Morphological aspects and therapeutic options in melanoma: a narrative review of the past decade

Abstract

Melanoma is a malignant cancer of the skin, the incidence of which has been increasing year by year. This neoplasm has high aggressivity as well as the potential for invasion and metastases. Multiple factors related to the proliferation of this type of tumor have been identified, such as exposure to ultraviolet (UV) radiation and specific genetic backgrounds. From a histological and cytological point of view, the most common cells that are found in melanoma are epithelioid or spindle cells. To confirm the diagnosis and the melanocytic origin of the tumor, specific and sensitive markers are used. Also, observation of the behavior of this cancer, including its proliferative properties, has led to the development of multiple therapies, each of which is characteristic of the pathological stage at the time of diagnosis. While surgery is the most important therapeutic and curative option in cases of melanoma in situ, chemotherapy has been the main treatment for advanced stages of melanoma for many years. However, recently, targeted therapy and immunotherapy have changed the approach to treatment. At present, multiple studies are attempting to obtain further data about the tumor microenvironment and investigating how targeting particular molecules can change the prognosis of patients

Introduction

Melanoma is one of the most aggressive types of malignancy. The prevalence and worldwide incidence of this skin neoplasm is increasing from year to year. Melanoma originates in melanin-producing cells known as melanocytes, which derive from the neural crest of the ectoderm. Due to the pathway of these cells during embryonic life, melanoma not only occurs in the skin but can also arise in other sites (e.g., mucosa, uvea) [1, 2, 3].

Knowledge of the aggressivity of this tumor is supported by data collected over a long period, dating as far back to the first description of melanoma in 1837. Even though melanoma represents only around 1% of malignant skin tumors, it is considered to be the deadliest form of cancer that can arise in the skin. Local aggressivity is shown by rapid growth and invasion of the deep layers of the skin, while systemic aggressivity is demonstrated by the capacity of the tumor to metastasize even in the early stages of disease. Currently, the treatment for melanoma is surgery followed by oncological therapy, depending on the tumor stage. Unfortunately, once the tumor becomes metastatic, the prognosis is very poor [2, 3, 4].

Melanoma can be classified according to the location of the tumor, which is linked to sun exposure. Therefore, we encounter cutaneous melanoma [related to ultraviolet (UV) radiation] and mucosal melanoma (unrelated to UV radiation). Nodular melanoma is a particular category of melanoma that can develop in either form, regardless of background or risk factors. This type is described as a rapidly growing melanoma by the World Health Organization (WHO) and has the worst prognosis of all types of melanomas currently known [1, 5, 6, 7].

Morphological aspects

According to WHO, melanoma can present in multiple forms, and it is classified based on parameters such as location (cutaneous or mucosal), UV exposure (cumulative solar damage), cellular type, histological type. Regarding the association with UV exposure, we can identify superficial melanoma, lentigo maligna, and desmoplastic melanoma. Malignant melanocytic tumors that are not associated with UV exposure are uveal melanoma, acral melanoma, mucosal melanoma, blue nevus melanoma and melanoma arising in congenital nevi. Nodular melanoma represents a special type of tumor, which is most likely the final form of evolution of all the types mentioned.

From a histological and cytological point of view, the most common cells that are found in melanoma are epithelioid (similar in shape and size to epithelial cells) or spindle cells. In many cases, a tumor can present both types of cells, situation in which we will classify them as “mixed”.

The tumor type with the worst prognosis is nodular melanoma. The histological appearance of this tumor consists in a solid tumor proliferation which presents multiple nests of tumoral cells. These cells are usually medium or large in size, with pale or eosinophilic cytoplasm and enlarged nuclei, that are sometimes irregular in shape and hyperchromic. One of the most distinctive features of melanoma is the presence of eosinophilic nucleoli. Mitoses are common and their number varies from case to case (Figure 1, A and B).

Figure 1.

