The classification of ameloblastoma in multicystic or unicystic variants is associated with its clinical behaviour. Recently, BRAF and SMO mutations have been reported in ameloblastomas. However, it is not clear if such mutations are shared by the multi- and unicystic variants of ameloblastoma or by odontogenic carcinomas. We assessed BRAFV600E and SMOF412E in multicystic, unicystic and desmoplastic ameloblastomas. In addition, we investigated whether the BRAFV600E mutation occurs in odontogenic carcinomas. A total of 28 formalin-fixed paraffin-embedded samples, comprising 17 ameloblastomas and 11 odontogenic carcinomas, were included. The BRAFV600E mutation was assessed by real-time PCR with a specific TaqMan probe and confirmed by Sanger sequencing. The SMOF412E mutation was assessed by Sanger sequencing. Fourteen out of 17 (82 %) ameloblastomas showed the BRAFV600E mutation, specifically, 5/6 (83 %) unicystic, 7/9 (78 %) multicystic and 2/2 desmoplastic ameloblastomas. BRAFV600E mutation was detected in 4/11 (36 %) malignant tumours, specifically, 3/8 (38 %) ameloblastic carcinomas and 1/1 clear cell odontogenic carcinoma, while the two ghost cell odontogenic carcinomas did not harbour this mutation. The SMOF412E mutation was not detected in ameloblastoma. The BRAFV600E-activating mutation is a common event in ameloblastomas, occurring regardless of site or histological type. This mutation is also detected in odontogenic carcinomas. SMO somatic mutation is a secondary genetic event in the ameloblastoma pathogenesis. Our findings support the possibility for personalised, molecular-targeted therapy for ameloblastomas and odontogenic carcinomas harbouring the BRAFV600E mutation.

Giant cell lesions of the jaw (GCLJ) are debilitating tumors of unknown origin with limited available therapies. Here, we analyze 58 sporadic samples using next generation or targeted sequencing and report somatic, heterozygous, gain-of-function mutations in KRAS, FGFR1, and p.M713V/I-TRPV4 in 72% (42/58) of GCLJ. TRPV4 p.M713V/I mutations are exclusive to central GCLJ and occur at a critical position adjacent to the cation permeable pore of the channel. Expression of TRPV4 mutants in HEK293 cells leads to increased cell death, as well as increased constitutive and stimulated channel activity, both of which can be prevented using TRPV4 antagonists. Furthermore, these mutations induce sustained activation of ERK1/2, indicating that their effects converge with that of KRAS and FGFR1 mutations on the activation of the MAPK pathway in GCLJ. Our data extend the spectrum of TRPV4 channelopathies and provide rationale for the use of TRPV4 and RAS/MAPK antagonists at the bedside in GCLJ.

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The BRAFV600E antibody (clone VE1) IHC may show non-specific staining, but molecular assays may be useful for the diagnosis of unicystic ameloblastoma, in conjunction with clinical, radiological and histopathological features.

Ameloblastoma is a locally destructive and invasive tumour that can recur despite adequate surgical removal. Molecular studies have offered interesting findings regarding ameloblastoma pathogenesis. In the present review, the following topics are discussed regarding its molecular nature: clonality, cell cycle proliferation, apoptosis, tumour suppressor genes, ameloblastin and other enamel matrix proteins, osteoclastic mechanism and matrix metalloproteinases and other signalling molecules. It is clear from the literature reviewed that translational studies are necessary to identify prognostic markers of ameloblastoma behaviour and to establish new diagnostic tools to the differential diagnosis of unicystic from multicystic ameloblastoma. Finally, molecular biology studies are also important to develop more effective alternative approaches to the treatment of this aggressive odontogenic tumour.

