Table 4.

Ex Vivo and Cultured IFN-γ ELISPOT Assay Responses for CMV-Seronegative Subjects

	Ex vivo ELISPOT Assay Results (SFU/106 PBMC)a			Cultured IFN-γ ELISPOT Assay Results (SFU/106 cultured cells)b
	pp65	gB	IE1	pp65		gB		IE1
Group 1	Wk 32			Wk 0	Wk 32	Wk 0	Wk 32	Wk 0	Wk 32
V01OC002	0	0	0	100	242	6	1475	75	14
V01OC005	8	10	12	0	42	50	83	42	25
V01OC008c	3	3	8	36	792	0	92	17	183
V01OC009	3	1	8	0	6	50	6	250	192
V01OC013	18	29	1	125	2258	92	125	0	42
V01OC014	29	31	4	11	1967	50	4467	0	0
V01OC016	32	22	8	11	2417	47	783	14	100
V01ST001	12	23	17	6	358	217	608	75	89
Group 2	Wk 32			Wk 0	Wk 32	Wk 0	Wk 32	Wk 0	Wk 32
V01ST002	0	3	0	11	0	8	33	0	0
V01ST004	52	25	15	75	5017	56	775	14	58
V01CO001	3	9	7	17	39	53	117	36	0
V01FH001	70	40	2	33	2433	6	542	50	17
V01OC021d	90	92	25	100	6067	217	5575	111	150
V01ST005	0	3	6	0	0	8	14	17	17
V01FH004	96	33	27	61	417	8	6375	19	33
V01OC022	27	53	0	3	4517	383	1600	42	50
Group 3	Wk 16e			Wk 0	Wk 16	Wk 0	Wk 16	Wk 0	Wk 16
V01OC023	14	n.d.f	0	33	58	333	538	8	11
V01OC026	56	n.d.	0	28	1900	542	1658	19	17
V01OC029	78	n.d.	0	53	1267	0	792	67	50
V01FH005	5	n.d.	5	17	1542	58	2100	25	108
V01OC030	8	n.d.	5	8	0	0	3	0	6
V01ST006	8	n.d.	0	11	583	17	950	33	92

^aA subject was considered to be a responder to the cytomegalovirus (CMV) proteins phosphoprotein 65 (pp65), glycoprotein B (gB) or immediate early antigen 1 (IE1) by ex vivo gamma interferon (IFN-γ) ELISPOT assay if the mean spot forming units per 10⁶ peripheral blood mononuclear cells (SFU/10⁶ PBMC) for replicate wells was greater than or equal to 50 and the mean SFU/well for replicate wells was greater than twice the mean background (without peptide) SFU/well. Positive responses are in bold type. Using the established cut-off, 63 CMV-seronegative subjects were all negative in the assay (100% specificity) and 50 CMV-seropositive subjects were all positive for at least one of the three antigens in the assay (100% sensitivity); positive responses for the CMV-seropositive subjects ranged from 50 SFU/10⁶ PBMC to the upper limit of the assay, 6000 SFU/10⁶ PBMC (authors’ unpublished data).

^bA subject was considered to be a responder to pp65, gB or IE1 by the cultured ELISPOT assay if the SFU/10⁶ cultured cells were >250 and the SFU/well was >2X the SFU of wells without peptide. Data from the analysis of PBMC from CMV-seronegative subjects prior to administration of the CMV DNA vaccine suggested that some subjects may have low levels of crossreactive T-cell responses to peptides in the pools [data not shown]; therefore, the criteria for a positive response to the vaccine included the requirement that the SFU/10⁶ cultured cells was >2X SFU/10⁶ cultured cells at baseline. Positive responses are in bold type. Using these criteria, no positive responses were detected for PBMC stimulated with the IE1 specificity control peptides (IE1 is not encoded in the vaccine, n=22 CMV-seronegative subjects vaccinated with VCL-CB01, 100% specificity for vaccine response). The lack of IE1 responses in PBMC from CMV-seronegative subjects up to Week 32 of the study suggests that none of those subjects acquired CMV during the course of the trial.

^cWk 0 PBMC were not available, Wk 2 PBMC were used in the assay

^dWk 32 PBMC were not available, Wk 24 PBMC were used in the assay

^eResults from initial evaluation in the ex vivo ELISPOT assay in which pp65 peptides are split into two pools each are reported. The data reported are the sums of SFU/10⁶ PBMC for pool I and pool II

^fn.d.,Not done