Quantification of Phytophthora infestans population densities and their changes in potato field soil using real-time PCR.

Tuber infection of Phytophthora infestans often occurs at harvest. However, it is difficult to accurately estimate the population densities of P. infestans in soil, especially Japanese soil. In the present study, P. infestans DNA was extracted from soil samples using a modified CTAB-bead method and quantified using real-time PCR to accurately, rapidly and easily estimate the P. infestans population densities in upland soils in Japan. P. infestans was well quantified in eleven types of soil samples, including nine types of upland soils in Japan, that were artificially inoculated with a zoosporangia suspension. The amounts of P. infestans DNA estimated by the real-time PCR were proportional to the inoculum densities. In the non-controlled experimental potato field, P. infestans population densities in soil corresponded to the development of symptoms and were correlated with the number of lesions on the potato foliage. These results imply that the proposed real-time PCR assay is suitable for the estimation or monitoring of P. infestans population densities in upland soils in Japan. The population densities at the ridge bottoms were larger than those at any other location in commercial potato fields. These results were similar to those of a previous report using a bioassay. Moreover, a correlation between DNA quantity and inoculum potential was observed. In conclusion, the real-time PCR assay developed in this study is suitable for indirect estimation of the inoculum potential of P. infestans.

Ability of plant pathogenic fungi Gibberella fujikuroi and Fusarium commune to react with airborne methyl jasmonate.

The pathogenic fungi Gibberella fujikuroi and Fusarium commune produce jasmonic acid. The application of volatile deuterium-labeled methyl jasmonate increased the amount of nonlabeled JA present in G. fujikuroi and F. commune. These results indicate that the fungi have the ability to react with airborne methyl jasmonate in a manner similar to a plant.

Tonsillectomy plus steroid pulse therapy is the most effective treatment in adult patients with C-Grade I IgA nephropathy, and the weight of the extracted palatine tonsils and Yamamoto scale have no significant correlation with the effects of this treatment.

OBJECTIVE: We studied patients who underwent tonsillectomy plus steroid pulse therapy (TSP) for immunoglobulin A nephropathy (IgAN), in order to investigate the clinical factors associated with a positive response to this treatment. METHODS: We analyzed 118 IgAN patients who underwent TSP. We collected patients' data retrospectively, including age, sex, blood pressure, onset of IgAN, pathological findings of a renal biopsy, serum concentration of creatinine, estimated glomerular filtration rate, serum concentration of protein, urinary protein, hematuria, past history of tonsillitis, the Yamamoto scale, the weight and pathological findings of the extracted palatine tonsils, and the presence or absence of anti-platelet drugs and renin-angiotensin system inhibitors (RAS-I) usage. This study included participants who were over 18 years of age, had undergone tonsillectomy within three months of steroid pulse therapy administered thrice, in whom renal biopsy was performed within a year before treatment, and with follow-up period of over 3 years. Clinical remission (CR) of urinary abnormalities was defined as remission of both proteinuria and hematuria: three consecutive negative results over a 6-month period, with a urinary sediment red blood cell count of <5/HPF, and a proteinuria qualitative reaction of (-) to (±). RESULTS: The CR rate of all cases was 56.8% and statistical significance was observed with respect to the C-Grade (P = 0.0003, P = 0.028) using both univariate and multivariate analysis. The CR rate of C-Grade І (73.4%) patients was significantly higher than that of C-Grade II patients (39.0%; P = 0.0004) and C-Grade III patients (30.8%; P = 0.003). We analyzed clinical factors in each C-Grade patient. No statistical significance was observed with respect to any of the factors using univariate analysis in C-Grade I patients. The weight of the extracted palatine tonsils and Yamamoto scale showed no statistical significance in every analysis. Fibrosis or hyalinization of the stroma of the palatine tonsils showed statistical significance (P = 0.026) only in the univariate analysis of C-Grade III patients. However, the patient number of C-Grade III was small. CONCLUSION: Our results indicate that TSP is mostly effective in patients with of C-Grade I IgAN and that the C-Grade reflects the clinical indication for TSP. The weight of the extracted palatine tonsils and Yamamoto scale did not show obvious correlations with the clinical effect of TSP.

Enhanced virulence of Fusarium species associated with spear rot of oil palm following recovery from osmotic stress.

