Verification of the Usefulness of an Assessment and Risk Control Sheet that Promotes Management of Cancer Drug Therapy.

Background: Proper management of adverse events is crucial for the safe and effective implementation of anticancer drug treatment. Showa University Hospital uses our interview sheet (assessment and risk control [ARC] sheet) for the accurate evaluation of adverse events. On the day of anticancer drug treatment, a nurse conducts a face-to-face interview. As a feature of the ARC sheet, by separately describing the symptoms the day before treatment and the day of treatment and sharing the information on the medical record, it is possible to clearly determine the status of adverse events. In this study, we hypothesized that the usefulness and points for improvement of the ARC sheet would be clarified by using and evaluating a patient questionnaire. Methods: This study included 174 patients (144 at Showa University Hospital (Hatanodai Hospital) and 30 at Showa University Koto Toyosu Hospital (Toyosu Hospital) who underwent pre-examination interviews by nurses and received cancer chemotherapy at the outpatient center of Hatanodai and Toyosu Hospital. In the questionnaire survey, the ARC sheet's content and quality, respondents' satisfaction, structural strengths, and points for improvement were evaluated on a five-point scale. Results: The patient questionnaire received responses from 160 participants, including the ARC sheet use group (132 people) and the non-use group (28 people). Unlike the ARC sheet non-use group, the ARC sheet use group recognized that the sheet was useful to understand the adverse events of aphthous ulcers (p = 0.017) and dysgeusia (p = 0.006). In the satisfaction survey questionnaire, there was a high sense of security in the pre-examination interviews by nurses using the ARC sheet. Conclusions: The ARC sheet is considered an effective tool for comprehensively evaluating adverse events. Pre-examination interviews by nurses using ARC sheets accurately determined the adverse events experienced by patients with anxiety and tension due to confrontation with physicians.

Diagnostic value of capsule-like rim enhancement on magnetic resonance imaging for distinguishing malignant from benign parotid tumours.

The purpose of this study was to clarify the diagnostic value of capsule-like rim enhancement (CLRE) on magnetic resonance imaging (MRI) for distinguishing malignant from benign tumours of the parotid gland. We retrospectively evaluated contrast-enhanced T1-weighted images of 100 patients with malignant and benign parotid tumours for the presence, completeness, and irregularity of CLRE and its maximum thickness. We investigated any correlation of imaging and histopathological findings for 51 cases showing CLRE with available histology. The presence and completeness of CLRE did not differ significantly between benign and malignant tumours. Malignant tumours had more irregular CLRE than benign tumours (P<0.05). The mean CLRE thickness was significantly greater for malignant (2.4 mm) than benign tumours (1.4 mm) (P<0.0001). The two types of tumour were most accurately distinguished using a cut-off value of 1.5 mm thickness. Histopathology demonstrated the general correspondence of thick CLRE on MRI in malignant tumours with thick but sparse fibrous tissue and infiltration of tumour cells and lymphocytes, whereas thin CLRE in benign tumours typically represented dense fibrous tissue without infiltration of tumour cells. CLRE was more irregular and thicker in malignant tumours than in benign tumours, which may be of help in differentiating them.

Bach1 is critical for the transformation of mouse embryonic fibroblasts by Ras(V12) and maintains ERK signaling.

Reactive oxygen species (ROS), by-products of aerobic respiration, promote genetic instability and contribute to the malignant transformation of cells. Among the genes related to ROS metabolism, Bach1 is a repressor of the oxidative stress response, and a negative regulator of ROS-induced cellular senescence directed by p53 in higher eukaryotes. While ROS are intimately involved in carcinogenesis, it is not clear whether Bach1 is involved in this process. We found that senescent Bach1-deficient mouse embryonic fibroblasts (MEFs) underwent spontaneous immortalization the same as did the wild-type cells. When transduced with constitutively active Ras (H-Ras(V12)), the proliferation and colony formation of these cells in vitro were markedly reduced. When transplanted into athymic nude mice, the growth and vascularization of tumors derived from Bach1-deficient cells were also decreased. Gene expression profiling of the MEFs revealed a new H-Ras(V12) signature, which was distinct from the previously reported signatures in epithelial tumors, and was partly dependent on Bach1. The Bach1-deficient cells showed diminished phosphorylation of MEK and ERK1/2 in response to H-Ras(V12), which was consistent with the alterations in the gene expression profile, including phosphatase genes. Finally, Bach1-deficient mice were less susceptible to 4-nitroquinoline-1-oxidide (4-NQO)-induced tongue carcinoma than wild-type mice. Our data provide evidence for a critical role of Bach1 in cell transformation and tumor growth induced by activated H-Ras(V12).

