Abstract and Introduction
Desmoplastic malignant melanoma is a distinct melanoma entity histologically subtyped into mixed and pure forms due to significantly reduced lymph node metastases in the pure form. Recent reports investigating common actionable driver mutations have demonstrated a lack of BRAF, NRAS, and KIT mutation in pure desmoplastic melanoma. In search for alternative driver mutations next generation amplicon sequencing for hotspot mutations in 50 genes cardinal to tumorigenesis was performed and in addition the RET G691S polymorphism was investigated. Data from 21 desmoplastic melanomas (12 pure and 9 mixed) were retrieved. Pure desmoplastic melanomas were either devoid of mutations (50%) or displayed mutations in tumor suppressor genes (TP53, CDKN2A, and SMAD4) singularly or in combination with the exception of a PIK3CA double-mutation lacking established biological relevance. Mixed desmoplastic melanomas on the contrary were frequently mutated (89%), and 67% exhibited activating mutations similar to common-type cutaneous malignant melanomas (BRAF, NRAS, FGFR2, and ERBB2). Separate analysis of morphologically heterogeneous tumor areas in four mixed desmoplastic malignant melanomas displayed no difference in mutation status and RET G691 status. GNAQ and GNA11, two oncogenes in BRAF and NRAS wild-type uveal melanomas, were not mutated in our cohort. The RET G691S polymorphism was found in 25% of pure and 38% of mixed desmoplastic melanomas. Apart from RET G691S our findings demonstrate absence of activating driver mutations in pure desmoplastic melanoma beyond previously investigated oncogenes (BRAF, NRAS, and KIT). The findings underline the therapeutic dichotomy of mixed versus pure desmoplastic melanoma with regard to activating mutations primarily of the mitogen-activated protein kinase pathway.
Desmoplastic malignant melanoma is a rare subtype of spindle cell malignant melanoma composed of spindled tumor cells interspersed in a characteristic, dense, paucicellular fibrous stroma, or fibromyxoid stroma. A variable admixture of an epithelioid or nondesmoplastic-spindled tumor cell component resembling conventional malignant melanoma is frequently seen. Presence of a conventional (nondesmoplastic) malignant melanoma component of >10% has been used as a cut-off to define mixed desmoplastic malignant melanoma as opposed to pure desmoplastic malignant melanoma. Desmoplastic malignant melanomas account for <4% of cutaneous melanoma cases and patients tend to present with advanced stage of disease due to lack of lesional pigmentation and occasional initial misdiagnosis. Clinically, desmoplastic malignant melanoma is frequently associated with lentigo maligna. When compared to conventional malignant melanoma of identical thickness, desmoplastic malignant melanoma has been associated with a reduced rate of lymph node metastases but higher local recurrence rates, most likely reflecting incomplete excisions of these tumors.[3,4] In addition, desmoplastic malignant melanoma exhibits a propensity for hematogenous spread primarily to the lung. Histological subclassification into pure and mixed desmoplastic malignant melanoma has been advocated on grounds of a purportedly negligible risk for lymph node metastases in pure desmoplastic malignant melanoma and restriction to sentinel node biopsy to patients with clinically conspicuous lymph nodes.[6,7] Targeted therapies (vemurafenib and imatinib) available for patients harbouring BRAF,NRAS, and KIT mutations have added therapeutic relevance to mutational subtyping of malignant melanoma. Increasing evidence demonstrated that mutational characteristics segregate with currently accepted subtyping based on histology and tumor location:[11–16] uveal melanomas present the prototypical case of a melanoma entity with a distinct mutation profile as they typically lack BRAF mutations, but instead frequently display GNAQ or GNA11 mutations.[18,19] Spitzoid melanomas demonstrate a reduced BRAF mutation frequency compared to general melanoma, and mucosal melanomas are frequently KIT mutated but BRAF wild type. Regarding desmoplastic malignant melanoma a complete lack of BRAF mutations was reported in one publication and a markedly decreased frequency of BRAF mutations in another. Subclassification of desmoplastic malignant melanoma into pure and mixed type is not only clinically relevant, but also apparently reflects different mutation patterns as well. Miller et al have recently shown that pure desmoplastic malignant melanoma is devoid of BRAF mutations as opposed to mixed desmoplastic malignant melanoma, which did exhibit BRAF mutations albeit at a low level (6% of cases). In addition, absence of BRAF mutations in an investigation including 10 desmoplastic melanomas (not further subclassified) has been reported by another group. To identify other, yet unrecognized oncogenic mutations in desmoplastic malignant melanoma we investigated 23 desmoplastic malignant melanomas by next generation amplicon sequencing using a mutation panel (IonAmpliSeq Cancer Hotspot Panel v2, Life Technologies), which covers 2855 COSMIC-annotated hotspot mutations of 50 cardinal tumor-related genes (for genes and respective amplicons included, as well as their genomic coordinates see Supplementary Table 1 https://www.nature.com/modpathol/journal/v28/n7/suppinfo/modpathol201539s1.html). This panel includes among others BRAF, CDKN2A, BRAF, TP53, PTEN, KIT as well as GNAQ, and GNA11, the latter reported in BRAF wild type uveal melanomas, but to our knowledge not yet investigated in desmoplastic malignant melanoma.
The mutation panel also includes several mutation hotspots of the receptor tyrosine kinase RET, a protooncogene with an amino-acid changing single-nucleotide polymorphism in the juxtamembrane region of RET, namely G691S (RETp). This polymorphism enhances the response of RET to glial cell line-derived neurotrophic factor, which activates both the RET-RAF-RAS-MEK-ERK pathway and the RET-PIK3-AKT pathway.[25,26] This activation has been linked to the observed neurotropism in desmoplastic melanoma and pancreatic ductal adenocarcinoma among others. Narita et al have described a significantly higher frequency of RETp in desmoplastic melanomas (61%) compared to nondesmoplastic melanomas (31%) analyzing RETp from tumor DNA, but Miller et al have only detected RETp in 33% of pure and in 24% of mixed desmoplastic malignant melanoma. In the light of occasional, somatic, de novo RETp occurrences reported in pancreatic adenocarcinoma, Bar et al have investigated germline DNA for RETp in melanoma patients and found germline RETp in 30% of patients with desmoplastic melanomas, compared to 21% in nondesmoplastic malignant melanoma patients, but this difference was not significant at the investigated sample size. As RET codon 691 is not covered by the abovementioned hotspot panel, tumor DNA was subjected to Sanger sequencing for additional evaluation of this RET codon in order to encompass germline and possible somatic RET G691S occurrences.
Mod Pathol. 2015;28(7):895-903. © 2015 Nature Publishing Group