Immune modulations during chemoimmunotherapy & novel vaccine strategies--in metastatic melanoma and non small-cell lung cancer.

This thesis describes the treatment of metastatic melanoma (MM) and non small-cell lung cancer (NSCLC) from an immunotherapeutic approach. The purpose of the first part of the thesis was to assess how treatment with Temozolomide (TMZ) chemotherapy affects the immune system in patients with metastatic MM. Our results showed that the number of T lymphocytes was significantly reduced after 3 treatment cycles. Furthermore, the induced lymphopenia was positive correlated to achievement of clinical benefit. We demonstrated that the proportion of CD4+ and Treg lymphocytes decreased whereas the CD8+ T cells increased. In particular, we demonstrated that mature CD8+ T cells increased during treatment. Analyses of peripheral blood before and after treatment showed that T cell responses against common viral epitopes were conserved despite chemotherapy. Surprisingly, we found a significant increase in T cell responses against well-known MM tumour specific antigens. Overall, we have verified that TMZ in addition to being an alkylating and cytotoxic chemotherapy, also possess immune modulatory effect in MM patients treated with standard dosage of TMZ. In the second part of the thesis we examined how treatment with Interferon alfa-2b and Interleukin 2 (IFNα/IL2) affects the immune system. We demonstrated a significant induced lymphocytosis during treatment. Furthermore, we showed that the percentage increase in lymphocytes was positively correlated to clinical outcome. Moreover, we have seen that IFNα/IL2 leads to significant increase in NK and Treg cells in both patients with and without clincal effect. In general, T cell responses against common viral epitopes and well-known melanoma tumour specific antigens were low. Furthermore, the study confirmed that elevated LDH is negatively correlated with both treatment response and median overall survival. Overall, we have characterized changes of immune cells and correlated them with clinical efficacy during the couse of IFNα/IL2 used in standard dosage. In the third part we investigated if vaccination with a peptide derived from IDO was feasible in patients with metastatic NSCLC. This "First in Man" trial was safe and showed modest side effects only. Since IDO was expressed in NSCLC tissues it was found to be a relevant target. One patient achieved significant regression of liver metastases (confirmed partial response) and another 6/15 patients achieved prolonged disease stabilization. Furthermore, median overall survival was 25.9 months demonstrating a better survival in vaccinated compared to non-vaccinated comparable NSCLC patients. The presence of IDO specific CD8+ T cells were detected by IFNy Elispot. In patients with clinical effect of the vaccine IDO-specific CD8+ T cells at pre-treatment was significanctly increased. Moreover, low-frequent IDO positive tetramer CD8+ T cells were detected and led to effective killing of an IDO+ HLA-A2 positive cancer cell line (SW480) in 1 patient. Moreover, flow cytometry was performed and in general no significant changes in CD8+ and CD4+ T cells were seen, although patients with clinical response showed a trend towards increased mature CD8+ T cells during treatment. In addition, we found lower levels of Tregs as well as an increased level of NK cells after 6 vaccinations. Elevated Kyn/Trp ratio is suggested to mirror IDO activity. In 8/11 patients the level after the 6th vaccine was stable compared to baseline. No differences between patients with clinical benefit (4/5) and patients with progressive disease (4/6) were demonstrated. Two patients had an increase in Kyn/Trp ration meanwhile demonstrating a high expression of IDO. In 2 patients with clinical response long-term stabilization of Kyn/Trp was observed. Overall, the vaccine was well tolerated with no adverse toxicity. Median overall survival was 25.9 months with long term disease stabilization achieved in 47% of the treated patients. Based on the promising clinical results achieved in the vaccine trial for NSCLC patients, we launched a new clinical trial for MM patients (ongoing patient recruitment) in June 2012. In order to enhance the immune response the vaccine comprises IDO plus Survivin peptide as well as the adjuvants Montanide, Aldara and GM-CSF. Finally, the vaccine is given in combination with TMZ. Patients are evaluated every 3rd month with PET-CT scan. Preliminary clinical data from the first 7/30 evaluable patients are presented. Two patients demonstrated a patial response with 57% and 45% tumour regression lasting for 10 months and 6+ months respectively, corresponding to a preliminary objective response rate of 29%. The vaccine has been manageable and without significant side effects.