Agustí Toll

Oncogenic PIK3CA mutations occur in epidermal nevi and seborrheic keratoses with a characteristic mutation pattern

Activating mutations of the p110 α subunit of PI3K (PIK3CA) oncogene have been identified in a broad spectrum of malignant tumors. However, their role in benign or preneoplastic conditions is unknown. Activating FGF receptor 3 (FGFR3) mutations are common in benign skin lesions, either as embryonic mutations in epidermal nevi (EN) or as somatic mutations in seborrheic keratoses (SK). FGFR3 mutations are also common in low-grade malignant bladder tumors, where they often occur in association with PIK3CA mutations. Therefore, we examined exons 9 and 20 of PIK3CA and FGFR3 hotspot mutations in EN (n = 33) and SK (n = 62), two proliferative skin lesions lacking malignant potential. Nine of 33 (27%) EN harbored PIK3CA mutations; all cases showed the E545G substitution, which is uncommon in cancers. In EN, R248C was the only FGFR3 mutation identified. By contrast, 10 of 62 (16%) SK revealed the typical cancer-associated PIK3CA mutations E542K, E545K, and H1047R. The same lesions displayed a wide range of FGFR3 mutations. Corresponding unaffected tissue was available for four EN and two mutant SK: all control samples displayed a WT sequence, confirming the somatic nature of the mutations found in lesional tissue. Forty of 95 (42%) lesions showed at least one mutation in either gene. PIK3CA and FGFR3 mutations displayed an independent distribution; 5/95 lesions harbored mutations in both genes. Our findings suggest that, in addition to their role in cancer, oncogenic PIK3CA mutations contribute to the pathogenesis of skin tumors lacking malignant potential. The remarkable genotype–phenotype correlation as observed in this study points to a distinct etiopathogenesis of the mutations in keratinocytes occuring either during fetal development or in adult life.

Postzygotic HRAS and KRAS mutations cause nevus sebaceous and Schimmelpenning syndrome

Nevus sebaceous is a common congenital cutaneous malformation. Affected individuals may develop benign and malignant secondary tumors in the nevi during life. Schimmelpenning syndrome is characterized by the association of nevus sebaceous with extracutaneous abnormalities. We report that of 65 sebaceous nevi studied, 62 (95%) had mutations in the HRAS gene and 3 (5%) had mutations in the KRAS gene. The HRAS c.37G>C mutation, which results in a p.Gly13Arg substitution, was present in 91% of lesions. Nonlesional tissues from 18 individuals had a wild-type sequence, confirming genetic mosaicism. The HRAS c.37G>C mutation was also found in 8 of 8 associated secondary tumors. Mosaicism for HRAS c.37G>C and KRAS c.35G>A mutations was found in two individuals with Schimmelpenning syndrome. Functional analysis of HRAS c.37G>C mutant cells showed constitutive activation of the MAPK and PI3K-Akt signaling pathways. Our results indicate that nevus sebaceous and Schimmelpenning syndrome are caused by postzygotic HRAS and KRAS mutations. These mutations may predispose individuals to the development of secondary tumors in nevus sebaceous.

Phacomatosis Pigmentokeratotica Is Caused by a Postzygotic HRAS Mutation in a Multipotent Progenitor Cell

Phacomatosis pigmentokeratotica (PPK) is a rare epidermal nevus syndrome characterized by the co-occurrence of a sebaceous nevus and a speckled lentiginous nevus. The coexistence of an epidermal and a melanocytic nevus has been explained by two homozygous recessive mutations, according to the twin spot hypothesis, of which PPK has become a putative paradigm in humans. However, the underlying gene mutations remained unknown. Multiple tissues of six patients with PPK were analyzed for the presence of RAS, FGFR3, PIK3CA, and BRAF mutations using SNaPshot assays and Sanger sequencing. We identified a heterozygous HRAS c.37G>C (p.Gly13Arg) mutation in four patients and a heterozygous HRAS c.182A>G (p.Gln61Arg) mutation in two patients. In each case, the mutations were present in both the sebaceous and the melanocytic nevus. In the latter lesion, melanocytes were identified to carry the HRAS mutation. Analysis of various nonlesional tissues showed a wild-type sequence of HRAS, consistent with mosaicism. Our data provide no genetic evidence for the previously proposed twin spot hypothesis. In contrast, PPK is best explained by a postzygotic-activating HRAS mutation in a multipotent progenitor cell that gives rise to both a sebaceous and a melanocytic nevus. Therefore, PPK is a mosaic RASopathy.

