The administration of ACT therapy and high-dose IL-2 therapy following lymphodepleting chemotherapy resulted in objective clinical responses in 18 of 35 patients with metastatic melanoma, including 34 patients who were refractory to treatment with high-dose IL-2. Thirteen of the 18 responding patients were also refractory to chemotherapy. Metastatic disease at numerous anatomic sites was susceptible to this therapy, including cutaneous lesions, bulky nodal disease, and visceral, brain, and bony metastases. The toxicities associated with this treatment included the expected toxicities of high-dose IL-2 therapy and the hematologic suppression associated with lymphodepleting chemotherapy; these were mostly transient and readily managed by standard supportive techniques.
Autoimmunity directed against normal melanocytes was observed in nine (75%) of the twelve patients who were treated with T-cell cultures that recognized melanocyte differentiation antigens and who experienced objective tumor regression. Melanocyte destruction in the skin commonly resulted in vitiligo in the responding patients. Similarly, anterior uveitis was detected in four responding patients, and was controlled with corticosteroid eye drops. A link between successful immunotherapy and the onset of autoimmunity has been noted in several clinical immune-based therapies,15,23,24
as well as preclinical immunotherapy models,25
suggesting a strong link between the mechanisms of tumor regression and autoimmune attack on normal tissues. The nature of the melanoma antigen targeted by TIL did not predict the outcome of treatment. Shared tumor antigens such as MART-1 that are expressed by nonmalignant tissues were as effective for targeting melanoma destruction as unique antigens expressed only by the tumor cells. This observation suggests that peripheral mechanisms of self-tolerance (ie, other than thymic deletion) can inhibit the endogenous immune response to tumor, and need to be overcome to induce clinically relevant antitumor responses.
The current protocol incorporated several changes compared to our prior efforts at ACT of patients with melanoma. Improved approaches were developed for the generation of tumor-reactive lymphocyte cultures that often contained multiple clonotypes and a mixture of CD4+
In the current trial, 31 treatments contained cells that were selected for tumor recognition and rapidly expanded with OKT3 only a single time, and five treatments (1 PR and 4 NR) contained cells expanded with OKT3 more than once. These cultures contrasted with the unselected TIL26
or the highly selected cloned CD8+
we administered in prior trials. In previous trials with cloned CD8+
lymphocytes for ACT, the generation of large numbers of cloned cells required at least three expansions mediated by OKT3. The decreased proliferative potential of the cells used in prior trials that were rapidly expanded multiple times in vitro may have accounted for the poor persistence of these cells after cell transfer and their failure to cause tumor regression. A second important change we incorporated into the current protocol was the administration of a conditioning chemotherapy regimen before the cell infusion. This chemotherapy regimen contained agents with little anti-melanoma activity and was designed to deplete endogenous lymphocytes before the cell transfer. The role of lymphodepletion in augmenting the antitumor function of adoptively transferred lymphocytes is an area of active investigation. Two potential mechanisms have been proposed based on studies in preclinical models: the elimination of suppressor T cells,27
or the decreased competition by endogenous lymphocytes for homeostatic regulatory cytokines such as IL-7 or IL-15.7,28,29
The observation of lymphocytosis in some responding patients (), together with the suggestion of antitumor T-cell persistence in patients with extended responses () offers hints as to the requirements for successful tumor treatment by ACT therapy. These observations suggest that the capacity for T-cell proliferation in vivo and the transfer into a lymphodepleted host are important determinants of efficacy. More research into these aspects of cell transfer therapy is warranted.
Several variations in administered treatments did not apparently impact the treatment efficacy. Although administration of G-CSF has been reported to be immunosuppressive in patients undergoing allotransplantation of stem cells,30,31
no statistical difference in the response rate was observed in the small, nonrandomized patient populations in this study. Similarly, there were no apparent immunologic or clinical differences between patients receiving a peptide vaccine after cell transfer and those patients who did not receive vaccination. Other aspects of treatment, which are active areas of investigation, include the route of cell administration (intra-arterial v
), and the dose and schedule of IL-2 administration.33
Some of the patients reported here had extensive tumor regression, but eventually recurred at pre-existing or new sites. Immunocytochemical analysis of pretreatment and recurrent lesions demonstrated a loss of antigen expression by the recurrent tumor in five (55%) of nine patients who relapsed after an objective response. In two patients (patients 2 and 28) the loss of HLA class I antigens observed by immunocytochemical analysis was confirmed by in vitro functional analysis. Melanoma cell lines derived from recurrent tumor biopsies of these patients demonstrated loss of HLA expression (due to mutation of the beta2-microglobulin gene in one patient) and failure to stimulate antigen-specific T cells (not shown). In one patient treated with MART-1 reactive cells, the expression of MART-1 protein was selectively decreased while other melanoma differentiation antigens including gp100 were not affected. These results demonstrate that the transferred T cells exerted a strong selective pressure on the tumor in vivo, and confirm that the antitumor effects of this treatment are antigen mediated.
In summary, the results presented here establish that ACT therapy can impact bulky metastatic melanoma that is refractory to other treatments including high-dose IL-2 therapy and chemotherapy. Fifty-one percent of the patients experienced objective tumor responses with durations from 2 months to more than 2 years, including three patients (9%) with an ongoing complete response. Many aspects of this treatment have the potential to be improved by additional studies of the basic scientific principles of immune-mediated tumor rejection and the optimization of their clinical application. Current studies in our laboratory are focused on the logistical aspects of generating autologous cell-based patient treatments, the genetic modification of lymphocytes with T-cell receptor genes and cytokine genes to change their specificity or improve their persistence, and the administration of antigen specific vaccines to augment the function of the transferred cells.