FOXO3 is essential for CD44 expression in pancreatic cancer cells.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most fatal types of cancer and the 5-year survival rate is only 5%. Several studies have suggested that cancer stem cells (CSCs) are thought to be involved in recurrence and metastasis and so it is essential to establish an approach targeting CSCs. Here we have demonstrated that cyclic guanosine monophosphate (cGMP) suppressed CD44 expression and the properties of CSCs in PDAC. Microarray analysis suggested that cGMP inhibited Forkhead box O3 (FOXO3), which is known as a tumor suppressor. Surprisingly, our data demonstrated that FOXO3 is essential for CD44 expression and the properties of CSCs. Our data also indicated that patients with high FOXO3 activation signatures had poor prognoses. This evidence suggested that cGMP induction and FOXO3 inhibition could be ideal candidates for pancreatic CSC.

Effect of highly lipolyzed goat cheese on HL-60 human leukemia cells: antiproliferative activity and induction of apoptotic DNA damage.

To establish cheese as a dairy product with health benefits, we embarked on examining the multifunctional role of cheeses, especially in the field of cancer prevention. The current study was designed to investigate whether different types of commercial goat cheeses may possess antiproliferative activity, using an HL-60 human promyelocytic leukemia cell line as a cancer cell model. Among 11 cheese extracts tested at 500µg/mL, 6 (Crottin de Chavignol, Pouligny Saint-Pierre, Chabichou du Poitou, Valencay, Kavli, and Sainte-Maure de Touraine) resulted in a significant decrease of cell viability, which is consistent with a decrease in viable cell number. Compared with the half-maximal inhibitory concentration (IC(50)) value of individual cheeses in cellular proliferation assays, the Pouligny Saint-Pierre extract showed strong inhibition. Incubation of cells in the presence of Pouligny Saint-Pierre extract resulted in induction of cellular morphological changes and apoptotic DNA fragmentation as well as expression of the active form of caspase-3 protein. Based on the quantification of the ratio of free fatty acids to triglycerides in different cheese samples, a significant correlation was detected between lipolytic ripeness and IC(50) values for antiproliferative capacity tested in HL-60 cells. Collectively, these results support a potential role of highly lipolyzed goat cheeses in the prevention of leukemic cell proliferation.

Predictive factors for radiation pneumonitis in oesophageal cancer patients treated with chemoradiotherapy without prophylactic nodal irradiation.

OBJECTIVE: The objective of this study was to identify clinical and dosimetric factors for the development of radiation pneumonitis (RP) among patients with oesophageal cancer treated with three-dimensional radiotherapy without prophylactic nodal irradiation. METHODS: 125 patients with oesophageal cancer had undergone dose-volume histogram (DVH) metrics and received chemoradiotherapy (CRT). Several clinical and dosimetric factors with regard to the lung were evaluated as predictive factors for the development of symptomatic RP. RESULTS: 26 patients (20.8%) developed symptomatic RP classified as greater than or equal to Grade 2. By univariate analysis, body weight loss, tumour length, Stage IV, response to treatment and all DVH parameters proved to be significant factors for the development of RP (p < 0.05). By multivariate analysis, Stage IV and all dosimetric factors were independent predictive factors for the development of symptomatic RP (p < 0.05). Recursive partitioning analysis indicated that V10 values of 24.8% or more and Stage IV were associated with higher development of RP (odds ratio 6.53). CONCLUSIONS: Our study demonstrated that severe RP was also developed in patients treated with the minimal radiation field. Stage IV and the dosimetric factors were identified as independent predictive factors for symptomatic RP in oesophageal cancer patients treated with CRT without prophylactic nodal irradiation.

SU-E-T-555: Weighted Four-Dimensional IMRT Planning for Dynamic MLC Tracking Using a Practical and Simple Framework.

