Optimization of intracerebral tumour protection by active-specific immunization against murine melanoma B16/G3.12

Development of brain metastases despite extracerebral response to systemic immunotherapy is a common problem in melanoma patients. We have previously described a murine melanoma vaccine of interferon-gamma (IFNgamma)-treated, irradiated syngeneic B16/G3.12 and allogeneic (Cloudman) melanoma cells, plus the adjuvant DETOX, that is protective against subcutaneous (93%) or intracerebral (69%) syngeneic challenge. This study aimed to optimize this vaccine. Groups of nine or 10 mice were immunized five times in 5 weeks with: (i) complete vaccine +/- IFNgamma (VAC+, VAC-); (ii) syngeneic 2 x 106 G3.12 cells plus DETOX (Syn+D), (iii) 2 x 106 allogeneic Cloudman cells plus DETOX (Allo+D); (iv) VAC+ without DETOX (no DETOX); (v) DETOX alone (DETOX); or (vi) phosphate buffered saline (PBS). Mice were challenged subcutaneously with 104 viable G3.12 (or Cloudman cells) and after 35 days intracerebrally with 104 G3.12 cells. Expression of H-2 antigens (measured using fluorescence-activated cell sorting), splenocyte cytotoxicity (measured using 51Cr release) and median overall survival (OAS) were analysed using the log-rank test. VAC+, VAC- and G3.12 mice were equally protected from subcutaneous (s.c.) and intracerebral (i.c.) melanoma challenge (OAS 65 days for s.c., 30 days for i.c.). Protection was less (P < 0.05) in DETOX mice (48 days for s.c.), PBS mice (47 days for s.c., 21 days for i.c.) or no DETOX mice (51 days for s.c.). Allo+D mice showed s.c. (59 days) but not i.c. protection (20 days). IFNgamma incubation did not increase the effect in either the challenge cells or the vaccine cells (P > 0.05). Specific cytotoxicity was seen with G3.12 targets in VAC+ (27%) but not PBS (2%; P < 0.05) mice with equal NK (YAC-1) lysis (10% versus 7%; P< 0.05). Optimal protection against s.c./i.c. experimental murine melanoma was yielded by irradiated syngeneic cells plus DETOX. DETOX alone was not active. Upregulation of H-2 antigens with IFNgamma under these conditions does not augment protection.