Possible role of ALDH1 and CD44 in lip carcinogenesis.

BACKGROUND: Lip squamous cell carcinoma (LSCC) accounts for 12% of all head and neck cancers. It is caused by chronic exposure to ultraviolet light solar radiation and related to previous actinic cheilitis (AC). This study aimed to investigate the immunostaining of the putative cancer stem cells (CSC) markers ALDH1 and CD44 in AC (n=30) and LSCC (n=20). ALDH1 positivity was found to be statistically higher in LSCC than in AC lesions (p=0.0045), whilst CD44 expression was statistically higher in AC than in LSCC lesions (p=0.0155). ALDH1+ cells in AC lesions were associated with specific clinical features: a younger age (<60 years old), the female gender, white skin, not smoking or consuming alcohol, and a fast evolution, and not associated with the chronic exposure to UV radiation (p<0.0001). CD44 positivity was associated with patients who were male, feoderm, smoked, consumed alcohol, underwent occupational exposure to UV-radiation, and demonstrated lesions with log-time evolution (p<0.0001). ALDH1 + cells were associated with mild dysplasia using a system from the World Health Organization (WHO), and with a low risk of malignant transformation, according to the binary system (p<0.0001). CD44+ cells were also associated with moderated dysplasia, according to the WHO system. In LSCC, ALDH1 + cells were positively associated with patients who were older (≥ 60 years old), smokers, and with those who consumed alcohol (p<0.0001). CD44 + cells in LSCC were associated with older (≥ 60 years old) patients as well, but also with female patients, white skin, non-smokers, and individuals who did not consume alcohol (p<0.0001), all of whom showed distinct patterns in pre- and malignant lesions of both markers. Additionally, in LSCC, both ALDH1 and CD44 staining were associated with smaller tumor sizes (T1/T2; p<0.0001). In summary, although both ALDH1 and CD44 were associated with the presence of dysplasia in AC lesions, the present findings suggest that ALDH1 and CD44 may be activated by different etiopathogenic pathways, predominantly in distinct steps of oral carcinogenesis. CD44 would thus be more significantly related to the potentially malignant lesion, while ALDH1 would be closely linked to malignancy.

Distinctive expression of DNA replication factors in squamous cell carcinomas of the lip, face and oral cavity.

OBJECTIVE: Uncontrolled proliferation and aberrations in cell-cycle progression are fundamental issues in cancer. In this study we aimed to determine and compare deoxyribonucleic acid (DNA) replication licensing factors at the mRNA and protein levels among squamous cell carcinomas (SCCs) of the lip, facial-skin and oral cavity. MATERIALS AND METHODS: A total of 103 lip, oral and face SCCs were immunohistochemically stained with MCM2 (mini-chromosome maintenance 2), geminin, and ki67, and their labeling-indices were calculated. Also, 57 SCCs from the same regions along with their adjacent normal tissues underwent quantitative reverse transcription-polymerase chain reaction analysis. RESULTS: All three proteins were overexpressed in the studied SCCs, but only geminin (P = 0.004) showed significant difference among the three regions, with higher levels in oral SCCs compared to lip (P = 0.005) and skin (P = 0.024) tumors. Geminin expression did not differ between skin- and lip-SCCs (P = 0.822). MCM2/ki67 ratio was higher in oral- compared to skin-neoplasms (P = 0.039), but no difference was found in geminin/ki67 among the SCC-subsites. There were significant differences in MCM2 and geminin mRNA between carcinomatous- and normal-tissues in all tumors, but not among the three locations. CONCLUSION: MCM2 and geminin are involved in the tumorigenesis of lip, face and oral SCC at both mRNA- and protein-levels. Geminin may have a role in the site-specific biologic behavior of SCC. Skin SCCs had the highest proportion of licensed non-proliferating cells, while actively proliferating cells were more prominent in oral tumors. Regarding DNA replication, lip SCCs seem to be closer to skin tumors compared to their oral counterparts.

EGCG inhibits growth of tumoral lesions on lip and tongue of K-Ras transgenic mice through the Notch pathway.