PURPOSE: We present a simple, practical framework for truly 4D lung IMRT planning based on a weighted individual-phase optimization paradigm. This strategy is specifically developed for use in real-time tumor tracking delivery systems so as to utilize respiratory motion as an additional degree of freedom rather than a constraint. METHODS: A 4D-CT scan from a lung SBRT patient was loaded into the Eclipse treatment planning system. The target and normal structures were manually contoured on each of the ten phases. For each phase, the total dose prescription was scaled by the number of phases and a seven-field plan was developed. An open-source deformable image and dose registration engine (DIRART) was used to deform the dose map at each phase to a reference phase. DVH data from the individually optimized phase plans were input into an in-house linear programming-based optimizer implemented in MATLAB, in order to determine dose-weighting factors for each phase. The objective function aimed to maintain PTV coverage while keeping normal structure dose as low as possible. This weighted-4D plan (W-4D) was compared to an ITV-based plan and a 4D plan with equal dose-weights to individual phases (E-4D). RESULTS: The W-4D dose fractions were determined to be 0.33, 0.01, 0.65 and 0.02 at phase 0%, 30%, 40%, and 90%, respectively (and zero elsewhere). PTV coverage (V95) was close to identical for all three strategies. The W-4D plan exhibited mean lung dose 18.8% and 8.5% lower and mean liver dose 23.3% and 5.7% lower than corresponding values from ITV-based and E-4D plans, respectively. CONCLUSIONS: By significantly improving normal structure sparing while maintaining PTV coverage, weighted 4D planning represents a more attractive solution than ITV-based planning for (currently investigational) real-time tumor tracking-based delivery systems.

A comparison of radiation treatment plans using IMRT with helical tomotherapy and 3D conformal radiotherapy for nasal natural killer/T-cell lymphoma.

The purpose of this study was to compare radiation treatment plans (RTPs) that used intensity-modulated radiation therapy (IMRT) with helical tomotherapy (HT) or three-dimensional conformal radiation therapy (3D-CRT) for nasal natural killer/T-cell lymphoma (NNKTL). We created RTPs that used IMRT with HT or 3D-CRT for eight NNKTL patients previously treated at our institution and conducted a pilot comparison between the two modalities using the parameters of the target coverage and homogeneity for the planning target volume (PTV) and the maximum and mean doses for organs at risk (OARs). The clinical target volume (CTV) included the gross tumour volume with an additional margin of 1.5 cm and the nasopharynx, palates and nasal cavity; the PTV with the CTV plus a 2 mm margin received a total dose of 50 Gy. IMRT achieved significantly better PTV coverage, with more than 99% of the PTV receiving 90% and 95% of the prescribed dose, whereas 3D-CRT could not provide adequate coverage of the PTV, with 89.1+/-2.6% and 84.5+/-2.7% of the PTV receiving 90% and 95% of the prescribed dose, respectively (both p <0.0001). The homogeneity index was 0.29+/-0.06 for IMRT and 0.046+/-0.022 for 3D-CRT, which was statistically significant (p <0.0001). IMRT tended to provide equivalent or slightly better OAR avoidance than 3D-CRT. In conclusion, 3D-CRT could not provide adequate coverage of the PTV because the PTV was close to many OARs. IMRT should be used for NNKTL because a lack of optimal RTPs could cause local failure.

False-positive positron emission tomography appearance with 18F-fluorodeoxyglucose after definitive radiotherapy for cancer of the mobile tongue.

(18)F-fluorodeoxyglucose-positron emission tomography (FDG-PET) is an effective tool for evaluating the results of radiotherapy. However, some false-positive appearances caused by physiological or pathological accumulation are reported. We report on three patients who showed a high accumulation of FDG in the lingual muscles but had no recurrent tumour after definitive radiotherapy for the mobile tongue. All patients had squamous cell carcinoma of the tongue and received interstitial radiotherapy with small sources. High uptake was seen in the lingual muscles without recurrence or inflammation, based on physical and MR examinations. This false-positive appearance is thought to relate to ill-balanced high activity of the lingual muscles after definitive radiotherapy.

Helical tomotherapy for brain metastases: dosimetric evaluation of treatment plans and early clinical results.

