The relevance of miRNAs as promising biomarkers in lip cancer.

OBJECTIVES: This study aimed to analyze the expression of miR-181b, miR-21, miR-31, and miR-345 in actinic cheilitis with and without epithelial dysplasia and lower lip squamous cell carcinomas, and to verify if the deregulated expression of these miRNAs would be indicative of malignant transformation. MATERIALS AND METHODS: The sample was selected from formalin-fixed paraffin-embedded tissues of 19 actinic cheilitis without epithelial dysplasia, 32 actinic cheilitis with epithelial dysplasia, 42 lower lip squamous cell carcinomas, and 10 nonaltered oral mucosa of the lip. The microRNA (miR, miRNA) expression was quantified by real-time RT-PCR and the expression of the selected miRNAs among the groups of actinic cheilitis and lower lip cancer was compared by chi-square. RESULTS: A higher expression of miR-181b, miR-31, and miR-345 was found in actinic cheilitis without epithelial dysplasia in comparison to that in actinic cheilitis with epithelial dysplasia and with lower lip cancer. There were no differences in miR-21 expression between actinic cheilitis and lower lip cancer. Hierarchical clustering analysis showed a tendency for a downregulation of miR-181b, miR-21, miR-31, and miR-345 in most patients with lower lip cancers. CONCLUSIONS: The upregulation of miR-181b, miR-31, and miR-345 expression in actinic cheilitis without epithelial dysplasia and the decrease in the expression of these miRNAs in actinic cheilitis with epithelial dysplasia and in lower lip cancer are potential biomarkers of malignant progression. CLINICAL RELEVANCE: This miRNA signature can help to identify actinic cheilitis with potential to progress to lip cancer.

PLACK syndrome is potentially treatable with intralipids.

We describe an 11-year-old girl with PLACK Syndrome (peeling skin, leukonychia, acral punctate keratosis, cheilitis, and knuckle pads), who was found to have a novel homozygous variant in CAST, the pathogenicity of which was confirmed using blood-derived RNA. There is no established treatment for PLACK syndrome. However, we demonstrate for the first time that this condition is associated with low levels of vitamin A and essential fatty acids, which prompted us to consider a potential treatment strategy. Indeed, we initiated this patient on intravenous lipid infusion (Vitalipid®; an emulsion of fat-soluble vitamins and lipofundin-MCT/LCT 20%) and the response was dramatic. Following the fourth monthly course of treatment, pruritis disappeared and the skin lesions showed remarkable objective improvement. PLACK syndrome is a very rare genodermatosis and only six families have been described to date with pathogenic CAST variants. This is the first report of an objective response to a therapeutic agent, which suggests that PLACK is a potentially treatable condition. The remarkable response we report and the relative safety of the intervention should prompt healthcare providers who care for PLACK syndrome patients to explore this as a potential treatment strategy in future studies.

Immunoexpression of glucocorticoid receptor alpha (GRα) isoform and apoptotic proteins (Bcl-2 and Bax) in actinic cheilitis and lower lip squamous cell carcinoma.

BACKGROUND: Actinic cheilitis (AC) is a potentially malignant disorder that can progress to squamous cell carcinoma (SCC), but this process is not fully understood. This study evaluated the immunoexpression of glucocorticoid receptor alpha (GRα) isoform and apoptotic proteins (Bcl-2 and Bax) in AC and lower lip SCC (LLSCC). METHODS: Twenty-two AC and 44 LLSCCs (22 with regional nodal metastasis and 22 without metastasis) were selected. The percentages of nuclear (GRα) and cytoplasmic (GRα, Bcl-2, and Bax) staining in epithelial cells were assessed and correlated with clinical (tumor size/extent and clinical stage) and histopathological parameters (risk of malignant transformation for AC and histopathological grade of malignancy for LLSCCs). RESULTS: Expression of GRα was observed in all cases studied, with relatively high median percentages of positive staining. When compared to AC, LLSCCs exhibited lower nuclear expression and higher cytoplasmic expression of GRα (P < 0.05). Regarding clinicopathological parameters, significant differences were only found for cytoplasmic expression of GRα according to the histopathological grade of LLSCCs (P = 0.036). Higher expression of Bax compared to Bcl-2 was observed in AC and LLSCCs (P < 0.05). In LLSCCs, there was a positive correlation between nuclear and cytoplasmic expressions of GRα (P = 0.006). CONCLUSION: Reduced nuclear translocation and increased cytoplasmic expression of GRα may be important events in lip carcinogenesis but are not involved in the progression of LLSCC. The role of GRα in lip cancer development does not seem to be primarily related to modulations in the expression of Bcl-2 or Bax.

