[Penile skin metastasis of gastric carcinoma associated with prostate carcinoma: a case report].

Skin metastasis of internal carcinoma is a rare situation and its risk is reported as 0.7-9%. The site of skin metastasis is more popular at upper part of the body such as neck and face. We report a case of perineal and penile skin metastases of gastric carcinoma associated with prostatic carcinoma. A 72-year-old man, who underwent total gastrectomy for gastric carcinoma 4 years ago, was found to have sclerotic change at perineal and penile skin. As his serum PSA level was 10.6 ng/ml, transrectal prostate biopsy and penile skin biopsy were performed. The prostate tissue pathologically demonstrated moderately differentiated adenocarcinoma and it was positive for both anti-PSA and anti-CEA antibody by immunohistochemical staining. The skin tissue was found to be infiltrative adenocarcinoma, negative for PSA and positive for CEA, which was compatible with the primary gastric carcinoma specimen. The patient had been treated for 7 months with administration of Doxifluridine and injection of LH-RH agonist, but died for progression of gastric carcinoma. A risk of skin metastasis of gastric carcinoma is reported as 6%, however, its metastasis to perineal and penile skin is the first case reported in the literature.

[Intermittent administration of 5-FU and isovorin to patients with advanced and recurrent colon cancer].

We attempted a new regimen of intermittent administration of 5-FU and low-dose Isovorin (F.I) to four patients with advanced and recurrent colon cancer. A partial response (PR) was achieved in two of four patients who had evaluable lesions for this treatment. We observed few side effects among these patients. Only one patient among four showed grade 1 neuropathy after two administrations of this chemotherapy. However, after a two-week pause in administration, the neuropathy disappeared and we could continue the therapy. This patient with multiple liver metastases achieved a partial response. Other patients had no side effects such as bone marrow suppression or digestive symptoms. This intermittent F.I treatment might be an effective and promising therapy with few side effects even for patients with poorer conditions.

Brachytherapy for prostate cancer in Japan.

The temporary implantation of iridium-192 (Ir-192) is available in Japan if the patient is isolated in a radiation control area during the treatment. In this study, brachytherapy for prostate cancer was preceded by using low-dose rate Ir-192. Twenty-six patients with localized prostate cancer were treated with brachytherapy since December 1997. The age of patients ranged from 61 to 82 (median 76) years and the initial serum prostate-specific antigen (PSA) level ranged from 4.3 to 141 (median 17.4) ng/ml. Ten to 14 needles with Teflon sheaths were introduced into the prostate via a transperineal approach and an Ir-192 wire was placed in each sheath. The dose-distribution curve based on computed tomography imaging was drawn and the irradiation time was calculated for the whole prostate to receive over 70 Gy. During the 1-22-month follow-up period, all cases with an initial PSA level of less than 25 ng/ml showed continuous PSA decrease without biochemical failure. Magnetic resonance imaging demonstrated tumor regression in all cases within 6 months after the treatment. One-year biopsy specimens from patients with PSA failure demonstrated viable tumor cells, and the specimens from patients with favorable progress revealed fibrosis replacement without any variable tumor cells. Characterization of nocturnal penile tumescence data and IIEF 15 score proved the minimal effect of the treatment on erectile function. No major side effects or complications were observed. Brachytherapy using Ir-192 is a less invasive and safe procedure, which is expected to become a standard treatment for localized prostate cancer patients with relatively low PSA levels.

[The brachytherapy with low dose-rate iridium for prostate cancer].

