**8. Immunotherapy**

The immune system recruitment may represent a powerful and innovative strategy in cancer therapy. Genetic mutations and alterations in regulatory processes of cancer cells lead to expression of various tumor-related antigens that can be presented to cytotoxic T-lymphocytes by antigen-presenting cells. A major understanding of immune activation, especially T-lymphocyte activation, has identified multiple co-stimulatory and co-inhibitory pathways regulating this process. The two most important targets of immunotherapy are co-inhibitory receptors, such as CTLA-4 (cytotoxic T-lymphocyte-associated antigen 4) and programmed cell death-1 (PD-1) receptor, expressed on the T-lymphocyte surface [26].

A molecule of IL-2 was first approved by the US FDA for immunotherapy of melanoma. It is of limited use due to the serious toxic side effect of this treatment [27]. The first approved checkpoint blocking antibody was ipilimumab.

*Ipilimumab* (Yervoy®) is a human monoclonal IgG1 antibody that binds the human antigen CTLA-4 located on the surface of T-lymphocytes and blocks its interaction with molecule on the surface of antigen presenting cells. CTLA-4 is a key negative regulator of adaptive immune response and works as a brake on the immune response. Blocking immune response to anticancer leads to a longer and stronger activation of T-lymphocytes and, ideally, to an attack and destruction of the tumor tissue, resulting in long term remission for 15–20% of patients [28]. Although its effectiveness is tested with many carcinomas, the best results were achieved just in the treatment of melanoma [29]. Randomized clinical studies show that the treatment with ipilimumab leads to a significant extension of the survival of patients with metastatic melanoma. Side effects of ipilimumab are related to the mechanism of its action. Typical side effects are accompanied by diarrhea, skin rash, pruritus, enteritis, vitiligo, endocrinopathies and hepatotoxicity. Ipilimumab is approved in the USA for the treatment of patients with advanced melanoma and in Europe for patients with previously treated advanced melanoma.

dacarbazine is characterized with low overall response rates (approximately 10–15%) and there is no valid evidence of survival benefit [23]. Temozolomide is a monofunctional alkylating agent of the imidotetrazine class. It is stable at the acid pH of the stomach and adminis-

For years, the cornerstones of cancer treatment have been surgery, chemotherapy, and radiation therapy. Significant changes occurred in antitumor therapy for disseminated melanoma during the last decade. Detailed knowledge in the molecular biology of melanoma and immune response lead to the two directions: immunotherapy and targeted therapy. Before 2011, two approved drugs were used to treat patients with metastatic melanoma in the USA: dacarbazine and recombinant human interleukin-2 (IL-2) [24]. The treatment landscape for advanced stage melanoma was revolutionized in 2011 with the approval of ipilimumab and vemurafenib, both of which improved overall survival in phase III clinical trials. More recently, the targeted inhibitors dabrafenib and trametinib have demonstrated similar therapeutic profiles [25]. The latest approved (PD-1)-blocking antibody pembrolizumab is indi-

cated for the treatment of patients with unresectable or metastatic melanoma [26].

cell death-1 (PD-1) receptor, expressed on the T-lymphocyte surface [26].

checkpoint blocking antibody was ipilimumab.

The immune system recruitment may represent a powerful and innovative strategy in cancer therapy. Genetic mutations and alterations in regulatory processes of cancer cells lead to expression of various tumor-related antigens that can be presented to cytotoxic T-lymphocytes by antigen-presenting cells. A major understanding of immune activation, especially T-lymphocyte activation, has identified multiple co-stimulatory and co-inhibitory pathways regulating this process. The two most important targets of immunotherapy are co-inhibitory receptors, such as CTLA-4 (cytotoxic T-lymphocyte-associated antigen 4) and programmed

A molecule of IL-2 was first approved by the US FDA for immunotherapy of melanoma. It is of limited use due to the serious toxic side effect of this treatment [27]. The first approved

