Melanoma

Primary Authors Layfield, Lester, MD. Wallander, Michelle, PhD.

Key Points

Disseminated melanoma is not generally responsive to traditional chemotherapy or radiotherapy. With the advent of targeted therapy, molecular testing in melanoma has gained importance.

  • Mutations by biology and targeted therapy (see table below)

Mutation

Biology

Targeted Therapy

Comments

BRAF

(v-raf murine sarcoma viral oncogene homolog B1)

ARUP Test(s)

BRAF Codon 600 Mutation Detection by Pyrosequencing 2002498

  • Downstream molecule in the mitogen activated protein kinase (MAPK) pathway which includes BRAF/MEK/ERK signaling
  • Activates pathway, mediates growth signaling, links with factor receptors such as tyrosine kinases leading to cell growth, and depresses immune regulation of cancer cells
  • >80% mutations substitute glutamic acid for valine at codon 600, resulting in V600E mutation
    • Other mutations – V600K, V600R, V600D

BRAF V600E inhibitors – vemurafenib (PLX4032) and dabrafenib (GSK2118436)

MEK inhibitors in phase I and II trials – AZD6244, GSK112012

  • 40-60% of melanomas
  • Molecular testing required if therapy directed at BRAF V600E mutation is considered
  • Associated with melanomas of sun-damaged skin, younger age (<55 yrs), and poorer prognosis compared to BRAF and NRAS wild type melanomas
  • Resistance develops in all patients
    • Resistance to BRAF inhibitors develops downstream, not at the initial mutation site
  • Combining BRAF with MEK inhibition may improve outcomes

RAS mutations

(NRAS, KRAS, HRAS)

ARUP Test(s)

NRAS Mutation Detection by Pyrosequencing   2003123

  • Molecule in the MAPK pathway
  • Most mutations are NRAS
  • Mutation leads to defective GTPase activity with uncontrolled cell proliferation
  • Most common mutations are located at codons 12, 13, and 61
  • Mutation is generally mutually exclusive of BRAF mutation

None available yet

Clinical trials combining BRAF with MEK, ATK, and  PI3K inhibitors

  • 15-20% of melanomas
  • Associated with thicker tumors, higher mitotic rate, and worse prognosis than BRAF mutation or BRAF/NRAS wild type

KIT

ARUP Test(s)

KIT Mutations, Melanoma 2002695

  • Encodes for type III transmembrane receptor tyrosine kinase which is involved in regulation of MAPK and PI3K pathways
  • Mutation functions as an oncogene
  • Most common mutations are located in exon 11; less commonly in 13, 17, and 18

Tyrosine kinase inhibitors (TKIs) – imatinib; dasatinib in clinical trials

  • Up to 15% of melanomas
    • Most common in acral lentiginous, sun-damaged skin or mucosal melanomas
  • Molecular testing should be performed if TKIs are being considered
    • KIT IHC staining does not predict mutation status nor sensitivity to TKIs
    • Exons 11, 13 most likely to have response to TKIs
  • Resistance develops in most patients with a secondary mutation in KIT
PTEN
  • Functions as the lipid phosphatase regulating PI3K/AKT pathways
  • Mutation causes high level activation of pathway with unregulated cell growth and proliferation
  • Mutation is generally mutually exclusive of NRAS  mutation, but usually inclusive for BRAF mutations
Multiple classes of inhibitors in phase I and II trials-PI3K, AKT, mTORC1, dual PI3K/mTOR inhibitors 

Diagnosis

Indications for Testing

  • Atypical melanocytic lesion (mole) requires histologic evaluation

Laboratory Testing

  • Refer to Key Points

Histology

  • Gold standard for diagnosis
    • Immunohistochemistry may be required to distinguish poorly differentiated lesions of carcinoma, sarcoma, or lymphoma (B-cell or T-cell) from melanoma
    • Stains to consider include cytokeratin 8,18 low molecular weight (CAM 5.2); melanoma antibody, HMB45; Ki-67 (Mib-1); melan A; p21 (Waf1/Cip 1); S-100 protein; and vimentin
  • Architectural and cytologic features of biopsy are interpreted in the context of clinical information
    • Architecture – large, asymmetric, poorly circumscribed lesion; pagetoid spread of melanocytes; effaced rete ridges and underlying dermal architecture; perineural invasion
    • Cytology – large nuclei with prominent nucleoli; atypical mitoses (especially in dermis); lack of melanocyte maturation in deep portion of lesion
  • Metaplastic differentiation (nonmelanocytic cells or tissues) may be identified that may not be melanoma

Imaging Studies

  • Routine cross-sectional imaging (CT, PET, MRI) is not recommended  in patients with localized melanoma due to low yield

