Factors Prognostic for Survival in Japanese Patients Treated with Sunitinib as First-line Therapy for Metastatic Clear Cell Renal Cell Cancer.

BACKGROUND: Factors predictive of survival have been identified in Western patients with metastatic clear cell renal cell carcinoma (mCCRCC) treated with sunitinib. Less is known, however, about factors predictive of survival in Japanese patients. This study evaluated factors prognostic of survival in Japanese patients with mCCRCC treated with first-line sunitinib. MATERIALS AND METHODS: This retrospective study evaluated 46 consecutive Japanese mCCRCC patients treated with sunitinib as first line therapy. Clinical and biochemical markers associated with progression-free survival (PFS) were analyzed, with prognostic factors selected by uni- and multivariate Cox regression analyses. RESULTS: Univariate analysis showed that factors significantly associated with poor PFS included Memorial Sloan-Kettering Cancer Center poor risk scores, International Metastatic RCC Database Consortium poor risk and high (>0.5 mg/dl) serum C-reactive protein (CRP) concentrations (p<0.001 each). Multivariate analysis showed that high serum CRP was independently associated with poorer PFS (p=0.040). Six month disease control rate (complete response, partial response and stable disease) in response to sunitinib was significantly higher in patients with normal (≤0.5 mg/dl) than elevated baseline CRP (p<0.001). CONCLUSIONS: CRP is a significant independent predictor of PFS for Japanese patients with mCCRCC treated with first-line sunitinib. Pretreatment CRP concentration may be a useful biomarker predicting response to sunitinib treatment.

Hepatic arterial infusion chemotherapy with cisplatin before radical local treatment of early hepatocellular carcinoma (JIS score 0/1) improves survival.

BACKGROUND: It has not yet been determined whether hepatic arterial infusion (HAI) chemotherapy improves survival in patients with early hepatocellular carcinoma (HCC). We evaluated the effectiveness of HAI with high-concentration cisplatin (DDP-H) for the treatment of HCC by comparing outcomes between patients who received HAI with DDP-H before radical local treatment of early-stage HCC [Japan Integrated Staging (JIS) score 0/1] and patients who did not receive HAI chemotherapy. PATIENTS AND METHODS: Survival was analyzed in 114 patients with early-stage HCC who underwent radical local treatment. The patients were divided into two groups: a HAI group (n = 79) who received DDP-H infusion into the whole liver via the proper hepatic artery, and a non-HAI group (n = 35) who did not receive HAI chemotherapy. RESULTS: The cumulative survival rates at 1, 3, and 5 years were 77.4%, 69.2%, and 55.3% in the non-HAI group and 97.4%, 87.0%, and 84.4% in the HAI group, respectively. Survival time prolonged significantly in the HAI group compared with the non-HAI group (log-rank test: P = 0.023; generalized Wilcoxon test: P = 0.012) Multivariate analysis using the Cox proportional hazards model identified HAI with DDP-H as the most important factor affecting survival. CONCLUSIONS: Whole-liver HAI with DDP-H before radical local treatment can improve the prognosis of patients with early-stage HCC.

Optimum design of a moderator system based on dose calculation for an accelerator driven Boron Neutron Capture Therapy.

An accelerator based BNCT has been desired because of its therapeutic convenience. However, optimal design of a neutron moderator system is still one of the issues. Therefore, detailed studies on materials consisting of the moderator system are necessary to obtain the optimal condition. In this study, the epithermal neutron flux and the RBE dose have been calculated as the indicators to look for optimal materials for the filter and the moderator. As a result, it was found that a combination of MgF2 moderator with Fe filter gave best performance, and the moderator system gave a dose ratio greater than 3 and an epithermal neutron flux over 1.0×10(9)cm(-2)s(-1).

Protection from non-alcoholic steatohepatitis and liver tumourigenesis in high fat-fed insulin receptor substrate-1-knockout mice despite insulin resistance.

