Telomerase peptide vaccination: a phase I/II study in patients with non-small cell lung cancer

Abstract

Purpose: A phase I/II study was conducted to investigate the safety, tolerability and clinical response to vaccination with a combination of telomerase peptides GV1001 (hTERT: 611-626) and HR2822 (hTERT: 540-548) in patients with non-small cell lung cancer.

Experimental design: Twenty-six patients with non-small cell lung cancer received intradermal administrations of either 60 nmole (112 microg) or 300 nmole (560 microg) GV1001 in combination with 60 nM (68.4 microg) HR2822 and granulocyte macrophage-colony stimulating factor. The treatment period was 10 weeks. Booster vaccinations with 300 nM GV1001 were offered as follow-up. Monitoring of blood samples, clinical examination and radiological staging were performed regularly. Immune responses were measured as delayed-type hypersensitivity skin reaction and in vitro T cell proliferation. Bone marrow function was monitored in long time survivors.

Results: The treatment was well tolerated with minor side effects. No bone marrow toxicities were observed in long time survivors with immune responses. Immune responses against GV1001 were detected in 11 of 24 evaluable patients during the primary regimen and in additional two patients following booster injections. Two patients responded to HR2822. Cloned GV1001-specific CD4+ T cells displayed a Th1 cytokine profile and recognized autologous antigen presenting cells pulsed with recombinant telomerase protein. A complete tumor response was observed in one patient who developed GV1001-specific cytotoxic T cells that could be cloned from peripheral blood.

Conclusion: The results demonstrate that GV1001 and HR2822 are immunogenic and safe to use in patients with NSCLC. Induction of GV1001-specific immune responses may result in objective tumor responses. Based on these initial encouraging results, further clinical studies of GV1001 in NSCLC patients are warranted.

Fig. 1

In vitro T cell response against GV1001 in patient 710. T cell proliferation of PBMC is given as SI at baseline, following one complete cycle of vaccination with low peptide dose (Week 10) and following booster vaccinations with high peptide dose. SI is defined as response with antigen divided by response without antigen. Arrows indicate booster vaccinations

Fig. 2

Recognition of GV1001 and recombinant hTERT by two CD4 + T cell clones derived from patient 710 after booster vaccination. HLA-class II restriction was determined by antibody blocking (Fig. 2a). Fifty thousand T cells were cultured with 25,000 irradiated autologous PBMC as APC and 1 μM GV1001 for 3 days in the presence or absence of blocking antibody as indicated. The antibodies used were SPVL3 (anti DQ) and B7/21 (anti DP). Final concentration of antibody was 10 μg/ml. In a parallel experiment (Fig. 2b), the same T cell clones were tested with recombinant hTERT (0.5 μM) as an antigen under the same conditions. GV1001 (0.1 μM) and medium alone were used as controls. Results are recorded as mean cpm of triplicate wells

Fig. 3

Specific killing of autologus Epstein–Barr virus transformed blasts pulsed with peptide p613 by the HLA-DQB*04 restricted GV1001 specific CD4+ T clone 710–76. Target cells were labeled with ⁵¹Cr for 1 h and 2,000 cells were added to each well before addition of peptide (1 μM) and effector cells at the ratios indicated. Supernatants containing released ⁵¹Cr were harvested after 6 h and counted. The peptides p613 (TSRLRFIPK), p614 (LTSRLRFIP) and p615 (LLTSRLRFI) are all nine mere fragments from the C-terminal part of GV1001. The vaccine peptide HR2822 was used as a negative control. Results are expressed as percentage of specific killing and represent mean of triplicate wells

Fig. 4

In vitro proliferative T cell response of PBMC against the N-terminal 9-mere fragment of GV1001 in patient 710. T cell proliferation is given as mean cpm of triplicate wells. Responses shown are from samples taken 1 h before booster vaccine 3 and 12 days after booster vaccination 3

Fig. 5

a Cytokine profile of the HLA-DPB*0401/0402 restricted CD4+ clone 710–13 following activation by GV1001. Supernatants were harvested 24 h after stimulation of 50,000 T cells with 25,000 APC and GV1001 (1 μM) in serum-free medium. Seventy-five microliters was harvested from duplicate wells and analyzed using a Bio-rad Bio-Plex instrument. Results are expressed as mean concentration of cytokines (pg/ml) in the supernatants based on calculation from standard curves established with recombinant cytokines. b GM-CSF production by two CD4+ clones from patient no. 710. The GM-CSF production by 50,000 T cells from the cytotoxic clone 710–76 corresponds to 500 ng/10⁶ T cells/24 h. Results were obtained as described in a

Fig. 6

Survival for patients that completed 10 weeks of vaccination. Patient 725 is stage IIB, patient 710 stage IIIA, patients 701,713 and 727 are stage IIIB. The remaining patients are all stage IV. Patients marked asterisk are still alive

Fig. 7

a At inclusion in the vaccination trial patient no. 710 had a 32 mm×32 mm tumor in right upper lobe. b CT scan of the patient’s mediastinal glands at inclusion in the trial. c, d right upper lobe and mediastinal glands 22 months after start of vaccination, respectively