Immunotherapy of established tumors in mice by intratumoral injection of an adenovirus vector harboring the human IL-2 cDNA: Induction of CD8(+) T-cell immunity and NK activity

Intratumoral (i.t.) injections of an adenovirus encoding the human interleu kin-2 (IL-2) under the control of the RSV (Ad-pRSV-IL-2) or CMV (Ad-pCMV-IL -2) promoter were performed in established mastocytoma P815 tumors in B6D2 mice. Both early and long-term survival were found increased in mice treate d with Ad-pCMV-IL-2 as compared with those obtained with Ad-pRSV- IL-2: tu mor regress ion was observed in 30-50% of mice for the former and 5-15 % fo r the latter. Difference in efficacy between the two vectors was directly c orrelated to the amount of IL-2 produced i.t. between 24 and 48 hours posti njection, which reached 10-20 ng/ tumor for Ad-pCMV-IL-2 and 0.3-0.5 ng/tum or for Ad-pRSV-IL-2. In both cases, expression in the tumor was clearly det ectable for a period of 7-10 days postinjection. Serum IL-2 was not detecta ble in mice treated with Ad-pRSV-IL-2, whereas expression peaked at a total of 1-2 ng at 24 hours but declined very rapidly in the Ad-pCMV-IL-2-treate d group. Constant production of IL-2 inside the tumor was necessary for suc cessful therapy because i.t. injections of recombinant IL-2 at levels up to 1 mug for five consecutive days did not lead to antitumoral activity. Evid ence of induced systemic immunity following Ad-pCMV-IL-2 injections was obt ained from re challenge experiments in which tumor-free mice after treatmen t rejected a subsequent centralateral injection of a lethal dose of P815 tu mor cells and from the observation that regression of nontreated tumors occ urred in animals bearing bilateral tumors that were treated i.t, in a singl e tumor with Ad-pCMV-IL-2. P815-specific cytotoxic T lymphocytes (CTL) were found specifically in spleen cells from cured mice or rechallenged mice bu t not in control mice. Interestingly, limiting dilution analysis of anti-P8 15 CTL precursor (CTLp) frequency revealed a significant increase in mice c u red of their tumor as com pared to that obtained in naive mice or control Mice treated or not with Ad-IL-2 but whose tumor was growing. In vivo depl etion of T-cell subsets, as well as natural killer cells at the time of i.t . injections with Ad-pCMV-IL-2, demonstrated that both CD8(+) T cells and n atural killer cells, but not CD4(+) T cells, were required for successful t herapy. Finally, mice preimmunized with Ad-null viruses were severely compr omised in their capacity to eradicate established P815 tumors after Ad-pCMV -IL-2 therapy, at least when neutralizing antibody titers reached a critica l level.