AIMS/HYPOTHESIS: Epidemiological studies have revealed that obesity and diabetes mellitus are independent risk factors for the development of non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma. However, the debate continues on whether insulin resistance as such is directly associated with NASH and liver tumourigenesis. Here, we investigated the incidence of NASH and liver tumourigenesis in Irs1 ( -/- ) mice subjected to a long-term high-fat (HF) diet. Our hypothesis was that hepatic steatosis, rather than insulin resistance may be related to the pathophysiology of these conditions. METHODS: Mice (8 weeks old, C57Bl/6J) were given free access to standard chow (SC) or an HF diet. The development of NASH and liver tumourigenesis was evaluated after mice had been on the above-mentioned diets for 60 weeks. Similarly, Irs1 ( -/- ) mice were also subjected to an HF diet for 60 weeks. RESULTS: Long-term HF diet loading, which causes obesity and insulin resistance, was sufficient to induce NASH and liver tumourigenesis in the C57Bl/6J mice. Obesity and insulin resistance were reduced by switching mice from the HF diet to SC, which also protected these mice against the development of NASH and liver tumourigenesis. However, compared with wild-type mice fed the HF diet, Irs1 ( -/- ) mice fed the HF diet were dramatically protected against NASH and liver tumourigenesis despite the presence of severe insulin resistance and marked postprandial hyperglycaemia. CONCLUSIONS/INTERPRETATION: IRS-1 inhibition might protect against HF diet-induced NASH and liver tumourigenesis, despite the presence of insulin resistance.

The G-protein on cholesterol-rich membrane microdomains mediates mucosal sensing of short- chain fatty acid and secretory response in rat colon.

AIM: Short-chain fatty acids (SCFA) stimulate colonic contraction and secretion, which are mediated by an enteric reflex via a mucosal sensing and cholinergic mechanisms. The involvement of G-protein signal transduction was examined in the secretory response to luminal propionate sensing in rat distal colon. METHODS: Mucosa-submucosa and mucosa preparations were used to measure short-circuit current (I(sc)) and acetylcholine (ACh) release respectively. Cholesterol-rich membrane microdomains, lipid rafts/caveolae, were fractionated using a sucrose gradient ultra-centrifugation after detergent-free extraction of the isolated colonic crypt. RESULTS: Luminal addition of methyl-ß-cyclodextrin (10 mm) and mastoparan (30 µm), lipid rafts/caveolae disruptors, significantly inhibited luminal propionate-induced (0.5 mm) increases in I(sc) , but did not affect increases in I(sc) induced by serosal ACh (0.05 mm) or electrical field stimulation (EFS). Luminal addition of YM-254890 (10 µm), a Gα(q/11) -selective inhibitor, markedly inhibited propionate-induced increase in I(sc) , but did not affect I(sc) responses to ACh and EFS. Both methyl-ß-cyclodextrin and YM-254890 significantly inhibited luminal propionate-induced non-neuronal release of ACh from colonocytes. Real-time PCR demonstrated that in mRNA expression of SCFA receptors, GPR 43 was far higher than that of GPR41 in the colon. Western blotting analysis revealed that the cholesterol-rich membrane microdomains that fractionated from colonic crypt cells were associated with caveolin-1, flotillin-1 and Gα(q/11) , but not GPR43. Uncoupling of Gα(q/11) from flotillin-1 in lipid rafts occurred under desensitization of the I(sc) response to propionate. CONCLUSIONS: These data demonstrate that the secretory response to luminal propionate in rat colon is mediated by G-protein on cholesterol-rich membrane microdomains, provably via Gα(q/11) .

17beta-Estradiol-induced enhancement of estrogen receptor biosynthesis via MAPK pathway in mouse skeletal muscle myoblasts.

