Influence of using history of immune checkpoint inhibitor therapy for neutropenia caused by combination therapy of ramucirumab and docetaxel.

Recently, pretreatment with immune checkpoint inhibitors (ICIs) has been shown to enhance the therapeutic effects of the combination therapy of ramucirumab (RAM) and docetaxel (DTX); however, its influence on the drug's side effects remains unclear. This study investigated the influence of pretreatment with ICIs on the incidence of neutropenia caused by RAM + DTX therapy in patients with non-small cell lung cancer (NSCLC). Patients with NSCLC who received RAM + DTX therapy at Gifu Prefectural General Medical Center between April 2016 and December 2020 were enrolled. Retrospective data regarding age, sex, performance status and detailed treatment history, among others, at treatment initiation were collected from the patients' electronic medical records. Additionally, data on the course number of RAM + DTX therapy, supportive therapy and blood biochemical parameters, including leukocyte and neutrocyte counts, during the treatment period were collected. We identified 41 patients receiving RAM + DTX therapy. Among the more than grade 3 adverse events caused by this therapy, neutropenia was the most common (78.1%). Despite the fact that all previous risk factors influencing this incidence rate had corresponded, the only factor influencing the incidence rate of neutropenia more than grade 3 was ICI treatment history. A difference in the incidence of neutropenia more than grade 3 in the Kaplan-Meier curve was observed between patients with and without ICI pretreatment history (p = 0.037). The pretreatment history of ICI therapy affects the incidence of neutropenia caused by RAM + DTX therapy in patients with NSCLC.

The importance of mRNA structure in determining the pathogenicity of synonymous and non-synonymous mutations in haemophilia.

INTRODUCTION: Mutational analysis is commonly used to support the diagnosis and management of haemophilia. This has allowed for the generation of large mutation databases which provide unparalleled insight into genotype-phenotype relationships. Haemophilia is associated with inversions, deletions, insertions, nonsense and missense mutations. Both synonymous and non-synonymous mutations influence the base pairing of messenger RNA (mRNA), which can alter mRNA structure, cellular half-life and ribosome processivity/elongation. However, the role of mRNA structure in determining the pathogenicity of point mutations in haemophilia has not been evaluated. AIM: To evaluate mRNA thermodynamic stability and associated RNA prediction software as a means to distinguish between neutral and disease-associated mutations in haemophilia. METHODS: Five mRNA structure prediction software programs were used to assess the thermodynamic stability of mRNA fragments carrying neutral vs. disease-associated and synonymous vs. non-synonymous point mutations in F8, F9 and a third X-linked gene, DMD (dystrophin). RESULTS: In F8 and DMD, disease-associated mutations tend to occur in more structurally stable mRNA regions, represented by lower MFE (minimum free energy) levels. In comparing multiple software packages for mRNA structure prediction, a 101-151 nucleotide fragment length appears to be a feasible range for structuring future studies. CONCLUSION: mRNA thermodynamic stability is one predictive characteristic, which when combined with other RNA and protein features, may offer significant insight when screening sequencing data for novel disease-associated mutations. Our results also suggest potential utility in evaluating the mRNA thermodynamic stability profile of a gene when determining the viability of interchanging codons for biological and therapeutic applications.

Genetic determinants of immunogenicity to factor IX during the treatment of haemophilia B.

Inhibitors are an impediment to the effective management of haemophilia B (HB), but there is limited understanding of the underlying genetic risk factors. In this study we aim to understand the role of F9 gene mutations on inhibitor development in patients with HB. Mutations in the F9 gene were identified and HLA typing performed for five boys with severe HB. Data from the CDC Haemophilia B Mutation Project (CHBMP) database were used to assess association between F9 gene mutation type and inhibitor development. Analysis of the CHBMP database showed that larger disruptions in the F9 gene are associated with a higher life-time prevalence of inhibitors. We detected the following mutations in the five subjects, including four novel mutations: Nonsense in three patients (c.223 C>T; p.Arg75* in two siblings, c.553 C>T; p.Glu185*); Splice site in two patients (c.723 + 1 G>A, c.278-27 A>G); Missense in one patient (c.580 A>G, p.Thr194Ala; c.723 G>T; p.Gln241His). Of the two siblings only one responded to immune tolerance induction (ITI). These siblings have identical F9 gene mutations but differ with respect to the HLA alleles. Interestingly, an analysis of peptide-MHC binding affinities shows a significantly higher (one-sided unpaired t-test, P = 0.0018) median affinity for FIX-derived peptides in the sibling that responded to ITI. We conclude that the nature of the F9 gene mutation may be an important risk factor for the development of inhibitors. In addition, the HLA alleles of the individual patients, in conjunction with the mutation type, could be a predictor for the development of inhibitors as well as the response to ITI.