Nodular melanoma: (A) Nests of tumoral cells with abundant eosinophilic cytoplasm are seen; the nuclear pleomorphism is severe; nuclei appear enlarged and present eosinophilic nucleoli; (B) The nests of tumoral cells occupy the dermis; the delimitation between the tumor and the epidermis can be seen. Hematoxylin–Eosin (HE) staining: (A) ×200; (B) ×100. (Collection of Pathology Department, Mureş Clinical County Hospital, Târgu Mureş, Romania)

Superficial spreading melanoma presents as a tumor located in the superficial dermis and the junction, usually associated with pagetoid migration (presence of tumoral cells in the epidermis, isolated or in nests). The tumor cells are usually organized in nests and present marked atypia, various sizes and shapes, eosinophilic cytoplasm and nuclear pleomorphism. The characteristic features (such as prominent eosinophilic nucleoli) are also present (Figure 2).

Figure 2.

Superficial spreading melanoma. The nests of atypical tumoral cells are seen at the dermo–epidermal junction and in the papillary dermis. HE staining, ×100. (Collection of Pathology Department, Mureş Clinical County Hospital, Târgu Mureş, Romania)

To confirm the diagnosis and the melanocytic origin of the tumor, specific and sensitive markers are used. Some of the most common immunohistochemistry (IHC) reactions used, as mentioned in the literature, are sex determining region Y (SRY)-box transcription factor 10 (SOX10), S100, Melan A, and human melanoma black 45 (HMB45) (Figure 3, A and B; Figure 4, A and B). One of the newest immunomarkers described is PReferentially expressed Antigen in MElanoma (PRAME), which has a high specificity in detecting melanoma metastases [1, 2].

Figure 3.

Immunostaining with nuclear markers in nodular melanoma: (A) S100, ×50; (B) SOX10, ×50. SOX10: Sex determining region Y (SRY)-box transcription factor 10. (Collection of Pathology Department, Mureş Clinical County Hospital, Târgu Mureş, Romania)

Figure 4.

Immunostaining with cytoplasmic markers in nodular melanoma: (A) Melan A, ×400; (B) HMB45, ×400. HMB45: Human melanoma black 45. (Collection of Pathology Department, Mureş Clinical County Hospital, Târgu Mureş, Romania).

Materials and Methods

We conducted a selective PubMed and PubMed Central (PMC) database search and selected the top 50 most relevant articles using the following keywords: ‘melanoma’, ‘genetics’, ‘epidemiology’, ‘treatment’, ‘chemotherapy’, and ‘prognosis’. All articles were published in the past 10 years. Many of them consisted of reviews of treatment options available up to the present.

Epidemiology

New cases of melanoma reported yearly have increased worldwide, mostly affecting Caucasians. The Canadian Cancer Society reported over 1200 deaths in 2017 and over 7000 new cases of cutaneous melanoma. From the number of cases reported, it is estimated that one in 34 males and one in 53 women will be diagnosed with melanoma. In 2021, melanoma was ranked as the 15th most common malignant tumor globally. However, incidence is highly variable from country to country. Australia reported over 16 000 cases each year, increasing from 10 000. In this country, however, the survival increased from 10% to 50%, as confirmed by Australia Melanoma Institute. European countries with the highest new occurrence rates are Denmark, the Netherlands and Sweden, reporting incidence associated with increased age (above 75 years) [8, 9, 10, 11, 12].

Genetic background

Melanoma is one of the forms of cancer in which genetics plays a crucial role. The most commonly encountered gene involved in the pathogenesis of this tumor is B-Raf proto-oncogene, serine/threonine kinase (BRAF), more specifically the mutated form BRAFV600E. This type of mutation causes mitogen-activated protein kinase (MAPK) to become activated, leading to uncontrolled and abnormal cellular proliferation. The BRAF mutation is most frequently identified in the types of melanomas related to UV exposure, meaning that sun damage is the main factor responsible for the appearance of the tumors. Because of this discovery, targeted therapy against BRAF, such as the inhibitor Vemurafenib, has been developed in recent years [13, 14, 15].