A case of cerebral aquaporinopathy.

A 35-year-old woman was hospitalized due to impaired consciousness. Magnetic resonance imaging (MRI) revealed multiple parenchymal lesions in supra and infratentorial brain regions, which were considered responsible for her declining consciousness level. She was treated with intravenous methylprednisolone. Neurological symptoms improved and she was discharged. She was readmitted 14 months later due to intractable hiccups. A follow-up brain MRI revealed an abnormal signal near the area postrema in the dorsal medulla. Serum aquaporin-4 antibody levels were positive, but there were no visual manifestations or myelitis. Spinal MRI was negative for longitudinally extended transverse myelitis throughout the clinical course.

Dimethyl sulphoxide relaxes rabbit detrusor muscle by decreasing the Ca2+ sensitivity of the contractile apparatus.

BACKGROUND AND PURPOSE: The intravesical administration of dimethyl sulphoxide (DMSO) is used to alleviate the symptoms of interstitial cystitis. We investigated the relaxant effect of DMSO and its underlying mechanism in the detrusor muscle. EXPERIMENTAL APPROACH: The effects of DMSO on contraction, on Ca2+ sensitivity of myofilaments, and on myosin light chain (MLC) phosphorylation were investigated in both intact and alpha-toxin-permeabilized strips of rabbit detrusor muscle. KEY RESULTS: In fura-PE3-loaded strips, DMSO (>1%) induced a significant relaxation during sustained contractions induced by 60 mM K+-depolarization or 10 microM carbachol, while having no effect on the [Ca2+](i) level. DMSO decreased the level of MLC phosphorylation during the contractions induced by 60 mM K+ and 10 microM carbachol. DMSO also inhibited both the contraction and MLC phosphorylation induced by calyculin-A in intact strips. In the alpha-toxin-permeabilized preparations, DMSO relaxed the Ca2+-induced contraction and also inhibited the tension development induced by a stepwise increment of Ca2+ concentrations. Such a relaxant effect of DMSO was enhanced in the presence of phosphate. CONCLUSIONS AND IMPLICATIONS: DMSO relaxes rabbit detrusor muscle by decreasing the Ca2+ sensitivity of myofilaments. Inhibition of the kinase activities involved in myosin phosphorylation may play a major role in DMSO-induced Ca2+ desensitization. Inhibition of the cross-bridge cycling at the step of phosphate release may also contribute to the relaxant effect of DMSO. Such relaxant effects of DMSO could be linked to the therapeutic effect of DMSO in interstitial cystitis.

Presence of Streptococcus infection in extra-oropharyngeal head and neck squamous cell carcinoma and its implication in carcinogenesis.

To investigate the involvement of S. anginosus infection in head and neck cancer in the extra-oropharyngeal cavity, we analyzed 3 DNA samples prepared from squamous cell carcinoma of the external auditory canal and one from squamous cell carcinoma of the skin using polymerase chain reaction (PCR) analysis and Southern blot analysis to detect the DNA sequence of S. anginosus. We also examined these four specimens by Gram's stain to detect the streptococcal bacterial bodies. By PCR analysis, the DNA sequence of S. anginosus was found in 4 out of 4 (100%) DNA samples obtained from these tumors. By Southern blot analysis, positive bands were detected in one out of the 3 (33%) samples from the tumor taken from the external auditory canal. We detected streptococcal bacterial bodies in one of the three specimens from the tumor obtained from cancer of the external auditory canal and in the one specimen from the skin cancer by the method of Gram's stain. Contrary to our expectations, these bacterial bodies were located in the middle of the tumor. Since S. anginosus is thought to exist in the mouth as a normal flora and to be located mainly in the gingiva and dental plaque, these data strongly indicate that S. anginosus infection is implicated in the carcinogenesis of head and neck squamous cell carcinoma.

A clinical study of oral tongue cancer.

Thirty-nine previously untreated patients with squamous cell carcinoma of the oral tongue treated with curative intent in our hospital from 1993 through 1998 are reviewed. Of these patients, those in the early stage (stages I and II) constituted 64%. The over all 5-year survival rate of all the patients was 60%. The 5-year survival rate of the patients with early stage cancers was unsatisfactory (stage I: 73%; II: 56%). This was thought to be related to the absence of elective neck dissection and the administration of chemotherapy in the patients with early stage cancer. We concluded that elective neck dissection for levels I, II and III is the first choice of treatment strategy for patients with stage II cancer. Our data indicate that chemotherapy in patients with early stage cancer was not beneficial and might have increased the risk of late lymph node metastasis in the clinically NO patients without neck dissection. There were 9 patients younger than 40 years of age and their survival rate at 5 years was 80%, which was better than that of the older patients. The treatment strategy for patients younger than 40 years of age was similar to that of older patients.

