The pathology of cutaneous melanoma - Rawa Muhsin

This is a slideshow about the pathology of cutaneous melanoma, including brief comments about its pathogenesis, clinical aspects, typical histology, histologic subtypes, immunohistochemistry, molecular findings, and some clinical aspects.

1. The pathology of cutaneous melanoma Dr. Rawa Muhsin Ali MBChB, ABHS- APath

2. Outline Introduction Histologic features Histologic subtypes Immunohistochemistry Molecular findings Clinical aspects

3. Introduction Most melanomas in white people are related to ultraviolet radiation exposure from cumulative sun damage (CSD) Majority of melanomas in non-white people are not related to CSD (and instead arise from sporadic or germline mutations, chronic inflammation, trauma, oxidative stress, etc ) Predisposing conditions include large number of nevi, xeroderma pigmentosum, NF-1, dysplastic nevus syndrome, and germline inactivating mutations in CDKN2A and BAP1 Nevi are more often stimulants (rather than precursors ) for melanoma (transformation is very rare, and more common in congenital and dysplastic types)

4. HISTOLOGIC FEATURES

5. Typical histology Junctional activity Prominent melanin pigmentation Invasion of surrounding tissue Marked cytologic atypia Nuclear grooves, folds, and pseudoinclusions Large eosinophilic nucleoli Abundant mitotic figures (including atypical forms)

6. Melanoma: The Great Mimicker Cells can be epithelioid, spindled , or extremely bizarre Cells can be small, large, or giant multinucleated Cytoplasm can be eosinophilic, basophilic, foamy, signet-ring shaped, rhabdoid, oncocytic , or clear (balloon cell) Melanin can be abundant, scanty, or absent

7. Melanoma: The Great Mimicker Growth pattern can be pseudoglandular , pseudopapillary, peritheliomatous , hemangiopericytoma-like, spitzoid , rosette-like, trabecular, verrucous (nevoid), carcinoid-like, or follicular Can be associated with marked fibroblastic response, myxoid changes, giant cells, and pseudoepitheliomatous hyperplasia Divergent differentiation includes osteocartilaginous , rhabdomyoblastic , neuroendocrine, and neuroid (MPNST-like)

8. Melanoma vs other malignancies Histologic clues Abundant eosinophilic, finely granular cytoplasm Pseudonuclear inclusions Combined epithelial and spindle cell patterns Fascicular and pseudoalveolar patterns Immunohistochemistry is of paramount importance

9. Melanoma vs benign melanocytic lesions Lack of cohesiveness and subepidermal cleft formation in the intraepidermal component Lateral trailing off of individual atypical melanocytes (poor circumscription) Pagetoid spread and pigmented parakeratosis Size and shape variation and confluence of nests Asymmetry in all aspects Lack of maturation of dermal melanocytes

10. Melanoma vs benign melanocytic lesions Prominent nuclei and nucleoli Deep dermal and atypical mitoses Abundant clear cytoplasm with a finely dispersed (dusty) chromatin Individual tumor cell necrosis Band-like dermal lymphocytic infiltrate

11. Purely intradermal lesions All primary melanomas have an intraepidermal or junctional component Melanomas without a junctional component can be due to: Regression of the junctional component Arising from an intradermal nevus Metastatic Dermal melanomas should be distinguished from clear cell sarcoma They have overlapping IHC profiles Clear cell sarcomas show t(12;22) ( EWS - ATF1 ) or t(2;22) ( EWS - CREB )

12. Clear cell sarcoma of skin

13. HISTOLOGIC SUBTYPES

14. Main clinicopathologic subtypes Superficially spreading Lentigo maligna Nodular Acral lentiginous Other types Nevoid, balloon (clear) cell, pleomorphic, sarcomatoid, spindle cell/desmoplastic/neuroid, small cell (neuroendocrine-like), signet-ring cell, myxoid, metaplastic, rhabdoid

15. Superficially spreading melanoma Most common form and can occur anywhere on the body ( low-CSD ) Radial growth phase followed by vertical growth phase (associated with formation of a nodule) Uniform atypical melanocytes forming nests with pagetoid spread

16. Superficially spreading melanoma

17. Superficially spreading melanoma

18. Lentigo maligna melanoma Flat, slow-growing lesion on the sun-exposed areas (especially cheek) of elderly white people ( high-CSD ) Basal proliferation of atypical melanocytes individually and in nests with pleomorphism and cytoplasmic retraction but less prominent upward migration Radial growth phase followed by vertical growth phase Can be associated with desmoplastic melanoma which has a higher risk of local recurrence and distant metastasis

19. Lentigo maligna

20. Lentigo maligna melanoma

21. Nodular melanoma Affects all body surfaces (can be low- or high-CSD ) and often younger patients No lateral flat component (vertical growth phase without recognizable radial growth phase)

