Phase I dose escalation trials using ex vivo expanded, anti-CD3 activated T cells (ATC) armed with anti-CD3 x anti-Her2/neu (Her2Bi) to evaluate toxicity and immune responses were conducted in 8 Stage IV breast cancer (BrCa), 5 high-risk Stage II BrCa, and 3 hormone refractory prostate cancer (HRPC) patients. Patients received 8 infusions (Inf) twice per week for 4 weeks totaling 40 billion (11/16 patients) or 80 billion (5/16 patients) HER2Bi-armed ATC. Phenotyping the expansion product showed 90.0±10.1% (mean±SD)CD3, 47.5±22.2 CD4, and 41.9±19.9 CD8. Specific lysis of SK-BR-3 targets by patients’ armed ATC (n= 16 patients) at E:T of 10, 5, and 2.5 averaged 54.1±17, 39.5±14, and 25.9±11, respectively. These results were comparable to 63.4±17 (p = 0.23), 57.7±20 (p = 0.09), and 33.9±16 (p = 0.38) specific lysis mediated by armed ATC from 6 normal donors at the respective E:T confirming the function of Her2Bi-armed ATC at the time of Inf. By ELISA, 4/8 (50%) of the patients developed anti-mouse antibody responses (HAMA) to OKT3, which peaked (median: 35.4 ng/ml; range: 4–64 ng/ml) between Inf#5 and Inf#8 (14–27 days post-Inf #1). There were no AEs associated with elevated HAMA titers. Th1/Th2 panels were obtained from patient sera using the Bio-Plex Protein Array System and analyzed as a function of number of Infs. The in vivo T cell response, calculated as the mean Th1[IL-2+IFN-g]/Th2[IL-4+IL-5] ratio of cytokines, showed a polarized Th1 response as a function of armed ATC Inf that increased from 89.1(Inf#1) to 538.6 (Inf #8). Levels of IL-12 also increased by Inf#5 and were constant from Inf#5 through Inf#8 (mean±SD = 34.0±17.2 pg/ml). These findings were consistent with increased CTL responses observed in ELISPOTs of patient post-Inf PBMC exposed to Her2+ tumor targets. Overall immune response, calculated as the average ratio of Th1[IL-2+IFN-g] /Th2 [IL-4+IL-5+IL-10+IL-13], remained Th1 polarized throughout treatment though the ratio decreased as a function of Inf#: going from 9.46 (Inf#1) to 1.42 (Inf #8). TNF-alpha levels were contant from Inf #1–8 (mean±SD = 9.5 ± 4.9 pg/ml), and GM-CSF peaked between Inf #5–8 (92.2 ± 36.4 pg/ml). Detection of MIgG2a+ (anti-CD3 coated) cells in the PBL from 8 patients was highest post-Inf#1 (18.4% ± 16.1) and decreased by Inf #5–8 (4.1% ± 6.1). In 2 patients, MIgG2a+ armed T cells could still be detected 1 week and 3 weeks after Inf#8. All patients received their targeted doses of armed ATC: no dose-limiting toxicities were observed in any patients, and with the exception of a few patients with grade II–III chills/grade II fever, infusions of Her2Bi-armed ATC were well-tolerated. These observations provide evidence that the endogenous immune system is being modulated by the infusion of armed ATC and further support the rationale for performing phase II trials using armed ATC in Her2+ advanced/high risk BrCa and HRPC patients.

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