Streptococcus anginosus in head and neck squamous cell carcinoma: implication in carcinogenesis.

It has been suggested that Helicobacter pylori (H. pylori) infection might be associated with not only gastric ulcers but also gastric malignancies. Recently, it was reported that the Streptococcus anginosus (S. anginosus) DNA sequence was found in DNA samples extracted from esophageal cancers. Because smoking and alcohol abuse are regarded as risk factors for both esophgeal cancer and head and neck cancer, infection of S. anginosus might be associated with carcinogenesis of head and neck cancer. To investgate the involvement of S. anginosus infection in head and neck cancer, we analyzed 217 DNA samples prepared from head and neck squamous cell carcinomas. We performed PCR analysis with S. anginosus-16S ribosomal DNA-specific primers, and Southern blot analysis. For detection of S. anginosus in the oral and pharyngeal cavities, we used oropharyngeal bacteriological culture and PCR analysis of gingival smears of the patients. By PCR analysis, the S. anginosus DNA sequence was found in 217 out of 217 (100%) DNA samples obtained from head and neck cancers. By Southern blot analysis, positive bands were detected in 41 out of 125 (33%) samples. We could find no S. anginosus colony in oropharyngeal bacteriological culture dishes of 53 patients with and without head and neck cancer. On the other hand, we found the S. anginosus DNA fragment in 8 out of 8 DNA samples obtained from gingival smears by PCR analysis. These data indicate that the upper aerodigestive environment of the patients permitting S. anginosus infection was implicated in the carcinogenesis of head and neck squamous cell carcinoma.

[An adult case of asymptomatic congenital tracheal stenosis].

We report an adult case of asymptomatic congenital tracheal stenosis. A 42-year-old woman was admitted to our hospital, whose chief complaint was tongue pain. Clinical examination revealed a tumor 35 x 20 mm in diameter along the middle, right edge of the tongue, and histopathological examination determined a diagnosis of squamous cell carcinoma of the tongue. She had no history of dyspnea, stridor nor recurrent pneumonia, and enjoyed playing table tennis in her daily life. Physical examination, electrocardiogram, spirogram and laboratory data showed no abnormal signs although chest roentgenogram revealed a narrowed trachea 6 mm in diameter, however, the tracheal stenosis was not noted at the time. On the 10th hospital day, radical operation for cancer of the tongue including right hemiglossectomy and supraomohyoid neck dissection, were performed. Oral intubation was not successful, and a tracheostomy was conducted. The trachea was incised, and revealed that complete tracheal rings existed without posterior membraneous tissue. The operation time was limited because of high airway pressure for a narrow tracheal tube (5 mm in diameter). The main postoperative complication was one crust formation in the tracheal lumen that resulted in CO2 narcosis due to check valve obstruction of the trachea on the 4th postoperative day. A nebulizer provided continuous moisture and was effective in preventing recurrence of the crust attachment to the tracheal membrane thus, the tracheostoma was closed by the 17th postoperative day. This case indicates that among healthy people there are extremely rare patients with congenital tracheal stenosis who survive their postnatal and infantile periods, and who experience no trouble in their daily lives. As physicians, we must be aware of this disease in adult patients in our clinics.

Mutation of the sterol 27-hydroxylase gene (CYP27) results in truncation of mRNA expressed in leucocytes in a Japanese family with cerebrotendinous xanthomatosis.

OBJECTIVES: A Japanese family with cerebrotendinous xanthomatosis (CTX) was investigated for a sequence alteration in the sterol 27-hydroxylase gene (CYP27). The expression of CYP27 has been mostly explored using cultured fibroblasts, prompting the examination of the transcripts from blood leucocytes as a simple and rapid technique. METHODS: An alteration in CYP27 of the proband was searched for by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis and subsequent sequencing. Samples of RNA were subjected to reverse transcription PCR (RT-PCR) and the product of the proband was amplified with nested primers and sequenced. RESULTS: A homozygous G to A transition at the 5' end of intron 7 was detected in the patient. In RT-PCR analysis, only a truncated transcript was detected in the patient, whereas both normal and truncated transcripts were detected in the siblings. The sequencing of the patient's cDNA fragment disclosed a direct conjuction of exon 6 and exon 8. CONCLUSION: The mutation at splice donor site and the truncation of mRNA were identical with those of a recently reported Italian patient, although different in symptomatology. The application of blood leucocytes can be a simple technique on analysing a constructive abnormality of CYP27 mRNA.