PURPOSE: Brachytherapy as an option for the treatment of prostate cancer has been commonly performed in USA. As the permanent seeding of the radioactive materials is strictly restricted by the law in Japan, brachytherapy must be performed by the temporary implant. This treatment has been performed at a few facilities in Japan mostly using high dose-rate iridium. Only our facility has been using low dose-rate iridium (LDR-Ir) for prostate cancer. This study evaluates the clinical results of the treatment. PATIENTS AND METHODS: Since December 1997 to December 1999, 26 patients with histologically diagnosed as prostate cancer (Stage B, 92%; Stage C, 8%) underwent brachytherapy. Twenty-two patients received brachytherapy alone, three were treated with a combination of brachytherapy and external beam radiotherapy (ERT) and one was treated with a combination of brachytherapy and neoadjuvant endocrine therapy. Patients ranged in age from 61 to 84 (median 76) years old. Treatment was initiated with perineal needle placement. From 10 to 14 needles were placed through the holes on the template which was fixed to the stabilizer of the transrectal ultrasound probe. After the needle placement. CT scan was performed to draw distribution curves for the treatment planning. LDR-Ir wires were introduced to the sheath and indwelled during the time calculated from dosimetry. Peripheral dose was 70 Gy for the monotherapy of brachytherapy. For the combination therapy, 40 Gy was given by brachytherapy and 36 Gy with ERT afterwards. LDR-Ir wires were removed after completion of the radiation and patients were followed with serum PSA level and annual biopsy. RESULTS: During 2 to 26 (median 12) months follow-up, 8 out of 9 patients with initial PSA level above 20 ng/ml showed PSA failure. All 13 patients with initial PSA level lower than 20 ng/ml were free from PSA failure. Eight out of 11 patients with Gleason's score 7 or higher showed PSA failure, and all 14 patients (including three patients with combined therapy) with Gleason's score 6 or less were free from PSA failure. Annual biopsy was performed in 8 patients, and 4 patients histologically revealed tumor free and 4 patients (two of them showed PSA failure) were tumor positive. No major complication was observed, however, some minor side-effect as irritability was seen in 65% of the patients. CONCLUSIONS: The results showed that brachytherapy with LDR-Ir was an acceptable treatment as long as the patients were selected strictly with PSA level and Gleason's score. A good candidate for this treatment is the patient whose PSA level is lower than 20 ng/ml and Gleason's score is 6 or less. The treatment is effective and safe, but further observation is necessary to reach the conclusion.

In vivo fluorescence imaging of the transport of charged chlorin e6 conjugates in a rat orthotopic prostate tumour.

Polymeric drug conjugates are used in cancer therapy and, varying their molecular size and charge, will affect their in vivo transport and extravasation in tumours. Partitioning between tumour vasculature and tumour tissue will be of particular significance in the case of photosensitizer conjugates used in photodynamic therapy, where this partitioning can lead to different therapeutic effects. Poly-l-lysine chlorin e6conjugates (derived from polymers of average Mr 5000 and 25000) were prepared both in a cationic state and by poly-succinylation in an anionic state. A fluorescence scanning laser microscope was used to follow the pharmacokinetics of these conjugates in vivo in an orthotopic rat prostate cancer model obtained with MatLyLu cells. Fluorescence was excited with the 454-528 nm group of lines of an argon laser and a 570 nm long pass filter used to isolate the emission. Results showed that the conjugates initially bound to the walls of the vasculature, before extravasating into the tissue, and eventually increasing in fluorescence. The anionic conjugates produced tissue fluorescence faster than the cationic ones, and surprisingly, the larger Mr conjugates produced tissue fluorescence faster than the smaller ones with the same charge. These results are consistent with differences in aggregation state between conjugates.

Transient absorption changes in vivo during photodynamic therapy with pulsed-laser light.

High intensity pulsed-laser light can be used to excite absorbing molecules to transient states in large proportions. The laser-induced spectral changes can be characterized by transient changes in light propagation; through the tissue provided the excited states of these molecules have altered absorption spectra. Characterization of these transient changes may then be used to exploit new mechanisms in photosensitization and/or to optimize photobiological effects. In this study, transmittance and reflectance were measured as a function of laser pulse energy, from tissue-simulating media as well as in rat muscle and liver slices, both with and without the photosensitizer benzoporphyrin derivative monoacid (BPD-MA) present. There was a transient decrease in absorption from the photosensitizer at peak pulse irradiance in the range of 100-1000 W cm(-2). The depth of photodynamic treatment-induced tissue necrosis was measured in a subcutaneous prostate cancer model in Copenhagen rats. A comparison between continuous wave irradiation and pulsed irradiation with the same average incident irradiance showed no statistically significant difference in the depth of necrosis at 48 h after irradiation. These results indicate that photosensitizer population-state changes are measurable in tissues and may provide a method for measuring triplet-state properties of photosensitizer in vivo, but for BPD-MA at clinically used concentrations these changes do not significantly affect the depth of photodynamically-induced tissue damage.