Factor IX oligomerization underlies reduced activity upon disruption of physiological conditions.

Coagulation factor IX (FIX) is a serine protease that plays a pivotal role in the blood coagulation cascade. FIX deficiency leads to a blood clotting disorder known as haemophilia B. FIX, synthesized as a prepro-peptide of 461 amino acids, is processed and secreted into plasma. The protein undergoes numerous modifications, including, but not limited to glycosylation, γ-carboxylation and disulphide bond formation. Upon processing and limited proteolysis, the protein is converted into an active protease. Under physiological conditions, the FIX zymogen is a monomer. The purpose of this work was to analyse the conditions that may affect FIX monomeric state and promote and/or reduce oligomerization. Using native gel electrophoresis and size exclusion chromatography, we found that under decreased pH and ionic strength conditions, the FIX zymogen can oligomerize, resulting in the formation of higher molecular weight species, with a concomitant reduction in specific activity. Similarly, FIX oligomers formed readily with low bovine serum albumin (BSA) concentrations; however, increased BSA concentrations impeded FIX oligomerization. We hypothesize that normal blood physiological conditions are critical for maintaining active FIX monomers. Under conditions of stress associated with acidosis, electrolyte imbalance and low albumin levels, FIX oligomerization is expected to take place thus leading to compromised activity. Furthermore, albumin, which is commonly used as a drug stabilizer, may enhance the efficacy of FIX biological drugs by reducing oligomerization.

Clinical practice at a multi-dimensional treatment centre for individuals with early psychosis in Japan.

Early intervention for psychosis in Japan has lagged behind that in western countries, but has rapidly begun to attract attention in recent years. As part of a worldwide trend, a multi-dimensional treatment centre for early psychosis consisting of a Youth Clinic, which specialises in young individuals with an at-risk mental state for psychosis, and Il Bosco, a special day-care service for individuals with early psychosis, was initiated at the Toho University Omori Medical Center in Japan in 2007. The treatment centre aims to provide early intervention to prevent the development of full-blown psychosis in patients with an at-risk mental state and intensive rehabilitation to enable first-episode schizophrenia patients to return to the community. We presently provide the same programmes for both groups at Il Bosco. However, different approaches may need to be considered for patients with an at-risk mental state and for those with first-episode schizophrenia. More phase-specific and need-specific services will be indispensable for early psychiatric interventions in the future.

Analysis of F9 point mutations and their correlation to severity of haemophilia B disease.

Haemophilia B is an X-linked recessive disorder caused by deficiency of functional coagulation factor IX, which results almost exclusively from mutations in the F9 gene. We sought to determine features, which could distinguish between mutations that cause severe disease symptoms from those that cause non-severe disease symptoms. Towards this objective, we have performed a statistical analysis of reported point mutations in F9. These include: potential local changes in mRNA free energy, codon usage, charge and type of mutated amino acid, location of the mutation with regard to protein secondary structure and functional domain and amino acids' evolutionary conservation scores. Wilcoxon signed-rank tests showed highly significant differences between severe and non-severe disease causing mutations in their effect on free energy of small mRNA fragments and evolutionarily conserved amino acids. Our results suggest that information at the mRNA level as well as conservation of the amino acid correlate well with disease severity. This study demonstrates that computational tools may be used to characterize the severity of haemophilia B associated with point mutations and suggests their utility in predicting the outcome of sequence changes in recombinant proteins.

Single administration of 1-benzyl-1,2,3,4-tetrahydroisoquinoline increases the extracellular concentration of dopamine in rat striatum.

