Long-term control of lung metastasis of tongue cancer treated effectively with cetuximab combination chemotherapy.

Generally, systemic chemotherapy is indicated for oral squamous cell carcinoma with distant metastasis and has a poor prognosis. Recently, the advent of molecular targeted drugs, such as cetuximab and immune checkpoint inhibitors, has dramatically improved prognosis, though controlling distant metastasis remains challenging. We report a case of tongue cancer in which lung metastases disappeared in the long term. A 60-year-old Japanese male with squamous cell carcinoma of the tongue underwent preoperative chemoradiotherapy and surgery including subtotal glossectomy, bilateral modified radical neck dissection, and immediate reconstruction with an anterolateral thigh flap. One month after surgery, multiple nodules less than 10 mm in diameter appeared in both lungs on CT imaging. Multiple lung metastases were diagnosed with no local recurrence or regional lymph node metastasis. The patient continues to receive a 4-week treatment course of chemotherapy that included cetuximab every 3 months and the lung metastases were markedly reduced in size or had disappeared. No local recurrence or newly emerged metastases were observed. The patient has been doing well for nine years since the appearance of the lung metastases.

Is the addition of extranodal extension and lymph node yield of pN0 to the lymph node ratio useful as a prognostic parameter for patients with oral squamous cell carcinoma?

We investigated the value of the weighted lymph node ratio (WLNR), a new marker in pN0 patients that incorporates the number of metastatic lymph nodes with extranodal extension and the lymph node yield, for the prognosis and postsurgical management of oral squamous cell carcinoma (OSCC). We designed a retrospective study and enrolled patients with OSCC who were treated by neck dissection (ND). The predictor variable was WLNR, and the outcome variable was overall survival (OS). The Cox proportional-hazards model was used to identify independent prognostic factors. In 133 patients with OSCC, the WLNR cut-off value for predicting OS was 0.0363 (area under the curve 0.723, p<0.001). When stratified according to WLNR, there was a significant difference in OS (88.4% for low WLNR and 63.0% for high WLNR, p<0.001). Univariate analyses showed close associations between OS and age, dissection area, postoperative management, extranodal extension, number of positive lymph nodes, pN stage, WLNR, and nodal disease area. Cox multivariate analysis identified the WLNR as an independent predictive factor for OS (HR 3.273, 95% CI 1.227 to 8.731, p=0.018). As a predictive factor, a high WLNR (≥0.0363) in patients with pN0 disease, which included the addition of extranodal extension and lymph node yield to the LNR, was associated with diminished survival.

Postoperative delirium after reconstructive surgery for oral tumor: a retrospective clinical study.

The aim of this study was to perform a statistical evaluation of the risk factors for postoperative delirium after oral tumor resection and reconstructive surgery. The records of 69 consecutive patients who underwent major head and neck tumor resection and reconstructive surgery, and who received postoperative management in the high care unit (HCU) or intensive care unit (ICU) of Tsukuba University Hospital between January 2013 and December 2017, were analysed retrospectively. Delirium was diagnosed in 23 patients (33.3%) after surgery. There were significant differences in age, sex, history of diabetes mellitus and chronic obstructive pulmonary disease, recent hospitalization history, sedation period, duration of ventilator use, length of ICU/HCU stay, postoperative blood tests (haemoglobin and potassium), and postoperative medication with a major tranquilizer between those with and without delirium. Logistic regression analysis of selected independent variables revealed a hazard ratio (95% confidence interval) of 1.42 (1.09-1.86) for the sedation period. Delirium was hyperactive type in 15 cases, hypoactive type in five, and mixed type in three. There was no obvious difference in postoperative day of onset or delirium period according to subtype. In conclusion, a history of diabetes and the sedation period were found to be related to postoperative delirium. However, this study was small and retrospective, so further investigation is necessary.

The effect of oral management on the severity of oral mucositis during hematopoietic SCT.

