology / Q000502:physiology; D019070:Cell Lineage; D002678:Chimera / Q000502:physiology*; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000502:physiology*; D006801:Humans; D008213:Lymphocyte Activation; D051379:Mice; D008822:Mice, Transgenic; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D011946:Receptors, Antigen / Q000235:genetics / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism*; D013950:Thymus Gland / Q000166:cytology*; D019076:Transgenes
D035683:MicroRNAs; D011946:Receptors, Antigen; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins; C529100:mirn181 microRNA, mouse