Isolation and characterization of the ATF2 gene encoding alcohol acetyltransferase II in the bottom fermenting yeast Saccharomyces pastorianus.

The ATF2 gene encodes alcohol acetyltransferase II, which catalyses the synthesis of isoamyl acetate from acetyl coenzyme A and isoamyl alcohol. To characterize the ATF2 gene from the bottom fermenting yeast Saccharomyces pastorianus, the S. pastorianus ATF2 gene was cloned by colony hybridization using the S. cerevisiae ATF2 gene as a probe. When an atf1 null mutant strain was transformed with a multi-copy plasmid carrying the S. pastorianus ATF2 gene, the AATase activity of this strain was increased by 2.5-fold compared to the control. The S. pastorianus ATF2 gene has 99% nucleic acid homology in the coding region and 100% amino acid homology with the S. cerevisiae ATF2 gene. Southern blot analysis of chromosomes separated by pulse-field gel electrophoresis indicated that the ATF2 gene probe hybridized to chromosome VII in S. cerevisiae and to the 1100 kb chromosome in S. pastorianus. As S. pastorianus is thought to be a hybrid of S. cerevisiae and S. bayanus, the S. bayanus-type gene, which has a relatively low level of homology with the S. cerevisiae-type gene, is also usually detected. Interestingly, an S. bayanus-type ATF2 gene could not be detected. These results suggested that the cloned ATF2 gene was derived from S. cerevisiae. Analysis using an ATF2-lacZ fusion gene in S. pastorianus showed that expression of the ATF2 gene was relatively lower than that of the ATF1 gene and that it is repressed by aeration but activated by the addition of unsaturated fatty acids. The S. pastorianus ATF1, Lg-ATF1 and ATF2 Accession Numbers in the DDBJ Nucleotide Sequence Database are D63449, D63450 and D86480, respectively.

In vivo fluence rate and fractionation effects on tumor response and photobleaching: photodynamic therapy with two photosensitizers in an orthotopic rat tumor model.

The effect of fluence rate and light fractionation on phototoxicity was investigated in vivo in an orthotopic rat bladder tumor model. Two photosensitizers, benzoporphyrin derivative monoacid ring A and 5-aminolevulinic acid-induced protoporphyrin IX, were studied. For a given cumulative light dose of 30 J/cm2, enhanced tumor destruction was observed from both photosensitizers when using either lower fluence rates or fractionated light delivery. Photobleaching experiments in vivo demonstrated that the photobleaching rate, however, was not fluence rate dependent. The fluence rate and light fractionation effects on tumor phototoxicity lead to rapid local depletion in oxygen concentration that inhibited subsequent photochemical reactions necessary for efficient photodestruction of tumor cells. Nicotinamide did not enhance photodynamic therapy efficacy, suggesting that the added increase of oxygen within the tumor was not sufficient to enhance photodestruction of hypoxic cell fractions. The independence of the photobleaching rate with fluence rate suggests distinct mechanisms, at least in part, of photodestruction of the tumor and the photosensitizer and that the rate of photosensitizer photo-bleaching may not always be an appropriate monitor for singlet oxygen availability and photodynamic therapy dosimetry.

Photodynamic therapy of orthotopic prostate cancer with benzoporphyrin derivative: local control and distant metastasis.