BRAF is not the only gene reported to play a major role in the pathogenesis of melanoma. Neuroblastoma RAS viral oncogene homolog (NRAS) is also associated with melanoma and increases the resistance to BRAF inhibitor therapy. The RAS family consists of specific types of proteins found in the cell membrane that also activate the MAPK pathway. Mutation of NRAS has been discovered to be the second most common type of mutation in melanoma.

Neurofibromatosis type 1 (NF1) is a tumor suppressor gene whose product acts as an inhibitor of RAS protein; therefore, loss of activity of NF1 leads to RAS activation. The absence of NF1 has been reported in almost one-third of melanoma cases associated with BRAF and NRAS, suggesting that it might also play a role in MAPK activation [16, 17].

Other genes involved in the pathogenesis of melanoma include phosphatase and tensin homolog gene (PTEN) and v-Kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog gene (c-KIT). PTEN is a tumor suppressor involved in multiple types of cancer, while c-KIT plays an important role in the proliferation of tumor cells. A further gene, melanocyte-inducing transcription factor (MITF), is involved in the development of sporadic melanoma and acts as an oncogene [18, 19].

Current treatment options

Treatment of melanoma depends on the stage of the tumor. The first step in diagnosis and treatment is surgical excision. After the sample has been collected, it is sent for processing and analysis according to the protocols in each pathology department. Treatment is then assessed according to diagnosis and stage, as well as several other characteristics (BRAF positivity and other genetic characteristics) [20, 21].

For melanoma in situ, surgery is the primary means of treatment. Standard excision with safety margins of 0.5 mm is recommended; biopsy of the sentinel node can also be helpful. For superficial invasive melanoma, sentinel node biopsy is mandatory and has been reported to be both accurate and important for the correct staging of the patient. It is also highly recommended in tumors of pathological T2 stage and above. Follow-up for all mentioned categories is mandatory for six months or yearly for the first five years, followed by an annual check-up [22, 23, 24].

For advanced stages of melanoma (stages III and IV), oncological therapy is required to provide patients with a better outcome. Although chemotherapy has been used for many years as the first option, nowadays it does not represent frontline therapy. However, there are cases where chemotherapy is still useful, e.g., melanomas that do not present specific mutations or that do not respond to other therapies. Since 1975, Dacarbazine has been one of the most popular drugs for advanced melanoma cases; this medication is administered intravenously and is associated with multiple side effects, such as nausea, myelosuppression, and vomiting. Although many studies failed to show any improvement in patient survival with this drug, Dacarbazine remains the most frequently used chemotherapeutic medication in patients with metastatic melanoma. Another drug used in the advanced stages of melanoma is Temozolomide, which has been reported to provide similar outcomes to Dacarbazine. Alkylating agents such as nitrosoureas have also been used, resulting in slightly increased efficiency in cases with brain metastases. Another chemotherapeutic administered to patients with advanced stages of melanoma is Carboplatin, which has reportedly been used in multiple tumors; its major role is to block the replication of tumor cells. Carboplatin can be used alone or in combination with other chemotherapeutic drugs. Combinations of taxanes and platinum-based drugs have also been used to treat advanced-stage melanoma; the results showed improved survival, compared with the previously used chemotherapeutic medications, in patients who had received prior treatment with other drugs as well as those who had not received any previous treatment [25, 26, 27, 28, 29, 30].

The discovery of BRAF in 2002 heralded a huge change in the techniques used by physicians and pathologists to manage melanoma. The identification of BRAF mutations in almost half of melanoma cases stimulated the development of targeted therapy, with one of the most important representatives in this category being Vemurafenib [31]. Furthermore, mitogen-activated protein kinase kinase (MEK) inhibitor therapy has the potential to induce apoptosis in both NRAS- and BRAF-mutated melanomas; such inhibitors have been developed in recent years and used in combination with targeted medication for BRAF [32, 33].

The most important result of targeted therapy has been improved outcomes for patients, compared with traditional chemotherapy. However, problems with targeted therapy can also develop [34].