The purpose of this study was to evaluate the feasibility and treatment plans of intensity-modulated radiation therapy using helical tomotherapy (HT) for brain metastases. Twenty-three patients with 1 to 4 brain metastases were treated with HT. In combination with whole-brain radiotherapy (simultaneous plans), metastatic lesions, and the whole brain were treated with 50 Gy and 30 Gy, respectively, in 10 fractions, with a simultaneous integrated boost technique. In patients treated for brain metastases alone (focal plans), metastatic lesions were treated with 35 or 37.5 Gy in 5 fractions. The treatment plans were compared regarding the conformation number (CN) and homogeneity index (HI), and differences in these indexes between simultaneous and focal plans were examined by Student's t-test. Seven and 16 patients were treated with simultaneous plans and focal plans, respectively. The mean +/- SD of CN and HI values were 0.75 +/- 0.13 and 0.063 +/- 0.042, respectively, for simultaneous plans, and 0.73 +/- 0.12 and 0.052 +/- 0.023, respectively, for focal plans. The CN and HI between the two plans were not significantly different. Response rates in 13 patients with follow-up imaging were approximately 90% for both plans and the local control rate at 1 year was 69%. One patient with a huge tumor (34.0 cc) and WHO performance status 3 treated with focal plans experienced severe headache, requiring prolongation of the treatment time, and died at 8 days after completion of treatment. The exact cause of deterioration was uncertain as no radiological investigation was performed in this patient. No late complications were observed during follow-up periods up to 20 months. HT is a viable non-invasive technique for treatment of brain metastases and achieves high accuracy in terms of dose conformity and homogeneity.

Identifying patients with peripheral-type early non-small cell lung cancer (T1N0M0) for whom irradiation of the primary focus alone could lead to successful treatment.

We investigated the indication of radiotherapy in operable patients with peripheral-type early non-small cell lung cancer (T1N0M0 (TNM staging in 1997)). The subjects comprised 396 patients with non-small cell lung cancer in whom the clinical stage was evaluated as IA. We examined age, gender, Brinkmann's index, histopathological type, the grade of histopathological differentiation, tumour diameter and the level of carcinoembryonic antigen as factors involved in lymph node metastasis. Lymph node metastasis was detected in 79 patients (20%). Factors such as the grade of histopathological differentiation and tumour diameter were involved in lymph node metastasis. In well-differentiated lesions, the probability of metastasis was <10% even when the tumour diameter exceeded 2 cm. However, the probability rapidly increased with tumour size in moderately and poorly differentiated lesions. Among the patients with peripheral-type early non-small cell lung cancer (T1N0M0), the risk of lymph node metastasis was low in those with well-differentiated carcinoma and those with moderately differentiated lesions measuring

Treatment results of boron neutron capture therapy using intra-arterial administration of boron compounds for recurrent head and neck cancer.

The effect of boron neutron capture therapy (BNCT) is correlated with the density of boron in the tumour. BNCT using intra-arterial administration of boron compounds was performed for recurrent head and neck cancer. Of the five patients treated, one achieved a complete response and four achieved a partial response. There was one case of transient headache but no severe adverse effects were observed. The advantages of using an intra-arterial administration route for BNCT, which causes the selective killing of tumour cells, might offer a new option in the treatment of recurrent head and neck malignancies. These promising results require further verification and optimization of the BNCT schedule; however, dose escalation would appear to be justified because the toxicity appears to be very low.

Intra-arterial chemoradiotherapy for locally advanced oral cavity cancer: analysis of therapeutic results in 134 cases.

The objective of this study was to investigate the therapeutic results of arterial injection therapy via the superficial temporal artery for 134 cases of stages III and IV (M0) oral cavity cancer retrospectively, and to clarify the prognostic factors. We administered intra-arterial chemoradiotherapy by continuous infusion of carboplatin in 65 cases from January 1993 to July 2002. Systemic chemotherapy was performed on 26 cases at the same time. We administered intra-arterial chemoradiotherapy by cisplatin with sodium thiosulphate in 69 cases from October 2002 to December 2006. Systemic chemotherapy was performed on 48 cases at the same time. The 3-year local control rate was 68.6% (T2-3: 77.9%; T4: 51.3%), and the 3-year survival rate was 53.9% (stage III: 62.9%; stage IV: 45.3%). Regarding the results of multivariate analysis of survival rates, age (<65), selective intra-arterial infusion, and the use of cisplatin as an agent for intra-arterial infusion were significant factors. The therapeutic results of intra-arterial chemoradiotherapy via the superficial temporal artery were not inferior to the results of surgery. In particular, the results of arterial injection therapy by cisplatin with sodium thiosulphate were excellent, so we believe that it will be a new therapy for advanced oral cavity cancer.