Evaluation of specific modified histones in lip carcinogenesis.

OBJECTIVE: Histones regulate chromatin density and therefore influence gene expression and cellular proliferation. These properties are modified by methylation, acetylation and phosphorylation of histones. The aim of this study was to investigate the variation of specific modified histones in actinic cheilitis (AC) and squamous cell carcinoma of the lip (SCCL). METHODS: Samples of non-neoplastic tissue of the lip (NNTL, n = 9), AC (n = 33), and SCCL (n = 27) were submitted to immunohistochemistry to detect the modified histones H3K36me3, H3K9ac, H4K12ac, and H3S10 ph. RESULTS: Reactivity for all of the modified histones was significantly decreased from NNTL to AC, but not from AC to SCCL. Dysplasia in AC or histological grade in SCCL were not related to the reactivity of any modified histones. CONCLUSIONS: Histone modifications are related to initial actinic damage, but not to malignant transformation in the lip.

Lip cancer and pre-cancerous lesions harbor TP53 mutations, exhibit allelic loss at 9p, 9q, and 17p, but no BRAFV600E mutations.

Molecular mechanisms of lip squamous cell carcinoma (LSCC) and actinic cheilitis (AC) are unclear. We aimed at assessing loss of heterozygosity (LOH) and TP53 and BRAF V600E mutations in these lesions. Formalin-fixed paraffin-embedded (FFPE) samples of 17 LSCC and 16 AC were included, with additional 5 fresh LSCC genotyped for TP53 mutations. LOH was assessed by six polymorphic markers located at 9p22, 9q22, and 17p13 and correlated with cell proliferation (Ki-67) and P53 immunostaining. Direct sequencing of TP53 exons 2-11 (fresh samples), and exons 5-9 (FFPE samples) was carried out. BRAF V600E mutation was genotyped in eight LSCC. LOH occurred in at least one marker in 15/17 LSCC and in 9/16 AC. The marker exhibiting the highest frequency of allelic loss (FAL) in LSCC was D9S157 (8/12 informative cases) and D9S287 in AC (4/11 informative cases). Cell proliferation was not correlated with LOH or with the FAL and no correlation between P53 IHC and 17p LOH was observed. We found TP53 missense mutations in both lesions and nonsense in LSCC, including CC>TT transition, which is a marker of UV damage. BRAF V600E mutation was not detected. LOH and TP53 mutations detected in LSCC and AC may be associated with tumorigenesis, whereas BRAF V600E mutation does not seem to significantly contribute to LSCC pathogenesis.

Loss-of-function mutations in CAST cause peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads.

Calpastatin is an endogenous specific inhibitor of calpain, a calcium-dependent cysteine protease. Here we show that loss-of-function mutations in calpastatin (CAST) are the genetic causes of an autosomal-recessive condition characterized by generalized peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads, which we propose to be given the acronym PLACK syndrome. In affected individuals with PLACK syndrome from three families of different ethnicities, we identified homozygous mutations (c.607dup, c.424A>T, and c.1750delG) in CAST, all of which were predicted to encode truncated proteins (p.Ile203Asnfs∗8, p.Lys142∗, and p.Val584Trpfs∗37). Immunohistochemistry shows that staining of calpastatin is reduced in skin from affected individuals. Transmission electron microscopy revealed widening of intercellular spaces with chromatin condensation and margination in the upper stratum spinosum in lesional skin, suggesting impaired intercellular adhesion as well as keratinocyte apoptosis. A significant increase of apoptotic keratinocytes was also observed in TUNEL assays. In vitro studies utilizing siRNA-mediated CAST knockdown revealed a role for calpastatin in keratinocyte adhesion. In summary, we describe PLACK syndrome, as a clinical entity of defective epidermal adhesion, caused by loss-of-function mutations in CAST.

STAG2 expression in oral cancer and potentially malignant lesions.

Oral cancer is a world health problem, and one of the highest incidence rates of oral cancer worldwide occurs in Brazil. STAG2 is part of the cohesin complex which is responsible for sister chromatid cohesion. STAG2 loss of expression was reported in a range of tumors, and STAG2 loss was found to cause chromosomal instability and aneuploidy in cancer cells. On the basis of these findings, we investigated STAG2 expression in oral cancer and potentially malignant lesions. We investigated STAG2 immunoexpression in oral cancer, lip cancer, oral leukoplakia, and actinic cheilitis, including complete clinical information. Normal oral mucosa samples were included as normal controls. STAG2 protein was highly expressed in all samples. We further tested STAG2 expression in gastric adenocarcinomas and glioblastomas, as these tumor types were previously shown to lose STAG2 expression. We found homogenous expression of STAG2 by these tumor cells. Our results suggest that STAG2 loss of expression is not a common event in oral carcinogenesis.