The main disadvantage of therapy against BRAF and MEK (which target the MAPK pathway) is that tumor cells develop resistance to it over time. Eroglu and Ribas emphasized the low percentage of cases where this type of therapy remains efficient, suggesting that a combination of pathways is involved in the evolution of melanoma, in addition to MAPK. The main purpose of the reported clinical trials was to highlight the superior effect of combined therapy, in comparison with monotherapy. Results obtained using Dabrafenib and Trametinib in combination indicated improved prognosis, compared with monotherapy using Dabrafenib. The survival rate in patients with advanced melanoma who received combined therapy was 79% at one year of follow-up, compared with 70% in those who received monotherapy. Vemurafenib and Cobimetinib were also used in the trials. The results obtained with this combination were similar to those obtained using Dabrafenib and Trametinib, showing a better outcome than monotherapy. However, toxicity was reportedly more frequent with combined therapy than with monotherapy, with side effects of diarrhea, photosensitivity, vomiting, retinopathy and elevated levels of aminotransferase and creatine kinase. The final combination of drugs to be tested was Binimetinib and Encorafenib; however, the trials were reported as still ongoing at the time of publication. A review conducted by Ziogas et al. showed supplementary data about Encorafenib and Binimetinib, concluding that combined therapy resulted in a partial or complete response in 64% of cases; lower percentages were found with monotherapy. In addition, a comparison of Encorafenib alone versus Vemurafenib alone showed a better outcome for Encorafenib, with a response in 52% of cases, compared with 41% for Vemurafenib [35, 36, 37].

Immune checkpoints are one of the most important targets in the oncological treatment of melanoma. They represent ligands and receptors that play a major role in the modulation of T-cell activity as part of the tumor microenvironment. Cytotoxic T-lymphocyte-associated protein 4 (CTL4), known as cluster of differentiation 152 (CD152), is one of the most important immune checkpoints to be targeted.

One of the greatest challenges for patients and oncologists is the treatment of metastatic melanoma. In the review conducted by Ziogas et al. (see above), which also included the management of patients with melanoma metastases, immune checkpoints are described as the future of melanoma therapy; furthermore, combined therapy (immunotherapy and targeted therapy) has been shown to lead to improvements in murine models [35].

Another checkpoint that can be targeted is programmed cell death protein 1 (PD-1), along with its ligand, PD-L1. PD-1 is expressed in both types of lymphocytes (T- and B-cells), as well as in natural killer (NK) cells, and is most commonly found in cells undergoing chronic stimulation. PD-L1 is expressed in multiple types of cells, including normal cells. The most critical role of PD-1 and its ligand is to prevent autoimmune reactions and the destruction of cells expressing self-antigens. Antibodies with affinity for either or both PD-1 and PD-L1 can activate the immune system and trigger tumor cells. Thus, therapy that targets PD-L1 and PD-1 has been developed for patients with melanoma; one of the most significant medications of this type is Nivolumab [38, 39, 40].

A study conducted by Terheyden et al. [41] demonstrated the efficacy of Nivolumab by performing a follow-up of 272 patients diagnosed with metastatic melanoma. The authors describe a median follow-up of 8.4 months, with partial remission in 28.3% of cases and total remission in 3.3% of patients. The same study describes the results obtained by Robert et al., who performed a follow-up of 210 patients also diagnosed with metastatic melanoma. Their results included total remission in 7.6% of cases and partial remission in 32.4%, while the follow-up was conducted for a maximum of 16.7 months [41, 42, 43].

These two follow-up studies provided important data regarding the effects of Nivolumab, proving the superiority of this medication, in comparison with Dacarbazine and Pembrolizumab. Another study, conducted by Wolkoch et al., showed results obtained by combining Nivolumab with another agent, Ipilimumab. They followed 314 patients for 38 months and demonstrated a patient survival rate of 39% after three years with combined therapy, compared with 32% for Nivolumab alone [44, 45].