Oral mucositis (OM) is a frequent adverse effect of allogenic or autologous hematopoietic SCT. It results from direct toxic injury to the mucosal epithelial cells by the immunosuppressive regimen. Here, we compared the incidence and severity of OM between a group of 24 patients who received proper oral management during hematopoietic SCT and a group of 24 who did not. The oral management group received pre-hematopoietic SCT instruction on oral care and an oral examination in the clean room. Differences in the incidence and severity of OM between the two groups were examined statistically. OM was observed in 14 (58.3%) patients in the oral management group and 22 (91.6%) in the control group. The median of the OM score was 1 for the oral management group (range 0 to 3) and 2 for the control group (range 0 to 3). There was a significant difference in the OM score (P<0.05) and in the incidence of OM between the two groups (P<0.01). This study shows that oral management may decrease the occurrence of OM. Our results also suggest that it is important to include an oral management provider on the hematopoietic SCT team.

Targeting fibroblast growth factor receptor 3 enhances radiosensitivity in human squamous cancer cells.

Conventional therapies including radiation therapy cannot cure squamous cell carcinoma (SCC), and new treatments are clearly required. Our recent studies have shown that SCC cell lines exhibiting radioresistance show significant upregulation of the fibroblast growth factor receptor 3 (FGFR3) gene. We hypothesized that inhibiting FGFR3 would suppress tumor cell radioresistance and provide a new treatment approach for human SCCs. In the present study, we found that RNA interference-mediated FGFR3 depletion in HSC-2 cells, a radioresistant cell line, induced radiosensitivity and inhibited tumor growth. Use of an FGFR3 inhibitor (PD173074) obtained similar results with suppression of the autophosphorylation extracellular signal-regulated kinase pathway in HSC-2 cells and lung cancer cell lines. Moreover, the antitumor growth effect of the combination of PD173074 and radiation in vivo was also greater than that with either drug alone or radiation alone. Our results provided novel information on which to base further mechanistic study of radiosensitization by inhibiting FGFR3 in human SCC cells and for developing strategies to improve outcomes with concurrent radiotherapy.

Clinical observations of postoperative delirium after surgery for oral carcinoma.

The aim of the present study was to clarify the clinical characteristics of postoperative delirium and to determine appropriate postoperative management for its prevention. The authors analysed 132 cases of primary surgery for oral carcinoma and observed 24 (18%) cases of postoperative delirium. Univariate analysis revealed that significant risk factors for postoperative delirium were older age, male gender, extensive surgery and morphine pain control. Logistic regression analysis showed that older age and male gender were significant risk factors for postoperative delirium, while patient-controlled analgesia with fentanyl was effective for prevention of postoperative delirium. There was a trend for postoperative delirium to be associated with extensive surgery. In those who had delirium, blood tests revealed that alkaline phosphatase, total protein, sodium, chlorine, red blood cell count, haemoglobin and haematocrit were significantly diminished after surgery. These results indicate that general condition is closely related to the onset of postoperative delirium, and suggest that appropriate postoperative management can reduce the incidence of this complication.

Inhibition of ICAM2 induces radiosensitization in oral squamous cell carcinoma cells.

We recently identified genes and molecular pathways related to radioresistance of oral squamous cell carcinoma (OSCC) using Affymetrix GeneChip. The current study focused on the association between one of the target genes, intercellular adhesion molecule 2 (ICAM2), and resistance to X-ray irradiation in OSCC cells, and evaluated the antitumor efficacy of combining ICAM2 small interfering RNA (siRNA) and X-ray irradiation. Downregulation of ICAM2 expression by siRNA enhanced radiosensitivity of OSCC cells with the increased apoptotic phenotype via phosphorylation (ser473) of AKT and activation of caspase-3. Moreover, overexpression of ICAM2 induced greater OSCC cell resistance to the X-ray irradiation with the radioresistance phenotype. These results suggested that ICAM2 silencing is closely related to sensitivity of OSCC cells to radiotherapy, and that ICAM2 may be an effective radiotherapeutic target for this disease.

Network-based analysis of calcium-binding protein genes identifies Grp94 as a target in human oral carcinogenesis.

To characterise Ca(2+) -binding protein gene expression changes in oral squamous cell carcinomas (OSCCs), we compared the gene expression profiles in OSCC-derived cell lines with normal oral tissues. One hundred Ca(2+) -binding protein genes differentially expressed in OSCCs were identified, and genetic pathways associated with expression changes were generated. Among genes mapped to the network with the highest significance, glucose-regulated protein 94 kDa (Grp94) was evaluated further for mRNA and protein expression in the OSCC cell lines, primary OSCCs, and oral premalignant lesions (OPLs). A significant (P<0.001) overexpression of Grp94 protein was observed in all cell lines compared to normal oral epithelium. Immunohistochemical analysis showed highly expressed Grp94 in primary OSCCs and OPLs, whereas most of the corresponding normal tissues had no protein immunoreaction. Real-time quantitative reverse transcriptase-PCR data agreed with the protein expression status. Moreover, overexpression of Grp94 in primary tumours was significantly (P<0.001) correlated with poor disease-free survival. The results suggested that Grp94 may have potential clinical application as a novel diagnosis and prognostic biomarker for human OSCCs.