Fusarium spp., which are common inhabitants of oil palm leaves, are weak pathogens of common spear rot (CSR). We investigated the influence of osmotic stress on the growth, virulence, and activity of cell wall-degrading enzymes of CSR fungi, using potato dextrose agar (PDA) supplemented with KCl or sucrose (hyperosmotic medium). Hyperosmotic stress significantly inhibited mycelial growth, but growth rapidly recovered when mycelia were transferred to control medium. When inoculated into oil palm spear leaflets, Fusarium sp., and F. incarnatum precultured on 1.0 and 1.5 M KCl-hyperosmotic medium induced lesions that were two to four times larger than those in non-stressed cultures, suggesting enhanced virulence of the weak pathogens. Lesion size was not greatly affected in hyperosmotic cultures of moderately virulent F. sacchari. No activity of pectin lyase was detected in liquid cultures of the Fusarium isolates. All isolates except F. incarnatum BT48 secreted polygalacturonase (PG), which was active in both liquid cultures and inoculated leaves. Significantly increased PG activity (5-32-fold) was observed on leaves inoculated with hyperosmotic cultures of Fusarium sp. and F. sacchari. These findings suggest that Fusarium sp., F. incarnatum, and F. sacchari exhibit an adaptive physiological plasticity to hyperosmotic stress that results in enhanced virulence.

Patients Over 60 Years of Age Have Poor Prognosis in Facial Nerve Decompression Surgery with Preserved Ossicular Chain.

OBJECTIVE: We report our retrospective study of the recovery rate of auditory ossicles preserved facial nerve decompression surgery via the transmastoid approach in cases of both an electroneurography score of < 10% and a Yanagihara score of ≤8 in Bell's palsy and Ramsay Hunt syndrome. MATERIALS AND METHODS: We retrospectively reviewed 47 patients who we were able to follow-up for more than 6 months following the onset of palsy. The recovery rate was defined by the Japan Society for Facial Nerve Research or the Yanagihara score. RESULTS: Twelve months after palsy onset, the recovery rate was 48.8% (20/41) for all patients, 65.2% (15/23) for patients with Bell's palsy, and 27.8% (5/18) for patients with Ramsay Hunt syndrome. Comparing the clinical efficacy of surgical treatment at 12 months after palsy onset, we observed a statistically significant effect of age. Comparing the Yanagihara scores of patients aged < 60 years with those of patients aged ≥60 years revealed that patients aged ≥60 years had significant poor prognosis, particularly in patients with Ramsay Hunt syndrome, which showed a very low recovery rate (14.3%). We also analyzed six other factors, but none showed statistical significance. CONCLUSION: The clinical efficacy of surgical treatment of Ramsay Hunt syndrome was inferior to that of Bell's palsy, which is consistent with previous reports. There was a statistically significant difference in the Yanagihara score between patients aged < 60 years and those aged ≥60 years. Particularly, patients with Ramsay Hunt syndrome aged ≥60 years have a very low recovery rate.

Treatment outcomes of sialendoscopy for submandibular gland sialolithiasis: The minor axis of the sialolith is a regulative factor for the removal of sialoliths in the hilum of the submandibular gland using sialendoscopy alone.

OBJECTIVE: To assess the general guidelines for the removal of sialoliths for submandibular gland sialolithiasis using sialendoscopy alone. METHODS: We analyzed 61 sialoliths treated using sialendoscopy in 42 patients with submandibular gland sialolithiasis. We evaluated the submandibular gland sialoliths and divided each case based upon the location: the Wharton's duct or the hilum. We measured the major and minor axes of the sialoliths using a soft tissue computed-tomography (CT) scan and evaluated the removal rate of the sialoliths using sialendoscopy alone. RESULTS: The removal rate of the sialoliths in the Wharton's duct (52.6%) was significantly higher than that in the hilum of the submandibular gland (26.1%) (P=0.042). The minor axis was significantly correlated to the treatment outcome of sialendoscopy alone for all cases (P=0.030). A significant correlation was observed for cases involving the hilum of the submandibular gland and the measurement of the minor axes of the sialoliths for the treatment outcome of sialendoscopy alone (P=0.009). The major axis showed no correlation with the treatment outcomes of sialendoscopy alone. CONCLUSION: The measurement of the minor axes of the sialoliths with a soft tissue CT scan was correlated with treatment outcome of sialendoscopy alone for all cases, particularly sialoliths in the hilum. The easurement of the major axis showed no correlation with outcomes of sialendoscopy alone.