A five-year study conducted by Larkin et al. also examined the effectiveness of Ipilimumab, an anti-CTL4 monoclonal antibody. This medication was used in combination with Nivolumab, as well as other drugs (including BRAF/MEK inhibitors) to treat a group of patients with stage III or IV melanoma and confirmed mutation of BRAF. The treatment was continued until the patients developed side effects or until the disease progressed. Out of a total of 1293 patients selected to participate in the trial, 945 were randomly assigned to treatment groups: 314 to the Nivolumab-plus-Ipilimumab group, 316 to the Nivolumab group and 315 to the Ipilimumab group. Follow-up revealed improved survival in patients treated with Nivolumab for both groups receiving this medication, compared with the group receiving Ipilimumab monotherapy. At the end of five years of follow-up, 52% of patients in the group receiving combined therapy had survived, compared with 44% in the group receiving Nivolumab alone and 26% in the group receiving Ipilimumab. Quality of life was also taken into consideration; the conclusion of the researchers in this regard was consistent with previous studies reporting the lack of deterioration in patients receiving either monotherapy or combined therapy [46].

Recently, a new concept regarding the treatment of patients with melanoma has emerged, which includes BRAF/MEK inhibitors combined with immunotherapy. It has been reported in multiple studies that BRAF inhibitors in association with MEK inhibitors provide a better prognosis for patients, regardless of the stage of the disease. Yet, as mentioned previously, the development of resistance to treatment is one of the problems that may occur in time. Immunotherapy provides a more sustained response in melanoma, which highlights the possibility of triple therapy [47, 48].

Long et al. published a study in 2021 describing six clinical trials comparing anti-PD-1 immunotherapy and BRAF/MEK targeted therapy. Out of the 192 patients in the study, 141 received immunotherapy (104 were treated with a combination of Ipilimumab and Nivolumab; 37 received anti-PD-1 monotherapy) and 51 patients received targeted therapy. They observed a full response in 40% of the cases treated with targeted therapy and 33% of cases treated with immunotherapy. However, they concluded that the benefits of neoadjuvant therapy in melanocytic malignant tumors remained unclear [49, 50].

In addition, Kakavand et al. studied the melanoma microenvironment. One of their most recent studies focused on V-domain immunoglobulin (Ig) suppressor of T-cell activation (VISTA), a homologue of PD-L1 belonging to the B7 family. They observed high expression of VISTA in inflammatory cells [tumor-infiltrating lymphocytes (TILs)] in cases where resistance to immunotherapy had been reported. These data suggest that combined therapy targeting both VISTA and PD-L1 may be beneficial for patients [51].

Multiple studies regarding this new protein have been made over the last few years, yet its role remains controversial. While some authors have associated the presence of VISTA with poor prognosis, other authors have described improved prognosis. There is also some uncertainty about the type of cells that express the molecule; reports of VISTA expression in TILs are numerous, but there are also studies showing expression in myeloid lineage cells. More precisely, it has been shown that immature cells, whether they are lymphocytes or myeloid cells, are more likely to express VISTA, compared with other types of cells. Tumor cells are also reported to express this protein; however, this requires further investigation. At present, there are several ongoing clinical trials targeting this pathway. For example, Curis, Inc., is holding a phase I trial with 50 patients presenting an advanced stage of melanoma (with metastases and tumors that cannot be removed by surgery). This study is trialing an inhibitor of PD-L1, PD-L2 and VISTA (CA-170), with results to be released in the near future [51, 52, 53].

Conclusions

Over the course of several years, multiple treatment options have been developed for patients with melanoma of various stages. The biggest challenge has been represented by advanced cases requiring treatment in addition to surgery. Chemotherapy has been the treatment of choice for many years, providing patients with extended survival rates. However, more recently, since targeted therapy (in particular, anti-BRAF medication) has gained more prominence and achieved better results, chemotherapy has ceased to be the first-line treatment. The latest hot topic regarding melanoma treatment is immunotherapy. One of the most important questions that we need to answer in the future is how we can produce efficient immunotherapy without causing exacerbated autoimmunity. For us to achieve this, we need to study the tumor microenvironment more closely.

Conflict of interests

The authors declare that they have no conflict of interests.