Bite force, occlusal contact area and masticatory efficiency before and after orthognathic surgical correction of mandibular prognathism.

The purpose of this study was to evaluate bite force, occlusal contact area and masticatory efficiency before and after sagittal split ramus osteotomy in 27 patients with mandibular prognathism, in comparison with 27 control subjects with normal occlusion. Bite force and occlusal contact area were simultaneously measured with a computerized occlusal analysis system, the Dental Prescale system. Masticatory efficiency was estimated by a low-adhesive colour-developing chewing-gum system. The data were collected at initial medical consultation, immediately before surgery, and at 6 weeks, 3 months, 6 months, 1 year and more than 2 years after surgery. Both bite force and occlusal contact area of the patients before surgery were significantly less than those of the controls. Although all three parameters had improved after orthognathic surgery, the bite force and occlusal contact area did not reach the values of the controls within 2 years postoperatively; masticatory efficiency at 2 years after surgery drew near to control levels. Bite force correlated with occlusal contact area in the patients postoperatively, whereas masticatory efficiency did not correlate with either of the other two parameters. These results suggest that further adjustment of occlusion and mechanical advantage should be considered before the end of treatment.

Relationship between the growth pattern of nasopharyngeal cancer and the cervical lymph nodes based on MRI findings: can the cervical radiation field be reduced in patients with nasopharyngeal cancer?

To identify patients with nasopharyngeal cancer in whom the cervical radiation field can be reduced, we classified the growth patterns of nasopharyngeal cancer based on MRI findings into 4 types and performed an evaluation. Based on MRI findings, we classified the growth patterns of primary cancer in 94 patients with nasopharyngeal cancer into Type 1 (superficial type), Type 2 (lateral invasive type), Type 3 (upward invasive type), and Type 4 (anterior extension type), and further classified Type 2, based upon nasopharyngoscopic findings, into Type 2a (unilateral invasive type) and Type 2b (bilateral invasive type). The cervical lymph node metastasis areas were evaluated according to these types. Type 2 showed a significantly higher incidence of cervical lymph node metastasis only on the ipsilateral side than the other types (p = 0.0024). In particular, all patients with Type 2a had cervical lymph node metastasis only on the ipsilateral side (p = 0.0212). This study suggests that the distribution of metastasised cervical lymph nodes depends on the pattern of tumour extent of the primary site.

Denervation atrophy of the masticatory muscles in a patient with nasopharyngeal cancer: MR examinations before and after radiotherapy.

We report on a patient with denervation atrophy of the masticatory muscles due to nasopharyngeal cancer who received therapeutic irradiation. Magnetic resonance imaging has significantly contributed to aid diagnosis of this pathology. Masticatory muscle atrophy should be a definitive finding of perineural invasion caused by head and neck tumors. Radiologists should be familiar with this appearance to avoid confusion with tumor invasion of the muscle.

Continuous selective intraarterial chemotherapy in combination with irradiation for locally advanced cancer of the tongue and tongue base.

We retrospectively evaluated the results of the concurrent combination therapy of selective continuous intraarterial chemotherapy and radiotherapy in 39 patients with locally advanced cancer of the tongue and tongue base between September 1992 and January 2000. Thirty patients were fresh cases (stage II, 10 patients; stage III, 15; stage IV, five) and nine were recurrent cases. The primary lesion was present in the mobile tongue in 33 patients and the tongue base in six. External irradiation (median dose, 48.6 Gy) was performed in all patients, and interstitial brachytherapy using an Au grain or Cs needle (median dose, 50 Gy) in 21. In intraarterial chemotherapy, a catheter was selectively inserted into the lingual artery via the superficial temporal artery, and carboplatin (CBDCA) was continuously infused (median dose, 460 mg/m(2)) concurrently with radiotherapy. In 13 patients with cervical lymph node metastasis, two courses of systemic chemotherapy with 5-FU (700 mg/m(2) x 5 days) and cisplatin (40-50 mg/m(2)x2 days) or its analog was also performed. In 37 (94.9%) of the 39 patients in whom this combination therapy was completed, the response rate was 94.6%. The 3-year local control rate, progression-free survival rate, and overall survival rate by Kaplan-Meier's method were 79.2, 53.2, and 58.9%, respectively. This combination therapy was effective for locally advanced cancer of the tongue and tongue base without causing severe adverse side effects, and a local control rate comparable to that by surgery can be expected.

