Please note: Below is an entry draft based on your previous paper, which is wrirren tightly around the entry title. Since it may not be very comprehensive, we kindly invite you to modify it (both title and content can be replaced) according to your extensive expertise. We believe this entry would be beneficial to generate more views for your work. In addition, no worry about the entry format, we will correct it and add references after the entry is online (you can also send a word file to us, and we will help you with submitting).

Definition

oncogene-directed therapy and immunotherapy represent the two most promising avenues for patients with metastatic melanoma. The recent oncogene-directed therapeutic, vemurafenib, usually produces high level of tumor shrinkage and survival benefits in many patients with B-RAF^V600E mutant melanomas, although the fast and high degrees of responses are likely short-lived. Conversely, the newly-approved immunotherapeutic, ipilimumab, produces durable responses in patients presenting CTLA-4 T-cell surface protein. Nevertheless, the possible synergy in combining these two therapeutic strategies primarily rely on the rational design of medical protocols (e.g., sequence and timing of agent administration; drug selectivity; compatibility of combined therapies i.e., adoptive T cell or agents, i.e., MEK inhibitor trametinib, PD-1 and PDL-1 blockers). Improved therapeutic protocols shall overcome therapeutic limitations such as the (i) tolerability and safety (i.e., minimal toxic side-effects); (ii) progression free survival (e.g., reduced relapse disease frequency); (iii) duration response (i.e., decreased drug resistance). Eventually, multidisciplinary approaches are still requested (e.g., genomics for personalized medicine, nanomedicine to overcome low free-drug bioavailability and targeting, systematic search of “melanoma stem cells” to enhance the prognosis and develop more valuable theranostics).

1. Introduction 

Melanoma (from Greek—melas: "“dark"”) is a tumor originated from malignant transformation of melanocytes (i.e., melanin pigment-producing cells) that can be found in the skin, bowel, and eye [1].

According to the estimations provided by the American Cancer Society (ACS) in 2010, 68.130 new cases of melanomas were diagnosed and approximately 8.700 people died from this cancer [2]. The incidence of melanoma in the US has increased of about three folds between the last three decades (i.e., from 7.89 per 100.000 in 1975 to 22.52 per 100.000 in 2008) [3]. Clinical and epidemiological data suggest that several risk factors can contribute to the increased incidence: (i) extensive or repeated exposure to sunlight [4]; (ii) individuals with family history of melanoma (5–12% of all reported cases) [5]; (iii) high nevi count and dysplastic nevus [6], thereby suggesting the need to perform a biopsy of the suspicious lesion. The biopsy permits to establish not only an accurate diagnosis but also to define the optimal staging and proceed earlier with the appropriate therapy (e.g., surgery, chemotherapy, and/or radiotherapy).

Metastatic melanoma (i.e., advanced malignant melanoma) is the most aggressive form of skin cancer with a median overall survival (OS) of only few months (8 to 18 months) [2]. This fact could be mainly explained by the modest results obtained with dacarbazine (DITC) and high-dose interleukin 2 (HD IL-2), the two unique FDA-approved therapies for metastatic melanoma until 2011 ^[7][8][9][7–9]. Indeed, DITC is limited by a low response rate (RR of 5% to 15%) and an insufficient OS (about 8 months) [7]. Besides, HD IL-2 is also limited by a low RR (6% to 10%), a short duration of responses in most patients as well as a severe toxicity ^[8][9][8, 9]. Since 2011, 3 new agents have been approved for the treatment of advanced melanoma by the Food and Drug Administration (FDA) ^[10][11][12][10–12]: (i) vemurafenib, a mutant

inhibitor, recommended for unresectable or metastatic melanoma [10]; (ii) ipilimumab, an anti-CTLA-4 monoclonal antibody, also preconized for the treatment of unresectable or metastatic melanoma [11]; (iii) pegylated interferon alpha-2b (PEG-IFN), a covalent conjugate of the polyethylene glycol (PEG) with the recombinant α-2b interferon (IFN), long-time used to treat chronic hepatitis patients infected with hepatitis c virus [12], and currently recommended as adjuvant treatment for stage III melanoma [13].

