Caries experience and oral health-related quality of life (OHRQoL) of children and adolescents with cerebral palsy in a low-resource setting.

BACKGROUND: Children with complex neurodevelopmental disabilities such as cerebral palsy (CP), have a higher risk of dental disease related at least in part to greater difficulties in performing and maintaining effective oral hygiene and oral care practices. However, to date, there are very few studies that have considered the impact of dental disease on the Oral Health-Related Quality of Life (OHRQoL) of children and adolescents with cerebral palsy. This study aimed to investigate the association between dental caries experience and oral health related quality of life (OHRQoL) among children and adolescents with cerebral palsy in a low-resource setting (Bangladesh). METHODS: A total of 90 children and adolescents with CP, 2-17 years old (median age 10 years; 37.8% female and 62.2% male) were randomly selected from the Bangladesh Cerebral Palsy Register (BCPR) The decayed, missing and filled teeth (dmft/DMFT) index was used to measure caries experience. Child Perceptions Questionnaire (CPQ) and Family Impact Scale (FIS) were used to assess oral health-related quality of life (OHRQoL). Binary logistic regression was used to investigate factors that may contribute to dental caries experience. RESULTS: Dental caries were observed among 55.6% of the participants. After adjusting for age and gender, binary logistic regression analysis showed that dental caries experience was significantly associated with those who had teeth/mouth pain (rate ratio 7.3; P = 0.02), food caught between teeth (rate ratio: 6.4; P = 0.02), difficulty in eating and drinking (rate ratio 5.9; p = 0.02) and those who felt frequently upset (rate ratio: 54.7; P = 0.02). CONCLUSION: In this study, we found that children and adolescents with CP in a low-resource setting had high dental caries experience and that dental caries had a negative impact on OHRQoL amongst these participants and their parents/caregivers. Health care professionals should be aware of the importance of dental health and oral hygiene in this population. These findings highlight the need for oral health promotion programs for children and adolescents with CP in these settings to reduce pain and to improve quality of life.

Ammonium tetrathiomolybdate enhances the antitumor effects of cetuximab via the suppression of osteoclastogenesis in head and neck squamous carcinoma.

Head and neck squamous cell carcinoma (HNSCC) poses a significant challenge clinically where one of the mechanisms responsible for the invasion into facial bones occurs via the activation of osteoclasts. Copper has been demonstrated to play a key role in skeletal remodeling. However, the role of copper in cancer-associated bone destruction is thus far unknown. Lysyl oxidase (LOX) is a copper-dependent enzyme that promotes osteoclastogenesis. In the present study, we investigated the effects of copper on HNSCC with bone invasion by the copper chelator, ammonium tetrathiomolybdate (TM) in vitro and in vivo. We demonstrate that TM blocks the proliferation of HNSCC cells, inhibits LOX activation and decreases the expression of the receptor activator of nuclear factor-κB ligand (RANKL) in osteoblasts and osteocytes, subsequently suppressing bone destruction. These findings suggest that copper is a potential target for the treatment of HNSCCs associated with bone destruction.

DKK3 Overexpression Increases the Malignant Properties of Head and Neck Squamous Cell Carcinoma Cells.

DKK3, a member of the dickkopf Wnt signaling pathway inhibitor family, is believed to be a tumor suppressor because of its reduced expression in cancer cells. However, our previous studies have revealed that DKK3 expression is predominantly observed in head and neck/oral squamous cell carcinoma (HNSCC/OSCC). Interestingly, HNSCC/OSCC patients with DKK3 expression showed a high rate of metastasis and poorer survival, and siRNA-mediated knockdown of DKK3 in HNSCC-derived cancer cell lines resulted in reduced cellular migration and invasion. From these data, it was hypothesized that DKK3 might exert an oncogenic function specific to HNSCC. In the present research, the DKK3 overexpression model was established, and its influences were investigated, together with molecular mechanism studies. The DKK3 expression profile in cancer cell lines was investigated, including HNSCC/OSCC, esophageal, gastric, colorectal, pancreatic, prostatic, and lung cancers. DKK3 overexpression was performed in HNSCC-derived cells by transfection of expression plasmid. The effects of DKK3 overexpression were assessed on cellular proliferation, migration, invasion, and in vivo tumor growth. The molecular mechanism of DKK3 overexpression was investigated by Western blotting and microarray analysis. DKK3 overexpression significantly elevated cellular proliferation, migration, and invasion, as well as increased mRNA expression of cyclin D1 and c-myc. However, reporter assays did not show TCF/LEF activation, suggesting that the increased malignant property of cancer cells was not driven by the Wnt/ß-catenin pathway. For the investigation of the pathways/molecules in DKK3-mediated signals, the Western blot analyses revealed that phosphorylation of Akt (S473) and c-Jun (Ser63) was elevated. The application of a PI3K kinase inhibitor, LY294002, on HSC-3 DKK3 cells significantly decreased tumor cell proliferation, migration, and invasion. From these results, we demonstrated that DKK3 might contribute to cellular proliferation, invasion, migration, and tumor cell survival in HNSCC cells through a mechanism other than the canonical Wnt signaling pathway, which might be attributed to PI3K-Akt signaling.

