Adjuvant Therapy for Melanoma in Dogs
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Vol. 5, 4249–4258, December 1999 Clinical Cancer Research 4249 Adjuvant Therapy for Melanoma in Dogs: Results of Randomized Clinical Trials Using Surgery, Liposome-encapsulated Muramyl Tripeptide, and Granulocyte Macrophage Colony-stimulating Factor1 E. Gregory MacEwen,2 Ilene D. Kurzman, colony-stimulating factor (rcGM-CSF) in dogs undergoing David M. Vail, Richard R. Dubielzig, surgery for oral melanoma. Ninety-eight dogs with histologically confirmed, clini- Karen Everlith, Bruce R. Madewell, cally staged, oral melanoma were entered into two random- 3 Carlos O. Rodriguez, Jr., Brenda Phillips, ized, double-blind, surgical adjuvant trials. In trial 1, 50 Courtney H. Zwahlen,4 Joyce Obradovich, dogs were stratified based on clinical stage and randomized Robert C. Rosenthal, Leslie E. Fox, to once a week L-MTP-PE or lipid equivalent (control). Mona Rosenberg, Carolyn Henry, and When all of the clinical stages were combined, no difference 5 in disease-free survival or in survival time (ST) were de- Janean Fidel tected. However, within stage I, dogs receiving L-MTP-PE University of Wisconsin, School of Veterinary Medicine, Madison, had a significant increase in ST compared with control, with Wisconsin 53706 [E. G. M., I. D. K., D. M. V., R. R. D., K. E., B. P.]; 80% of the dogs treated with L-MTP-PE still alive at >2 University of California, School of Veterinary Medicine, Davis, years. Within each stage II and stage III, there was no California 95616 [B. R. M., C. O. R., B. P., C. H. Z.]; Oakland Veterinary Referral Services, Bloomfield Hills, Michigan 48302 difference detected between the treatment groups. In trial 2, [J. O.]; Veterinary Specialists of Rochester, Rochester, New York 48 dogs were stratified on the basis of clinical stage and 14623 [R. C. R.]; University of Florida, College of Veterinary extent of surgery (simple resection or radical excision), Medicine, Gainesville, Florida 32610 [L. E. F.]; Veterinary Cancer treated with L-MTP-PE two times a week, and randomized Referral Group, Los Angeles, California 90064 [M. R.]; University of to rcGM-CSF or saline (placebo) given s.c. daily for 9 weeks. Missouri, College of Veterinary Medicine, Columbia, Missouri 65211 Within each stage and when all of the stages were combined, [C. H.]; and Special Veterinary Services, Berkeley, California 94704 [J. F.] there was no difference between the treatment groups. In both studies, stage I COM is associated with a better prog- nosis. No effect on survival was observed with regard to ABSTRACT tumor location in the oral cavity, sex, type/extent of surgery, Spontaneous canine oral melanoma (COM) is a highly or age. In a subset of dogs tested, pulmonary alveolar metastatic cancer, resistant to chemotherapy, and can serve macrophage cytotoxicity was enhanced with combined as a model for cancer immunotherapy. Liposome-encap- rcGM-CSF and L-MTP-PE but not in dogs treated with sulated muramyl tripeptide-phosphatidylethanolamine (L- L-MTP-PE alone. MTP-PE) can activate the tumoricidal activity of the mono- The present study indicates that after surgery, cyte-macrophage system following i.v. injection. The L-MTP-PE administered alone or combined with rcGM- objective of these studies was to evaluate the therapeutic CSF showed no significant antitumor activity in treating effectiveness of L-MTP-PE administered alone and com- advanced stage COM. In early stage COM, L-MTP-PE was bined with recombinant canine granulocyte macrophage shown to result in a prolongation of ST. Furthermore, this study provides additional rationale for the use of the dog model for human malignant melanoma. INTRODUCTION Received 3/4/99; revised 8/30/99; accepted 9/21/99. There is ample evidence that the immune system can The costs of publication of this article were defrayed in part by the modulate the progression and metastasis of malignant mela- payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to noma. Several biological agents and approaches have demon- indicate this fact. strated antitumor activity (1–10). The most widely used treat- 1 This work was supported by a grant from the Morris Animal Foun- ments include IFN-a (5, 6), IL-26 and adoptive immunotherapy dation, Englewood, CO, and Ciba-Geigy Ltd., Basel, Switzerland. 2 To whom requests for reprints should be addressed, at Department of Medical Sciences, University of Wisconsin, 2015 Linden Drive West, Madison, WI 53706. 3 Present address: University of Texas, M. D. Anderson Cancer Center, 6 The abbreviations used are: IL, interleukin; DFS, disease-free survival; 1515 Holcombe Boulevard., Houston, TX 77054. ST, survival time; PAM, pulmonary alveolar macrophage; MTP-PE, 4 Present address: Southwest Surgery and Oncology, 17672 Cowan muramyl tripeptide-phosphatidylethanolamine; L-MTP-PE, liposome- Avenue, Irvine, CA 92614. encapsulated MTP-PE; MLV, multilamellar vesicle; OSA, osteosar- 5 Present address: University of Zurich, Winterthurerstrasse 260, CH- coma; GM-CSF, granulocyte macrophage colony-stimulating factor; 8057 Zurich, Switzerland. rcGM-CSF, recombinant canine GM-CSF; ADCC, antibody-dependent Downloaded from clincancerres.aacrjournals.org on September 28, 2021. © 1999 American Association for Cancer Research. 