Epigallocatechin-3-gallate (EGCG), the main active ingredient of green tea, exhibits low toxic side effect and versatile bioactivities, and its anti-cancer effect has been extensively studied. Most of the studies used cancer cell lines and xenograft models. However, whether EGCG can prevent tumor onset after cancer-associated mutations occur is still controversial. In the present study, Krt14-cre/ERT-Kras transgenic mice were developed and the expression of K-RasG12D was induced by tamoxifen. Two weeks after induction, the K-Ras mutant mice developed exophytic tumoral lesions on the lips and tongues, with significant activation of Notch signaling pathway. Administration of EGCG effectively delayed the time of appearance, decreased the size and weight of tumoral lesions, relieved heterotypic hyperplasia of tumoral lesions, and prolonged the life of the mice. The Notch signaling pathway was significantly inhibited by EGCG in the tumoral lesions. Furthermore, EGCG significantly induced cell apoptosis and inhibited the proliferation of tongue cancer cells by blocking the activation of Notch signaling pathway. Taken together, these results indicate EGCG as an effective chemotherapeutic agent for tongue cancer by targeting Notch pathway.

A rare case of high-grade intraductal carcinoma of the upper lip: immunohistochemical and genetic analyses.

Although intraductal carcinoma (IDC) of the salivary glands was previously called low-grade cribriform cystadenocarcinoma, it was newly categorized in the 4th version of the World Health Organization classification. We report a case of IDC of the upper lip and examined it immunohistochemically and genetically. The patient was a 48-year-old Japanese female, who noticed a tiny nodule on her left upper lip. Histologically, the tumor cells, which had eosinophilic cytoplasm, exhibited papillary and solid growth patterns, and regions of suspected microinvasion or intraductal spread were also seen at the periphery of the tumor. Small necrotic foci were noted. Immunohistochemically, the tumor cells were diffusely positive for the androgen receptor, CK19, CK5/6, EGFR, and SOX10, whereas they were focally positive for GCDFP-15, S-100 protein, and mammaglobin. The tumor nests were surrounded by alpha-smooth muscle actin-p63-/calponin-/CK14-positive myoepithelial cells. The Ki-67 labeling index was 51.2%. Genetic analysis showed no evidence of the TRIM27-RET or NCOA4-RET fusion gene. We finally diagnosed the tumor as a high-grade mixed intercalated duct/apocrine-type IDC of the upper lip. IDC of the minor salivary glands is exceedingly rare. We discuss diagnostic problems associated with minor salivary gland lesions, and the "basal-like" phenotype of this case.

Low-dose radiotherapy for extranodal marginal zone B lymphoma of the lip: Case report and literature review.

Non-Hodgkin lymphoma (NHL) of the lip is extremely rare. It is usually indolent and in early stages a local approach is often indicated. We present a case report of a patient with extranodal NHL of the lip treated with chemotherapy and low-dose radiation treatment (RT). The patient was affected by B-cell NHL of the marginal zone, Stage IAE. After a few months of observation with progressive disease, the patient was submitted to two cycles of chemotherapy with no response. Therefore, he was treated with very low-dose RT consisting of two fractions of 2 Gy. Complete response was observed and after 1-year follow-up, persistent complete response was recorded. In cases of localized disease, especially in patients with comorbidities of poor performance status (PS), low-dose RT can be an appropriate approach with excellent outcomes in terms of effectiveness and low risk of toxicity.

Solitary Lip Extranodal Natural Killer/T-Cell Lymphoma on FDG PET/CT.

A 54-year-old man presented with a history of upper lip pain for 4 weeks. Biopsy of the lip lesion revealed extranodal natural killer/T-cell lymphoma. F-FDG PET/CT scan showed the solely high uptake in the right upper lip without any other nodal or extranodal involvements.

Non- Hodgkin lymphoma of the lips: A rare entity.