Diagnostic and treatment effects of sialendoscopy for patients with swelling of the parotid gland when sialoliths are undetected with computed tomography.

Between August 2009 and May 2016, 74 patients underwent sialoendoscopic surgery. 32 patients had parotid gland disease and 9 patients had intermittent swelling of the parotid gland and sialoliths were not detected with CT imaging. 4 patients were diagnosed with idiopathic Stensen's duct stenosis. Sialendoscopy directly confirmed Stensen's duct stenosis in 2 patients. However, the sialendoscope was unable to be inserted in the other 2 patients, who had stenosis of the orifice of the Stensen's duct. Balloon expansion of the duct was performed in these 2 patients and a steroid drug was injected into the duct in one patient. Complete remission was archived in one patient treated with sialendoscopy. Three patients had sialolithiasis. Microsialoliths and/or white floating matter was observed and removed using sialendoscopy. All patients experienced complete remission. In cases of Sjögren syndrome and recurrent parotitis, sialendoscopic surgery was performed, but the symptoms showed no improvement. For patients with microsialoliths, sialendoscopy may be most useful for diagnosis and treatment when the sialoliths are not detected with CT imaging. At present, sialendoscopic surgery have limitation in the treatment of Stensen's duct stenosis and may similarly have limitation in the treatment of Sjögren's syndrome and recurrent parotitis.

The landmark for removal of sialoliths using sialendoscopy alone in parotid gland sialolithiasis.

OBJECTIVE: To assess the general guidelines for removal of sialoliths in parotid gland sialolithiasis using sialendoscopy alone. METHODS: We analyzed 34 sialoliths treated using sialendoscopy in 26 patients with parotid gland sialolithiasis. We divided the Stensen's duct and parotid gland into for parts using computed tomography findings: (A) front of the masseter, (B) anterior and lateral to the center (anterolateral) of the masseter, (C) posterior and lateral to the center (posterolateral) of the masseter, (D) behind of the masseter. The location and size of each sialolith was assessed. RESULTS: The removal rates of sialoliths in the different locations by sialendoscopy alone were as follows: front of the masseter, 68.8%; anterolateral of the masseter, 60.0%; posterolateral of the masseter, 0%; and behind of the masseter, 33.3%. The removal rate using sialendoscopy alone was significantly higher in the sections anterior to the center of the masseter than in those posterior to the center of the masseter (66.7% [14/21] vs. 20.0% [2/10]; P=0.019). The size of the sialolith was not correlated to the removal rate by sialendoscopy alone. CONCLUSION: Sialoliths of the parotid gland located in positions anterior to the center of the masseter are significantly easier to remove by sialendoscopy alone. The center of the masseter is a general landmark for removal of sialoliths from the parotid gland using sialendoscopy alone. The size of the sialolith is not correlated with removal, except rare huge sialoliths.

Elucidation of the biosynthetic pathway of cis-jasmone in Lasiodiplodia theobromae.

In plants, cis-jasmone (CJ) is synthesized from α-linolenic acid (LA) via two biosynthetic pathways using jasmonic acid (JA) and iso-12-oxo-phytodienoic acid (iso-OPDA) as key intermediates. However, there have been no reports documenting CJ production by microorganisms. In the present study, the production of fungal-derived CJ by Lasiodiplodia theobromae was observed for the first time, although this production was not observed for Botrytis cinerea, Verticillium longisporum, Fusarium oxysporum, Gibberella fujikuroi, and Cochliobolus heterostrophus. To investigate the biosynthetic pathway of CJ in L. theobromae, administration experiments using [18,18,18-2H3, 17,17-2H2]LA (LA-d5), [18,18,18-2H3, 17,17-2H2]12-oxo-phytodienoic acid (cis-OPDA-d5), [5',5',5'-2H3, 4',4'-2H2, 3'-2H1]OPC 8:0 (OPC8-d6), [5',5',5'-2H3, 4',4'-2H2, 3'-2H1]OPC 6:0 (OPC6-d6), [5',5',5'-2H3, 4',4'-2H2, 3'-2H1]OPC 4:0 (OPC4-d6), and [11,11-2H2, 10,10-2H2, 8,8-2H2, 2,2-2H2]methyl iso-12-oxo-phytodienoate (iso-MeOPDA-d8) were carried out, revealing that the fungus produced CJ through a single biosynthetic pathway via iso-OPDA. Interestingly, it was suggested that the previously predicted decarboxylation step of 3,7-didehydroJA to afford CJ might not be involved in CJ biosynthesis in L. theobromae.