We performed a combined neurochemical and behavioral study to determine the effects of 1-benzyl-1,2,3,4-tetrahydroisoquinoline (1-BnTIQ) on the extracellular dopamine concentrations in the striatum. Single dose administration of 1-BnTIQ (20, 40, and 80 mg/kg i.p.) increased striatal dopamine extracellular levels in a dose-dependent manner when an in vivo microdialysis technique was used to assess dopamine levels in the striatum of rats. Enhancement of striatal dopamine levels by systemic administration of a single dose of 1-BnTIQ was suppressed by perfusion of tetrodotoxin and a calcium ion-free solution into the striatum. This 1-BnTIQ-induced increase in extracellular dopamine concentration was also inhibited by pre-treatment with a dopamine uptake inhibitor, GBR12909 (1-(2-[bis(4-Fluorophenyl)-4-(3-phenylpropyl)piperazine dihydrochloride). Local application of 1-BnTIQ into the striatum via a dialysis probe failed to enhance the extracellular concentration of dopamine. However, microinjection of 1-BnTIQ into the substantia nigra pars compacta increased the extracellular dopamine levels in the striatum. Locomotor activity was increased by systemic administration of a single dose of 1-BnTIQ in a dose-dependent manner. This 1-BnTIQ-induced locomotor activity was attenuated by pre-treatment with SCH23390 (R(+)-7-Chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrochlodride) and raclopride, D(1) and D(2) dopaminergic receptor antagonists, respectively. Moreover, 1-BnTIQ induced ipsilateral rotational behavior in 6-hydroxydopamine-lesioned rats. These results suggest that systemic administration of a single dose of 1-BnTIQ increases striatal extracellular dopamine concentration through activation of dopaminergic nigra striatal neurons via the dopamine transporter.

Dyke-Davidoff-Masson Syndrome Demonstrated by Current MR images. A Case Report.

Current magnetic resonance imaging techniques demonstrated MR findings of Dyke-Davidoff-Masson syndrome in a 44-year-old man. Statistical parametric mapping analysis of the T1-weighted images showed focal atrophy in the basal ganglia. Three-dimensional white matter fibers of corticospinal tracts, corpus callosum and cingulate bundle were demonstrated using diffusion tensor data correlated to the patient's clinical conditions.

A novel beta-N-acetylglucosaminidase of Clostridium paraputrificum M-21 with high activity on chitobiose.

A beta- N-acetylglucosaminidase gene ( nag3A) from Clostridium paraputrificum M-21 was cloned in Escherichia coli. The nag3A gene consists of an open reading frame of 1,239-bp, encoding 413 amino acids with a deduced molecular weight of 45,531 Da. Nag3A is a single domain enzyme containing a family 3 glycoside hydrolase catalytic domain. Nag3A was purified from recombinant E. coli and characterized. The enzyme hydrolyzed chitooligomers such as di- N-acetylchitobiose, tri- N-acetylchitotriose, tetra- N-acetylchitotetraose, penta- N-acetylchitopentaose, hexa- N-acetylchitohexaose, ball-milled chitin, and synthetic substrates such as 4-methylumbelliferyl N-acetyl beta- D-glucosaminide [4-MU-(GlcNAc)], but had no activity at all against p-nitrophenyl-beta- D-glucoside, p-nitrophenyl-beta- D-xyloside, or p-nitrophenyl-beta- D-galactosamine. The enzyme was optimally active at 50 degrees C and pH 7.0, and the apparent K(m) and V(max) values for 4-MU-(GlcNAc) were 7.9 micro M and 21.8 micro mol min(-1) mg protein(-1), respectively. SDS-PAGE, zymogram, and immunological analyses suggested that this enzyme is induced by ball-milled chitin.

Effect of spermidine on the in vivo degradation of ornithine decarboxylase in Saccharomyces cerevisiae.

As part of our studies on the regulation of polyamine biosynthesis in Saccharomyces cerevisiae, we have investigated the effect of spermidine on the degradation of ornithine decarboxylase in this organism. We have found that in S. cerevisiae, as in other eukaryotic cells, the rate of degradation of ornithine decarboxylase, measured either enzymatically or immunologically, is increased by the addition of spermidine to a yeast culture. It is noteworthy that this effect of added spermidine is found even when the experiments are conducted with strains in which the ornithine decarboxylase is overexpressed several hundred-fold more than the wild-type level. The effect of added spermidine in the overexpressed SPE1 strains is best seen in spe2 mutants in which the initial intracellular spermidine is very low or absent. Experiments with cycloheximide show that new protein synthesis is required to effect the breakdown of the ornithine decarboxylase. These results indicate that S. cerevisiae contains an antizyme-like mechanism for the control of the level of ornithine decarboxylase by spermidine, even though, as contrasted with other eukaryotic cells, no specific antizyme homologue has been detected either in in vitro experiments or in the S. cerevisiae genome.