AIM: To investigate clinico-pathological features of lymphoma of the lips, and review the literature. MATERIALS AND METHODS: Retrospective analysis and review of English literature, 1996-2016. RESULTS: Analysis included 23 cases, 7 new cases and 16 from literature, 12 M: 11 F, age 7-82 years. Four occurred in children, mean age 10.1; 19 in adults, mean 61.1 years. The lower lip was involved in the majority of cases (16, 69.56%). 14 (60.87%) were isolated to the lips, 8 (34.78%) were multifocal. Nine (39.13%) occurred in association with Sjogren's syndrome, of which one also had Hashimoto thyroiditis. IgG4-related disease and HIV were reported in one case each. The lip salivary glands were involved in most cases (19, 82.6%); 3 (13.6%) showed only cutaneous involvement. The typical presentation was single or multiple nodules (15, 65.21%), with surface ulceration in only two (8.69%). Constituent symptoms were absent in all cases, paresthesia was reported in one (4.34%). The majority (18, 78.26%) was extranodal marginal zone B-cell lymphoma - mucosa-associated lymphoid tissue lymphoma (EMZB-MALT), and one case each was mantle cell, NK-T cell, CD30 positive and plasmablastic lymphoma. CONCLUSION: The lips seem to have a unique pattern of non-Hodgkin lymphoma dominated by EMZB-MALT lymphoma, rarely other types. In more than half, neither Sjogren's syndrome nor other chronic inflammation was identified. Lesions tend to present as asymptomatic slowly progressing, non-ulcerated submucosal masses. Lymphoma should be considered even in the absence of constituent symptoms, as most cases showed none. Although the number of reported cases is rather small, disease course is usually prolonged and prognosis seems to be good.

Multiple keratin granulomata: A potential histologic clue to cutaneous squamous cell carcinoma responding to programmed cell death protein 1 inhibitor therapy.

We present histologic features of a locally advanced cutaneous squamous cell carcinoma (cSCC) treated with the programmed cell death protein-1 (PD-1) antagonist, pembrolizumab, with a partial response. This contributes to a growing body of literature supporting the efficacy of pembrolizumab in treatment of surgically unresectable cSCC. We also provide a detailed description of the histologic features, particularly keratin granulomata with adjacent lymphocytic aggregates and fibrosis, observed in cSCC under treatment with a PD-1 antagonist.

The stem cell markers expression CD44v6 and podoplanin in lip cancer: clinical significance.

This study aimed to analyze the immunoexpression of cancer stem cell markers, CD44v6, and podoplanin in 91 patients with lip squamous cell carcinomas (LSCC). The immunostaining of podoplanin and CD44v6 was evaluated in ten high-power fields (× 400 magnification) at the invasive front of LSCC, using a semi-quantitative score method. Chi-square test or Fisher's exact test was used to verify the association of podoplanin and CD44v6 expressions with clinicopathologic variables. Spearman's correlation test was used to analyze the correlation between the two antibodies in lip cancer. Disease-free survival probabilities in 5 and 10 years were estimated according to the Kaplan-Meier method and compared using the log-rank test. The independent effects of the significant variables were analyzed by Cox proportional hazards regression model. A strong podoplanin expression was observed in the membrane and cytoplasm of most lip tumor cells, and this was inversely associated with locoregional recurrence (p = 0.028) and with histopathological grade of malignancy (p = 0.026). Additionally, CD44v6 immunostaining was strongly expressed in the membrane of tumor cells in 95.4% of the LSCC. Patients with strong membranous (p = 0.016) or strong cytoplasmic (p = 0.030) podoplanin-positive tumors resulted in significantly better disease-free survival than those who had podoplanin weak/negative tumors, confirming podoplanin expression as a favorable independent prognostic factor. Podoplanin and CD44v6 were strongly expressed by tumor cells and podoplanin immunoexpression can help to determine lip cancer patients with lower risk for disease recurrence.

Twist and E-cadherin deregulation might predict poor prognosis in lower lip squamous cell carcinoma.

