Annals of Surgical Oncology

, Volume 20, Issue 4, pp 1128–1135 | Cite as

Plasma Cytokine Analysis in Patients with Advanced Extremity Melanoma Undergoing Isolated Limb Infusion

  • Gina Shetty
  • Georgia M. Beasley
  • Sara Sparks
  • Michael Barfield
  • Melanie Masoud
  • Paul J. Mosca
  • Scott K. Pruitt
  • April K. S. Salama
  • Cliburn Chan
  • Douglas S. Tyler
  • Kent J. Weinhold
Regional Cancer Therapies



Preprocedure clinical and pathologic factors have failed to consistently differentiate complete response (CR) from progressive disease (PD) in patients after isolated limb infusion (ILI) with melphalan for unresectable in-transit extremity melanoma.


Multiplex immunobead assay technology (Milliplex MAP Human Cytokine/Chemokine Magnetic Bead Panel, Millipore Corp., Billerica, MA; and Magpix analytical test instrument, Luminex Corp., Austin, TX) was performed on pre-ILI plasma to determine concentrations of selected cytokines (MIP-1α, IL-1Rα, IP-10, IL-1β, IL-1α, MCP-1, IL-6, IL-17, EGF, IL-12p40, VEGF, GM-CSF, and MIP-1β) on a subset of patients (n = 180) who experienced CR (n = 23) or PD (n = 24) after ILI. Plasma from normal donors (n = 12) was also evaluated.


Of 180 ILIs performed, 28 % (95 % confidence interval 22–35, n = 50) experienced a CR, 14 % (n = 25) experienced a partial response, 11 % (n = 21) had stable disease, 34 % (n = 61) had PD, and 13 % (n = 23) were not evaluable for response. Tumor characteristics and pharmacokinetics appeared similar between CR (n = 23) and PD (n = 24) patients who underwent cytokine analysis. Although there were no differences in cytokine levels between CR and PD patients, there were differences between the melanoma patients and controls. MIP-1α, IL-1Rα, IL-1β, IL-1α, IL-17, EGF, IL-12p40, VEGF, GM-CSF, and MIP-1β were significantly higher in normal controls compared to melanoma patients, while IP-10 was lower (p < 0.001) in controls compared to melanoma patients.


Patients with unresectable in-transit melanoma appear to have markedly decreased levels of immune activating cytokines compared to normal healthy controls. This further supports a potential role for immune-targeted therapies and immune monitoring in patients with regionally advanced melanoma.


Melanoma Progressive Disease Melanoma Patient Complete Response Rate Isolate Limb Infusion 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



Supported in part by T32 Grant CA093245-10 from NIH (G.M.B.) and Duke Translational Research Institute CTSA Grant (UL1RR024128). The ADH-1 trial was supported by a grant from Adherex Technologies, Inc. Bayer Healthcare Pharmaceuticals provided study drug (sorafenib, Nexavar) for the phase I trial of systemic sorafenib and regional melphalan. D.S.T. is on the speaker’s bureau of Novartis, has been a Scientific Advisory Board Member for Roche/Genetech, and has received clinical trial support from Merck/ScheringPlough Corporation.


