This study confirms the feasibility of routinely acquiring, processing, and preparing clinical-grade HSPPC-96 in a timely manner from fresh tumor weighing as little as 2 g for use in patients with metastatic melanoma. The current study was limited to weekly dosing for 4 consecutive weeks by an imposed 2-month shelf life for HSPPC-96, which has since been extended [6
]. Based on the experience in the 36 patients reported here, a dose of 25 μg was technically feasible for ≥ 4 consecutive treatments, whereas we had difficulty filling the 100 μg cohort (400 μg total dose). With the widespread adoption of sentinel node mapping at the time of primary diagnosis and with early detection of recurrence, accrual to future trials of autologous tumor-derived HSPPC-96 will be more limited because of a presumably smaller pool of patients with advanced regional disease.
This trial showed that HSPPC-96 treatment was safe with no unacceptable toxicities or detected autoimmune reactions. A common exclusion criterion in active immunotherapy trials is a negative response to recall antigens by skin testing (Multitest Mérieux; Imtix, Milan, Italy). In the current pilot study, however, we did not exclude anergic patients as we had in earlier whole-cell vaccine trials [15
], preferring to remain open-minded regarding an immune response to HSPPC-96 being independent of a DTH reaction. Three of the 5 patients with increasing SFCs by the IFNγ ELISPOT assay had a negative (grade 0-1) DTH response to DNCB at baseline. Nevertheless, future trials should probably exclude anergic patients since anergy is proving to be an active signaling process which can interfere with the induction of an effective systemic cellular immune response [16
SFC counts against foreign antigens in patients who are exposed to blood borne-infection are generally orders of magnitude higher than those observed against altered self-antigens in patients with malignancy [17
]. The relatively weak and transient changes in SFC (overall SFC range for the entire study, 10.7 - 22) during the course of treatment with HSPPC-96 did not show a dose: response relationship. The IFNγ ELISPOT assay may not have been a reliable biomarker, especially since antitumor immune effecter cells which rapidly traffic to the tissue compartment can elude detection by a peripheral blood assay. Alternatively, in the range of doses tested, HSPPC-96 treatment may have been ineffective in mounting a consistent detectable immune response. At the time this study was conducted (1998-2000), not enough fresh material was available to perform the more sensitive tetramer assays.
Among 16 patients with indicator lesions, there was no major objective response, and 81% had clear progression of disease within 8 weeks. Among patients with stage IV disease treated in the adjuvant setting, a median time to progression of 30.4 months with 82% alive at 10 years is encouraging, but it is impossible to determine whether HSPPC-96 treatment contributed directly to this outcome. Immune responses to HSPPC-96 treatment as measured by IFNγ ELISPOT assay were in general weak and transient and correlated poorly with clinical outcomes.
The results presented here can also be compared with those of a subsequent study of HSPPC-96 treatment in metastatic melanoma patients, as reported by Belli et al [9
]. Among 28 melanoma patients with indicator lesions there were two reported complete responses, one each at the 5 and 50 μg dose level, lasting >36 and 20 months, respectively. However, there were no partial responses, and the overall median time to progression for the 28 patients was 29 days. There was also no difference in the frequency of immune response by dose level or by route of administration (subcutaneous or intradermal). Eleven patients were treated in the adjuvant setting, and the longest disease-free intervals, reported in 3 patients, were 8, 12, and 21 months, considerably shorter than in the current report.
The progression-free and overall survival data for advanced melanoma patients treated in the adjuvant setting can now be analyzed in context with data from a large phase III trial that included similar patients treated either with Canvaxin, an allogeneic vaccine, or placebo after being rendered surgically free of disease. In that study by Morton et al, 496 patients (out of a planned total enrollment of 670) with stage IV melanoma were treated in the adjuvant setting. The median DFS was 7-8 months with a 5-year DFS of 20.9-27.4 percent. The median overall survival was 32-39 months, with 5-year overall survival of 40%-45% [18
]. Although the results of that study do not support the continuation of Canvaxin in clinical development, they do confirm that highly selected stage IV patients who can be rendered surgically free of disease and who remain disease-free long enough after surgery to enroll in stage IV adjuvant cancer vaccine trials can have a prolonged duration of survival. These data contrast the median survival of 8-12 months, with 5-year survival of <5%, for patients who are typically enrolled in stage IV melanoma trials.
The current pilot study excluded patients with evidence of 25% progression of disease in visceral organs or appearance of new disease during the recuperative month after surgery, as these patients were considered to be candidates for other clinical trials at MD Anderson Cancer Center. Had patients with symptomatic or rapidly progressive disease been enrolled, many would have been inevaluable for biomarker response, a primary endpoint of this study. In favoring the recruitment of patients with relatively indolent or no clinical evidence of disease after surgery, the relative contribution of HSPPC-96 treatment to the long term survival of several of the patients with advanced melanoma cannot be ascertained. Furthermore, six of 11 patients with stage IV disease treated with HSPPC-96 in the adjuvant setting may have been favorably affected by prior or subsequent systemic treatment; for example, five of these patients each received at least 6 cycles of biochemotherapy.
Because of the limitations reported in this study, the hypothesis that immunoprotection is best achieved against unique tumor antigens can neither be confirmed nor refuted. The question also remains whether the doses and schedule tested in the current study were adequate to mount an effective immune response. Nonetheless, HSPPC-96 retains conceptual appeal as a tailored active immunotherapy approach.
Persistence of a robust CTL response is likely required to cause tumor shrinkage and this persistence is dependent on continuous antigenic stimulation [19
], administration of costimulatory cytokines, inhibition of negative costimulatory molecules (such as cytotoxic T-lymphocyte antigen 4 [20
].), and inhibition of immune-suppressing regulatory T cells [21
]. A benefit of adding IL2 to HSPPC-96 treatment was not demonstrated in a study by Amato et al. in renal cell carcinoma [22
There has been substantial advancement in the understanding of the critical attributes of autologous tumor-derived HSPPC-96 as a pharmaceutical product. Increased purity, yield and manufacturing success rate have been achieved through improvements in inhibition of tumor-associated proteases using a protease inhibitor cocktail and in mechanical tissue homogenization. In addition, process steps have been eliminated and chromatographic steps have been optimized. Allowing greater flexibility in the clinic, formal validation studies now support an extended 18-month shelf life, with further improvements expected. (Personal communication, Antigenics, Inc. Lexington, MA).
Kinetics studies are unraveling how peptides which are tightly bound to HSP-96 can nevertheless transfer to MHC class I molecules in an ATP-binding and ATPase dependent step [23
]. In an effort to limit acquisition of extraneous targets with HSPPC-96, synthetic immunogenic melanoma peptides -- such as MART-1, gp100, or MAGE-- have been bound to cloned HSP-96 and other chaperones, such as HSP-70 [25
]. Such engineered constructs are being evaluated for their ability to immunize more effectively than peptide alone [27
In summary, despite the lack of demonstrable efficacy of HSPPC-96 as treatment of patient with advanced melanoma in this pilot study, we have demonstrated that clinical-grade HSPPC-96 can be produced from even small amounts of metastatic melanoma tumor tissue and that treatment with HSPPC-96 is safe and well tolerated. HSPs acting as peptide chaperones continue to be an important area of development in the in vivo stimulation of professional APCs against autologous tumor cells.