22. Nodular melanoma

23. Nodular melanoma

24. Acral lentiginous melanoma More common in blacks and Asians on the palms, soles, subungual areas, anus, and mucocutaneous junctions ( non-CSD-related ) Radial growth phase followed by vertical growth phase Lentiginous component is dendritic and bizarre, with hyperplastic epidermis, and inflamed papillary dermis

25. Acral lentiginous melanoma

26. Desmoplastic melanoma Subtype of spindle cell melanoma occurring on the head and neck ( high-CSD or non-CSD-related ) Short fascicles of spindle cells with heavy desmoplastic reaction, deep invasion, infiltration of nerves, peripheral lymphoid aggregates, and overlying junctional melanocytic proliferation If more cellular with less stroma, consider as spindle cell melanoma Neurotropic melanoma is a variant with a peripheral nerve sheath pattern (distinguish from superficial MPNST by diffuse S100 expression and association with nevus or history of melanoma)

27. Desmoplastic melanoma

28. IMMUNOHISTOCHEMISTRY

29. Histochemistry and EM Histochemistry Silver-based (argentaffin) based on reducing properties of melanin granules Fontana-Masson is the most common To detect finely dispersed granules or distinguish from hemosiderin Electron microscopy Melanosomes and premelanosomes Both techniques no longer widely used since they are replaced by immunohistochemistry

30. S100 protein Involved in intracellular calcium trafficking and/or microtubular assembly Melanocytes only synthesize the alpha-alpha dimeric form Expression is both nuclear and cytoplasmic Expressed in both normal and neoplastic melanocytes Most labs use antibodies that detect all three isotypes, with a 98% sensitivity (at least focally), regardless of histologic subtype Also expressed by various carcinomas (including breast), some histiocytic tumors, gliomas, peripheral nerve sheath tumors, and Langerhans cell histiocytosis Fix skin tissue for 6-48 hours for optimal performance

31. S100 expression in melanoma

32. Loss of S100 vs retained MART1 in overfixed biopsy of melanoma

33. SOX10 Transcription factor in cells from the neural crest (expressed in Schwann cells and melanocytes) Positive in majority of nevi and melanomas (including spindle cell/desmoplastic type) Also expressed in benign neural lesions , schwannomas, neurofibromas, and half of MPNSTs Proliferating Schwann cells and epithelial cells from eccrine and apocrine glands may be confused for desmoplastic melanoma in the scar of a prior procedure

34. gp100 and PMel-17-related monoclonal antibodies Premelanosome membrane proteins (gp100 transcripts present in other tissue types, but only translated in melanocytic elements) MART-1/ melan -A and HMB-45 are the most widely used (others include HMB-50 and NKI/C3) Sensitivity of 60-80% depending on commercial preparation Both are also expressed by the PEComa family of tumors as they also contain premelanosomes Melan-A cross-reacts with steroidogenic tumors such as adrenocortical carcinomas and sex-cord tumors of the ovary <10% of spindle cell/desmoplastic/neuroid melanomas express these markers due to a shift to a fibroblastic/ schwannian phenotype

35. Melan-A expression in melanoma

36. Tyrosinase Tyrosinase converts tyrosine to dopa (first step in melanogenesis), and its gene transcripts are confined to melanin-producing cells (hence can use PCR to detect isolated tumor cells) Sensitivity of >80% and specificity of 100% for non-spindle type Nevi and nonneoplastic melanocytes often nonreactive (hence possibly related to maturation and differentiation)

37. Tyrosinase expression in melanoma

38. Microphthalmia transcription factor Controls activity of melanogenic enzymes by upregulating cAMP Sensitive and specific for melanocytic differentiation Lower expression in spindle cell/desmoplastic melanomas, but supports the diagnosis of melanoma over MPNST Also expressed by PEComas , cellular neurothekeomas, and cutaneous granular cell tumors

39. MITF expression in melanoma

40. Ki-67 Nuclear marker in proliferating cells Common and dysplastic nevi show <1% staining , mostly at the dermal-epidermal junction or superficial dermal component Melanomas have an average rate of >10% staining throughout the dermal component, especially the deep edge Desmoplastic melanomas also show a much higher rate than desmoplastic nevi

41. Intermediate filament proteins Include keratins, vimentin, desmin , neurofilament proteins, and GFAP Melanomas only express vimentin intensely Vimentin may only be useful to determine whether tissue antigenicity is preserved, but all the other markers have built-in internal controls in the skin, rendering its use of limited value Rarely express low-molecular-weight keratins but negative for EMA, CEA, TAG-72, BER-EP4, and MOC-31 Very rarely express other filaments aberrantly in specific subtypes