*Ipilimumab* (Yervoy®) is a human monoclonal IgG1 antibody that binds the human antigen CTLA-4 located on the surface of T-lymphocytes and blocks its interaction with molecule on the surface of antigen presenting cells. CTLA-4 is a key negative regulator of adaptive immune response and works as a brake on the immune response. Blocking immune response to anticancer leads to a longer and stronger activation of T-lymphocytes and, ideally, to an attack and destruction of the tumor tissue, resulting in long term remission for 15–20% of patients [28]. Although its effectiveness is tested with many carcinomas, the best results were achieved just in the treatment of melanoma [29]. Randomized clinical studies show that the treatment

tered orally with 100% bioavailability [22].

114 Human Skin Cancers - Pathways, Mechanisms, Targets and Treatments

**7. New approaches**

**8. Immunotherapy**

*Tremelimumab*, another drug of this group, is human therapeutic monoclonal antibody IgG2, with the same mechanism of action as ipilimumab. This antibody is currently in progress in phase II/III clinical study [30].

Ipilimumab, in combination with high dose IL-2, and tremelimumab, in combination with interferon alfa provide increased overall response rate, progression-free survival, or higher percentage of complete responses. *Interferon alfa* is FDA approved in adjuvant treatment for patients with high-risk melanoma and it has significant immunomodulatory effects [31–33]. Interferon alfa monotherapy has limited utility in the treatment of stage IV melanoma; therefore, its antitumor activity has led to profound investigation of its use in combination with other therapies [34].

Cancer immunotherapy can be achieved by inhibition of the PD-1/PD-Ll axes, which affect the overall survival in an important fraction of patients. PD-1 is an inhibitory receptor that is upregulated on activated lymphocytes. PD-1 has two known ligands, PD-L1 and PD-L2, which can be expressed on tumor and stromal cells; PD-L1 expression can be induced by cytokines produced by tumor-infiltrating lymphocytes [35].

*Pembrolizumab* (Keytruda®, Merck & Co) is the first anti-PD-1 immunotherapeutic agent approved by FDA. Keytruda® was granted FDA approval on September 4, 2014 for the treatment of patients with unresectable or metastatic melanoma. This molecule is a potent and highly selective humanized monoclonal antibody of IgG4-kappa isotype, designed to directly block the interaction between PD-1 receptor, expressed on T-cells, and its ligands, PD-L1 and PD-L2, without antibody-dependent cell-mediated or complement-dependent cytotoxicity. In practice, blocking PD-1 activity is believed to prevent inhibition of T-cell immune surveillance of tumors and, in some models, has resulted in decreased tumor growth [26]. The recommended dose of pembrolizumab is 2 mg/kg administered as an intravenous infusion over 30 min every 3 weeks until disease progression or unacceptable toxicity. Most common adverse reactions (reported in ≥20% of patients) included fatigue, cough, nausea, pruritus, rash, decreased appetite, constipation, arthralgia, and diarrhea [36].

Another approach, which has already been tested, is to combine anti-PD-1 and anti-CTLA-4 treatment and is represented by *nivolumab* (Opdivo®, Bristol-Meyers Squibb). Nivolumab is used alone or in combination with ipilimumab [37, 38]. Combination therapy with anti-CTLA-4 and anti-PD-1 monoclonal antibodies has recently led to remarkable antitumor effects, long-term survival and potential cures [39].