  1. 1.
    Pawlik TW, Ross MI, Johnson MM, et al. Predictors and natural history of in-transit melanoma after sentinel lymphadenectomy. Ann Surg Oncol. 2005;11:1612–61.Google Scholar
  2. 2.
    van Poll D, Thompson JF, McKinnon JG, et al. A sentinel node biopsy procedure does not increase the incidence of in-transit recurrence in patients with primary melanoma. Ann Surg Oncol. 2005;12:597–608.PubMedCrossRefGoogle Scholar
  3. 3.
    Raymond AK, Beasley GM, Broadwater G, et al. Current trends in regional therapy for melanoma: lessons learned from 225 regional chemotherapy treatments between 1995 and 2010 at a single institution. J Am Coll Surg. 2011;213:306–16.PubMedCrossRefGoogle Scholar
  4. 4.
    Beasley GM, Caudle A, Petersen RP, et al. A multi-institutional experience of isolated limb infusion: defining response and toxicity in the United States. J Am Coll Surg. 2009;208:706–16.PubMedCrossRefGoogle Scholar
  5. 5.
    Lindner P, Doubrovsky A, Kam PCA, et al. Prognostic factors after isolated limb infusion with cytotoxic agents for melanoma. Ann Surg Oncol. 2002;9:127–36.PubMedGoogle Scholar
  6. 6.
    Kroon HM, Moncrieff M, Kam PCA, et al. Outcomes following isolated limb infusion for melanoma: a 14-year experience. Ann Surg Oncol. 2008;15:3003–13.PubMedCrossRefGoogle Scholar
  7. 7.
    Beasley GM, Petersen RP, Yoo JS, et al. Isolated limb infusion for in-transit malignant melanoma of the extremity: a well tolerated but less effective alternative to hyperthermic isolated limb perfusion. Ann Surg Oncol. 2008;15:2195–205.PubMedCrossRefGoogle Scholar
  8. 8.
    Thompson JF, Hunt JA, Shannon KF, et al. Frequency and duration of remission after isolated limb perfusion for melanoma. Arch Surg. 1997;132:903–7.PubMedCrossRefGoogle Scholar
  9. 9.
    Augustine CK, Jung SH, Sohn I, et al. Gene expression signatures as a guide to treatment strategies for in-transit metastatic melanoma. Mol Cancer Therapy. 2010;9:779–90.PubMedCrossRefGoogle Scholar
  10. 10.
    Hong M, Puaux AL, Huang C, et al. Chemotherapy induces intratumoral expression of chemokines in cutaneous melanoma, favoring T cell infiltration and tumor control. Cancer Res. 2011;71:6997.PubMedCrossRefGoogle Scholar
  11. 11.
    Sznol M. Molecular markers of response to treatment for melanoma. Cancer J. 2011;17:127–33.PubMedCrossRefGoogle Scholar
  12. 12.
    Boni A, Cogdill AP, Dang P, et al. Selective BRAFV600E inhibition enhances T-cell recognition of melanoma without affecting lymphocyte function. Cancer Res. 2010;70:5213–9.PubMedCrossRefGoogle Scholar
  13. 13.
    Galluzzi L, Senovilla L, Zitvogel L, et al. The secret ally: immunostimulation by anticancer drugs. Nat Rev Drug Discov. 2012;11:215–33.PubMedCrossRefGoogle Scholar
  14. 14.
    Nardin A, Wong WC, Tow C, et al. Dacarbazine promotes stromal remodeling and lymphocyte infiltration in cutaneous melanoma lesions. J Invest Dermatol. 2011;131:1896–905.PubMedCrossRefGoogle Scholar
  15. 15.
    Donepudi M, Jovasevic VM, Raychaudhuri P, Mokyr MB. Melphalan-induced up-regulation of B7-1 surface expression on normal splenic B cells. Cancer Immunol Immunother. 2003;52:162–70.PubMedGoogle Scholar
  16. 16.
    Sabatino M, Kim-Schulze S, Panelli MC, et al. Serum vascular endothelial growth factor and fibronectin predict clinical response to high-dose interleukin-2 therapy. J Clin Oncol. 2009;27:2645–52.PubMedCrossRefGoogle Scholar
  17. 17.
    Yurkovetsky ZR, Kirkwood JM, Edington HD, et al. Multiplex analysis of serum cytokines in melanoma patients treated with interferon-alpha 2b. Clin Cancer Res. 2007;13:2422–28.PubMedCrossRefGoogle Scholar
  18. 18.