Prognosis

  • Depth of tumor invasion – used to determine prognosis
    • Breslow thickness – most important factor contributing to T status in tumor, nodes and metastases staging
      • Measured from top of epidermal granular cell layer to deepest malignant melanocyte
      • 5-year survival – 95.3% with Breslow thickness ≤1.0 mm (corresponds to T1 lesion), 45.1% with Breslow thickness >4.0 mm
    • Clark level – important in T1 lesions (<1 mm thick) (corresponds to 95% or better survival)
      • Clark I – melanoma in situ; does not cross basement membrane
      • Clark II – invasion of papillary dermis
      • Clark III – expansion of papillary dermis
      • Clark IV – invasion of reticular dermis
      • Clark V – invasion of subcutaneous fat
  • Additional histologic features used to determine prognosis
    • Involvement of regional lymph nodes (regional nodal dissection or sentinel node testing)
    • Metastases
    • Growth phase
    • Mitotic count
    • Regression
    • Ulceration
  • Additional serum testing
    • Lactate dehydrogenase – prognostic factor in late-stage melanoma
      • Elevated levels may indicate metastases
    • S-100B, serum – elevated level associated with poor prognosis

Differential Diagnosis

  • Clinical – melanocytic nevi
  • Histologic
    • Well differentiated – melanocytic nevi
    • Poorly differentiated – carcinoma, sarcoma, lymphoma (T-cell, B-cell)

Screening

  • American Academy of Dermatology (AAD) and American Cancer Society (ACS) recommend regular skin exams in all patients
    • U.S. Preventive Services Task Force finds insufficient evidence to recommend regular skin exams

Monitoring

  • NCCN recommends skin examination at least once per year for life for melanoma patients (including those with stage 0, in situ melanoma)
    • Patients with stage IA-IIA melanoma, no evidence of disease
      • Comprehensive exam every 3-12 months for five years and annually thereafter as clinically indicated
    • Patients with stage IIB-IV melanoma, no evidence of disease
      • Comprehensive exam every 3-6 months for two years, then every 3-12 months for three years and annually thereafter as clinically indicated
    • Although not recommended at baseline, x-ray, CT/MRI, and/or PET/CT every 6-12 months can be considered to screen for recurrent or metastatic disease at discretion of physician
      • Most recurrences manifest within first 5 years – routine imaging not recommended beyond this period
  • S-100B, serum
    • Rising concentrations after treatment indicate disease recurrence

Clinical Background

Melanoma is a malignancy of the melanocytes. This disease is experiencing a rising incidence worldwide, ranking second to adult leukemia in loss of years of potential life per death.

Epidemiology

  • Incidence – 18/100,000
    • Estimated 76,250 new melanomas and 9,180 deaths in U.S. in 2012 (American Cancer Society, 2012)
    • Tripled over the last 20 years
    • Highest incidence worldwide is in Australia
  • Age – median is 57 years; rare in children
  • Sex – M>F, 1.5:1
  • Ethnic – tenfold increased incidence in Caucasians
    • Acral lentiginous melanoma has equal distribution across all ethnic groups

Inheritance

  • Familial atypical multiple mole melanoma – associated with mutations in the CDKN2A (also known as p16) gene
    • Mutations in CDKN2A and CDK4 associated with high risk of developing melanoma

Classification

  • Superficial spreading melanoma (~70%)
  • Nodular melanoma (10-15%)
  • Lentigo maligna melanoma (10%)
  • Acral lentiginous melanoma (~5%)
  • Mucosal lentiginous melanoma (3%)

Risk Factors

  • Sunlight – sunburns in childhood, intermittent UV exposure associated with higher risk
  • Blue or green eyes; red or blond hair
  • Melanocytic nevus – increased risk by number and size of nevi
  • Family history – twofold risk if first-degree relative had melanoma
    • Clustering of melanoma in familial retinoblastoma and Li-Fraumeni syndrome
    • Increased risk of familial melanoma in the presence of family history of pancreatic cancer or astrocytoma
  • Immunosuppression

Genetics

  • BRAF mutations
    • 40-60% of melanomas
    • Most common in cutaneous melanomas derived from intermittent sun-exposed skin
  • KIT mutations
    • As many as 15% of melanomas
    • Most common in mucosal and acral lentiginous, sun-damaged skin; small presence in sun-damaged melanomas
    • Most common mutations located at exons 9,11,13, and 17
    • Predicted to respond favorably to the tyrosine kinase inhibitor imatinib
  • RAS mutations
    • 15-20% of melanomas
    • Associated with thicker tumors, high mitotic rate, and worse prognosis than BRAF mutation or BRAF/NRAS wild type
    • Most common mutations located at codons 12,13, and 61
    • Generally mutually exclusive of BRAF mutation
  • PTEN mutations
    • Generally mutually exclusive of NRAS mutations but usually inclusive for BRAF mutations

Clinical Presentation

  • 82-85% of melanoma patients present with localized disease; 10-30% present with regional disease; 2-5% with distant metastatic disease
  • Nevus with Asymmetry, Border irregularity, Color variation, Diameter >6 mm and Evolving changes (ABCDE criteria)
  • Head and neck most common sites
  • Ulceration, pigment loss
  • Rare sites – eye (iris, ciliochoroidal), mucosa, unknown primary
  • Mortality rates have plateaued in recent years (attributed to early diagnosis)