This is the first report of photodynamic therapy (PDT) in an orthotopic prostate tumor model and shows that PDT combined with surgery (tumor bed sterilization) gave significant local control of the primary tumor and significant reduction in distant metastases. By contrast, either treatment alone (surgery or PDT) gave relatively poorer local control, and PDT gave a significant increase in the mean number of lung metastases. The MatLyLu variant of the Dunning 3327 rat prostate cancer cell line, which has been selected to be metastatic to lymph nodes and lungs, was injected into the ventral lobe of the rat prostate. After 7 days, tumors were either treated by surgical removal of the ventral lobe, PDT with liposomal benzoporphyrin derivative monoacid ring A, or a combination of surgery, followed by PDT of the tumor bed. Results after 21 days showed a reduction in prostate tumor weight in all groups compared with controls, which became highly significant only for the combination group (17% of control mean tumor weight; P < 0.001; 7 of 13 clinical complete responses). The combination treatment also led to a reduction in lymph node metastasis and reductions in both the frequency and mean number of lung metastases compared with other treatment groups. The PDT-alone group, however, had a mean number of lung metastases per animal, which was nine times the control group and 34 times the combination group. These findings suggest that a tumor bed sterilization approach may be promising for locally advanced prostate cancer and suggest that factors other than local control may need to be evaluated when considering PDT for primary prostate cancer.

Co-existence of two types of chromosome in the bottom fermenting yeast, Saccharomyces pastorianus.

The bottom fermenting yeasts in our collection were classified as Saccharomyces pastorianus on the basis of their DNA relatedness. The genomic organization of bottom fermenting yeast was analysed by Southern hybridization using eleven genes on chromosome IV, six genes on chromosome II and five genes on chromosome XV of S. cerevisiae as probes. Gene probes constructed from S. cerevisiae chromosomes II and IV hybridized strongly to the 820-kb chromosome and the 1500-kb chromosome of the bottom fermenting yeast, respectively. Five gene probes constructed from segments of chromosome XV hybridized strongly to the 1050-kb and the 1000-kb chromosomes. These chromosomes are thought to be S. cerevisiae-type chromosomes. In addition, these probes also hybridized weakly to the 1100-kb, 1350-kb, 850-kb and 700-kb chromosome. Gene probes constructed from segments including the left arm to TRP1 of chromosome IV and the right arm of chromosome II hybridized to the 1100-kb chromosomes of S. pastorianus. Gene probes constructed using the right arm of chromosome IV and the left arm of chromosome II hybridized to the 1350-kb chromosome of S. pastorianus. These results suggested that the 1100-kb and 1350-kb chromosomes were generated by reciprocal translocation between chromosome II and IV in S. pastorianus. Three gene probes constructed using the right arm of chromosome XV hybridized weakly to the 850-kb chromosome, and two gene probes from the left arm hybridized weakly to the 700-kb chromosome. These results suggested that chromosome XV of S. cerevisiae was rearranged into the 850-kb and 700-kb chromosomes in S. pastorianus. These weak hybridization patterns were identical to those obtained with S. bayanus. Therefore, two types of chromosome co-exist independently in bottom fermenting yeast: one set which originated from S. bayanus and another set from S. cerevisiae. This result supports the hypothesis that S. pastorianus is a hybrid of S. cerevisiae and S. bayanus.

[Prostate cancer with high serum level of CEA and CA19-9: a case report].

We report a case of primary adenocarcinoma of the prostate cancer producing carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9). An 80-year-old man admitted to our hospital with the complaint of appetite loss. Serum CEA and CA19-9 levels were increased to 28.9 ng/ml (normal < 3.5) and 271 U/ml (normal < 37), respectively. Serum PSA level was also high (33 ng/ml; normal < 3.6). Computed tomography (CT) demonstrated para-aortic lymph node swelling and bone scan revealed multiple bone metastasis. Prostate biopsy was performed and the specimen showed no evidence of malignancy. However endocrine therapy was started because of the strong suspicion of prostate cancer. In spite of the treatment, the patient died 2 months after the treatment. Histology of autopsy specimen demonstrated primary prostate cancer (poorly differentiated adenocarcinoma) and metastases to multiple organs, such as lungs, liver, thyroid, bone marrow and adrenals. Immunohistochemical staining for CEA, CA19-9 and PSA demonstrated the existence of each protein at both primary and metastatic sites.