Monophasic epithelial synovial sarcoma arising in the temporomandibular joint.

Synovial sarcoma is a mesenchymal spindle-cell tumour that occurs infrequently in the head and neck. It originates from unknown stem cells differentiating into mesenchymal and/or epithelial structures. Most synovial sarcomas are biphasic in character, consisting of epithelial and spindle-cell elements. Here is reported a case of monophasic epithelial synovial sarcoma arising in the temporomandibular joint. The tumour was of a predominantly epithelial pattern, although a minute area of sarcomatous cells was found. The primary mode of treatment was wide en-bloc excision. Two years after surgery, the patient died of hepatocellular carcinoma, but there was no evidence of synovial sarcoma recurrence.

Ubiquitous mitochondrial creatine kinase downregulated in oral squamous cell carcinoma.

In this study, we performed two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionisation time of fly mass spectrometry to identify the protein(s) associated with the development of oral squamous cell carcinomas (OSCCs) by comparing patterns of OSCC-derived cell lines with normal oral keratinocytes (NOKs), and found that downregulation of ubiquitous mitochondrial creatine kinase (CKMT1) could be a good candidate. Decreased levels of CKMT1 mRNA and protein were detected in all OSCC-derived cell lines examined (n=9) when compared to those in primary normal oral keratinocytes. Although no sequence variation in the coding region of the CKMT1 gene with the exception of a nonsense mutation in exon 8 was identified in these cell lines, we found a frequent hypermethylation in the CpG island region. CKMT1 expression was restored by experimental demethylation. In addition, when we transfected CKMT1 into the cell lines, they showed an apoptotic phenotype but no invasiveness. In clinical samples, high frequencies of CKMT1 downregulation were detected by immunohistochemistry (19 of 52 (37%)) and quantitative real-time RT-PCR (21 of 50 (42%)). Furthermore, the CKMT1 expression status was significantly correlated with tumour differentiation (P<0.0001). These results suggest that the CKMT1 gene is frequently inactivated during oral carcinogenesis and that an epigenetic mechanism may regulate loss of expression, which may lead to block apoptosis.

Overexpression of stathmin in oral squamous-cell carcinoma: correlation with tumour progression and poor prognosis.

Stathmin is an intracellular phosphoprotein that is overexpressed in a number of human malignancies. Our previous study using proteomic profiling showed that significant upregulation of stathmin occurs in oral squamous-cell carcinoma (OSCC)-derived cell lines. In the current study, to determine the potential involvement of stathmin in OSCC, we evaluated the state of stathmin protein and mRNA expression in OSCC-derived cell lines and human primary OSCCs. A significant increase in stathmin expression was observed in all OSCC-derived cell lines examined compared to human normal oral keratinocytes. In immunohistochemistry, 65% of the OSCCs were positive for stathmin, and no immunoreaction was observed in corresponding normal tissues. Real-time quantitative reverse transcriptase-polymerase chain reaction data were consistent with the protein expression status. Moreover, stathmin expression status was correlated with the TNM stage grading. Furthermore, we found a statistical correlation between the protein expression status and disease-free survival (P=0.029). These results suggest that expression of stathmin could contribute to cancer progression/prognosis, and that stathmin may have potential as a biomarker and a therapeutic target for OSCC.

Monitoring of circulating tumour-associated DNA as a prognostic tool for oral squamous cell carcinoma.