Keratinocytic epidermal nevi are associated with mosaic RAS mutations

Background Activating RAS mutations in the germline cause rare developmental disorders such as Costello syndrome. Somatic RAS mutations are found in approximately 30% of human cancers. Keratinocytic epidermal nevi (KEN) represent benign congenital skin lesions arranged along Blaschkos lines. A subgroup of KEN is caused by hotspot oncogenic FGFR3 and PIK3CA mutations in mosaicism, but the majority lack these mutations. Methods This study screened 72 KEN for activating mutations in RAS genes and other oncogenes. Results Activating RAS mutations were identified in 28/72 (39%) of KEN. HRAS was the most commonly affected oncogene (86%), with the HRAS p.G13R substitution representing a new hotspot mutation. Conclusion These results indicate that activating RAS somatic mutations leading to mosaicism result in benign KEN of the skin. Given the prevalence of KEN, mosaic HRAS mutations appear to be more common in patients than germline ones. These findings identify KEN as a mosaic RASopathy and lend further support to the notion that genetic mosaicism is an important contributor to disease.

Multiple oncogenic mutations and clonal relationship in spatially distinct benign human epidermal tumors

Malignant tumors result from the accumulation of genetic alterations in oncogenes and tumor suppressor genes. Much less is known about the genetic changes in benign tumors. Seborrheic keratoses (SK) are very frequent benign human epidermal tumors without malignant potential. We performed a comprehensive mutational screen of genes in the FGFR3-RAS-MAPK and phosphoinositide 3-kinase (PI3K)-AKT pathways from 175 SK, including multiple lesions from each patient. SK commonly harbored multiple bona fide oncogenic mutations in FGFR3, PIK3CA, KRAS, HRAS, EGFR, and AKT1 oncogenes but not in tumor suppressor genes TSC1 and PTEN. Despite the occurrence of oncogenic mutations and the evidence for downstream ERK/MAPK and PI3K pathway signaling, we did not find induction of senescence or a DNA damage response. Array comparative genomic hybridization (aCGH) analysis revealed that SK are genetically stable. The pattern of oncogenic mutations and X chromosome inactivation departs significantly from randomness and indicates that spatially independent lesions from a given patient share a clonal relationship. Our findings show that multiple oncogenic mutations in the major signaling pathways involved in cancer are not sufficient to drive malignant tumor progression. Furthermore, our data provide clues on the origin and spread of oncogenic mutations in tissues, suggesting that apparently independent (multicentric) adult benign tumors may have a clonal origin.

Epidermal growth factor receptor gene numerical aberrations are frequent events in actinic keratoses and invasive cutaneous squamous cell carcinomas

Please cite this paper as: Epidermal growth factor receptor gene numerical aberrations are frequent events in actinic keratoses and invasive cutaneous squamous cell carcinomas. Experimental Dermatology 2010; 19: 151–153.

Molecular diagnosis of dermatofibrosarcoma protuberans: a comparison between reverse transcriptase-polymerase chain reaction and fluorescence in situ hybridization methodologies.