[Superselective intra-arterial infusion chemotherapy of high-dose cisplatin for advanced paranasal sinus carcinomas].

Eighteen patients with advanced paranasal sinus carcinomas were treated by "two-route" intra-arterial chemotherapy using cis-diamminedichloroplatinum (CDDP) and sodium thiosulfate (STS) to preserve the hard palate and the eye. In these patients, 100 mg/m2 of CDDP was administered weekly through each feeding artery of the tumor superselectively at 5 mg/min. During infusion of CDDP, STS at a two-hundred fold dose of CDDP was injected through a catheter placed in the brachiocephic vein introduced via the subclavian vein. The complete and partial response rates were 14/18 (78%) and 4/18 (22%), respectively. None of the nine patients following operation showed residual tumors histologically. The peak of the mean total plasma platinum concentration was 5.5 micrograms/ml, and this concentration was rapidly reduced to 1.5 micrograms/ml in 5 hours. The peak of the plasma protein unbound platinum was 3.9 micrograms/ml. This concentration rapidly decreased to almost zero within 5 hours after IA infusion. The mean tumor platinum content achieved by superselective IA infusion was as high as 6.0 micrograms/g tumor, and this decreased rapidly to 2.4 micrograms/g tumor on the 5th day after the 1st intra-arterial infusion. All patients were free from chemotoxicity such as renal, hematological dysfunctions, or gastrointestinal symptoms. Each chemotherapy treatment could be done weekly on schedule. All but one patient was alive for 5-40 months. This new method of chemotherapy appears very effective for advanced paranasal sinus carcinomas.

[Loss of heterozygosity of 3p21 and 9p21 in head and neck squamous cell carcinomas and its prognostic implication].

To examine whether genetic factors influence the prognosis of cancer patients, several microsatellite markers were used to determine the allelic loss of certain areas of the genome. Three microsatellite markers, D3S1067, IFNA and D9S171 were used to study the loss of heterozygosity (LOH) of 3p21 and 9p21 in 93 head and neck squamous cell carcinomas. Of 57 informative cases, LOH was detected in 27 of 57 (47%) DNA samples obtained from cancer specimens when at least one marker was used. The frequency of LOH was not correlated with the clinical factors. However, the frequency of LOH was significantly higher in the recurrent cases than in the non-recurrent cases, and patients with 3p21 and/or 9p21 LOH tended to survive for a shorter period of time. These results suggested that the allelic loss at 3p21 and/or 9p21 could be correlated with the prognosis of the patients, and that it was a novel prognostic factor independent of other clinical factors concerning head and neck cancers. LOH at 3p21 and/or 9p21 may help to identify head and neck cancer patients with a poor prognosis, who need an intensive postoperative follow-up protocol, or who are suitable for novel investigational therapeutic approaches.

Loss of heterozygosity of chromosome 9p21 and 7q31 is correlated with high incidence of recurrent tumor in head and neck squamous cell carcinoma.

To examine whether genetic factors influence the prognosis of cancer patients, several polymorphic markers were used to determine the allelic loss of certain areas of the genome. Two polymorphic markers, IFNA and D9S171 were used to study the loss of heterozygosity (LOH) of chromosome 9p21 in 75 head and neck squamous cell carcinomas. LOH was detected in 14 out of 64(22%) DNA samples obtained from cancer specimens when at least one marker was used. The frequency of LOH was not correlated with the localization of the tumor, clinical stage of the patient, tumor size and lymph node involvement. However, the frequency of LOH was significantly higher in the recurrent tumors than in the non-recurrent tumors, suggesting that the allelic loss at 9p21 can be correlated with the short term prognosis of the patients. LOH was identified in only three out of 19(16%) samples when D7S522 was used as a marker. However, all of these three cases were recurrent, and two of the three showed the allelic loss at 9p21 at the same time. These results indicate that LOH of 9p21 and/or 7q31 is a novel prognostic factor independent of other clinical factors for head and neck squamous cell carcinoma. Replication error (RER) was observed in 4 cancers, implicating genetic instability in the carcinogenesis of a subset of head and neck squamous cell carcinoma.

In vitro assembly of FAD, AMP, and the two subunits of electron-transferring flavoprotein: an important role of AMP related with the conformational change of the apoprotein.