Frequent allelic imbalances (AIs) including loss of heterozygosity and microsatellite instability on a specific chromosomal region have been identified in a variety of human malignancies. The objective of our study was to assess the possibility of prognostication and monitoring of oral squamous cell carcinoma (SCC) by microsatellite blood assay. DNA from normal and tumorous tissues and serum DNA obtained at three time points (preoperatively, postoperatively, and 4 weeks postoperatively) from 64 patients with oral SCC was examined at nine microsatellite loci. In all, 38 (59%) DNA samples from tumorous tissues and 52% from serum showed AIs in at least one locus. Patterns of AIs in the serum DNA were matched to those detected in tumour DNA. Of them, AIs were frequently detected preoperatively (44%, 28 of 64), and postoperatively (20%, 13 of 64). Moreover, among 12 cases with AIs during the postoperative period, six had no evidence of an AI 4 weeks postoperatively, and they had no recurrence and were disease free. In contrast, six patients with AI-positive DNA 4 weeks postoperatively have died with distant metastasis within 44 weeks. Thus, our results suggest that the assessment of microsatellite status in the serum DNA could be a useful predictive tool to monitor disease prognosis.

Aberrant expression of RAB1A in human tongue cancer.

This study was designed to identify specific gene expression changes in tongue squamous cell carcinomas (TSCCs) compared with normal tissues using in-house cDNA microarray that comprised of 2304 full-length cDNAs from a cDNA library prepared from normal oral tissues, primary oral cancers, and oral cancer cell lines. The genes identified by our microarray system were further analysed at the mRNA or protein expression level in a series of clinical samples by real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) analysis and imuunohositochemistry. The microarray analysis identified a total of 16 genes that were significantly upregulated in common among four TSCC specimens. Consistent with the results of the microarray, increased mRNA levels of selected genes with known molecular functions were found in the four TSCCs. Among genes identified, Rab1a, a member of the Ras oncogene family, was further analysed for its protein expression in 54 TSCCs and 13 premalignant lesions. We found a high prevalence of Rab1A-overexpression not only in TSCCs (98%) but also in premalignant lesions (93%). Thus, our results suggest that rapid characterisation of the target gene(s) for TSCCs can be accomplished using our in-house cDNA microarray analysis combined with the qRT-PCR and immunohistochemistry, and that the Rab1A is a potential biomarker of tongue carcinogenesis.

Novel OK-432-conjugated tumor vaccines induce tumor-specific immunity against murine tongue cancer.

Priming with tumor antigens is one of the most important strategies in cancer immunotherapy. To enhance tumor antigenicity, OK-432, a streptococcal preparation, was coupled to squamous cell carcinoma (KLN-205) by means of a 0.2% glutaraldehyde method. The purpose of this study was to investigate whether OK-432-conjugated tumor vaccines could induce tumor-specific immunity. Our originally developed mouse tongue cancer model was used throughout this work for the analysis of antitumor effects. Prepared OK-432-conjugated KLN-205 vaccines were immunized 3 times to DBA/2 mice. The results showed that the KLN-205 vaccines induced cytolytic activity and strongly suppressed both KLN-205 tumor incidence and growth, and survival of the mice was improved. Moreover, the histological results showed that a greater number of lymphocytes had infiltrated around tumor cells by 24 hours after tumor inoculation in the vaccine group. These results suggest that immunizations with KLN-205 vaccines increase the antitumor effects against tongue cancer.

Rapid tumor necrosis induced by electrochemotherapy with intratumoral injection of bleomycin in a hamster tongue cancer model.

This study evaluated the effects of electrochemotherapy (ECT) with intratumoral injection of bleomycin (BLM) on the chemically induced tongue cancer model in the hamster. Intratumoral injection of BLM followed by high-voltage electrical treatment induced rapid necrosis of the tumor within 48 hours and subsequent rapid tumor volume reduction. Three weeks after the ECT with BLM, 3 of the 6 animals showed no palpable tumor, while no antitumor effects were observed in the control groups. Because of the remarkable antitumor effect with no major observed side effects, we concluded that ECT combined with intratumoral injection of BLM has the potential to enhance treatment results for tongue cancer.

Rectal and vaginal immunization with a macromolecular multicomponent peptide vaccine candidate for HIV-1 infection induces HIV-specific protective immune responses.