BRAF-mutant and BRAF-wild type patients, who progressed after ipilimumab therapy, were included into the Phase III study. One group of patients received nivolumab (3 mg/kg every 2 weeks), and in comparator group patients were treated with chemotherapy. Patients treated with nivolumab demonstrated higher response rate compared to the chemotherapy group—32% vs. 11% [40]. There was no statistical difference in two contemporary oncology median overall survivals between the study arm with nivolumab, 15.7 months, vs. the comparator group with chemotherapy, 14.4 months. The limitation of the clinical benefit of nivolumab could be related to the fact that control group patients (40%) received pembrolizumab, when progressed during chemotherapy. Furthermore, the number of patients with elevated lactate dehydrogenase levels and brain metastases was imbalanced, favoring the chemotherapy [41]. Adverse events are less frequent in patients treated with nivolumab than in those treated with ipilimumab or chemotherapy [4, 10]. The most frequently observed adverse events included fatigue, pruritus, diarrhea, rash, and nausea. The most commonly observed immune-related adverse events were pruritus, rash, diarrhea, vitiligo, hypothyroidism, and elevated aminotransferase activities [42].

Vaccines specific for cancer antigens exert antitumor effects by inducing cytotoxic T lymphocytes (CTLs) that recognize and attack antigenic cancer-derived peptides comprising 8–10 amino acid residues presented on major histocompatibility complex molecules on the cancer cell surface. Peptides, proteins, mRNA, DNA, and viral vectors can be used as cancer vaccines. Various peptides derived from Wilms' tumor gene-1, glycoprotein 100 (gp100), and melanoma-associated antigen 3 (MAGE-A3) have been used as vaccines

Possibilities for the Therapy of Melanoma: Current Knowledge and Future Directions

http://dx.doi.org/10.5772/intechopen.70368

117

Tumor antigen encoded by genes of the MAGE-A family, MAGE-A3 is of importance because they are expressed in a wide array of malignancies including melanoma, brain, breast, lung and ovarian cancer. Its ability to elicit spontaneous humoral and cellular immune responses has been shown in cancer patients. As antigen-specific immune responses can be stimulated by immunization with MAGE-A3, several clinical trials have used MAGE-A3 vaccines to observe clinical responses. The frequent expressions of this antigen in various tumors and its immunogenicity in cancer patients have led to application of this antigen in cancer immunotherapy. Indeed, the initial trials performed with MAGE-A3 peptides showed no significant toxicity [48]. Vaccination with a tumor-specific MAGE-A3 peptide, even without adjuvant, has been shown to induce a CTL response in a melanoma patient followed by reduction in tumor mass. MAGE-A3 protein produced by recombinant technology is more popular in clinical trials as a result of its potential to activate a wide range of T-cell responses as well as its potential application in a larger population of patients with MAGE-A3 expressing tumors [49]. As MAGE-A3 specific therapies have not reached their final goals to cause a significant improvement in the survival of patients. For future perspective of MAGE-A3 therapy, studies are needed to find the most effective vaccine formulations, the most immunogenic adjuvants

Over the past several years, oncolytic viruses for treating various cancers have been investigated. Oncolytic viruses play a role in cancer vaccination because antigen-specific immunity is effectively evoked against components released by destroyed cancer cells due to virusinduced production of type I interferon. The effects of oncolytic virus therapy are mediated not only by direct cell disruption, but also by indirect induction of cancer-specific immune

Oncolytic viruses selectively replicate within and lyse cancer cells without damaging normal cells. On October 27, the FDA approved the first oncolytic virus therapy, *talimogene laherparepvec (Imlygic™, T-VEC)*. The agency approved T-VEC for the treatment of some patients with unresectable cutaneous, subcutaneous, and nodal lesions in melanoma recurrent after initial surgery [50]. T-VEC is a modified herpes simplex virus, type 1 (HSV-1) that has undergone genetic modifications to promote selective tumor cell replication, while reducing viral pathogenicity and promoting immunogenicity. T-VEC improves overall response rate and durable response rate as a single agent, shows promise in combination therapy with immunotherapy, and is well tolerated. Ongoing trials will determine if T-VEC has a role in early

as well as the most applicable criteria for selection of patients.

**10. Oncolytic virus therapy**

responses [48].

against melanoma [47].