Hormonal modulation of the accumulation of 5-aminolevulinic acid-induced protoporphyrin and phototoxicity in prostate cancer cells.

The effect of hormonal modulation on the response of human prostate cancer cell lines to photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) was studied. Two cell lines, one responsive to androgens (LNCaP) and the other non-responsive (PC-3), were used. Fetal bovine serum was depleted of steroid hormones by stripping with charcoal-dextran, and then resupplied with known concentrations of the androgen 5alpha-dihydrotestosterone (DHT) or the estrogen estradiol. It was found that LNCaP cells alone increased growth rate in response to both added hormones, but only androgen had an effect on PDT treatment. LNCaP cells pretreated with 10(-7) M DHT and then 1 mM ALA accumulated 70% more PpIX, compared to cells pre-treated with 10(-12) M DHT. Exposure of the cells treated with high DHT and ALA to 630 nm light led to an 85% decrease in the number of surviving cells compared to the low DHT treated group. The uptake of 14C-ALA was increased with high DHT treatment of the cells, consistent with the above data. No effect of hormones was seen with PC-3 cells or with either cell line and the exogenous photosensitizer benzoporphyrin derivative.

A cDNA clone encoding a glycinin A1a subunit precursor of soybean.

A cDNA clone covering the whole coding region for a glycinin subunit precursor containing the A1a acidic subunit, one of the A2 family, has been identified from a library of soybean cotyledonary cDNA clones using a mixed oligonucleotide probe. Analysis of the cDNA insert revealed that it contained 1746 nucleotides of mRNA sequence with a 5'-terminal nontranslated region of 54 nucleotides, a signal peptide region corresponding to 19 amino acids, an acidic subunit region (A1a) corresponding to 291 amino acids followed by a basic subunit region corresponding to 185 amino acids, and a 3'-terminal nontranslated region of 207 nucleotides. By comparing the predicted protein sequence of this precursor with that of the legumin A precursor of pea, it was found that glycinin A2 subunit family appeared to be more closely related to the legumin than to the A3 subunit family, and that the evolutional rearrangement of glycinin genes has occurred.

Glycinin A5A4B3 mRNA: cDNA cloning and nucleotide sequencing of a splitting storage protein subunit of soybean.

The complete nucleotide sequence of a cloned cDNA, designated pGA5A4B3822, corresponding to glycinin A5A4B3 mRNA was determined. Analysis of the cDNA insert revealed that it contained 1899 nucleotides of mRNA sequence with a 5'-terminal non-translated region of 31 nucleotides, a signal peptide region corresponding to 23 amino acids, an acidic subunit region (A5) corresponding to 97 amino acids, an acidic subunit region (A4) corresponding to 257 amino acids followed by a basic subunit region (B3) corresponding to 185 amino acids, and a 3'-terminal non-translated region of 182 nucleotides. These results show that the glycinin A4 subunit, which is not found to be linked to a basic subunit via a disulfide bond, is synthesized as a full-sized precursor, i.e. the A5A4B3 subunit complex, from a single mRNA, followed by post-translational processing to generate an intermediary subunit complex (A5-B3), covalently linked by a disulfide bond, and the mature A4 subunit, which may associate with the above subunit complex by non-covalent interactions. From the results obtained by the Chou-Fasman rules we speculated that the two post-translational cleavage sites of this subunit precursor might be processed by the same proteolytic enzyme.

Glycinin A3B4 mRNA. Cloning and sequencing of double-stranded cDNA complementary to a soybean storage protein.