OBJECTIVE: The aim of this study was to evaluate the expression of Twist and E-cadherin in lower lip squamous cell carcinoma (LLSCC) and their association with clinicopathologic parameters. STUDY DESIGN: Fifty-nine cases of LLSCC were analyzed by applying immunohistochemistry techniques in a semiquantitative manner. The systems proposed by Bryne etal., Brandwein-Gensler etal., and Almangush etal. were applied for analysis of the histopathologic malignancy grading system. RESULTS: Higher E-cadherin expression (general and membrane) was observed in cases presenting with disease-free survival after 5years of follow-up (P < .05). Higher Twist expression was observed in lesions classified as being in advanced stages, displaying recurrence, and having a high degree of malignancy. A significant negative correlation was detected between cytoplasmic Twist expression and membrane E-cadherin expression (P = .028). A statistically significant relationship was detected between high total Twist expression in tumors classified as high risk by Brandwein-Gensler etal., and no significant difference was observed among total, membrane, and cytoplasmic E-cadherin expressions in LLSCC cases and the 3 applied grading systems (P > .05). CONCLUSIONS: The results of the present study suggest the potential involvement of Twist and E-cadherin in the modulation of events related to worse prognoses in LLSCC cases.

Oral fibropapillomatosis and epidermal hyperplasia of the lip in newborn lambs associated with bovine Deltapapillomavirus.

Congenital fibropapillomatosis of the gingiva and oral mucosa and epidermal hyperplasia of the lip are described, for the first time, in two newborn lambs. Expression of the E5 oncoprotein of bovine deltapapillomavirus types 2 (BPV-2) and -13 (BPV-13) was detected in both fibropapillomas and the hyperplastic epidermal cells suggesting the BPV infection was the cause of the proliferative lesions. No DNA sequences of BPV-1 and BPV-14 were detected. Both BPV-2 and BPV-13 DNA were also amplified from peripheral blood mononuclear cells (PBMCs) of the newborn lambs' dams. The concordance between BPV genotypes detected in the blood of dam and the oral and skin pathological samples of their offspring suggests that a vertical hematogeneous transmission was most likely source of BPV infection. Immunoblotting revealed the presence of E5 dimers allowing the viral protein to be biologically active. E5 dimers bind and activate the platelet derived growth factor ß receptor (PDGFßR), a major molecular mechanism contributing to disease. The detection of E5 protein within the proliferating cells therefore adds further evidence that the BPV infection was the cause of the proliferative lesions seen in these lambs. This is the first evidence of vertical transmission of BPVs in sheep resulting in a clinical disease.

Participation of hypoxia-inducible factor-1α and lymphangiogenesis in metastatic and non-metastatic lower lip squamous cell carcinoma.

This study evaluated the lymphatic density and HIF-1α immunoexpression in lower lip squamous cell carcinoma (LLSCC) and their correlation with clinicopathological (nodal metastasis, clinical stage, histological grade, recurrence and disease outcome) and survival parameters in 20 metastatic and 20 non-metastatic LLSCCs. Lymphatic density was established by counting microvessels (D2-40+) at the tumor core (intratumoral lymphatic density, ILD) and at the invasive front (peritumoral lymphatic density, PLD) and percentages of immunopositive cells for HIF-1α were established. No statistically significant differences in lymphatic densities in relation to clinicopathological parameters were observed (P > 0.05). All cases exhibited nuclear and cytoplasmic HIF-1α immunoexpression, with relatively high percentages of positivity, but this expression was not statistically different in relation to clinicopathological variables (P > 0.05). Positive correlations were observed between ILD and PLD (P = 0.002), and between nuclear HIF-1α immunoexpression at the tumor core and ILD (P = 0.001). The results suggest ILD and PLD are not directly related to the development of lymph node metastasis in LLSCC. The striking expression of HIF-1α suggests the involvement of this protein in the etiopathogenesis of LLSCCs, possibly stimulating lymphangiogenesis at the tumor core. However, this protein does not seem to exert a determining influence on the biological aggressiveness of these tumors.

Expression of cancer stem cell markers CD44, ALDH1 and p75NTR in actinic cheilitis and lip cancer.