Telomerase-specific oncolytic adenovirus: antitumor effects on radiation-resistant head and neck squamous cell carcinoma cells.

BACKGROUND: Radioresistance remains a critical issue in the use of radiotherapy for the treatment of head and neck squamous cell carcinoma (HNSCC). This study evaluated the efficacy of combination treatment with OBP-301, a telomerase-specific replication-selective adenovirus, and radiotherapy in overcoming radioresistance by examining its effect on radiation-resistant HNSCC cells. METHODS: Radiation-resistant HNSCC cells were treated with OBP-301 and radiation in vitro and in an orthotopic nude mouse model in vivo and synergism was assessed. Apoptosis and expression of MRN complex, which plays a key role in DNA repair machinery, were also analyzed. RESULTS: Infection with OBP-301 was found to enhance the antitumor efficacy of radiation both in vitro and in vivo by inhibiting MRN complex expression and increasing apoptosis induction. CONCLUSION: Combined OBP-301 and radiation therapy seems to overcome radioresistance in HNSCC cells by inhibiting DNA repair machinery, and may thus be a novel therapeutic strategy for treating HNSCC.

DNA markers linked to Pga1, an adzuki bean gene that confers resistance to Cadophora gregata race 1.

Brown stem rot (BSR) caused by Cadophora gregata f. sp. adzukicola (syn. Phialophora gregata) is a serious soilborne disease of adzuki bean (Vigna angularis) in Japan. Cultivation of resistant cultivars is the most effective disease control method, therefore the selection of resistant lines is a priority for breeders. BSR-resistant adzuki bean lines have been screened in pathogen-infected fields. However, field selection using the pathogen and artificial inoculation methods is time-consuming and labor-intensive. In the present study, we used 105 F3 lines derived from a cross between a BSR-resistant cultivar 'Syumari' and a susceptible cultivar 'Buchishoryukei-1' for BSR inoculation tests. Amplified fragment-length polymorphism (AFLP) analyses with 1024 primer sets revealed that six fragments were polymorphic between resistance and susceptible bulked groups. Five DNA markers (Pg77, Pg118, Pg138, Pg139 and Pg126) were developed from the nucleotide sequences of polymorphic AFLP markers and their flanking regions. Pg118, which was derived from E-ACT/M-ACT-118, was tightly linked to the resistance gene Pga1 and was converted into a codominant marker for its easier use in marker-assisted selection for adzuki bean BSR resistance. Finally, the applicability of the developed markers for BSR resistance was tested on 32 adzuki bean accessions or cultivars.

Olfactory neuroblastoma: 15 years of experience.

BACKGROUND: Olfactory neuroblastoma is a rare malignant neoplasm of the nasal cavity and of the paranasal sinus. In the treatment of patients with advanced olfactory neuroblastoma, the combination of craniofacial resection with radiotherapy and/or chemotherapy has significantly improved the survival rate. However, locoregional recurrence and distant metastasis frequently occur, irrespective of the aggressiveness of therapy. We report our experience on the outcomes of olfactory neuroblastoma in patients treated with concurrent chemoradiotherapy (CCRT). PATIENTS AND METHODS: We retrospectively analyzed the cases of seven patients with olfactory neuroblastoma, treated in the past 15 years. Six patients were treated with CCRT, consisting of cisplatin (60 mg/m(2), day 4), 5-fluorouracil (600 mg/m(2), given over 24 h for 5 days, days 1-5), methotrexate (30 mg/m(2), day 1) and leucovorin (20 mg/m(2), days 1-5) (PFML). One patient was treated with radiotherapy and neoadjuvant chemotherapy, which consisted of a combination of three drugs: cyclophosphamide (360 mg/m(2), day 1), cisplatin (60 mg/m(2), day 4), 5-fluorouracil (600 mg/m(2), given over 24 h for 5 days, days 1-5) (PFC). Salvage surgery was performed in the primary remaining site. RESULTS: The overall 5-year survival rates were 3/6 for patients treated with CCRT using PFML and 2/5 for patients with Kadish stage C olfactory neuroblastoma. The locoregional response rate was 4/6. CONCLUSION: In our limited experience, CCRT with PFML had therapeutic efficacy as a primary treatment, while surgical treatment and postoperative radiotherapy have been the main treatment modalities.