Vemurafenib has emerged as a highly selective mutant inhibitor with little effect on wild type B-RAF, thereby demonstrating significant tumor regression (> 40%) while minimizing side effects in a large number of patients with metastatic melanoma ^[14][15][16][14–16]. Nevertheless, the single use of this oncogene-targeted agent presents the following main disadvantages: (i) short median duration of response (MDR) and progression-free survival (PFS) (i.e., about 6 months only) [16]; (ii) low RR in patients who harbor mutations other than V₆₀₀E. The proportion of these patients ranges between 10% and 30% (e.g., V₆₀₀K is present in 5% to 20% of patients with melanoma) ^[17][18][17, 18].

Ipilimumab was designed and developed to block the cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4), thereby increasing the T-cell activity and promoting antitumor activity in patients with cancers [19]. Thereby, in patients with unresectable or metastatic melanoma, ipilimumab plus DITC versus DITC alone significantly improved the OS (about 11 months versus 9 months, resp.) and RR (about 15% versus 10%, resp.) ^[20][21][20, 21].

PEG-IFN, similarly to high-dose interferon (HDI or Intron A) ^[1][22][1, 22], has been approved for the adjuvant treatment (after surgical resection) of stage III melanoma patients. This approval was mainly based on final results of a recent randomized phase III trial organized by the European Organization for Research and Treatment of Cancer (EORTC) 18991 that showed greater relapse-free survival (RFS of 45.6%) in comparison to observation (38.9%), although no significant effect on OS was noticed ^[12][23][12, 23]. Previously, in an open-label phase 2 study, the efficacy and safety of PEG-IFN in combination with temozolomide were investigated in patients with metastatic melanoma without brain metastases [24]. The RR of this combination reached 31% of the patients, the median OS was 12 months, and no patient developed brain metastases while receiving study treatment, which was besides well tolerated. Up to date, and in the best of my knowledge, it remains unknown whether PEG-IFN can provide better efficacy and safety results than the biochemotherapy combining cisplatin, vinblastine, DTIC plus IL-2 (Proleukin), and interferon. Indeed, a recent phase 3 trial (SWOG S0008) only assessed the efficacy and safety of this biochemotherapy versus HDI in patients with high-risk melanoma [25]. The results showed major improvement in the median RFS in favor of biochemotherapy (4.3 years versus 1.9 years with HDI). OS, however, was exactly the same (56% at 5 years), and acute grade 4 toxicity was more frequent with biochemotherapy. All together, the biochemotherapy can be considered as a better adjuvant treatment than HDI. Currently, both HDI and PEG-IFN are considered as category 2B due to their limited benefits, and so will not be further detailed in this paper.

Eventually, in one hand, these new exciting chemotherapies represent a great hope for the physicians and patients with advanced melanoma. In the other hand, their respective limitations clearly emphasize the importance of developing novel treatment strategies (e.g., cell-based therapies, advanced and rational combinatorial therapeutic approaches, nanodrug formulations). These alternative therapeutic options might help to improve OS, PFS, RFS, RR, and MDR while minimizing toxic adverse events, thereby contributing in fine to the quality of the patient'’s life.

2. Latest FDA-Approved Drugs for Advanced/Metastatic Melanoma

Most patients with unresectable stage III or stage IV disease require systemic treatment rather than metastasectomy.