Electron-transferring flavoprotein from pig kidney is composed of four non-covalently bound components: alpha and beta subunits, flavin adenine dinucleotide (FAD), and adenosine monophosphate (AMP). This paper reveals the pathway of assembly of the electron-transferring flavoprotein. The holoprotein can be formed by two different pathways. (i) alpha + beta <==> (alpha-beta)*, (alpha-beta)* + AMP <==> (alpha-beta-AMP)*, (alpha-beta-AMP)* <==> alpha-beta-AMP, alpha-beta-AMP + FAD <==> holoprotein. (ii) alpha + beta <==> alpha-beta, alpha-beta + FAD <==> alpha-beta-FAD, alpha-beta-FAD + AMP <==> holoprotein. Here the presence and absence of asterisks distinguish different conformations with the same composition. The monomeric forms of alpha and beta showed no significant binding with FAD and AMP. AMP and FAD associated with different heterodimer forms which were formed as a result of weak binding between alpha and beta. The binding of alpha + beta + AMP was much faster than that of alpha + beta + FAD because the rate of alpha + beta --> (alpha-beta)* was much faster than that of alpha + beta --> alpha-beta. The alpha-beta-AMP complex associated with FAD rapidly. As a result, the binding of FAD with the subunits is promoted by AMP. The alpha-beta-FAD complex associated with AMP much more slowly than the mixture of alpha and beta. Thus the AMP binding with the subunits is inhibited by the preceding FAD binding.

Preparation of poly(D,L-lactide) and copoly(lactide-glycolide) microspheres of uniform size.

In an attempt to prepare monodisperse poly(D,L-lactide) and copoly(lactide-glycolide) microspheres, a novel emulsification technique (membrane emulsification) was employed and the preparation conditions which might affect the monodispersity were evaluated. With this technique nearly monodisperse poly(D,L-lactide) and copoly(lactide-glycolide) microspheres were successfully prepared and their sizes were controllable only by making use of microporous glass membranes of different pore sizes. However, in the present system of emulsion (methylene chloride/water) the surfactant used was limited to ionic ones and the amount of polymers available for the formation of microspheres was inevitably too small in concentration to entrap sufficient amounts of drug. As for the drug release, the effect of particle size was not appreciable but the method of solvent removal gave a great influence; the solvent extraction method showed a more drug-sustaining effect than did the solvent evaporation method. The present results suggest the possibility of making drug-loaded and biodegradable monodisperse microspheres.

In vitro refolding and unfolding of subunits of electron-transferring flavoprotein: characterization of the folding intermediates and the effects of FAD and AMP on the folding reaction.

Electron-transferring flavoprotein (ETF) from pig kidney is composed of two subunits (alpha and beta, molecular weights of 33,000 and 29,000) and two small molecules, FAD and AMP. In this study, in vitro refolding and unfolding of the subunits of ETF were carried out with urea as the denaturing reagent. The refolding reaction of alpha and beta was revealed to proceed kinetically in two steps: D in equilibrium with I-->N, where D,I, and N denote the denatured, intermediate, and native forms, respectively. The features of the I forms of alpha and beta, described below, are consistent with the concept of the so-called "molten globule state," which is frequently observed in protein refolding. (i) The conversion between D and I was very rapid. (ii) The I form showed as much secondary structure as the N form as judged from the far-UV circular dichroism. (iii) The solvent accessibility of the I form, estimated by the analysis of equilibrium unfolding experiments, was intermediate between those of the D and N forms. (iv) The standard free energy of the I form is almost the same as that of the D form. The refolding reaction progressed more slowly and the environment of the tryptophan chromophore was changed more drastically in beta refolding that in alpha refolding. We previously reported that the reconstitution of holoETF from denatured subunits is speeded up by increasing the AMP concentration. In this study, the effects of AMP, FAD, and the other subunit on the single subunit folding were examined, but no effect was detected. This result suggests that AMP plays a role in a later process, namely, assembly of the four components (refolded alpha and beta, FAD, and AMP).

Abnormalities and the implication of retinoblastoma locus and its protein product in head and neck cancers.

Abnormalities in the Retinoblastoma tumor suppressor gene (Rb) have been observed in a large number of human cancers. Loss of heterozygosity (LOH) is a common mode of allelic inactivation of Rb and other tumor suppressor genes. We investigated DNA from 45 primary human head and neck cancers to determine LOH at the Rb locus using a polymerase chain reaction-based restriction fragment length polymorphism assay. Of informative cases, we found LOH in 4 of 28 (14%) head and neck cancers. Of immunohistochemical staining of Rb protein, we found that in one of ten LOH negative cases the nuclei of fibroblasts were stained with anti-Rb antibody but there was no nuclear staining tumor cells. These results suggest that inactivation of Rb protein is involved in the carcinogenesis of head and neck cancer at all levels of the process of protein expression: DNA, mRNA and protein.