Solar cheilosis: an ominous precursor: part I. Diagnostic insights.

Squamous cell carcinoma (SCC) of the lower lip is a deadly nonmelanoma skin cancer. Its precursor, a distinctive cutaneous neoplasia analogous to cervical dysplasia, is known by the confusing term actinic cheilitis. Solar cheilosis (SC) is a more appropriate designation. It represents incipient SCC in situ. SC is widely recognized as an ultraviolet light-induced precancer of the lower lip that is typically seen in light-skinned individuals and others with poorly pigmented lower lips. Lip SCC is one of the most common malignancies of the oral cavity. SCC is much more likely to metastasize from the lip than cutaneous surfaces, with a 5-year overall survival rate of less than 75%. SC results from long-term exposure to ultraviolet radiation. The occurrence of SC is dose-dependent and is influenced by the patient's solar exposure, age, genetic predisposition, geographic latitude of residence, occupation, leisure activities, and use of lip protective agents. Molecular abnormalities of SC are similar to those of actinic keratosis and facilitate the evolution to SCC. A high degree of clinical suspicion should be maintained, given the malignant nature of this condition. Ulceration and nodularity often indicate progression to SCC. We performed a Medline and Google Scholar search for all articles related to actinic cheilitis, actinic cheilosis, SC, actinic keratosis, solar keratosis, premalignant oral disease, and lip SCC, and have also evaluated many other articles and book chapters. One hundred forty-two peer-reviewed articles were identified as being of particular value. Pertinent facts were selected and analyzed.

Identification of novel fibroblast growth factor receptor 3 gene mutations in actinic cheilitis and squamous cell carcinoma of the lip.

OBJECTIVE: Activating mutations in the fibroblast growth factor receptor 3 (FGFR3) gene are responsible for several craniosynostosis and chondrodysplasia syndromes as well as some human cancers, including bladder and cervical carcinoma. Despite a high frequency in some benign skin disorders, FGFR3 mutations have not been reported in cutaneous malignancies. Actinic cheilitis (AC) is a sun-induced premalignancy affecting the lower lip that frequently progresses to squamous cell carcinoma (SCC). The objective of this study was to determine if FGFR3 gene mutations are present in AC and SCC of the lip. STUDY DESIGN: DNA was extracted and purified from microdissected, formalin-fixed, paraffin-embedded tissue sections of 20 cases of AC and SCC arising in AC. Exons 7, 15, and 17 were PCR amplified and direct sequenced. RESULTS: Four novel somatic mutations in the FGFR3 gene were identified: exon 7 mutation 742C-->T (amino acid change R248C), exon 15 mutations 1850A-->G (D617G) and 1888G-->A (V630M), and exon 17 mutation 2056G-->A (E686K). Grade of dysplasia did not correlate with presence of mutations. CONCLUSION: The frequency of FGFR3 receptor mutations suggests a functional role for the FGFR3 receptor in the development of epithelial disorders, and perhaps this change may contribute to the pathogenesis of some AC and SCC.

p53 and MDM2 protein expression in actinic cheilitis.

Actinic cheilitis is a potentially malignant lip lesion caused by excessive and prolonged exposure to ultraviolet radiation, which can lead to histomorphological alterations indicative of abnormal cell differentiation. In this pathology, varying degrees of epithelial dysplasia may be found. There are few published studies regarding the p53 and MDM2 proteins in actinic cheilitis. Fifty-eight cases diagnosed with actinic cheilitis were histologically evaluated using Banóczy and Csiba (1976) parameters, and were subjected to immunohistochemical analysis using the streptavidin-biotin method in order to assess p53 and MDM2 protein expression. All studied cases expressed p53 proteins in basal and suprabasal layers. In the basal layer, the nuclei testing positive for p53 were stained intensely, while in the suprabasal layer, cells with slightly stained nuclei were predominant. All cases also tested positive for the MDM2 protein, but with varying degrees of nuclear expression and a predominance of slightly stained cells. A statistically significant correlation between the percentage of p53 and MDM2-positive cells was established, regardless of the degree of epithelial dysplasia. The expression of p53 and MDM2 proteins in actinic cheilitis can be an important indicator in lip carcinogenesis, regardless of the degree of epithelial dysplasia.

