Posts Tagged ‘Clark’s multi-step model’

Author, Editor: Tilda Barliya, PhD

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Although melanoma accounts for only 4 percent of all dermatologic cancers, it is responsible for 80 percent of deaths from skin cancer; only 14 percent of patients with metastatic melanoma survive for five years (1). The incidence of melanoma is increasing worldwide, with a growing fraction of patients with advanced disease for which prognosis remains poor despite advances in the field (2). Treatment options are limited despite advances in immunotherapy and targeted therapy. For patients with surgically resected, thick (≥2 mm) primary melanoma with or without regional lymph node metastases, the only effective adjuvant therapy is interferon-α (IFN-α). However, because of the limited benefit upon disease-free survival and the smaller potential improvement of overall survival, the indication for IFN-α treatment remains controversial (2). A better understanding of melanoma immunosurveillance is therefore essential to enable the design of better, targeted melanoma therapies (4).

Risk factors (2):

  • Family history of melanoma, multiple benign or atypical nevi, and a previous melanoma
  • Immunosuppression
  • Sun sensitivity
  • Exposure to ultraviolet radiation

Each of these risk factors corresponds to a genetic predisposition or an environmental stressor that contributes to the genesis of melanoma and each factor is understood to various degrees at a molecular level. The Clark model of the progression of melanoma emphasizes the stepwise transformation of melanocytes to melanoma. The model depicts the proliferation of melanocytes in the process of forming nevi and the subsequent development of dysplasia, hyperplasia, invasion, and metastasis.


This Clark’s multi-step model, and predict that the acquisition of a BRAF mutation can be a founder event in melanocytic neoplasia. While mutations of the BRAF gene are frequent in melanomas on non-chronic sun damaged skin which are prevalent in Caucasians, acral and mucosal melanomas harbor mutations of the KIT gene as well as the amplifications of cyclin D1 or cyclin-dependent kinase 4 gene.

The choice of target antigens is key to the success of tumour vaccination or tumour immunotherapy. Melanoma candidate antigens include: (A) mutated or aberrantly expressed molecules (e.g. CDK4, MUM-1, beta-catenin) (B) cancer/testis antigens (e.g. MAGE, BAGE and GAGE) and (C) melanoma- associated antigens (MAA).

MAAs are self-antigens normally expressed during the differentiation of melanocytes and play a role in different enzymatic steps of melanogenesis. However, in transformed melanocytes (melanoma cells), MAAs are often overexpressed (4).

The main MAAs are tyrosinase, an enzyme that catalyses the production of melanin from tyrosine by oxidation, the tyrosinase-related proteins (TRP-1) and 2 (TRP-2), the glycoprotein (gp)100 (silver-gene) and MelanA/MART. It is thought that the specialized cell biology of melanin synthesis may favour the loading of MAA peptides into the antigen presentation pathway. 50% of melanoma patients have tumour-infiltrating lymphocytes (TILs) recognising tyrosinase and Melan A, indicating that these antigens are important in the natural melanoma immunosurveillance. Moreover, MAAs are well characterized in mice and humans, allowing the development of tetramers to detect antigen-specific immune responses.

Tα1 Mechanism of action

Tα1, a 28 amino acid peptide of ∼3.1 kDa, is endogenously produced by the thymus gland by the cleavage of its precursor pro-Ta1.  Although the fine immunologic mechanism(s) of action of T1 have not fully been elucidated, experimental evidence points to its strong immunomodulatory properties. In particular, it was reported that Ta1 enhances T cell–mediated immune responses by several mechanisms, including increased T cell production (i.e., CD4+, CD8+, and CD3+ cells), stimulation of T cell differentiation and/or maturation, reduction of T cell apoptosis, and restoration of T cell–mediated antibody production (5).

Furthermore, it was demonstrated that T1 acts on the immune system by modulating the release of proinflammatory cytokines (i.e., interleukin-2 (IL-2), interferon-gamma (IFN-)),12–14 and through the activation of natural killer and dendritic cells.12 In addition, T1 was also demonstrated to have direct effects on cancer cells by increasing the levels of expression of different tumor antigens and of components of the major histocompatibility complex class I, as well as by reducing cancer cell growth.

Together, these experimental findings bear relevance for cancer immunotherapy and suggest that T1 can activate innate and adaptive immune responses and modulate the immunophenotype of cancer cells, improving their immunogenicity and their recognition by the immune system.