Semaphorin 4D promotes bone invasion in head and neck squamous cell carcinoma.

Head and neck squamous cell carcinomas (HNSCCs) frequently invade the bones of the facial skeleton. Semaphorin 4D (Sema4D) is an axon guidance molecule produced by oligodendrocytes. Sema4D was also identified in the bone microenvironment and many cancer tissues including HNSCC. To date, however, the role of Sema4D in cancer-associated bone disease is still unknown. This is the first study to demonstrate the role of Sema4D in bone invasion of cancer. In the clinical tissue samples of bone lesion of HNSCC, Sema4D was detected at high levels, and its expression was correlated with insulin-like growth factor-I (IGF-I) expression. In vitro experiments showed that IGF-I regulates Sema4D expression and Sema4D increased proliferation, migration and invasion in HNSCC cells. Sema4D also regulated the expression of receptor activator of nuclear factor κß ligand (RANKL) in osteoblasts, and this stimulated osteoclastgenesis. Furthermore, knockdown of Sema4D in HNSCC cells inhibited tumor growth and decreased the number of osteoclasts in a mouse xenograft model. Taken together, IGF-I-driven production of Sema4D in HNSCCs promotes osteoclastogenesis and bone invasion.

Risk factors for dental caries among children with cerebral palsy in a low-resource setting.

AIM: To describe the oral health status and investigate factors affecting dental caries experience among children with cerebral palsy (CP) in rural Bangladesh. METHOD: A cross-sectional study was conducted among children with CP who are part of the Bangladesh Cerebral Palsy Register (BCPR) study. Caries experience was measured by identifying decayed, missing, and filled teeth for deciduous and permanent teeth (dmft/DMFT). Clinical periodontal index, body mass index, oral hygiene behaviour, masticatory ability, and dietary habits were recorded. CP motor types and severity of functional mobility (Gross Motor Function Classification System [GMFCS]) were assessed. RESULTS: Of 90 children with CP (mean age 9y 7mo, range 2-17y, 37.8% female and 62.2% male), 35% of 2 to 6 year olds, and 70% of 7 to 11 year olds (p=0.014) experienced caries (dmft+DMFT>0). The mean values (standard deviation [SD]) of dmft and DMFT were 2.46 (3.75) and 0.72 (1.79) respectively. After adjusting for age and sex, binary logistic regression analysis showed a significant relationship with dental caries for children who had quadriplegia (odds ratio [OR] 5.56, p=0.035), tooth cleaning less than one time/day (OR 0.08, p=0.016), using toothpowder or charcoal for cleaning (OR 7.63, p=0.015), and snacking between meals more than one time/day (OR 6.93, p=0.012). INTERPRETATION: Early oral health preventive care is required for children with CP because dental caries is highly prevalent in these children.

Synthetic Terrein Inhibits Progression of Head and Neck Cancer by Suppressing Angiogenin Production.