Prevention

  • Decrease UV exposure
    • Use sunscreen
    • Avoid midday sun
    • Cover skin with hat and clothing
  • Skin self-examination
  • Regular skin evaluation by primary care physician or dermatologist

Pediatrics

Clinical Background

Epidemiology

  • Prevalence – 300-425 cases annually in the U.S. (NCI, 2006)
  • Age – 15-19 years most common
  • Sex – M>F (minimal)

Risk Factors

  • Family history – twofold risk if first-degree relative had melanoma
    • Clustering of melanoma in familial retinoblastoma and Li-Fraumeni syndrome
    • Increased risk of familial melanoma in the presence of family history of pancreatic cancer or astrocytoma
  • History of severe sunburns (>3 before 20 years)
  • Xeroderma pigmentosa
  • Immunosuppression
  • Dysplastic nevi (three- to sixfold increased risk)

Clinical Presentation

  • Lesions frequently amelanotic or nodular
  • ABCDE criteria (Asymmetry, Border irregularity, Color variation, Diameter >6 mm, and Evolving changes) less reliable than in adults

Diagnosis

Indications for Testing

  • Refer to Diagnosis tab

Laboratory Testing

  • Refer to Diagnosis tab

Histology

  • Congenital nevus has slightly different histologic appearance
  • Refer to Histology section in Diagnosis tab

Imaging Studies

  • Refer to Diagnosis tab

Differential Diagnosis

  • Spitz nevus
  • Blue nevus
  • Congenital nevus

Indications for Laboratory Testing

  • Tests generally appear in the order most useful for common clinical situations
  • Click on number for test-specific information in the ARUP Laboratory Test Directory
Test Name and Number Recommended Use Limitations Follow Up
BRAF Codon 600 Mutation Detection by Pyrosequencing 2002498
Method: Polymerase Chain Reaction/Pyrosequencing

Detect activating BRAF mutations associated with anti-EGFR therapy resistance

Analytic sensitivity/specificity – 99%

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codon 600 will not be detected

 
KIT Mutations, Melanoma 2002695
Method: Polymerase Chain Reaction/Sequencing

Provide prognostic and predictive information for  kinase inhibitor (TKI) therapy planning

Analytical sensitivity – 25% mutant alleles (50% tumor)

Mutations outside of targeted exons are not detected

 
NRAS Mutation Detection by Pyrosequencing 2003123
Method: Polymerase Chain Reaction/Pyrosequencing
Screen for tumors that may respond to therapy targeted to downstream genes in the MAPK signalling pathway

Limit of detection – 10% mutant alleles

Oncogenic mutations outside of codons 12, 13, and 61 are not detected

Presence or absence of mutations does not guarantee a positive response to anti-EGFR therapies

 
Cytokeratin 8,18 Low Molecular Weight (CAM 5.2) by Immunohistochemistry 2003493
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
Melanoma Antibody, HMB45 by Immunohistochemistry 2003935
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
Ki-67 with Interpretation by Immunohistochemistry 2007182
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and resulted by ARUP

   
Melan A by Immunohistochemistry 2003996
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
S-100 Protein by Immunohistochemistry 2004127
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
Vimentin by Immunohistochemistry 2004181
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
p21 (Waf1/Cip 1) by Immunohistochemistry 2004067
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
BRAF V600E by Immunohistochemistry 2008710
Method: Immunohistochemistry

Aid in histologic diagnosis of melanoma

Stained and returned to client pathologist; consultation available if needed

   
Lactate Dehydrogenase, Serum or Plasma 0020006
Method: Quantitative Enzymatic

Monitor treatment and determine prognosis of patients with malignant melanoma

Not used for diagnosis

 
S-100B Protein, Serum 2001766
Method: Quantitative Enzyme-Linked Immunosorbent Assay

Monitor treatment and determine prognosis of patients with malignant melanoma

Not used for diagnosis; S-100B is not specific for malignant melanoma; increased serum concentrations are found in patients with liver and renal injury, inflammation, infection, and brain injury

 
Additional Tests Available
 
Click the plus sign to expand the table of additional tests.
Test Name and NumberComments
BRAF V600E Mutation Detection by Allele-Specific PCR, Fine Needle Aspirate 2006516
Method: Polymerase Chain Reaction

Diagnosis, prognosis, and therapeutic decisions in a variety of tumors

Assess for responsiveness to BRAF inhibitor therapy

For tissue block or formalin-fixed, paraffin-embedded (FFPE) cell blocks prepared from FNA, BRAF codon 600 Mutation Detection by Pyrosequencing may be used

Solid Tumor Mutation Panel by Next Generation Sequencing 2007991
Method: Massively Parallel Sequencing

Prognosis/treatment of individuals with solid tumor cancers at initial diagnosis or with refractory disease

48-gene panel assay for detection of mutations, including KRAS, EGFR, BRAF, and ERBB2 genes