Native-like tertiary structure formation in the alpha-domain of a hen lysozyme two-disulfide variant.

Structure formation in two species of the two-disulfide variant of hen lysozyme was investigated by means of CD spectroscopy, disulfide exchange measurement, and 1H-NMR spectroscopy. One species, 2SS [6-127, 30-115], which contained the two disulfide bonds found in the alpha-domain of authentic lysozyme, had amounts of secondary and tertiary structures, and bacteriolytic activity comparable to those of authentic lysozyme, and showed a cooperative thermal unfolding. By contrast, the other species, 2SS [64-80, 76-94], which contained the beta-domain disulfide bond as well as the inter-domain one, had a limited amount of secondary structure and little tertiary structure. Disulfide-exchange did not occur for 2SS [6-127, 30-115], whereas it occurred for 2SS [64-80, 76-94], indicating that the protein main-chain fold coupled with the formation of two disulfide bonds is relatively stable for the former variant, while unstable for the latter. 1H-NMR spectra of 2SS [6-127, 30-115] showed that native-like local environment is present within the region that corresponds to the alpha-domain, while it is absent within the region that corresponds to the beta or inter-domain. These results indicate that the alpha-domain of hen lysozyme can be an independent folding domain at equilibrium. Although the bipartite nature in the structure formation of hen lysozyme is similar to that reported for alpha-lactalbumin, differences exist between the disulfide-intermediates of the two proteins in terms of the structural domain that accomplishes tertiary structure.

Efficacy of transthoracic needle aspiration in community-acquired pneumonia.

OBJECTIVE: To evaluate the indications, efficacy, and safety of transthoracic needle aspiration (TNA) in diagnosing community-acquired pneumonia (CAP). METHODS: TNA procedure was performed using an ultrathin needle with ultrasonography and/or computed tomography. The aspirate samples were Gram-stained and sent for cultures. The results were compared with those from conventional microbiological studies. PATIENTS: Sixty patients with CAP who were admitted to the hospital and were studied prospectively between July 1994 and June 1999 were included in the study. RESULTS: TNA culture was positive in 30 cases (50.0%). Streptococcus pneumoniae was the most frequently isolated pathogen, followed by the Streptococcus milleri group, and anaerobes. The results of TNA were consistent with those of quantitative sputum cultures in 9 patients and with those of blood cultures in 4. Complications arose in 3 patients who developed small to moderate pneumothorax. CONCLUSIONS: TNA is a safe procedure with a good diagnostic yield. In particular, anaerobes or microaerophils such as the S. milleri group were highly detectable by TNA. The results obtained by TNA were highly consistent with those obtained by the gold standard methods. Combined with conventional methods, TNA is considered highly useful for determining the etiology of CAP.

Recombinant expression of perchloric acid-soluble protein reduces cell proliferation.

Perchloric acid-soluble protein (PSP) may play an important role in the regulation of cellular physiological functions because it has been highly conserved throughout evolution; however, this role has not been well elucidated. In previous reports, we suggested that PSP regulates cell proliferation. In this study, we examined the effect of PSP expression on proliferation of the normal rat kidney cell line NRK-52E, the rat hepatocyte cell line RLN-10, and the rat hepatoma cell line dRLh-84. Cells transfected with pcDNA-sense-PSP (pcDNA-S-PSP) over-expressed PSP mRNA and protein, and cell proliferation of the transfected cells was suppressed compared with that of cells transfected with pcDNA-empty (pcDNA-E). Cell viability of pcDNA-S-PSP-transfected cells was similar to that of pcDNA-E-transfected cells. Thus, over-expression of PSP suppresses cell proliferation without any influence on cell viability. These findings are the first to report an inhibitory activity of PSP on cell proliferation.

Inhibition of environmental estrogen-induced proliferation of human breast carcinoma MCF-7 cells by flavonoids.