PURPOSE: The aim of this work was to evaluate the expression of the cancer stem cell (CSC) markers CD44, ALDH1 and p75NTR in the ultraviolet-induced lesions actinic cheilitis (AC) and lip squamous cell carcinoma (LSCC), and to correlate it with p53 expression. METHODS: Immunohistochemistry was performed in 4 cases of normal lip (NL), 43 of AC and 20 of LSCC. RESULTS: All cases were positive for CD44, showing a membranous staining without differences between the groups. ALDH1 showed cytoplasmic staining and it was invariable amongst the grades of epithelial dysplasia and between AC and LSCC. p75NTR presented membranous/cytoplasmic staining in the basal and parabasal layer of NL and AC, while LSCC presented cytoplasmic staining in the peripheral layers of the tumor islands. p75NTR showed different expression amongst the dysplasia grades (p < 0.001) but no differences between AC and LSCC. p53 expression was similar amongst the dysplasia grades and between AC and LSCC. CD44, ALDH1 and p75NTR were unrelated amongst themselves and to p53 expression. CONCLUSIONS: CSC markers are expressed in potentially malignant and malignant lesions of the lip. Their expressions were invariable between AC and LSCC and unrelated to p53. p75NTR expression increased with the worsening of epithelial dysplasia grade.

Expression of stem cell markers Nanog and Nestin in lip squamous cell carcinoma and actinic cheilitis.

Cancer stem cell (CSC) proteins have been observed in several lesions and are associated with tumor beginning, evolution, and resistance to treatment. OBJECTIVES: To investigate the presence of NANOG, NESTIN, and ß-tubulin in lip squamous cell carcinoma (LSCC), actinic cheilitis (AC), and normal epithelium (NE). MATERIALS AND METHODS: Thirty cases of LSCC, thirty cases of AC (both analyzed according to the WHO classification and AC according to the binary classification), and twenty cases of NE were submitted to an immunohistochemical study. RESULTS: NANOG was more expressed in the nuclei of AC compared to NE (p = 0.007), as well as in high-risk AC cases (p = 0.017) and well-differentiated LSCCs (no significance). There was an accumulation of nuclear NANOG from mild to moderate and severe ACs. NESTIN was significantly less present in NE compared to AC (p = 0.001) and LSCC (p = 0.003). There was a higher expression in severe dysplasia or high-risk AC and well-differentiated LSCC. These results indicate an upregulation of NANOG and NESTIN in the early stages of carcinogenesis. ß-tubulin was intensely present in all lesions. CONCLUSION: The results suggest an upregulation of NANOG and NESTIN in the biological behavior these diseases, mainly in the transformation from AC to LSCC.

Thioredoxin and metallothionein: Homeostasis-related proteins in lip carcinogenesis.

OBJECTIVE: Thioredoxin (Trx) and metallothionein (MT) are involved in the development of some carcinomas; however, the role of these proteins in labial carcinogenesis has not yet been tested. The aims of the study were to evaluate and to correlate the immunoexpression of Trx and MT in actinic cheilitis, lip squamous cell carcinoma, and normal vermillion lip mucosa. DESIGN: Immunohistochemistry was undertaken for Trx and MT in samples of actinic cheilitis, lip squamous cell carcinoma, and normal lip mucosa. Qualitative and semi-quantitative evaluations were conducted. The proportion of stained cells, intensity of staining, and the cell compartment labeled were evaluated. A quickscore index was also calculated by multiplying the values of extension and intensity of nuclear and cytoplasmic staining, respectively, giving a maximum value of 9. Statistics were performed. RESULTS: A remarkable nuclear Trx staining was seen in normal lip mucosa and cheilitis, not in carcinoma (p<0.05). Cytoplasmic Trx expression was widely detected in all lesions (p>0.05). MT was broadly expressed in nuclei and cytoplasm of carcinoma, but not in normal lip mucosa and cheilitis (p<0.05). Quickscores were in accordance with the qualitative results. CONCLUSIONS: The current study showed a different immunopattern of Trx and MT between normal lip mucosa, actinic cheilitis and lip squamous cell carcinoma. The cellular compartment-based analyses evidenced differences that can be related to the proteins function. Considering the relevant roles of these proteins in cellular homeostasis, they seem to have an important role in lip carcinogenesis.

Altered Expression of Sphingosine-1-Phosphate Metabolizing Enzymes in Oral Cancer Correlate With Clinicopathological Attributes.

