LB01

We report the results of a single arm Phase II clinical trial of autologous dendritic cells (DC) pulsed ex vivo with tumor lysate from established melanoma cell lines (Dandrit Biotech) pre-treated with a demethylating agent to enhance immunogenicity. Twenty patients (9 males, 11 female) with advanced colorectal cancer who express one or more MAGE antigens were accrued and DC vaccine administered intradermally every 2 weeks for 10 injections.
 The median age was 70 years (n = 20) and all patients had progressive disease (PD) prior to accrual. The commonest MAGE gene expressed by RT-PCR was MAGE A4 (60%) and 75% expressed only one MAGE gene. The patients had an average of 3 metastatic tumor sites each, 55% had baseline ECOG status of > 2 and the cohort had received a median of 3 prior chemotherapy and/or biologic agents. The average dose per vaccination was 4.20 x 106 DC vaccine and 12/20 completed all 10 injections. DC production analysis by a novel patented tumor-necrosis-factor-α-based method confirmed a high quality mature phenotype with median expression of HLA-D at 99.79%, CD86 at 99.76%, CCR7 at 60.24% and CD14 at 17.08%. There were no > Grade 2 toxicities.
 At last follow up, the disease control rate was 35% with 6 stable disease (SD > 3 months) and 1 partial response (PR) by RECIST criteria. At 1 year post-DC vaccine, 40% of patients were alive. The overall median duration of response (including SD) was 201 days. In the single patient who achieved PR, time to response was 79 days, and further tumor regression was seen at 281 days. In a patient with prolonged SD, tumor shrinkage of 17.8% occurred after 464 days.
 A significantly positive delayed-type hypersensitivity (DTH) reaction was demonstrated in more patients after X10 DC vaccinations (p<0.0001) than after only X5 DC vaccinations (p=0.466). A positive DTH correlation with disease control was not seen. Enzyme-linked immunospot assay for patient 1-10 showed a positive response only in patient 4 who had a sustained SD of 470 days (β). Comparing foxp3+CD4+ regulatory T cells (Treg) by FACS of baseline and after the 9th DC vaccine showed a significant decrease from a median of 10.35% to 4.36% of CD3+ T cells (p = 0.001). All patients demonstrated a decrease in foxp3+ Treg except two patients with progressive disease. Serial analysis of the programme death-1 (PD-1) gene by quantitative real-time PCR on MACS-isolated CD8+ T cells in 6 consecutive patients showed that in the 3 patients with PD, all 3 had rising PD-1 levels (up to 12X after 7th DC vaccine in patient 1). In the 2 patients with SD, a downward trend in PD-1 were noted for both (up to 8X decrease after 7th DC vaccine in patient 7). Patient 9 who achieved a PR had the lowest level of PD-1 at 0.0162, and remained low at month 7 (0.0557) post-vaccine (β).
 In conclusion, we have shown that DC vaccination pulsed with an allogeneic tumor lysate in pre-treated MAGE-gene-expressing advanced colorectal cancer patients can induce both durable disease-control and immunological responses in selected patients. Treg levels and PD-1 expression on T cells may be important biomarkers in cancer vaccine studies.
 (β) Full results will be available for Elispot assay and PD-1 analysis by meeting time.
 Grant funded by the Singapore Cancer Syndicate

First AACR Centennial Conference on Translational Cancer Medicine-- Nov 4-8, 2007; Singapore