Dermatofibrosarcoma protuberans (DFSP) is characterized by the presence of the t(17;22)(q22;q13) that leads to the fusion of the COL1A1 and PDGFB genes. This translocation can be detected by multiplex reverse transcriptase‐polymerase chain reaction (RT‐PCR) or fluorescence in situ hybridization (FISH) techniques. We have evaluated the usefulness of a dual color dual fusion FISH probe strategy for COL1A1/PDGFB detection in a series of 103 archival DFSPs and compared the obtained results with RT‐PCR analyses. FISH and RT‐PCR were carried out on paraffin embedded tissue samples. Regarding the RT‐PCR approach, all COL1A1 exons and exon 2 of PDGFB were evaluated. Sensitivity, specificity, positive and negative predictive values were assessed considering the histological diagnosis as the gold standard. We also analyzed the relationship between the genetic findings and the clinicopathological variables of the tumors. The COL1A1/PDGFB translocation was detected in 93% of DFSP. Both techniques showed a similar specificity (100%), but FISH was more sensitive than RT‐PCR (90% vs. 72%). Regarding, clinicopathological features, a higher percentage of positive cells detected by FISH was significantly associated with the fibrosarcomatous DFSP variant (P < 0.001). Interestingly, all CD34 negative DFSP (n = 5) were positive for COL1A1/PDGFB translocation by both techniques. In conclusion, the majority of DFSP harbor the COL1A1/PDGFB translocation and FISH technique should be recommended as a routine diagnostic tool, especially in cases showing unusual histopathological subtypes and/or immunohistochemical features.

HRAS Mutation Mosaicism Causing Urothelial Cancer and Epidermal Nevus

To the Editor: Mosaicism of an oncogenic AKT1 mutation causes Proteus syndrome, which is associated with epidermal nevi and an increased risk of cancer.1 The occurrence of oncogenic mutations in mo...

CD30 antigen expression in cutaneous inflammatory infiltrates of scabies: a dynamic immunophenotypic pattern that should be distinguished from lymphomatoid papulosis

Background: Expression of CD30 antigen is a distinct marker of lymphocyte activation that was originally described in the Reed–Sternberg cells of Hodgkins disease. The observation of CD30+ cells has been considered a diagnostic feature of cutaneous CD30 lymphoid proliferations. However, CD30 expression has also been reported in some cutaneous benign inflammatory infiltrates.

Epithelial to mesenchymal transition markers are associated with an increased metastatic risk in primary cutaneous squamous cell carcinomas but are attenuated in lymph node metastases

BACKGROUND Cutaneous squamous cell carcinoma (cSCC) is the second most common malignancy in humans and approximately 5% metastasize, usually to regional lymph nodes. Epithelial to mesenchymal transition (EMT) is a process involving loss of intercellular adhesion, acquisition of a mesenchymal phenotype and enhanced migratory potential; epithelial markers, such as E-cadherin, are down-regulated and mesenchymal proteins (Vimentin), increased. OBJECTIVE To investigate the expression of EMT markers in metastatic SCC (MSCC) and their corresponding metastases, and to correlate them with clinico-pathological factors associated with an increased risk of metastasis. METHODS We performed a retrospective study that included 146 cSCC samples (51 primary non-metastatic, 56 primary metastatic, 39 lymphatic metastases). Immunohistochemistry for E-cadherin, Vimentin, Snail, beta-catenin, Twist, Zeb1 and Podoplanin was performed. RESULTS Loss of membranous E-cadherin was observed in 77% cSCCs, with no differences between MSCC and non-MSCC. Among the transcriptional factors controlling EMT, no significant Snail1 expression was detected. Twist, Zeb1, Vimentin, beta-catenin and Podoplanin were significantly overexpressed in MSCCs. Twist ectopic expression in SCC13 cells induced Zeb1, Vimentin and Podoplanin expression and E-cadherin delocalization. These changes resulted in a scattered migration pattern in vitro. Expression of EMT markers was decreased in the metastases when compared with the corresponding primary tumors. CONCLUSION These results suggest that a partial EMT, characterized by the expression of Twist but without a total E-cadherin depletion, is involved in the acquisition of invasive traits by cSCC, but the process is downregulated in lymph node metastases.