In the study of Pyo and Kang the effects of various immunotherapeutic agents and chemotherapy for unresected or metastatic melanomas were compared. They performed a network meta-analysis using a Bayesian statistical model to compare objective response rate of various immunotherapies from 12 randomized controlled studies. The estimated overall response rates of immunotherapy and chemotherapy were 0.224 and 0.108, respectively. The overall response rates of immunotherapy in untreated and pretreated patients were 0.279 and 0.176, respectively. In network meta-analysis, the odds ratios for overall response rate of nivolumab (1 mg/ kg)/ipilmumab (3 mg/kg), pembrolizumab (10 mg/kg) and nivolumab (3 mg/kg) were 8.54, 5.39 and 4.35, respectively, compared with chemotherapy alone. Their results showed that various immunotherapies had higher overall response rates rather than chemotherapy alone [43].

Except immunological checkpoint blockades the approach of *adoptive T cell therapy* seems to be a highly promising in use against cancer including melanoma. The ability of T cells to specifically lyse tumor cells and secrete cytokines to recruit and support immunity against cancer make them an attractive proposition for therapy. Since the first idea in 1989 to genetically redirect T cells, a lot of experiments have been performed. Recent methods of generating tumor-specific T cells include the genetic modification of patient's lymphocytes with receptors to endow them with tumor specificity. These T cells are then expanded *in vitro* followed by infusion of the patient in adoptive cell transfer protocols. Genes used to modify T cells include those encoding T-cell receptors and chimeric antigen receptors. Several trials with gene-modified T cells are ongoing and some remarkable responses have been reported [44]. In fact, current adoptive T cell therapy response rates are 80–90% for hematological malignancies and 30% for metastatic melanoma refractory to multiple lines of therapy. Although these results are encouraging, there is still much to be done to fulfill potential of adoptive T cell therapy, specifically with regard to improving clinical efficacy, expanding clinical indications and reducing toxicity [45].

## **9. Cancer vaccines**

Melanoma vaccines have the goal to induce long lasting immunity against melanoma to prevent the development of metastases. However, melanoma cells express many different tumorassociated antigens. Ideally, vaccines need to contain all these different tumor-associated antigens for antigen-presenting-cells (APC) to induce an adequate immune response [46].

Vaccines specific for cancer antigens exert antitumor effects by inducing cytotoxic T lymphocytes (CTLs) that recognize and attack antigenic cancer-derived peptides comprising 8–10 amino acid residues presented on major histocompatibility complex molecules on the cancer cell surface. Peptides, proteins, mRNA, DNA, and viral vectors can be used as cancer vaccines. Various peptides derived from Wilms' tumor gene-1, glycoprotein 100 (gp100), and melanoma-associated antigen 3 (MAGE-A3) have been used as vaccines against melanoma [47].

Tumor antigen encoded by genes of the MAGE-A family, MAGE-A3 is of importance because they are expressed in a wide array of malignancies including melanoma, brain, breast, lung and ovarian cancer. Its ability to elicit spontaneous humoral and cellular immune responses has been shown in cancer patients. As antigen-specific immune responses can be stimulated by immunization with MAGE-A3, several clinical trials have used MAGE-A3 vaccines to observe clinical responses. The frequent expressions of this antigen in various tumors and its immunogenicity in cancer patients have led to application of this antigen in cancer immunotherapy. Indeed, the initial trials performed with MAGE-A3 peptides showed no significant toxicity [48]. Vaccination with a tumor-specific MAGE-A3 peptide, even without adjuvant, has been shown to induce a CTL response in a melanoma patient followed by reduction in tumor mass. MAGE-A3 protein produced by recombinant technology is more popular in clinical trials as a result of its potential to activate a wide range of T-cell responses as well as its potential application in a larger population of patients with MAGE-A3 expressing tumors [49]. As MAGE-A3 specific therapies have not reached their final goals to cause a significant improvement in the survival of patients. For future perspective of MAGE-A3 therapy, studies are needed to find the most effective vaccine formulations, the most immunogenic adjuvants as well as the most applicable criteria for selection of patients.