BACKGROUND/AIM: Head and neck cancers are the fifth most common cancer type worldwide, affecting more than half a million patients annually. Development of effective therapeutic drugs is, therefore, required for this type of disease. This study assessed the effects of synthetic terrein on head and neck cancer. MATERIALS AND METHODS: Synthetic terrein was prepared by using the modified Altenhach's procedure. The effect of synthetic terrein on cell proliferation of head and neck cancer cells and HUVECs was assessed. Angiogenin secretion and ribosome biogenesis were examined by ELISA and silver staining of the nucleolar organizer region. A mouse xenograft model was prepared by inoculating mice with suspensions of cells of the human head and neck cancer cell line OSC-19 subcutaneously into the dorsal region of each mouse. Ki-67, CD31 and angiogenin expression in xenografted tumors was examined by immunohistochemistry. RESULTS: Synthetic terrein inhibited the growth of various head and neck cancer cells. In addition, an in vivo experiment revealed that synthetic terrein inhibited a xenograft tumor growth in athymic mice. Immunohistochemical analysis revealed that expression of Ki-67, CD31 and ANG was down-regulated in synthetic terrein-treated tumors, compared to controls. Synthetic terrein suppressed the ANG secretion and ribosome biogenesis in cancer cells, and cell proliferation in vascular endothelial cells. CONCLUSION: The mechanism underlying the anti-tumor effects of synthetic terrein against head and neck cancer consists of the inhibition of both tumor cell proliferation and angiogenesis via the suppression of ANG production.

Brm inhibits the proliferative response of keratinocytes and corneal epithelial cells to ultraviolet radiation-induced damage.

Ultraviolet radiation (UV) from sunlight is the primary cause of skin and ocular neoplasia. Brahma (BRM) is part of the SWI/SNF chromatin remodeling complex. It provides energy for rearrangement of chromatin structure. Previously we have found that human skin tumours have a hotspot mutation in BRM and that protein levels are substantially reduced. Brm-/- mice have enhanced susceptibility to photocarcinogenesis. In these experiments, Brm-/- mice, with both or a single Trp53 allele were exposed to UV for 2 or 25 weeks. In wild type mice the central cornea and stroma became atrophic with increasing time of exposure while the peripheral regions became hyperplastic, presumably as a reparative process. Brm-/-, Trp53+/-, and particularly the Brm-/- Trp53+/- mice had an exaggerated hyperplastic regeneration response in the corneal epithelium and stroma so that the central epithelial atrophy or stromal loss was reduced. UV induced hyperplasia of the epidermis and corneal epithelium, with an increase in the number of dividing cells as determined by Ki-67 expression. This response was considerably greater in both the Brm-/- Trp53+/+ and Brm-/- Trp53+/- mice indicating that Brm protects from UV-induced enhancement of cell division, even with loss of one Trp53 allele. Cell division was disorganized in Brm-/- mice. Rather than being restricted to the basement membrane region, dividing cells were also present in the suprabasal regions of both tissues. Brm appears to be a tumour suppressor gene that protects from skin and ocular photocarcinogenesis. These studies indicate that Brm protects from UV-induced hyperplastic growth in both cutaneous and corneal keratinocytes, which may contribute to the ability of Brm to protect from photocarcinogenesis.

Self-reported aural symptoms, headache and temporomandibular disorders in Japanese young adults.

BACKGROUND: To investigate the associations of aural symptoms, headache and depression with the presence of temporomandibular disorder (TMD) symptoms in a young adult population in Japan. METHODS: A personal interview survey was conducted on first-year university students (n = 1,930) regarding symptoms of TMD, aural problems, headache, shoulder pain and depression. Logistic regression was applied to assess the associations of these problems with the presence of TMD symptoms after controlling for age and gender. RESULTS: Among the 1,930 students, 543 students exhibited TMD symptoms and were classified into 7 groups: clicking only (Group I, n = 319), pain in the TMJ only (Group II, n = 21), difficulty in mouth opening only (Group III, n = 18), clicking and pain (Group IV, n = 29), clicking and difficulty in mouth opening (Group V, n = 48), difficulty in mouth opening and pain (Group VI, n = 11), and combination of three symptoms (Group VII, n = 97). The control group (n = 1,387) were subjects without any TMD symptoms. After adjusting for age and gender, a strong association was observed between TMD symptoms (Group II and IV) and tinnitus (OR = 12.1 and 13.2, respectively). TMD symptoms (Group I, II and III) were also associated with vertigo and headache. Otalgia and depression were significantly associated with the presence of clicking only. CONCLUSIONS: TMD symptoms were significantly correlated to aural symptoms and headache. A functional evaluation of the stomatognathic system should be considered in subjects with unexplained aural symptoms and headache.