2.1. Oncogene-Directed Therapy: Mutant B-RAF Inhibitors

Melanoma is a molecularly heterogeneous disease with approximately half (40%–60%) of the cutaneous melanoma cells harboring an activating mutation in the B-RAF gene, which encodes a serine/threonine kinase protein kinase, and most of the mutations (>70%) are V₆₀₀E (i.e., substitution of valine for glutamate at amino acid position 600) ^{[17][26][27][28]}[17, 26–28]. Because mutated B-RAF leads to constitutive activation of the mitogen-activated protein kinase pathway (MAPK) that, in turn, increases the cellular proliferation and drives the oncogenic activity ^[29][30][29, 30], intensive research has consisted to selectively inhibit mutated B-RAF in patients with melanoma (e.g., studies with sorafenib, a multitargeted kinase inhibitor), but the results were globally disappointing due to off-target side effects mainly induced through inhibition of wild type B-RAF ^{[14][31][32][33][34][35]}[14, 31–35].

Among highly selective B-RAF inhibitors, only the recent FDA-approved vemurafenib (formerly PLX4032, currently marketed as Zelboraf and initially developed by Genentech Roche) is capable of silencing mutant B-RAF^V600E without interfering with wild type B-RAF. Indeed, in a phase 2 clinical trial involving patients with metastatic melanoma harboring B-RAF^V600E mutation (n=132), vemurafenib demonstrated substantial tumor regression in 81% of the cases, a RR of 52%, and a MDR of 6.8 months ^[14][15][16][14–16]. Further, in a phase 3 clinical trial (BRIM3) involving previously untreated patients (n=675), vemurafenib was much better than DITC in terms of RR (48% versus 5%, resp.), PFS (5.3 months versus 1.6 months, resp.), and percent of patients alive at six months (OS of 84% versus 64%, resp.) [10]. Also, in a recent open-label pilot study, it was stated that vemurafenib could be beneficial for previously treated metastatic melanoma patients with brain metastases [36]. Besides, common adverse events associated with vemurafenib included accelerated growth of cutaneous squamous cell carcinomas (SCCs) and keratoacanthomas ^{[10][37][38][39][40]}[10, 37–40], most probably through paradoxical activation of MAPK signaling (about 20–25% of the patients with advanced melanoma) ^{[37][38][39][40]}[37–40].

Eventually, vemurafenib represents an excellent model for successful targeted anticancer therapy (i.e., high RR and low toxicity) in patients with B-RAF^V600E mutations [41]. Nevertheless, these clinical benefits are counterbalanced by the relatively short MDR, high selectivity for the B-RAF^V600E mutation, and related toxicities of the drug. Owing to consideration that 10% to 30% of patients have a non-B-RAF^V600E mutation (e.g., B-RAF^V600K mutation is present in 5% to 20% of melanoma patients) ^[17][18][17, 18], further studies are required to examine the efficacy of vemurafenib, alone or in combination, in patients with a non-B-RAF^V600E mutation. These studies are important to avoid useless administration of vemurafenib in a subset of patients, who might otherwise become resistant to the drug. Alternatively, rational combination of vemurafenib with other agents (e.g., ipilimumab) might circumvent an eventual drug resistance and/or further improve the clinical outcome of the patients (e.g., MDR, OS).

2.2. Immunotherapy: CTLA-4 Inhibitors

Melanoma is one of the most immunogenic tumors due to the presence of tumor infiltrating lymphocytes (TIL) in resected melanoma, clinical responses to immune stimulation, and occasional spontaneous regressions. CTLA-4 expression is necessary for activation of self-regulation of T cells, and so CTLA-4 inhibitors could represent serious therapeutic options to generate T-cell hyperresponsiveness and overcome tumor immune escape [19].