In the present study, we evaluated the individual and combined effects of environmental estrogens and flavonoids on the proliferation of human breast carcinoma MCF-7 cells. These compounds are as follows: (1) pharmaceutical chemicals such as diethylstilbestrol, 17alpha-ethynylestradiol (17ES), tamoxifen, mestranol, and clomiphene, (2) industrial chemicals such as bisphenol A (BisA), 4-octylphenol (OP), 4-nonylphenol (NP), and p,p'-biphenol, and (3) flavonoids such as daidzein (D), genistein (G), quercetin (Q), and luteolin (L). We found that nanomolar concentrations of 17ES, BisA, OP, and NP were sufficient to stimulate the proliferation of MCF-7 cells. Among them, 1 microM BisA exhibited cell proliferation-stimulating activity as strong as 10 nM 17beta-estradiol; and D and G exhibited cell proliferation-stimulating activity at 10 nM. On the other hand, Q and L exhibited cell proliferation-inhibiting activity. We also found that 10 nM flavonoids, such as D, G, Q, and L, were able to inhibit the proliferation-stimulating activity in MCF-7 cells by 1 microM environmental estrogens.

Intermediate subunit of the Gal/GalNAc lectin of Entamoeba histolytica is a member of a gene family containing multiple CXXC sequence motifs.

Killing by Entamoeba histolytica requires parasite adherence to host galactose- and N-acetyl-D-galactosamine (Gal/GalNAc)-containing cell surface receptors. A 260-kDa heterodimeric E. histolytica Gal/GalNAc lectin composed of heavy (Hgl) and light (Lgl) subunits has been previously described. Here we present the cloning and characterization of Igl, a 150-kDa intermediate subunit of the Gal/GalNAc lectin. Igl, Hgl, and Lgl colocalized on the surface membrane of trophozoites. Two unlinked copies of genes encoding Igl shared 81% amino acid sequence identity (GenBank accession no. AF337950 and AF337951). They encoded cysteine-rich proteins with amino- and carboxy-terminal hydrophobic signal sequences characteristic of glycosylphosphatidylinositol (GPI)-anchored membrane proteins. The igl genes lacked carbohydrate recognition domains but were members of a large family of amebic genes containing CXXC and CXC motifs. These data indicate that Igl is part of the parasite's multimolecular Gal/GalNAc adhesin required for host interaction.

Enhanced cardiac L-type calcium current response to beta2-adrenergic stimulation in heart failure.

The beta2-adrenergic receptor (beta2-AR)-mediated increase in cardiac L-type Ca2+ current (I(Ca,L)) has been documented in normal subjects. However, the role and mechanism of beta2-AR activation on I(Ca,L) in heart failure (HF) are unclear. Accordingly, we compared the effect of zinterol (ZIN), a highly selective beta2-AR agonist, on I(Ca,L) in isolated left ventricular cardiomyocytes obtained from normal control and age-matched rats with HF induced by left coronary artery ligation (4 months). I(Ca,L) was measured by using the whole-cell voltage-clamp technique. In normal myocytes, superfusion of ZIN (10(-5) M) caused a 21% increase in I(Ca,L) (9.21 +/- 0.24 versus 7.59 +/- 0.20 pA/pF) (p < 0.05). In HF myocytes, the same concentration of ZIN produced a significantly greater increase (30%) in I(Ca,L) (6.20 +/- 0.24 versus 4.75 +/- 0.17 pA/pF) (p < 0.01). This ZIN-induced increase in I(Ca,L) was further augmented in both normal and HF myocytes (normal: 59 versus 21%; HF: 71 versus 30%) after the incubation of myocytes with pertussis toxin (PTX, 2 microg/ml, 36 degrees C, 6 h). These effects were not modified by the incubation of myocytes with CGP-20712A (3 x 10(-7) M), a beta1-AR antagonist, but were abolished by pretreatment of myocytes with ICI-118551 (10(-7) M), a beta2-AR antagonist. In addition, all of the effects induced by ZIN were completely prevented in the presence of an inhibitory cAMP analog, Rp-cAMPS (100 microM, in the patch-pipette solution). In conclusion, beta2-AR activation stimulates L-type Ca2+ channels and increases I(Ca,L) in both normal and HF myocytes. In HF, beta2-AR activation-induced augmentation of I(Ca,L) was increased. These effects are likely to be mediated through a cAMP-dependent mechanism and coupled with both stimulatory G protein and PTX-sensitive G protein.