Anti-tumor effect of a novel FAK inhibitor TAE226 against human oral squamous cell carcinoma.

OBJECTIVES: Focal adhesion kinase (FAK) overexpression is frequently found in invasive and metastatic cancers, but its role in oral squamous cell carcinoma is not yet well understood. In order to seek therapies targeting oral squamous cell carcinoma, we developed the novel FAK Tyr(397) inhibitor TAE226 and investigated its anti-tumor effects and mechanisms. MATERIALS AND METHODS: Expression of phosphorylated FAK Tyr(397) was examined by immunohistochemical and immunoblot analysis. The effect of TAE226 on in vitro and in vivo studies were confirmed by proliferation, cell cycle, apoptosis and angiogenesis analysis. RESULTS: We found that phosphorylated FAK was highly expressed in human tongue oral squamous cell carcinoma in patients. Importantly, TAE226 greatly suppressed the proliferation, migration and invasion of human oral squamous cell carcinoma SAS cells with an apparent structural change of actin fiber and a loss of cell adhesion. In addition, TAE226 inhibited the expression of phospho-FAK Tyr(397) and phospho AKT Ser(473), resulting in caspase-mediated apoptosis. Furthermore, oral administration of TAE226 in mice suppressed the growth and angiogenesis of oral squamous cell carcinoma xenografts in vivo. CONCLUSIONS: Our results provide compelling evidence that FAK is critically involved in oral squamous cell carcinoma and that the FAK inhibitor TAE226 can potentially be effectively used for the treatment of oral squamous cell carcinoma.

Sonic hedgehog signaling promotes growth of oral squamous cell carcinoma cells associated with bone destruction.

Sonic hedgehog (Shh) and its signaling have been identified in several human cancers, and increased levels of its expression appear to correlate with disease progression and metastasis. However, the role of Shh in bone destruction associated with oral squamous cell carcinomas, which frequently invade the maxilla or the mandible, is still unclear. In this study we show that the use of siRNA for Shh to block SHH secreted by SAS oral squamous cell carcinoma cells suppressed the tumor growth and tumor angiogenesis of subcutaneous SAS xenografts in vivo. Moreover, blockade of Shh in SAS cells decreased tumor growth and osteoclast number in a tibial metaphysis mouse model. Significantly, we clearly show that SHH stimulated osteoclast formation in a co-culture system consisting of murine bone stromal ST2 cells and murine CD11b(+) bone marrow cells. These findings suggest that Shh signaling is a potential target for the treatment of oral squamous cell carcinoma associated with bone destruction.

Increased expression of the PRL-3 gene in human oral squamous cell carcinoma and dysplasia tissues.

Phosphatase of regenerating liver (PRL) belongs to a class of the protein tyrosine phosphatase family, which is known so far to consist of 3 members, PRL-1, PRL-2, and PRL-3. The aim of this study was to uncover the role of PRL genes in development of oral malignancy. We analyzed expression levels of the 3 PRL genes in 50 human oral squamous cell carcinomas (OSCCs), 11 dysplasia and 12 normal mucosa tissues by a real-time RT-PCR method. PRL-3 but not PRL-1 or PRL-2 expressions were significantly higher in OSCC and dysplasia than in normal mucosa tissues. Additionally, PRL-3 expressions were significantly higher in OSCC tissues harboring dominant-negative p53 or recessive p53 mutation than in those harboring wild-type p53. These results suggest that PRL-3 plays a role in oral cancer development and can be useful as a marker of pre-malignant and malignant lesion of oral mucosa.

Antitumor effect of novel HSP90 inhibitor NVP-AUY922 against oral squamous cell carcinoma.