We have determined the gene expression of sphingosine-1-phosphate (S1P) metabolizing enzymes (SphK1, SphK2, SGPL1, SGPP1, SGPP2, PPAP2A, PPAP2B, and PPAP2C) by quantitative real-time polymerase chain reaction in tumor tissues and adjacent normal tissues of 50 oral squamous cell carcinoma (OSCC) patients. Expression of SphK1 and SGPP1 genes was up-regulated significantly in 70% and 75% OSCC tumors respectively. Importantly, expression of SphK2 and PPAP2B was down-regulated in the tumor tissues of 70% OSCC patients. Expression of SphK2 and PPAP2B negatively correlated with tumor-node-metastasis (TNM) staging and tumor volume respectively. Furthermore, LPP1 is an independent predictor of TNM staging and lymph node ratio.

PDGFRa amplification in multiple skin lesions of undifferentiated pleomorphic sarcoma: A clue for intimal sarcoma metastases.

A 62-year-old human immunodeficiency virus-positive man was admitted for multiple cutaneous and subcutaneous nodules on his lower limbs, corresponding to an undifferentiated proliferation of spindle and pleomorphic cells, with irregular nuclei and numerous mitoses. The tumor cells were negative for a large panel of immunohistochemical markers, except CD10. MDM2 immunohistochemical staining was also negative, leading to the diagnosis of Fédération Nationale des Centres de Lutte contre le Cancer grade III undifferentiated pleomorphic sarcoma (UPS). Array-comparative genomic hybridization showed a highly complex karyotype, with amplification of the 4q12 region, an area that contains only the platelet-derived growth factor receptor α (PDGFRa) gene. This amplification of PDFGRa, molecular hallmark of intimal sarcoma (IS), led to the diagnosis of skin IS metastasis. A positron emission tomography showed a hypermetabolic mass protruding in the preaortic area, consistent with the diagnosis of aortic IS. Our study shows that a rare differential diagnosis in peripheral UPS can be IS skin metastasis, and underlines the importance of molecular analyses in UPS.

Benign vascular lesions of the lips: Diagnostic approach.

BACKGROUND: Although not rare, vascular lesions occurring in the lips sometimes poses a difficult in properly diagnosing and handling them. In this study, vascular lesions occurring in the lips were retrieved from an Oral Pathology Service. METHODS: Among 5600 biopsies, 131 cases were found. The following diagnoses were attributed: caliber-persistent artery, infantile hemangioma, vascular malformation, venous lake, thrombus, papillary endothelial hyperplasia and pyogenic granuloma. Clinical data were obtained from patients' records. RESULTS: The lesions' frequency were: pyogenic granuloma (48%), followed by venous lake (17.5%), thrombus (14.5%), papillary endothelial hyperplasia (9.1%), infantile hemangioma (6.1%), caliber-persistent artery (3%) and vascular malformation (1.5%). Glucose transporter protein of the erythrocyte type was positive only in infantile hemangioma. The other markers (CD34 and smooth muscle action) were positive in all lesions, except for podoplanin, which was negative. CONCLUSION: It is important to be aware of the occurrence of different vascular lip lesions and their histomorphologies in order to properly handle them. Despite most lesions do not represent any risk to the patient, some of them can reach large dimensions and thus cause aesthetical trouble. Immunohistochemistry may help when the vascular character of the lesion is not promptly determined and to differentiate among some lesions.

Mismatch repair system proteins in oral benign and malignant lesions.

Different environmental agents may cause DNA mutations by disrupting its double-strand structure; however, even normal DNA polymerase function may synthesize mismatch nucleotide bases, occasionally demonstrating failure in its proofreading activity. To overcome this issue, mismatch repair (MMR) system, a group of proteins specialized in finding mispairing bases and small loops of insertion or deletion, works to avoid the occurrence of mutations that could ultimately lead to innumerous human diseases. In the last decades, the role of MMR proteins in oral carcinogenesis and in the development of other oral cavity neoplasms has grown, but their importance in the pathogenesis and their prognostic potential for patients affected by oral malignancies, especially oral squamous cell carcinoma (OSCC), remain unclear. Therefore, in this manuscript we aimed to review and critically discuss the currently available data on MMR proteins expression in oral potentially malignant lesions, in OSCC, and in other oral neoplasms to better understand their relevance in these lesions.