42. Patchy positivity with an anti-keratin cocktail in melanoma

43. Other markers p16 Inhibits melanocyte growth in culture Expression preserved in most nevi but lost in some melanomas Useful as screening for p16 homozygous loss to trigger FISH for 9p21 PNL2 Stains junctional nevi and melanomas with adequate specificity Soluble adenylyl cyclase ( sAC ) Positive in lentigo maligna but negative in benign melanocytes, thus may be useful to distinguish between the two in sun-damaged skin

44. Other markers Neuroectodermal associations Neurosecretory granules (synaptophysin and chromogranin) are occasionally present in melanocytic lesions Melanocytic markers expressed in some neuroendocrine and neuroectodermal tumors NSE, CD56, CD57 and CD99 preferentially expressed in nevoid melanocytic lesions (except CD99) Some express CD10 (also expressed by atypical fibroxanthoma), CD68 (but not CD163), and CD117 CD117 and BRAF V600E expression may guide targeted therapy

45. Caveats No marker consistently separates benign from malignant melanocytic lesions HMB-45 in nevi shows a graded pattern that decreases from top to bottom (except in blue nevi, cellular blue nevi, penetrating nevi, and some Spitz and Reed nevi) while melanomas are diffusely positive Ki-67 shows a similar pattern to HMB-45 in nevi and melanomas PRAME is positive in the majority (>80%) of melanomas and negative in the majority (>85%) of nevi All markers may be lost in metastatic cases due to phenotypic dedifferentiation

46. Differential expression of HMB-45 in nevi and melanomas

47. Differential expression of Ki-67 in nevi and melanomas

48. PRAME expression in melanoma vs nevus

49. MOLECULAR FINDINGS

50. Molecular findings Benign lesions have one genetic abnormality, intermediate ones (melanocytomas) have more than one, and melanomas have multiple (highlighted by cytogenetics and CGH) Activation of MAPK and PTEN/AKT pathways MAPK pathway includes KIT, NRAS, BRAF, and MiTF BRAF p.V600 mutations are the most frequent in low-CSD melanomas NRAS, KIT, and non-p.V600E BRAF alterations are more common in high-CSD melanomas Targeted therapies to override these pathways

51. Molecular findings CDKN2A inactivation in some hereditary and rarely sporadic melanomas Combined TERT and BRAF mutations more aggressive FISH can help distinguish melanoma from nevi Probes for 6p gain, 6q loss, 11q gain, 9p21, 8q24 Not all melanomas have these aberrations Different sensitivity and specificity in ambiguous lesions Genetic signature assay with similar accuracy exists

52. CLINICAL CONSIDERATIONS

53. Regression Sudden onset of irregular halo around the tumor Microscopically, there is dense lymphocytic infiltrate in the early stage, and vascular scar with melanophages in the late stage Partial regression is common Complete regression is less common May occur when there is already metastasis

54. Atypical melanocytic lesion Descriptive terms used when the biologic potential of an intraepidermal melanocytic lesion is uncertain (criteria fall short of melanoma) Reproducibility is poor among pathologists for intermediate lesions of low malignant potential Recommended to excise completely but conservatively MPATH-Dx schema offers a management-oriented classification based on histologic features

55. Biopsy and frozen section Incisional and punch biopsies are suitable for diagnosis When proven malignant, excision with safe margins (1-3 cm) is required Frozen section Not recommended for diagnosis or for sentinel lymph nodes Can be used for margins, but permanent sections are better

56. Spread and metastasis Radial and vertical growth phases in different types Deeper invasion has higher risk of metastasis Cutaneous metastases close to the tumor are called satellite nodules , and those between the site of the tumor and the regional lymph nodes are called in-transit metastases Cutaneous metastases may develop a secondary intraepidermal component, but they have more vascular invasion, obvious mitotic figures, patchy HMB-45, and high Ki-67 rate Nodes may be involved even if clinically negative

57. Sentinel lymph node biopsy Recommended for melanomas with Thickness ≥1 mm Dermal mitoses Ulceration If positive, lymph node dissection is carried out Examine at least three step sections with H&E and one IHC (preferably HMB-45 or a cocktail) Distinguish from benign intranodal nevus cells (present in 20% of lymphadenectomies) and histiocytes by cytology and IHC (mainly HMB-45, as other markers may be expressed by them)

58. Metastatic melanoma and nodal nevus in sentinel lymph nodes

59. Prognosis Tumor thickness and level of invasion Ulceration Dermal mitotic rate (in melanomas ≤1 mm thick) Tumor-infiltrating lymphocytes Microscopic satellites (distinct >50 micrometer nests) Sentinel lymph node involvement (including tumor burden and extranodal extension)

60. AJCC (8 th Edition)

61. AJCC (8 th Edition)

62. The End