Heat-shock protein 90 (HSP90) is a major cellular chaperone protein. HSP90 supports the correct conformation, stabilization, activation, and localization of 'client' oncoproteins, many of which are involved in tumor progression. Therefore, the use of HSP90 inhibitors has become a new strategy in antitumor therapy. However, the effects of an HSP90 inhibitor on oral squamous cell carcinoma are still unclear. NVP-AUY922 (Novartis) is a novel 4,5-diaryloxazole adenosine triphosphate-binding site HSP90 inhibitor. In this study, we investigated the antitumor effect of novel HSP90 inhibitor NVP-AUY922 against oral squamous cell carcinoma. NVP-AUY922 inhibited the proliferation of oral squamous cell carcinoma cells in vitro. NVP-AUY922 caused degradation of client protein inducing ErbB2, p-Akt, p-S6, hypoxia-inducible factor 1-α (HIF1-α) and vascular endothelial growth factor (VEGF) and up-regulation of HSP70 in HSC-2 oral squamous cell carcinoma. NVP-AUY922 increased the expression of cleaved caspase-3 and induced apoptosis in HSC-2 cells. Treatment of NVP-AUY922 induced a robust antitumor response and suppressed p-Akt and VEGF expression in an HSC-2 xenograft model. In summary, NVP-AUY922 exhibits in vitro and in vivo efficiency against oral squamous cell carcinoma, representing a promising therapeutic approach for oral squamous cell carcinoma.

Induction of MMP-13 expression in bone-metastasizing cancer cells by type I collagen through integrin α1ß1 and α2ß1-p38 MAPK signaling.

BACKGROUND: Breast cancer cells frequently metastasize to the skeleton and produce and secrete proteinases, such as matrix metalloproteinase-13 (MMP-13), which promote destruction of the bone matrix. However, the mechanism of MMP-13 expression induced in areas of bone metastasis is unknown. Here, the interaction between tumors and type I collagen in bone metastasis was investigated. MATERIALS AND METHODS: A mouse model of bone metastasis was prepared by inoculating mice with suspensions of cells of the human metastatic breast cancer cell line MDA-MB-231 via the left cardiac ventricle. MMP-13 expression was examined by immunohistochemical, Western blot, and real-time RT-PCR analyses. RESULTS: MMP-13 expression was highly up-regulated in MDA-MB-231 cells, and attachment of these cells to type I collagen and the induction of MMP-13 were down-regulated by treatment with integrin α1, α2 or ß1 neutralizing antibodies. The attachment of MDA-MB-231 cells to type I collagen induced the activation of focal adhesion kinase (FAK) and p38 mitogen-activated protein kinase (MAPK). Inhibition of FAK and p38 MAPK down-regulated type I collagen-induced MMP-13 expression. CONCLUSION: Our study indicates that metastatic breast cancer cells in the bone microenvironment attached to type I collagen, which stimulated integrins α1ß1 and α2ß1, via FAK and p38 MAPK pathways, to induce MMP13 expression and further osteolysis.

Anti-tumor effect in human breast cancer by TAE226, a dual inhibitor for FAK and IGF-IR in vitro and in vivo.

Focal adhesion kinase (FAK) is a 125-kDa non-receptor type tyrosine kinase that localizes to focal adhesions. FAK overexpression is frequently found in invasive and metastatic cancers of the breast, colon, thyroid, and prostate, but its role in osteolytic metastasis is not well understood. In this study, we have analyzed anti-tumor effects of the novel FAK Tyr(397) inhibitor TAE226 against bone metastasis in breast cancer by using TAE226. Oral administration of TAE226 in mice significantly decreased bone metastasis and osteoclasts involved which were induced by MDA-MB-231 breast cancer cells and increased the survival rate of the mouse models of bone metastasis. TAE226 also suppressed the growth of subcutaneous tumors in vivo and the proliferation and migration of MDA-MB-231 cells in vitro. Significantly, TAE226 inhibited the osteoclast formation in murine pre-osteoclastic RAW264.7 cells, and actin ring and pit formation in mature osteoclasts. Moreover, TAE226 inhibited the receptor activator for nuclear factor κ B Ligand (RANKL) gene expression induced by parathyroid hormone-related protein (PTHrP) in bone stromal ST2 cells and blood free calcium concentration induced by PTHrP administration in vivo. These findings suggest that FAK was critically involved in osteolytic metastasis and activated in tumors, pre-osteoclasts, mature osteoclasts, and bone stromal cells and TAE226 can be effectively used for the treatment of cancer induced bone metastasis and other bone diseases.

A multiple primary carcinoma consisting of leukoplakia and SCC: a case report with p53 mutation analysis.