Combined molecular targeted drug therapy for EGFR and HER-2 in head and neck squamous cell carcinoma cell lines.

The epidermal growth factor receptor (EGFR) and related family member, HER-2, are often overexpressed simultaneously in patients with a variety of malignant tumors, and the combination may cooperatively promote cancer cell growth and survival. The purpose of this study was to examine antitumor effects of the combination treatment of cetuximab and trastuzumab on head and neck squamous cell carcinoma (HNSCC) using 16 HNSCC cell lines in terms of antiproliferative effect and antibody-dependent cell-mediated-cytotoxicity (ADCC). Previously we have reported the expression levels of EGFR mRNA on 16 HNSCC cell lines. All cell lines expressed mRNA for EGFR, HER-2 and HER-3; 12 cell lines expressed mRNA for HER-4; and 4 cell lines did not express mRNA for HER-4. In in vitro proliferation assay, the combination treatment of cetuximab and trastuzumab significantly lowered cell viability compared to either drug alone. The mRNA expression levels of EGFR and HER-2 were not correlated with the efficacy of the combination treatment of cetuximab and trastuzumab and the expression levels of HER-3 and HER-4 also showed no correlation with the efficacy of the combination treatment. We evaluated the gene status of HER-2 exons 23 and 24 in 16 HNSCC cell lines, but there was no mutation of HER-2 in any of the cell lines. Either drug showed ADCC in the 3 cell lines using peripheral blood mononuclear cells (PBMCs), however, a significant combination effect was not observed. Combined molecular targeted antibody drug therapy for EGFR and HER-2 may be useful in the treatment of HNSCC.

Diagnostic sensitivity of ¹8fluorodeoxyglucose positron emission tomography for detecting synchronous multiple primary cancers in head and neck cancer patients.

We assessed the sensitivity of positron emission tomography (PET) for detecting synchronous multiple primary cancers, particularly synchronous esophageal cancers in head and neck cancer patients. We retrospectively reviewed 230 head and neck cancer patients. All the patients routinely underwent the following examinations: urinalysis, occult blood, tumor marker detection [squamous cell carcinoma (SCC), cytokeratin fragment (CYFRA), and carcinoembryonic antigen (CEA)], esophagogastroduodenoscopy, colonoscopy (when CEA was high or occult blood was positive), abdominal ultrasonography, plain chest computed tomography (CT), and PET. Bronchoscopy was performed when CT revealed lung shadow of central region. Synchronous multiple primary cancers were detected in 42 (18.2%) patients. The diagnostic sensitivity of PET for synchronous primary cancers was as follows: esophagus, 7.6% (1/13); stomach, 25.0% (2/8); lung, 66.7% (4/6); head and neck, 75.0% (3/4); colon, 0% (0/1); kidney, 0% (0/1); and subcutaneous, 100% (1/1). The sensitivity of PET for detecting synchronous esophageal cancers is low because these are early-stage cancers (almost stage 0-I). Therefore, it is necessary to perform esophagogastroduodenoscopy for detecting synchronous esophageal cancers. PET is an important additional tool for detecting synchronous multiple primary cancers because the diagnostic sensitivity of PET in synchronous head and neck cancer and lung cancer is high. But PET has the limitation of sensitivity for synchronous multiple primary cancers because the diagnostic sensitivity of PET in synchronous esophageal cancer is very low.

Common Spear Rot of Oil Palm in Indonesia.

Common spear rot (CSR), which is also known as crown disease, was first reported in Indonesia in the 1920s. It has caused considerable losses in young oil palm plantings, and yet the pathogenic agent has remained elusive. Symptomatic spear leaves were collected from oil palm plantations and farm plots in South Sumatra, North Sumatra, and Bangka-Belitung, Indonesia. Of the 14 different fungi isolated, Fusarium incarnatum, F. solani, an undescribed Fusarium sp., and Ceratocystis paradoxa were isolated most frequently from diseased leaf tissue. F. incarnatum and the undescribed Fusarium sp. were also frequently isolated from healthy leaf tissue, along with Pestalotiopsis microspora and Curvularia affinis. Ceratocystis paradoxa was never isolated from healthy leaf tissue. Koch's postulate experiments showed that C. paradoxa was able to infect wounded oil palm leaves causing a symptom of extensive rotting similar to that found in the field. Although isolated less frequently and less virulent than C. paradoxa, F. sacchari was also capable of causing lesions on succulent wounded, inoculated leaves. For both C. paradoxa and F. sacchari, the disease severity index was greater when the oil palm leaves appeared to have more succulent growth. Likewise, other Fusarium species and other nonfusarial fungi that were usually not pathogenic were weakly virulent on palms with more succulent growth. These findings confirm that C. paradoxa is one pathogen that is associated with CSR of oil palm in Indonesia.

