Table 2.

Recognition of WT1 peptide and transfectants by PBL retrovirally transduced to express a WT1 reactive TCR^a

		Donor 1		Donor 2		Donor 3
	DMF5b	GFP	WT1 TCR	GFP	WT1 TCR	GFP	WT1 TCR
media	28	30	29	28	28	30	30
T2 + MART peptidec	9980g	37	37	34	29	52	40
T2 + WT1 peptidec	56	35	>20000	32	>20000	45	>20000
COS-A2 + MARTd	4710	31	37	36	33	83	43
COS-A2 + WT1d	27	31	11449	34	8482	77	7254
293 SP + A2/MARTe	872	29	28	28	28	29	29
293 SP + A2/WT1e	28	29	52	28	66	31	61
293 IP + A2/MARTf	79	29	29	33	30	34	41
293 IP + A2/WT1f	28	29	1963	32	3606	36	1947

^aPBL from 3 HLA-A*0201+ donors were transduced with retroviral particles encoding GFP as a negative control or with the WT1 reactive TCR (TRAV12-1*01; CDR3: CVVNTPPNTDKLIF; TRBV7-2*01; CDR3: CASTPFTSGSGWDEQFF). Transduced T cells were cocultured overnight with the indicated target cells, and IFNγ in coculture supernatants was measured by ELISA.

^bDMF5 is a MART-1 reactive T cell population included as a positive control for the transfections.

^cT2 cells were pre-loaded with 1 μg/ml of a control peptide from HBV with high binding affinity for HLA-A*0201 or WT1:126-134 ~1 hour prior to the coculture.

^dCOS-7 cells stably expressing high levels of HLA-A*0201 by means of retroviral transduction and antibiotic selection were transiently transfected with GFP cDNA as a negative control or full-length WT1 cDNA ~24 hours prior to the coculture.

^e293 cells expressing components of the standard proteasome were transiently co-transfected with HLA-A*0201 cDNA and either full-length MART-1 or WT1 cDNA ~24 hours prior to the coculture.

^f293 cells expressing inducible subunits 1, 2, and 5 of the immunoproteasome were transiently co-transfected with HLA-A*0201 cDNA and either full-length MART-1 or WT1 cDNA ~24 hours prior to the coculture.

^gunderlined values indicate IFNγ secretion >100 pg/ml and >2X background with any negative control target cell.