Redistribution in I-123 N-isopropyl-p iodoamphetamine single-photon emission computed tomography in cerebrovascular disease and the effects of rehabilitation.

We performed N-isopropyl-p (I-123) iodoamphetamine (IMP) single-photon emission computed tomography (SPECT) on 28 patients with severe cerebrovascular disease before rehabilitation, and compared the degree of redistribution and the assessment of activities of daily living (ADL). We calculated a redistribution (RD) ratio in the central and peripheral parts of the lesions: RD ratio (c) and RD ratio (p). We classified the patients into four groups based on the degree of redistribution: complete: both RD ratio (c) and (p) > or = 75; peripheral: RD ratio (c) < 75, RD ratio (p) > or = 75; incomplete: both RD ratio (c) and (p) < 75 and at least one of RD ratio (c) or (p) > or = 25; no redistribution: both RD ratio (c) and (p) < 25. We assessed the ADL using the modified Barthel index (BI). deltaBI was defined as BI after rehabilitation-BI before rehabilitation (BIpost-BIpre). The deltaBI of the four groups were as follows: complete-redistribution group (40.8 +/- 22.8), peripheral-redistribution group (40.0 +/- 15.8), incomplete-redistribution group (27.2 +/- 22.6), no-redistribution group (8.8 +/- 12.3). The deltaBI of the complete and peripheral redistribution groups were significantly higher than that of the no-redistribution group. However, deltaBI was almost the same in the complete- and peripheral-redistribution groups. This suggests that the effect of rehabilitation might be closely related to the viability of the peripheral part of the lesion.

Demonstration of a rhodopsin-retinochrome system in the stalk eye of a marine gastropod, Onchidium, by immunohistochemistry.

The stalk eye of Onchidium sp. (Gastropoda, Mollusca) is the principal photoreceptor in a multiple photoreceptive system that consists of the stalk and dorsal eyes, dermal photoreceptor cells, and photosensitive neurons. To examine the localization of photopigments, the stalk eyes were immunostained with specific antibodies to rhodopsin, retinochrome, and retinal-binding protein (RALBP), which had been generated against squid retinal proteins. The retina of the stalk eye was divided into villous, pigmented, somatic, and neural layers. It was comprised mainly of two types of visual and pigmented supportive cells. The type 1 visual (VC1) cell was characterized by well-developed microvilli on its apical protrusion and photic vesicles in the cytoplasm. The photic vesicles were specifically blackened by prolonged osmification. The type 2 visual (VC2) cell had less numerous, shorter microvilli on its concave apical surface and lacked photic vesicles. The anti-squid rhodopsin antiserum was localized specifically to the villous layer that corresponded to the VC1 microvilli. With the anti-retinochrome peptide antibody, the somatic layer showed specific but patchy, positive staining that corresponded to the cytoplasm of the VC1 cells. Because the photic vesicles are known to contain retinochrome, these results indicate that this retinochrome is localized in the VC1 cytoplasm. Anti-RALBP antibody stained the supranuclear cytoplasm to the distal cytoplasm of VC1 cells. This is the first demonstration of the localization of RALBP in the Gastropoda Onchidium stalk eye. In squid retina that were immunostained as positive controls, the anti-rhodopsin antibody stained rhabdomeric microvilli, the anti-retinochrome antibody stained the inner segment and the basal region of the outer segment, and the anti-RALBP antibody stained the outer and inner segments, respectively. These results suggest that the rhodopsin-retinochrome system that has been established in cephalopod eyes is present in the Onchidium stalk eye.

Preprimed artificial lung for emergency use.