4250 Adjuvant Immunotherapy for Canine Melanoma free of disease at 24 weeks (33). In a follow-up report on these same 18 patients, 4 remain free of disease more than 5 years after surgical resection and L-MTP-PE therapy (40). Undoubt- edly, the greatest antitumor activity of L-MTP-PE has been demonstrated in children with metastatic OSA leading to a Phase III randomized trial presently underway (41, 42). Our group has focused on spontaneous canine tumors as models for immunotherapy (43). We have previously reported the antimetastatic activity of L-MTP-PE, administered alone or in combination with chemotherapy, in surgical adjuvant ran- domized clinical trials in dogs with spontaneous OSA (44, 45) and hemangiosarcoma (46). In over 125 dogs with OSA, L- MTP-PE was found to prolong metastasis-free and overall STs when given alone or after systemic chemotherapy. In dogs with Fig. 1 Schematic diagram of the two oral melanoma clinical trials. hemangiosarcoma, L-MTP-PE administered concurrently with doxorubicin and cyclophosphamide, resulted in prolonged me- tastasis-free and overall STs compared with dogs receiving chemotherapy alone (46). (11), and active immunization using tumor vaccines (12–15). GM-CSF is a cytokine with a unique ability to stimulate Advances in melanoma treatment over the past decade have differentiation of hematopoietic progenitor cells into dendritic been directed at the role of adjuvant systemic therapy for pa- cells, a potent antigen-presenting cell (47, 48). GM-CSF has tients with melanoma at high risk of recurrence or metastasis (1, received particular attention for enhancing the antitumor activity 9, 10, 16). in transduced tumor cell vaccines (14, 15, 49). In vitro, GM- Canine melanoma is a highly metastatic spontaneous tumor CSF has been shown to slightly enhance cytotoxic activity of arising most commonly in the oral cavity (17–19). COM has a peripheral blood monocytes and lymphocytes and to markedly biological behavior that is similar to oral melanoma in humans. increase ADCC (50, 51) and enhanced monocyte cytotoxicity Melanomas are the most common neoplasm seen in the oral against a melanoma cell line (52). In vivo administration of cavity in dogs (20–22). These tumors arise in many breeds but GM-CSF in human cancer patients enhanced monocyte ADCC are more frequently identified in the dark pigmented breeds (53) and increased secretion of both TNF-a and IFN (54). In (e.g., Scottish terrier, Boston terrier, Airedale, and Cocker span- another study in cancer patients, GM-CSF enhanced monocyte iel). COM occurs most often in dogs that are 8–12 years of age, cytoxicity against colon HT29 tumor cells, although no antitu- roughly corresponding to the ages of highest prevalence in mor responses were seen (55). humans (40–60 years; Ref. 22). Metastasis is common and can This study was designed in a randomized manner to be seen in regional lymph nodes, lung, liver, brain, and kidney. evaluate the efficacy of L-MTP-PE when used in a surgical The treatment of choice is surgical excision, and, depending on adjuvant setting and administered alone or in combination with clinical stage, the median ST is 8–10 months, with death due to GM-CSF. metastasis (18, 19, 23, 24). The systemic activation of macrophages using a variety of techniques has been shown to inhibit the proliferation of or to MATERIALS AND METHODS destroy a wide range of syngeneic, allogeneic, and xenogeneic Patient Population. Ninety-eight dogs with previously tumors (25, 26). In rodents, dogs, and humans, a liposome- untreated, histologically confirmed spontaneous oral melanoma, encapsulated lipophilic derivative of muramyl dipeptide, known without radiographic evidence of distant metastasis, were stud- as MTP-PE, when administered in vivo can activate monocytes ied. The pretreatment evaluation included a complete physical and macrophages resulting in antitumor activity (27–38). It has examination, three-dimensional measurements of the primary been shown that L-MTP-PE can result in the regression of tumor, a complete blood count, serum chemistry profile, and established metastasis in the B16 murine melanoma model (39). urinalysis. For evaluation of metastatic disease, regional lymph In humans with melanoma, two Phase II trials using L-MTP-PE nodes were evaluated with fine needle aspiration or biopsy and have been reported (32, 33). In one study, 30 patients with radiographs of the thorax. Only dogs without evidence of overt measurable metastatic melanoma were treated, and, although no lung metastasis, in overall good health, and a tumor (including responses were detected, peripheral blood monocyte cytotoxic- regional lymph nodes) amenable to complete surgical removal ity was increased (32). In the other study, 18 stage III melanoma were eligible for entry into this study.