New Sources of Resistance to Cadophora gregata f. sp. adzukicola and Fusarium oxysporum f. sp. adzukicola in Vigna spp.

Adzuki bean brown stem rot (BSR), caused by Cadophora gregata f. sp. adzukicola, and adzuki bean Fusarium wilt (AFW), caused by Fusarium oxysporum f. sp. adzukicola, are serious problems in Hokkaido, Japan, and have been managed using cultivars with multiple resistance. However, a limited number of adzuki bean varieties are resistant to these pathogens because of the frequent appearance of new races; thus, new sources of resistance have been sought in related Vigna spp., particularly in the section Angulares to which adzuki bean belongs. An analysis of selected Vigna accessions (JP81231 to JP235420) conserved in the Genebank of the National Institute of Agrobiological Sciences, Tsukuba, Japan, revealed wide variation in resistance spectra and resistance combinations, and eight disease response groups (A to H) were identified. Four of eight were newly detected response groups, suggesting the existence of novel resistance genes. Of 252 accessions from 26 species, 28 accessions in Vigna angularis var. nipponensis, V. hirtella, V. minima, and V. tenuicaulis (section Angulares) from group D, which were cross-compatible with adzuki bean, are expected to be potential sources of multiple resistance genes. They were resistant to all races of BSR and AFW pathogens examined.

Gene status of head and neck squamous cell carcinoma cell lines and cetuximab-mediated biological activities.

Cetuximab is a chimeric IgG1 monoclonal antibody that targets epidermal growth factor receptor (EGFR). Cetuximab binds to EGFR and prevents phosphorylation of EGFR. Moreover, preclinical results have shown the ability of cetuximab to induce either complement-mediated tumor cell killing (CDC) or antibody-dependent cell-mediated-cytotoxicity (ADCC). We previously reported mutation in EGFR regarding head and neck squamous cell carcinoma (HNSCC) cell lines. In the present study, we analyzed the same 16 HNSCC cell lines for mutations in KRAS, PIK3CA, BRAF and PTEN. Furthermore, we evaluated cetuximab-mediated biological activities (antiproliferative effect by the MTT assay and ADCC) regarding these cell lines. Mutations in PIK3CA and PTEN were observed in two cell lines (2/16, 12.5%), but no mutation was observed in KRAS and BRAF. The antiproliferative effect of cetuximab by the MTT assay was not strong, and no correlation was observed between the antiproliferative effect of cetuximab and mutations in EGFR, KRAS, PIK3CA, BRAF and PTEN in these cell lines. Therefore, the mutation status of EGFR and downstream molecules were not useful for predicting the antitumor effects of cetuximab on HNSCC. Cetuximab-mediated ADCC was observed in these cell lines and might have been influenced by the expression of EGFR. Therefore, cetuximab-mediated ADCC seems to be an important part of the antitumor mechanisms of cetuximab and the expression levels of EGFR might influence the antitumor activity of cetuximab. Therefore, besides the antiproliferative effect of cetuximab by the MTT assay, it appeared important to evaluate cetuximab-mediated ADCC as well as EGFR expression in HNSCC cells.

Examination of the optimal condition on the in vitro sensitivity to telomelysin in head and neck cancer cell lines.

OBJECTIVE: Telomelysin (OBP-301) is a telomerase-specific replication-competent adenovirus with a human telomerase reverse transcriptase (hTERT) promoter. Telomelysin has a strong antitumor effect on a variety of cancers, including head and neck squamous cell carcinoma (HNSCC), and combining telomelysin treatment with paclitaxel or cisplatin enhances the antitumor effect on HNSCC. In the present study, we investigated the relationship between the antitumor activity of telomelysin and tumor cell doubling time(DT), S-phase fraction, and E1A expression. We also investigated whether the antitumor effects of OBP-301-resistant tumor cells are enhanced by cisplatin, paclitaxel, or streptolysin O. METHODS: The tumor cell DT of 17 human HNSCC cell lines was examined. Antitumor activities of telomelysin (OBP-301) for each HNSCC cell line were examined by MTT assay. Cell cycle analysis was conducted by flowcytometry. E1A gene expressions after infection with telomelysin, hTERT, CAR (Cocksackie Adenovirus Receptor), and c-Myc were examined by quantitative PCR, and E1A expressions were examined again after pretreatment with cisplatin, paclitaxel, or streptolysin O. Correlations were analyzed by Spearman's correlation coefficient. RESULTS: There was a significant relationship between telomelysin sensitivity and DT, S-phase fraction and early E1A expression, and pretreatment with cisplatin, paclitaxel, and streptolysin O increased infectivity of telomelysin-resistant HNSCC cell lines. CONCLUSION: These findings are useful for advancing clinical trials, and suggest that adjuvant telomelysin treatment would be effective even in telomelysin-resistant HNSCC cell lines.