    Balch CM, Gershenwald JE, Soong SJ, et al. Final version of 2009 AJCC melanoma staging and classification. J Clin Oncol. 2009;27:6199–206.PubMedCrossRefGoogle Scholar
  19. 19.
    Therasse P, Arbuck SG, Eisenhauer EA, et al. New guidelines to evaluate the response to treatment in solid tumors. J Natl Cancer Inst. 2000;92:205–16.PubMedCrossRefGoogle Scholar
  20. 20.
    McMahon N, Cheng TY, Beasley GM, et al. Optimizing melphalan pharmacokinetics in regional melanoma therapy: does correcting for ideal body weight alter regional response or toxicity? Ann Surg Oncol. Apr. 2009;16:953–61.PubMedCrossRefGoogle Scholar
  21. 21.
    Beasley GM, Riboh JC, Augustine CK, et al. A prospective multi-center phase II trial of systemic ADH-1 in combination with melphalan via isolated limb infusion (M-ILI) in patients with advanced extremity melanoma. J Clin Oncol. 2011;29:1210–5.PubMedCrossRefGoogle Scholar
  22. 22.
    Beasley GM, Coleman AP, McMahon N, et al. A phase I multi-institutional study of systemic sorafenib in conjunction with regional melphalan for in-transit melanoma of the extremity. Ann Surg Oncol. 2012;19:3896–905.PubMedCrossRefGoogle Scholar
  23. 23.
    Ehrsson H, Eksborg S, Lindfors A. Quantitative determination of melphalan in plasma by liquid chromatography after derivatization with N-acetylcysteine. J Chromatogr. 1986;380:220.Google Scholar
  24. 24.
    Freedman JA, Tyler DS, Nevins JR, Augustine CK. Use of gene expression and pathway signatures to characterize the complexity of human melanoma. Am J Pathol. 2011;178:2513–22.PubMedCrossRefGoogle Scholar
  25. 25.
    Yao KA, Hsueh EC, Essner R, et al. Is sentinel lymph node mapping indicated for isolated local and in-transit recurrent melanoma? Ann Surg. 2003;238:743–7.PubMedCrossRefGoogle Scholar
  26. 26.
    Tjin EP, Konijnenberg D, Krebbers G, et al. T cell immune function in tumor, skin and peripheral blood of advanced stage melanoma patients: implications for immunotherapy. Clin Cancer Res. 2011;17:5736–47.PubMedCrossRefGoogle Scholar
  27. 27.
    Balachandran VP, Cavnar MJ, Zeng S, et al. Imatinib potentiates antitumor T cell responses in gastrointestinal stromal tumor through the inhibition of Ido. Nat Med. 2011;17:1094–100.PubMedCrossRefGoogle Scholar
  28. 28.
    Rivoltini L, Carrabba M, Huber V, et al. Immunity to cancer: attack and escape in T lymphocyte–tumor cell interaction. Immunol Rev. 2002;188:97–113.PubMedCrossRefGoogle Scholar
  29. 29.
    Harlin H, Meng Y, Peterson AC, et al. Chemokine expression in melanoma metastases associated with CD8+ T-cell recruitment. Cancer Res. 2009;69:3077–85.PubMedCrossRefGoogle Scholar
  30. 30.
    Erdag G, Schaefer JT, Smolkin ME, et al. Immunotype and immunohistologic characteristics of tumor-infiltrating immune cells are associated with clinical outcome in metastatic melanoma. Cancer Res. 2012;72:1070–80.PubMedCrossRefGoogle Scholar
  31. 31.
    Obeid M, Tesniere A, Ghiringhelli F, et al. Calreticulin exposure dictates the immunogenicity of cancer cell death. Nat Med. 2007;13:54–61.PubMedCrossRefGoogle Scholar

Copyright information

© Society of Surgical Oncology 2013

Authors and Affiliations

  • Gina Shetty
    • 1
  • Georgia M. Beasley
    • 2
  • Sara Sparks
    • 2
  • Michael Barfield
    • 2
  • Melanie Masoud
    • 1
  • Paul J. Mosca
    • 2
  • Scott K. Pruitt
    • 2
  • April K. S. Salama
    • 4
  • Cliburn Chan
    • 3
  • Douglas S. Tyler
    • 2
  • Kent J. Weinhold
    • 1
    • 2
  1. 1.Department of ImmunologyDuke UniversityDurhamUSA
  2. 2.Department of SurgeryDuke UniversityDurhamUSA
  3. 3.Department of Biostatistics and BioinformaticsDuke UniversityDurhamUSA
  4. 4.Department of MedicineDuke UniversityDurhamUSA

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