Presentation on theme: "Clinical Experience at Baylor College of Medicine with T Lymphocytes Genetically Modified Using Retroviral Vectors Gianpietro Dotti G Center for Cell and."— Presentation transcript:
1 Clinical Experience at Baylor College of Medicine with T Lymphocytes Genetically Modified Using Retroviral VectorsGianpietro Dotti GCenter for Cell and Gene TherapyBaylor College of Medicine, Houston TX
2 Type of vector and packaging cell lines Moloney-derived retroviral vector(SFG from Mulligan R.)Packaging cell line Phoenix-Ecooriginal cells from ATCCgeneration of a master cell bank under GMP conditionsPackaging cell line PG13
3 Generation of stable packaging cell line Multiple infection of PG13 cells using Eco-pseudotyped retroviral particles (transient supernatant)Generation of single cell clone from infected PG13 cellsExpansion of the single cell clone to make a master cell bank (MCB)Production and collection of supernatant from the MCB
4 ManufacturingManufacturing of the single cell clone under GLP conditionsExpansion of the master cell bank (MCB) and production of the retroviral supernatant under GMP conditions
5 Packaging cell lines 9 Packaging cell lines generated and tested RCR detection negative in all cases for both Ecotropic and Galv envelopes (co-culture assay)18 lots of supernatant produced and testedRCR detection negative in all cases for GalV envelope (co-culture assay)
6 Clinical protocols IND Transgene Cell product Disease # CAR.GD2z Activated T cells vs NeuroblastomaEBV-CTLs19 patients treated.38 T-cell/CTL products infused all negative by co-culture assay.
7 Clinical protocols IND Transgene Cell product Disease # CAR.CD19.CD28z vs Activated T cells CD19+ LymphomaCAR.CD19z# CAR.CD19.CD28z vs Activated T cells vs CD19+ LymphomaCAR.CD19z EBV-CTLs# CAR.CD19.28z Virus-specific CTLs CD19+ Lymphoma8 patients treated (6 in # and 2 in #).16 T-cell/CTL products infused. 6 negative by co-culture assay. 8 pending and 2 not needed (frozen within 4 days of culture).
8 Clinical protocols IND Transgene Cell product Disease # CAR.HER2.CD28z Activated T cells Osteosarcoma# CAR.HER2.CD28z and EBV-CTLs Lung cancerTGFbRIIdcyt (DNRII)# CAR.HER2.CD28.z CMV-specific CTLs Glioblastoma4 patients treated (2 in #14050 and 2 in #14036).4 T-cell/CTL products infused. 1 negative by co-culture assay. 3 pending.
9 Clinical protocols IND Transgene Cell product Disease # TGFbRIIdcyt (DNRII) LMP-2a-CTLs EBV+ Lymphoma4 patients treated.4 CTL products infused. All negative by co-culture assay.
10 Clinical protocols IND Transgene Cell product Disease # icaapase9.2A.DCD allodepleted T cells hematological malignancies/haplo HSC transplant4 patients treated.4 T-cell products infused. All negative by co-culture assay.
11 T/CTL line infused summary 92 lines infused to 73 patients2 did not require assay8 results pending82 lines negativeRCR co-culture assay for GalV envelope
12 Follow up patient samples - 73 patients infused (follow up from 1 month to 15 years)- 371 samples analyzed- All RCR negative by Q-PCR (GalV envelope)
13 Previous experience at St. Jude Children’s Research Hospital (Gene marking)34 patients infused with gene marked EBV-specific CTLsAll 34 CTL lines negative by co-culture assay- 271 follow up samples all negative by PCR assay (samples continue to be archived from 2002 but no longer tested)
14 Conclusions and Suggestions T cell/CTLs modified with retroviral vectors are consistently RCR negativeRCR assay based on co-culture is time consuming and extremely expensive limiting the possibility to extend the use of promising immunotherapy approachesBased on the current experience samples can be stored and analyzed for RCR contamination in case of severe adverse events