Danielli R and colleagues have very nicely outlined the use of the Thymosin a1 in the clinical setting for treating melanoma (5) titled :”Thymosin a1 in melanoma: from the clinical trial setting to the daily practice and beyond”.  A large body of available preclinical in vitro and in vivo evidence points to thymosin alpha 1 (Ta1) as a useful immunomodulatory peptide,with significant therapeutic potential in metastatic melanoma in the absence of clinically meaningful toxicity.  The results emerging frominitial trials provide support of the ability of T1 to improve the clinical outcome of advanced melanoma patients through the activation of the immune system.

Ta1 and Clinical Trials in Melanoma

A large scale, randomized, phase II study was conducted at 64 European centers between 2002 and 2006 to investigate the efficacy of Ta1 administered in combination with DTIC (Dacarbazine) or with DTIC + IFNa, versus only DTIC + IFNa, in 488 previously untreated patients with cutaneous metastatic melanoma. The study was designed to evaluate the ability of Ta1 to potentiate the therapeutic efficacy of DTIC.

Patients were randomly assigned to five treatment groups: DTIC + IFNa and 1.6 mg of Ta1; DTIC + IFNa and 3.2 mg of T1; DTIC + IFN-a and 6.4 mg of Ta1; DTIC + 3.2 mg of Ta1; and DTIC + IFNa


The clinical rate (CBR), defined as the proportion of patients with a complete response, partial response, or stable disease, was significantly higher in patients who received Ta1 + DTIC than in those who received control therapy. Results in patients who received T1 (all groups combined) compared with those who received the control treatment

  • Improved progression-free survival (hazard ratio (HR): 0.80;
  • 95%confidence interval (CI): 0.63–1.01; P = 0.06) and
  • OS (median: 9.4 vs. 6.6 months)

These outcomes suggested to addition of Thymosin a1 to the treatment resulted in the reduction in the risk of mortality and disease progression in patients with metastatic malignant melanoma, and pointed to a poor effect of IFN- in the combination. More so, the poor results of the IFN group is not surprising due to the limited therapeutic activity of IFN observed in phase III clinical trials.

This study however have some limitations as standard assessment criteria, such as RECIST and WHO indications,  conventionally applied to cytotoxic agents, do not adequately capture some patterns of response observed in the course of immunotherapy; stemming from these considerations, immune-related response criteria (irRC) were developed to measure primary and secondary endpoints in immunotherapy clinical trials.

Therefore the above study might underestimate the therapeutic efficacy of Thymosin a1 since irRC criteria were not used.

In Summary:

A large scale phase III clinical trial should be designed to further explore the therapeutic activity of Thymosine a1 in melanoma patients with defined endpoints and irRC criteria. Moreover, combination studies should explore the activity of T1 in association with other approved agents, such as ipilimumab and vemurafenib or as maintenance therapy in melanoma patients who experience clinical benefit after treatment with these agents.

Also, because of the pleiotropic immunemechanism(s) of action of T1, including the upregulation of T cell–driven immune responses against specific tumor antigens, priming of immune responses and potentiation of antitumor T cell–mediated immune responses through the activation of Toll-like receptor 9 on dendritic cells, coupling Ta1 to cancer vaccines should be an additional useful therapeutic strategy to pursue. T1 could, in fact, prove helpful in overcoming the limited immunogenicity and the short-lived persistency of adequate immunologic antitumor responses frequently reported as potential causes of failure of therapeutic vaccines.


1. Arlo J. Miller, M.D.,., and Martin C. Mihm, Jr. Mechanisms of disease: Melanoma. N Engl J Med 2006 (6); 355:51-65.



2. Garbe C., Eigentler TK., Keilholz U., Hauschild A and Kirkwood JM. Systematic review of medical treatment in melanoma: current status and future prospects. Oncologist 2011;16(1):5-24.


3. http://flipper.diff.org/app/items/info/1983

4.  Träger U, Sierro S, Djordjevic G, Bouzo B, Khandwala S, et al. (2012) The Immune Response to Melanoma Is Limited by Thymic Selection of Self-Antigens. PLoS ONE 7(4): e35005. doi:10.1371/journal.pone.0035005.


5. Riccardo Danielli, Ester Fonsatti, Luana Calabr` o, Anna Maria Di Giacomo, and Michele Maio. Thymosin 1 in melanoma: from the clinical trial setting to the daily practice and beyond. Ann. N.Y. Acad. Sci. 1270 (2012) 8–12.



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