We investigated the possibility of preprimed storage of an artificial lung (AL), aiming at facilitating its emergency use. Test ALs, consisting of a special microporous hollow fiber membrane made of polyolefin in which direct blood-gas contact was completely eliminated, were preprimed with saline solution, sterilized by gamma-ray irradiation, and evaluated after 1-3 months of storage at room temperature. A small amount of bubble was noted in the priming solution after storage in some ALs, which most likely originated from the air dissolved in the priming solution or persisted in the liquid compartment at priming. Although the preprimed solution contained several polyolefin-breakdown products due to irradiation, including ethyl alcohol, n- and t-butyl alcohol, acetone, and carbon dioxide, the levels of these substances were at concentrations known to be not toxic. Endotoxin concentration was negligible. In SEM observation, no perceptible microstructural change was observed in the hollow fibers after preprimed storage. Maximum tensile stress and ultimate elongation of the hollow fiber in the test ALs were reduced by approximately 20% and 3%, respectively, from those of the control AL. The influence of preprimed storage on gas-exchange function was examined in a venoarterial bypass animal study using a goat. Oxygen transfer function was well preserved whereas carbon dioxide removal function was slightly lowered according to the storage term in the stored ALs compared with those of a nonpreprimed control AL. On the basis of these results, we conclude that preprimed storage of the AL with gamma-ray sterilization is basically feasible and realistic.

Low-dose GnRH agonist therapy for the management of endometriosis.

OBJECTIVE: In order to examine whether treatment with a GnRH agonist alone can maintain estrogen levels within the "estrogen window" that inhibits endometriosis without influencing bone-mineral density, we studied the effects of GnRH agonist therapy and changes in bone-mineral density. METHODS: Buserelin acetate nasal spray was administered 3 times a day for 8 weeks (daily dose, 900 micrograms) to 21 women with endometriosis. The drug was then given twice a day for 16 weeks (daily dose, 600 micrograms). The total duration of treatment was 24 weeks. The bone-mineral density of the lumbar vertebrae was measured by dual-energy X-ray absorptiometry before treatment (baseline), at the end of treatment, and 24 weeks after the end of treatment. RESULTS: The bone-mineral density of the lumbar vertebrae at the end of treatment was 2.44% +/- 0.46% (mean +/- standard error) lower than the baseline value. The value at 24 weeks after the end of treatment was 1.10% +/- 0.64% lower than the baseline value. More than 80% of the patients had serum-estradiol levels of 45 pg/ml or less. During treatment, more than 90% of the patients had serum-estradiol levels of 60 pg/ml or less. Genital bleeding was inhibited in 90% of the patients. After 8 weeks of treatment, the clinical symptoms improved in 75% of the patients; such improvement persisted for the duration of the treatment. CONCLUSION: Decreasing the dose of GnRH agonist during treatment can minimize the loss of bone-mineral density without lessening the beneficial effects on endometriosis. This technique might be useful in the management of endometriosis.

Early postmenopausal bone loss is prevented by estrogen and partially by 1alpha-OH-vitamin D3: therapeutic effects of estrogen and/or 1alpha-OH-vitamin D3.

A total of 79 Japanese women who were within 5 years of menopause were randomly assigned 1alpha-hydroxyvitamin D3 [1alpha(OH)D3] 1.0 microg/day, conjugated estrogens 0.625 mg/day, a combination of both, or control (no treatment). Lumbar spine and proximal femur bone mineral density (BMD) and biochemical indices were monitored over 2 years. In the 1alpha(OH)D3-treated group, there was a nonsignificant decrease in lumbar spine BMD compared with controls, and no significant loss in the femoral neck compared with controls. In the estrogen-treated group, there was a nonsignificant increase in spine BMD (+2.17% in the first year and +1.71% in the second year), and no loss in femoral neck BMD. The combination of conjugated estrogens +1alpha(OH)D3 was more effective in increasing BMD in the spine (+3. 68% in the first year and +3.63% in the second year) and femur (+2. 56% in the first year and +4.44% in the second year) BMD. There was a significant difference in lumbar spine BMD in both the first and second years between the combination-treated group and the 1alpha(OH)D3-treated and control groups (P < 0.01). Serum osteocalcin (OC) significantly decreased in the combination-treated group (-23.8% in the first year) and the estrogen-treated group (-37. 6% and -41.2% at 6 and 18 months, respectively), and serum alkaline phosphatase (Alp) decreased significantly in the first year in the combination-treated (-31.5%), estrogen-treated (-27.3%), and 1alpha(OH)D3-treated (-7.9%) groups, whereas serum OC increased (+45. 4% in the first year) in women without treatment. The results of this study indicate that early postmenopausal bone loss in the femoral neck is prevented by conjugated estrogens, 1alpha(OH)D3, or both, whereas bone loss in the spine is not prevented by 1alpha(OH)D3. Estrogen proves effective in preventing early postmenopausal bone loss by markedly inhibiting bone turnover. Moreover, a synergistic bone-sparing effect can be expected when estrogen is administered concomitantly with 1alpha(OH)D3 rather than when used alone.