Patients with an oral squamous cell carcinoma (OSCC) often develop multiple malignant lesions. This report examined whether individual tumours developed in a patient show the same genetic alteration, such as p53 mutations. This case study describes three SCCs and three leukoplakias which developed simultaneously in a single 67-year-old Japanese man. A p53 mutation was detected in two of the three SCCs and one of the three leukoplakias. One SCC had a missense mutation at codon 285 (GAG>AAG, Glu>Lys) and the other a nonsense mutation at codon 336, and the leukoplakia had a missense mutation at codon 273 (CGT>CAT, Arg>His). This case showed that individual oral tumours may have different genetic changes even when they develop in a single patient. Therefore, this report provided strong evidence that in cases of multiple tumours it is necessary to design tailor-made therapies for each individual tumour rather than a single standardised therapy for all multiple tumours.

Parathyroid hormone-related peptide regulates matrix metalloproteinase-13 gene expression in bone metastatic breast cancer cells.

BACKGROUND: Breast cancer (BC) cells often metastasize to bone where they express large amounts of parathyroid hormone-related protein (PTHrP). In this study, we investigated the possibility that PTHrP may have roles in breast cancer bone metastasis independently of, or in addition to, its roles in osteoclastic function. MATERIALS AND METHODS: A mouse model of bone metastasis was prepared by inoculating mice with suspensions of the human BC cell line MDA-MB-231 tumor cells via the left cardiac ventricle. Matrix metalloproteinase-13 (MMP-13) expression in the bone microenvironment was examined by Western blot and Real-time RT-PCR (RT-PCR) analysis, as well as by confocal microscopy. RESULTS: The invading MDA-MB-231 cells contained conspicuous amounts of both PTHrP and MMP-13, an important matrix-degrading enzyme; and treatment of the cells in culture with exogenous PTHrP markedly stimulated MMP13 gene expression. Analysis of signaling mechanisms showed that PTHrP treatment led to rapid increases in the levels of phosphorylated protein kinase C (PKCα) and extracellular signal-regulated kinase (ERK1/2). Pharmacologic inhibition of ERK1/2 and PKC as well as of PKA activities counteracted the PTHrP-dependent stimulation of MMP13 expression. Indeed, pharmacologic activation of PKA or PKC was sufficient for stimulation of MMP13 expression. CONCLUSION: Consistent with these findings, the inhibition of PKC prevented PTHrP-induced activation of ERK1/2, whereas 12-O-tetradecanoylphorbol-13-acetate (TPA), a stimulator of PKC, up-regulated the PTHrP-induced activation of ERK1/2. Taken together, our data indicate that the MDA-MB-231 breast cancer cells may carry out bone destruction and favor their own metastatic behavior by producing MMP-13. Given that the cells expressed PTHrP and that this factor stimulated MMP-13 expression, metastatic bone destruction may result from a PTHrP autocrine loop involving a PKC-ERK1/2 signaling pathway.

Mutant p53 R248Q but not R248W enhances in vitro invasiveness of human lung cancer NCI-H1299 cells.

More than half of all human cancers are associated with mutations of the TP53 gene. In regard to the functional interaction with the remaining wild-type (WT) p53 allele, p53 mutations are classified into two types, recessive and dominant-negative (DN) mutations. The latter mutant protein has a DN activity over the remaining WT allele. We previously showed that the DN p53 mutant was useful as a predictor of poor outcome or a risk factor for metastatic recurrence in patients with some types of cancers, regardless of the presence or absence of loss of heterozygosity (LOH) of WT p53, suggesting that the DN p53 had 'gain-of-function (GOF)' activity besides the transdominance function. In this study, we investigated GOF activity of two DN p53 mutants which had a point mutation at codon 248 (R248Q and R248W), one of the hot spots, by transfecting them respectively into H1299 cells which originally expressed no p53 protein. Growth activity of the transfectants with the two mutants was not different from that of parent or Mock transfectants. Meanwhile, in vitro invasions of Matrigel and type I collagen gel by R248Q-transfectants were significantly higher than those by R248W-transfectants or the control cells. However, there were no differences in cell motile activities, expressions of extracellular matrix-degradative enzymes such as matrix metalloproteinases, urokinase-type plasminogen activator and heparanase, and their inhibitors, between R248Q- and R248W-transfectants. These findings indicate that the p53 mutants have a different quality in GOF activities even if the mutations occurred at the same codon. And detailed information of the status of p53, including transdominancy and GOF activity, is expected to be useful for diagnosis and therapeutic strategy fitting the individual patients.