Antitumor effects of lapatinib (GW572016), a dual inhibitor of EGFR and HER-2, in combination with cisplatin or paclitaxel on head and neck squamous cell carcinoma.

The epidermal growth factor receptor (EGFR) and a related family member, HER-2, are often overexpressed simultaneously in patients with a variety of malignant tumors, and the combination may cooperatively promote cancer cell growth and survival. Heterodimerization of EGFR and HER-2 has been known to create intense proliferative signals. Lapatinib (GW572016) is a small molecule that is administrated orally and functions as a reversible inhibitor of both EGFR and HER-2 tyrosine kinases. In the present study, we evaluated the antitumor effect of lapatinib on head and neck squamous cell carcinoma (HNSCC) cell lines in vitro and in vivo. In vivo we examined the antitumor effects of combined treatment with lapatinib and either cisplatin or paclitaxel. In vitro lapatinib displayed antiproliferative effects on HNSCC cells. The IC50 of lapatinib ranged between 13.6 and 60.2 microM after 24-h exposure to lapatinib. A correlation was not observed between results of in vitro proliferation assays for lapatinib and the expression of EGFR or HER-2. In vivo lapatinib displayed antitumor activity, and induced apoptosis in nude mice bearing an established xenograft of YCU-H891 cells. Lapatinib did not significantly inhibit angiogenesis. Combination treatment of lapatinib with cisplatin or paclitaxel enhanced antitumor activity mainly by inducing apoptosis. Inhibition of antiangiogenesis was observed only for combination treatment of lapatinib with paclitaxel (compared to vehicle control). These results suggest that: i) lapatinib has antitumor effects in vitro and in vivo; ii) lapatinib may be more effective in combination with cisplatin or paclitaxel; and iii) lapatinib might provide useful clinical benefits to HNSCC patients.

Antitumor effects of telomelysin in combination with paclitaxel or cisplatin on head and neck squamous cell carcinoma.

Telomelysin (OBP-301) is a telomerase-specific replication-component adenovirus. Telomelysin has a human telomerase reverse transcriptase (hTERT) promoter element which efficiently kills human cancer cells, but not normal cells. The present study investigated the correlation between the antitumor effect of telomelysin and mRNA expression of hTERT and coxsackievirus and adenovirus receptor (CAR) in head and neck squamous cell carcinoma (HNSCC) in vitro and whether telomelysin enhances the antitumor effect of paclitaxel or cisplatin, in vivo using a HNSCC xenograft model. We also determined the optimal order for combining telomelysin treatment and chemotherapy as concurrent treatment, telomelysin treatment first and chemotherapy later, chemotherapy first and telomelysin treatment later for achieving the best anticancer effect. The mRNA expression of hTERT and CAR genes was examined by quantitative RT-PCR in 17 HNSCC cell lines. There was no significant correlation between the growth inhibition of telomelysin (ID50 for day 3, 5 and 7) in vitro and mRNA expression levels of hTERT and CAR. Regarding the correlation between CAR expression and telomelysin ID50 for day 3, all cell lines that showed a relative amount of CAR/beta-actin mRNA >0.4 had a low telomelysin ID50. This may indicate that CAR expression contributes to the efficacy of adenovirus infection and the antitumor activity of telomelysin in early stages of treatment. In our in vivo study, combining telomelysin and paclitaxel had an additive effect regardless of treatment order. On the other hand, combining telomelysin and cisplatin had additive effect only when cisplatin treatment preceded telomelysin treatment. These results suggest that paclitaxel is considered innocuous for replication of telomelysin, however cisplatin may influence replication of telomelysin.