The cDNA clones encoding the precursor form of glycinin A3B4 subunit have been identified from a library of soybean cotyledonary cDNA clones in the plasmid pBR322 by a combination of differential colony hybridizations, and then by immunoprecipitation of hybrid-selected translation product with A3-mono-specific antiserum. A recombinant plasmid, designated pGA3B41425, from one of six clones covering codons for the NH2-terminal region of the subunit was sequenced, and the amino acid sequence was inferred from the nucleotide sequence, which showed that the mRNA codes for a precursor protein of 516 amino acids. Analysis of this cDNA also showed that it contained 1786 nucleotides of mRNA sequence with a 5'-terminal nontranslated region of 46 nucleotides, a signal peptide region corresponding to 24 amino acids, an A3 acidic subunit region corresponding to 320 amino acids followed by a B4 basic subunit region corresponding to 172 amino acids, and a 3'-terminal nontranslated region of 192 nucleotides, which contained two characteristic AAUAAA sequences that ended 110 nucleotides and 26 nucleotides from a 3'-terminal poly(A) segment, respectively. Our results confirm that glycinin is synthesized as precursor polypeptides which undergo post-translational processing to form the nonrandom polypeptide pairs via disulfide bonds. The inferred amino acid sequence of the mature basic subunit, B4, was compared to that of the basic subunit of pea legumin, Leg Beta, which contained 185 amino acids. Using an alignment that permitted a maximum homology of amino acids, it was found that overall 42% of the amino acid positions are identical in both proteins. These results led us to conclude that both storage proteins have a common ancestor.

Measurement of urinary estrogen level with the urinary estrogen micrometry method in in vitro fertilization and embryo transfer therapy.

We have evaluated a new micrometering kit using hemagglutination inhibition reaction for determination of total estrogens in urine of normal fertile women and women undergoing in vitro fertilization and embryo transfer (IVF-ET). The micrometering kit was sensitive enough to monitor follicular maturation, and its technique was simple and rapid. Results were obtained in 2.5 hours. By the analysis of the individual estrogen levels in urine collected every 4 hours from 20 IVF-ET patients, the following results were found, with a few exceptions: once the level had exceeded 60 ng/ml, the onset of the luteinizing hormone surge was induced within the next 72 hours; when the level had been under 60 ng/ml, the onset was not induced within the next 24 hours

Mutagenicity of Nagara river sediment.

The sediments of Nagara river and its bystreams were extracted with ether. The mutagenicities of the extracts were determined by the Ames test with Salmonella typhimurium TA100 with S9 mix. The extracts were fractionated on an activated silica-gel column with four kinds of organic solvent, namely iso-octane, iso-octane:benzene (1:1), benzene:ethyl acetate (1:1) and benzene:methanol (1:1). The highest mutagenicity was observed in the iso-octane-benzene fraction of Arata river by using TA100 with S9 mix. Chemical substances in this fraction were identified by GC-MS and GC, and several kinds of polycyclic aromatic hydrocarbons (PAH) were found. Several peaks were unidentified, but these PAH, especially benzo [b] fluoranthene and benzo [a] pyrene, may contribute substantially to the mutagenicity of this fraction.

Studies on the outbreak of hepatitis A in an institute for mentally retarded children.

The authors encountered an outbreak of acute hepatitis in a public institute for mentally retarded children in Aomori Prefecture, Japan. Studies revealed that the probable vehicles of transmission of infection were contaminated water, contaminated meals, and close contact. From the clinical manifestations and epidemiological investigations of 41 affected children and staff members, an outbreak of hepatitis A was strongly suspected. Immune electron microscopy disclosed hepatitis A virus antigen particles in the stool specimens collected during the few days before and after peak transaminase elevation. Hepatitis A antigen was further extracted and purified. The antigen was the first reported recovery of the virus from a natural outbreak of hapatitis A in Asia. Subsequently, with the immune adherence hemagglutination test, using this extracted antigen, an increase in titer of antibody to hepatitis A antigen was demonstrated. Thus, this epidemic was serologically established as an outbreak of hepatitis A. Human immune serum globulin for the protection against hepatitis A was administrated to the 80 individuals concerned, and it was effective in preventing the clinical manifestation of hepatitis.