Up to date, ipilimumab (formerly MDX-010, MDX-101, or MDX-CTLA-4, currently marketed as Yervoy and initially developed by Bristol-Myers Squibb) is a fully human IgG1 monoclonal antibody that blocks CTLA-4, subsequently increasing the T-cell activity and promoting an antitumor activity and represents the only approved immunotherapeutic agent for systemic treatment [20]. In the first phase 3 randomized trial involving patients with previously treated unresectable stage III or IV melanoma (n=676), ipilimumab compared to the glycoprotein 100 peptide (gp100) vaccine demonstrated an improved median OS (10.1 months versus 6.4 months, resp.) and a much better RR (10.9% versus 1.5%, resp.), albeit the occurrence of toxicities with ipilimumab, including grade 3 or 4 immune-related adverse events (e.g., enterocolitis, hepatitis, and dermatitis) and deaths, was higher than with gp100 (10–15% versus 3%, resp.) [11]. Ipilimumab plus gp100, compared to gp100, did not improve the OS observed with ipilimumab alone (10.0 months versus 10.1 months, resp.) [11]. In the second phase 3 randomized trial involving previously untreated patients with metastatic melanoma (n=502), ipilimumab combined with DITC demonstrated a modest but statistically significant improvement in OS compared to DITC plus placebo (11.2 months versus 9.1 months, resp.) as well as a better overall RR (15.2% versus 10.3%, resp.) ^[20][21][20, 21]. Interestingly, survival rates over the years were always significantly higher in the ipilimumab-DITC group than in the group treated with the single agent DITC (at 1 year: 47.3% versus 36.3%; at 2 years: 28.5% versus 17.9%; at 3 years: 20.8% versus 12.2%, resp.), clearly demonstrating that ipilimumab is able to confer a durable response (MDR of 19.3 months versus 8.1 months, resp.). Nevertheless, median PFS was barely improved (2.8 months versus 2.6 months, resp.). Also, grade 3 or 4 adverse events (e.g., hepatitis) occurred more frequently in patients treated with ipilimumab plus DITC than in patients treated with DITC (plus placebo) (56.3% versus 27.5%, resp.), although low rates of gastrointestinal events and no drug-related deaths occurred in the ipilimumab-DITC group ^[20][21][20, 21].

Eventually, although the OS and MDR noticed with ipilimumab are higher than that one observed with vemurafenib, the most important limitation of this drug tested alone or in combination remains the modest RR. This strongly suggests a need for rational combination between ipilimumab and other commercially available free- or nanoencapsulated drugs (e.g., vemurafenib and bevacizumab, resp.) that might provide complementary clinical benefits.

3. Conclusions

The incidence of metastastic melanoma is increasing worldwide. Vemurafenib and ipilimumab, based on their respective success rates, are bringing hopes to physicians and patients. Vemurafenib has emerged as a highly selective B-RAF^V600E mutant melanoma inhibitor and could display good response rates in patients with unresectable or metastatic melanoma. Nevertheless, its main disadvantage remains the short median duration response as well as its limited use to patients who harbor mutations other than B-RAF^V600E (e.g., B-RAF^V600K). Besides, ipilimumab was developed to block the cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) in patients with unresectable or metastatic melanoma. Interestingly, ipilimumab presents the opposite main advantage and disadvantage than vemurafenib. Several clinical trials are underway to address the question of rational combination of those two approved drugs, together and/or separately with other potential therapeutic targets (PD-1, PD-L1, c-KIT, MEK1, VEGF-A…). Cell therapy such as adoptive T cell is encouraging. Nanoencapsulation of the recent FDA-approved free drugs (e.g., vemurafenib, ipilimumab), using proper nanocarriers, or rational combination of these free drugs with available adjuvant nanotherapeutics might be beneficial as they might enhance the overall pharmacological features (e.g., bioavailability and targeting). The importance of direct targeting of potential melanoma initiating/propagating cells within a given patient was not detailed in this paper but might be important in order to avoid immune resistance, immune escape, and disease relapse. Owing to consideration that the incidence of advanced/metastatic melanoma in the younger population is increasing, clinical trials in pediatric patients appear necessary. Eventually, the rational molecular or cellular combo therapy is a key strategy, as it shall beneficiate a larger number of patients. The recent results reported in this paper are quite exciting and, undeniably, constitute a new hope for patients and healthcare professionals.