p53 and MDM2 protein expression in actinic cheilitis

Actinic cheilitis is a potentially malignant lip lesion caused by excessive and prolonged exposure to ultraviolet radiation, which can lead to histomorphological alterations indicative of abnormal cell differentiation. In this pathology, varying degrees of epithelial dysplasia may be found. There are few published studies regarding the p53 and MDM2 proteins in actinic cheilitis. Fifty-eight cases diagnosed with actinic cheilitis were histologically evaluated using Banóczy and Csiba (1976) parameters, and were subjected to immunohistochemical analysis using the streptavidin-biotin method in order to assess p53 and MDM2 protein expression. All studied cases expressed p53 proteins in basal and suprabasal layers. In the basal layer, the nuclei testing positive for p53 were stained intensely, while in the suprabasal layer, cells with slightly stained nuclei were predominant. All cases also tested positive for the MDM2 protein, but with varying degrees of nuclear expression and a predominance of slightly stained cells. A statistically significant correlation between the percentage of p53 and MDM2-positive cells was established, regardless of the degree of epithelial dysplasia. The expression of p53 and MDM2 proteins in actinic cheilitis can be an important indicator in lip carcinogenesis, regardless of the degree of epithelial dysplasia.

Follicular cheilitis. A distinctive histopathologic finding in actinic prurigo.

Actinic prurigo (AP), a chronic skin disease caused by an abnormal reaction to sunlight, is commonly associated with cheilitis and conjunctivitis. Characteristic ethnic, genetic, environmental (occurs at high altitudes, above 1,500 m), clinical, and histopathologic features have been reported. AP occurs in American Indians of Canada and the United States and most commonly in Latin American countries, where Mestizos (mixed ancestry) are predominantly affected. The present study investigates AP involving the lips, where it is characterized by a dense lymphocytic infiltrate, often with well-formed lymphoid follicles; the latter feature we refer to as "follicular cheilitis" (FC). The histopathologic findings of FC are characteristic of and helpful in diagnosing actinic prurigo involving the lips.

Chronic granulomatous disease in an adult female with granulomatous cheilitis. Evidence for an X-linked pattern of inheritance with extreme lyonization.

We describe in this paper a female patient affected by chronic granulomatous disease with all the features of the classic X-linked form of the disease and presenting a mild form of the disease, the major clinical manifestation being a granulomatous cheilitis. The capability of the patient's phagocytes to undergo a respiratory burst in response to different stimuli was markedly depressed and only 10% of the patient's neutrophils were able to reduce nitroblue tetrazolium when stimulated with phorbol myristate acetate to an extent similar to normal cells. With this test, the neutrophils of the patient's mother showed a clear mosaicism, only 40% being able to reduce the dye. Activation of NADPH oxidase in cell-free systems showed that the phagocyte defect was at the level of a membrane component. Difference in spectra revealed that the observed membrane defect was due to a lack of cytochrome b558, the terminal component of NADPH oxidase. Incubation for 2 or 24 h of the patient's neutrophils with human recombinant interferon-gamma and granulocyte macrophage colony-stimulating factor did not correct their defective capability to undergo a respiratory burst However, cultivation of the patient's monocytes with interferon-gamma for prolonged times substantially enhanced their capability to produce hydrogen peroxide.

Actinic prurigo of the lower lip. Review of the literature and report of five cases.

Actinic prurigo (AP) is a chronic, familial, photodermatitis that primarily affects American Indians. It is more prevalent in young females and may be evident clinically as a pruritic lower lip cheilitis that typically does not respond to conventional therapy. Other clinical features associated with AP include conjunctivitis, alopecia of the eyebrows, and formation of pterygia. The histologic features of AP have been described as nonspecific. We are presenting five cases of AP in which the major clinical manifestation was a pruritic, unsightly lower lip cheilitis. All of our patients were American Indians. Three patients were from the same family. The youngest patient was 10 years old and the oldest was 69 years old. All five cases were characterized histologically by numerous germinal centers within the lamina propria and a dense perivascular plasma cell infiltrate. In four of the five cases, there was a moderate to dense infiltrate of eosinophils. All five patients had been generally unresponsive to conventional therapy. With the exception of the youngest patient, all had had the disease for at least several years. One patient, now deceased, had the lower lip lesion surgically removed for cosmetic reasons. To our knowledge, this is the first report in the dental literature that describes the clinical and histologic features of AP of lower lip.