The first step towards the prevention of cancer is to develop an in-depth understanding of tumourigenesis and the molecular basis of malignant transformation. What drives tumour initiation? Why do most benign tumours fail to metastasize? Oncogenic mutations, previously considered to be the hallmark drivers of cancers, are reported in benign cysts and tumours, including those that have an odontogenic origin. Despite the presence of such alterations, the vast majority of odontogenic lesions are benign and never progress to the stage of malignant transformation. As these lesions are likely to develop due to developmental defects, it is possible that they harbour quiet genomes. Now the question arises - do they result from DNA replication errors? Specific candidate genes have been sequenced in odontogenic lesions, revealing recurrent BRAF mutation in the case of ameloblastoma, KRAS mutation in adenomatoid odontogenic tumours, PTCH1 mutation in odontogenic keratocysts, and CTNNB1 (Beta-catenin) mutation in calcifying odontogenic cysts. Studies on these benign and rare entities might reveal important information about the tumorigenic process and the mechanisms that hinder/halt neoplastic progression. This is because the role of relatively common oncogenic mutations seems to be context dependent. In this review, each mutation signature of the odontogenic lesion and the affected signalling pathways are discussed in the context of tooth development and tumorigenesis. Furthermore, behavioural differences between different types of odontogenic lesions are explored and discussed based on the molecular alteration described. This review also includes the employment of molecular results for guiding therapeutic approaches towards odontogenic lesions.

Letter to the editor Recurrent KRAS G12V pathogenic mutation in adenomatoid odontogenic tumours Dear Editor, The adenomatoid odontogenic tumour (AOT) is a non-aggressive encapsulated tumour, being usually diagnosed in association with an unerupted permanent maxillary canine [1,2]. There are scarce reports of multiple AOTs [3-5] and a patient with Schimmelpenning syndrome (SS) with AOT was reported [6]. SS is characterized by sebaceous nevi, associated with ipsilateral abnormalities of the central nervous system, resulting from postzygotic autosomal dominant HRAS or KRAS lethal mutations that survive by somatic mosaicism [7]. RAS mutations were previously reported in lesional tissue (including nevus sebaceous) of a patient, but not in normal skin or blood leukocytes, consistent with a somatic mosaicism [7]. We evaluated one AOT sample from a SS patient having multiple AOTs (index patient) and two sporadic AOTs (samples #1 and #2) for mutations in a panel of 50 oncogenes and tumour suppressor genes, including RAS family, by using Ion AmpliSeq TM Cancer Hotspot Panel v2 (Life Technologies, Carlsbad, USA). After filtering by missense variants, candidate variants from the panel were defined as those pathogenic variants in regions with a depth greater than X500 and frequency greater than 5%. Only KRASc.35G > T (KRASG12V) fit this criteria, and was validated by TaqMan Ò Mutation Detection Assay using the probes KRAS_476_mu and KRAS_rf (Applied Biosystems, Foster City, USA). We further interrogated the KRASG12V mutation in six extra AOTs (samples #3-8) by the TaqMan Ò Assay. This KRAS mutation was detected in the three samples, as well as in four (samples #3, #4, #5 and #7) out of six additional samples. The mutation was validated by Sanger sequencing (Fig. 1). No other pathogenic mutation interrogated was detected. Blood leukocytes from the index patient were negative for KRASG12V mutation. To determine the specificity of the KRASG12V mutation in the context of odontogenic tumours, we evaluated three ameloblastomas, two dentinogenic ghost cell tumours and two normal oral mucosa samples using the TaqMan Ò Assay, being all negative for the mutation. Constitutively activation of the MAPK pathway by BRAFV600E mutation was reported in ameloblastoma [8-10], and in ameloblastic carcinoma [11]. We describe a recurrent oncogenic mutation in an upstream activator of MAPK, KRAS, in AOT. RAS mutations are found in 30% of human cancers and 80% of KRAS mutations occur at codon 12, being highly frequent in lung adenocarcinoma, pancreatic and colon carcinomas [12]. In our series, seven out of nine AOTs exhibited the KRASG12V mutation. The KRAS mutation was identified in the index patient sample and in sporadic AOTs, a candidate to driver mutation in these lesions. Driver mutations confer growth advantage to tumour cells and are positively selected during cancer evolution [13].

Our data show methylation of the promoter of P21 gene in OKCs. In addition, methylation of the P27 and RB1 genes are commonly found in dental follicles. Further studies are necessary to determine the functional relevance of these alterations.