Antitumor effect of temsirolimus against oral squamous cell carcinoma associated with bone destruction.

The mammalian target of rapamycin (mTOR) is engaged in the molecular pathogenesis of oral squamous cell carcinoma, which frequently invades the maxilla or the mandible. However, the effects of a mTOR inhibitor on bone destruction associated with oral squamous cell carcinoma are still unclear. In this study, we investigated the antitumor effect of temsirolimus-mediated mTOR inhibition against advanced oral squamous cell carcinoma. Temsirolimus inhibited the proliferation and migration of HSC-2 oral squamous cell carcinoma cells in vitro and suppressed the growth of oral squamous cell carcinoma xenografts in vivo. Significantly, we clearly show that temsirolimus inhibited osteoclast formation both in vitro and in vivo. Reverse transcriptase-PCR analysis showed that temsirolimus decreased the mRNA expression of receptor activator for nuclear factor-κB ligand, known as an osteoclast differentiation factor in bone stromal ST2 cells. Moreover, temsirolimus normalized blood-free calcium concentration in mouse models for humoral hypercalcemia. These findings suggest that mTOR signaling is a potential target of oral squamous cell carcinoma associated with bone destruction, and hence we describe the efficacy of temsirolimus for the treatment of advanced oral squamous carcinoma.

Bone destruction by invading oral squamous carcinoma cells mediated by the transforming growth factor-beta signalling pathway.

BACKGROUND: Gingival squamous cell carcinoma (SCC) cells frequently invade mandibular bone, and this destruction is associated with a worse prognosis. However, the relationship between bone destruction and associated factors is unclear. In this study, the role and diagnostic utility of transforming growth factor-beta (TGF-beta) type I receptor (TbetaRI) in bone destruction of the mandible was investigated. PATIENTS AND METHODS: The expression of TbetaRI was explored by using an immunohistochemical method on paraffin-embedded tissues from 21 cases of mandibular SCC. An inhibitor of the kinase activity of the TbetaRI (TbetaRI-I) was used to assess the role of TbetaRI in bone destruction by a human oral SCC cell line (HSC-2) that highly expresses TbetaRI. RESULTS: TbetaRI-positive signals were closely associated with destructive invasion of the mandible by oral SCC cells. Consistent with these results, TbetaRI-I greatly reduced HSC-2 cell-induced bone destruction and osteoclast formation in vivo and in vitro. TbetaRI-I treatment reduced the expression of TNF-alpha, RANKL and connective tissue growth factor (CTGF/CCN2), all of which were up-regulated by TGF-beta in HSC-2 cells. CONCLUSION: These data demonstrated an important role for TGF-beta signalling in bone invasion by oral SCC cells, and suggest that the bone destruction is mediated by RANKL, TNF-alpha and CCN2.

PTHrP regulates angiogenesis and bone resorption via VEGF expression.

BACKGROUND: Parathyroid hormone-related protein (PTHrP) is a key regulator of osteolytic metastasis of breast cancer (BC) cells, but its targets and mechanisms of action are not fully understood. This study investigated whether/how PTHrP (1-34) signaling regulates expression of vascular endothelial growth factor (VEGF) produced by BC cells. MATERIALS AND METHODS: A mouse model of bone metastasis was prepared by inoculating mice with tumour cell suspensions of the human BC cell line MDA-MB-231 via the left cardiac ventricle. VEGF expression was examined by Western blot and real-time RT-PCR analysis, as well as by confocal microscopy in the bone microenvironment. RESULTS: PTHrP was expressed in cancer cells producing PTH/PTHrP receptor and VEGF that had invaded the bone marrow, and PTHrP was up-regulated VEGF in MDA-MB-231 in vitro. The culture medium conditioned by PTHrP-treated MDA-MB-231 cells stimulated angiogenesis and osteoclastogenesis compared with control medium, giving a response that was inhibited by VEGF-neutralizing antibody treatment. Inhibition of protein kinase C (PKC) prevented PTHrP-induced extracellular signal-regulated kinase (ERK1/2) and p38 activation, and PTHrP-induced VEGF expression. CONCLUSION: PTHrP plays an important role in modulating the angiogenic and bone osteolytic actions of VEGF through PKC-dependent activation of an ERK1/2 and p38 signaling pathway during bone metastasis by breast cancer cells.