Synthesis of carbocyclic nucleosides bearing a cyclopropane ring.

A carbocyclic cyclopropane fused nucleoside, 9-(c-4-hydroxymethylbicyclo[3.1.0]hex-r-2-yl)-9H-adenine, has been efficiently synthesized from 2-azabicyclo-[2.2.1]hex-5-en-3-one (ABH) in 6 steps, namely cyclopropanation, -reductive amide cleavage (RAC) reaction and adenine ring construction.

Case of acrania associated with congenital medulloblastoma.

A case of acrania associated with medulloblastoma, agenesis of the cerebellum, and nasoshizis, is reported. The diagnosis of acrania was made at the 20th gestational week by sonographic examination. To our knowledge, this is the first report of fetal acrania associated with congenital brain tumor.

Deamination of cyclaradine by adenosine deaminase under high pressure.

The deamination of cyclaradine corresponding to a carbocyclic analogue of ara-A having anti-HSV activity by adenosine deaminase was examined under various pressure. The deamination of (+)- and (+/-)-cyclaradine was remarkably facilitated by high pressure, and the rate was increased with increasing of pressure. However, (-)-cyclaradine was not deaminated even under high pressure.

Spermine is not essential for growth of Saccharomyces cerevisiae: identification of the SPE4 gene (spermine synthase) and characterization of a spe4 deletion mutant.

Spermine, ubiquitously present in most organisms, is the final product of the biosynthetic pathway for polyamines and is synthesized from spermidine. In order to investigate the physiological roles of spermine, we identified the SPE4 gene, which codes for spermine synthase, on the right arm of chromosome XII of Saccharomyces cerevisiae and prepared a deletion mutant in this gene. This mutant has neither spermine nor spermine synthase activity. Using the spe4 deletion mutant, we show that S. cerevisiae does not require spermine for growth, even though spermine is normally present in the wild-type organism. This is in striking contrast to the absolute requirement of S. cerevisiae for spermidine for growth, which we had previously reported using a mutant lacking the SPE3 gene (spermidine synthase) [Hamasaki-Katagiri, N., Tabor, C. W., Tabor, H., 1997. Spermidine biosynthesis in Saccharomyces cerevisiae: Polyamine requirement of a null mutant of the SPE3 gene (spermidine synthase). Gene 187, 35-43].

Reduction of glycogen in eggs of the silkworm, Bombyx mori, by use of a trehalase inhibitor, trehazolin, and diapause induction in glycogen-reduced eggs.

A new trehalase inhibitor, trehazolin, caused a potent inhibition of ovary trehalase in the silkworm, Bombyx mori. A single injection of trehazolin into pupae (40&mgr;g/animal) did not interfere with the accumulation of proteins and lipids, but markedly reduced glycogen content in eggs accompanied by a remarkable increase in hemolymph trehalose levels. The most potent effect of trehazolin was expressed in eggs that developed at the mid-stage of pupal-adult development. In these eggs glycogen content was reduced to a trace level, less than 3% of that of the control. The reduced glycogen content was almost restored to the control level by injection of glucose but not by trehalose. Trehazolin treatment influenced oviposition and larval hatching, whereas embryogenesis went on normally in glycogen-reduced eggs. Injection of synthetic diapause hormone into non-diapause type hosts induced an incidence of 45% diapause in the eggs and increased their glycogen content. Surprisingly, injection of trehazolin never affected diapause induction by the hormone, despite considerably reduced glycogen content in these eggs. Thus, our findings provide a new method for production of eggs containing various amounts of glycogen, and a novel system for analyzing diapause-associated metabolism besides the well-known glycogen-sorbitol metabolism.