An effective vaccine for human immunodeficiency virus (HIV) is needed to stimulate the immune response of the genital mucus to prevent mucosal transmission of the virus. We have developed a macromolecular multicomponent peptide vaccine candidate, VC1. Both rectal and vaginal immunization of VC1 mixed with cholera toxin (CT) induced HIV-1-specific IgA antibody in mouse fecal extract solution and vaginal wash. These antibody productions were enhanced by the combination with IL-4 or GM-CSF expressing plasmids. Either fecal extract or vaginal wash solution from immunized mice inhibited production of HIV-1IIIB p24 protein. The mononuclear cells from spleen, intestinal lymph nodes, or Peyer's patches from VC1- and CT-immunized mice released IFN-gamma or IL-4, when these cells were co-cultured with VC1 antigen. In addition, the regional lymphoid cells from rectal and vaginal region of mice immunized with VC1 and CT also elicited a substantial level of HIV-1-specific cytotoxic T cell (CTL) response. This CTL response was enhanced by the addition of IL-12 expressing plasmid. Our results clearly demonstrated that both rectal and vaginal immunization could induce systemic and mucosal immunities specific for HIV-1.

In vivo antitumor effects of electrochemotherapy in a tongue cancer model.

PURPOSE: This study investigated the in vivo antitumor effects of electrochemotherapy (ECT) using electroporation and bleomycin in a hamster tongue cancer model to assess its clinical applicability. MATERIALS AND METHODS: Twenty animals with chemically induced tongue cancer were divided into four experimental groups designated B-E-, B-E+, B+E-, and B+E+. The B+E+ and B+E- groups received an intraperitoneal injection of 100 microg bleomycin. Fifteen minutes after the injection, the B+E+ animals received electric pulses. The B-E+ group received only electric pulses. The B-E- group received neither bleomycin nor electric pulses. Each group received the same treatment twice. The antitumor effects were assessed based on tumor volume reduction and histologic findings. RESULTS: The B+E+ group showed remarkable tumor volume reduction, decreasing an average to 8.8% of its original volume 14 days after the treatment. Complete loss of the protruding tumor was observed in two of the five animals. Histologically, the tumors of the B+E+ group consisted of severely degenerated tumor cells and desquamative keratinizing cells. No living cancer cells were detected in three animals. The B+E-, B-E+, and B-E- groups showed progressive tumor growth, exceeding 200% of initial tumor volume during the experimental period. CONCLUSION: The current study showed remarkable antitumor effects of ECT with bleomycin in the hamster tongue cancer model. ECT with bleomycin may be clinically applicable to the treatment of oral cancer.

Comparison between hockey stick and reversed hockey stick incision: gently curved single linear neck incisions for oral cancer.

Hockey stick incision (HSI) and reversed-HSI are known to be useful incisions for lymph node dissections of the neck. Both are gently curved single linear incisions without three-point suture line junctions, but are different at the base of the skin flap. The HSI allows the elevation of a superiorly-based single cervical skin flap and the reversed-HSI allows for an inferiorly-based flap. We compared the viability of the skin flaps, exposure of the operation field and cosmetic results to evaluate the characteristics of each incision. HSI appeared to be the suitable incision for radical neck dissection due to adequate exposure of the operation field while rendering excellent cosmetic results. Reversed-HSI was applied in combination with block resection of parts of the oral cavity because it provided much better exposure of the operation field than HSI, while still achieving acceptable cosmetic results. Using this technique, a small area of marginal necrosis was occasionally seen at the apex of the skin flap due to poor blood supply.

DNA vaccination followed by macromolecular multicomponent peptide vaccination against HIV-1 induces strong antigen-specific immunity.

The induction of a strong and long-lasting immunity characterized by both a humoral and cell-mediated immune (CMI) response is one of the most important considerations in developing an effective HIV vaccine. In previous studies, we have independently developed both DNA vaccine and macromolecular multicomponent peptide vaccine (VC1) candidates. In the present study, we attempted to optimize the vaccination protocol using mice, guinea pigs, rabbits and Macaca fuscata monkeys. Repeated vaccination with VC1 induced a substantial level of multivalent antibodies which neutralized various HIV-1 strains, as determined using a p24 inhibition assay. On the other hand, repeated immunization with DNA vaccine induced and sustained high levels of cytotoxic T lymphocytes (CTLs). In addition, when DNA vaccination was followed by multicomponent peptide vaccination, levels of both humoral immunity and CMI increased, and this effect continued for at least 10 months. These data clearly demonstrate that for inducing HIV-1 specific immunity, immunization with DNA vaccine followed by VC1 boosting produces better results than immunizing with either vaccine alone.