The skeletal muscle is one of the important target tissues for the actions of estrogen via both nuclear and extranuclear (non-genomic) pathways. However, there is a paucity of information about the receptor (ER) involved. The aim of this study was thus to explore the ER expression in skeletal muscle, and the influence of estrogen on it, by using C2C12 myoblasts derived from mouse skeletal muscle. Significant expression of a approximately 66-kD protein immunoreactive to ER type alpha (ERalpha) monoclonal antibody, which was comparable to that in ovary, was detected in the whole-cell (total) and nucleus-free (nonnuclear) fractions of C2C12 myoblasts. The expression level of these ER proteins increased in several hours with treatment with 17beta-estradiol (E2), which was preceded by the elevation of the ER mRNA level. This increase appeared to reflect the acceleration of de novo synthesis of ER protein, as proved by the (35)S-methionine immunoprecipitation method. A similar extent of fast increase in ER expression was also induced by a membrane-impermeable, BSA-conjugated estradiol (E2-BSA). Unexpectedly, the E2-induced increases in total and nonnuclear ER were further enhanced by the classic ER antagonists tamoxifen and ICI182,780 in a wide concentration range, implying some structural difference of the involved ER from the classical one. Treatment with the ERK1/2 inhibitor, PD98059 (10 microM), or the p38 MAPK-specific inhibitor, SB203580 (10 microM), greatly inhibited the E2-induced ER increase, while the protein kinase C (PKC) activator TPA (1 microM) enhanced it. These results collectively suggest that C2C12 skeletal myoblasts express a high level of ER, a considerable part of which is extranuclear. Further, the expression of ER in these cells may be significantly upregulated by estrogen itself via increased biosynthesis linked to membrane-bound ER and downstream MAPK-mediated signaling pathways.

Maxillary haemangioma successfully resected by endoscopic approach.

OBJECTIVE: We report an extremely rare case of maxillary haemangioma. METHOD: Case report and review of the literature concerning haemangioma arising from the nasal cavity and paranasal sinuses. RESULTS: Maxillary haemangioma is rare and sometimes requires wider resection than nasal haemangioma if a large tumour is found. We present a case of maxillary haemangioma in a 37-year-old Japanese woman, which was completely resected by pre-operative embolisation and endoscopic sinus surgery. CONCLUSION: Our findings suggest that if a large maxillary haemangioma is diagnosed pre-operatively, the treatment of choice is pre-operative embolisation followed by endoscopic sinus surgery, in order to avoid the surgical complications associated with wide resection.

Single nucleotide polymorphisms in DNA repair genes might be prognostic factors in muscle-invasive bladder cancer patients treated with chemoradiotherapy.

DNA repair enzymes repair DNA damaged by platinum agents and ionising radiation. Single nucleotide polymorphisms (SNPs) in DNA repair genes modulate the repair capacity and might affect response and prognosis following platinum-based chemoradiotherapy (CRT). We investigated associations between the functional SNPs in DNA repair genes and response and survival in muscle-invasive bladder cancer patients treated with CRT to determine the predictive value of the SNPs in patient selection for bladder conservation therapy. The study group comprised 78 patients who underwent CRT for transitional cell carcinoma of the bladder. Single nucleotide polymorphisms in xeroderma pigmentosum complementation groups C (Lys939Gln, A/C), D (XPD; Lys751Gln, A/C), and G (Asp1104His, G/C), and X-ray repair cross-complementing groups 1 (XRCC1; Arg399Gln, G/A) and 3 (Thr241Met, T/C) genes were genotyped. Combined genotypes with at least one variant allele in XPD or XRCC1 were significantly associated with improved cancer-specific survival compared with remaining groups (P=0.009). In multivariate analysis, only the combined XPD and XRCC1 genotypes were independently associated with cancer-specific survival (P=0.04). The association was stronger in stage T3/T4 patients (P=0.0008). These results suggest that combined XPD and XRCC1 genotypes might be prognostic factors in muscle-invasive bladder cancer patients treated with CRT.

Infectious delivery of the 132 kb CDKN2A/CDKN2B genomic DNA region results in correctly spliced gene expression and growth suppression in glioma cells.

The expression of genes from genomic loci can be relatively complex, utilizing exonic, intronic and flanking sequences to regulate tissue and developmental specificity. Infectious bacterial artificial chromosomes (iBACs) have been shown to deliver and express large genomic loci (up to 135 kb) into primary cells for functional analyses. The delivery of large genomic DNA inserts allows the expression of complex loci and of multiple splice variants. Herein, we demonstrate for the first time that an iBAC will deliver and correctly express in human glioma cells the entire CDKN2A/CDKN2B genomic region, which encodes for at least three important cell-cycle regulatory proteins (p16(INK4a), p14(ARF) and p15(INK4b)). Two of these proteins are expressed from overlapping genes, utilizing alternative splicing and promoter usage. The delivered locus expresses each gene at physiological levels and cellular responses (apoptosis versus growth arrest) occur dependent on cellular p53 status, as expected. The work further demonstrates the potential of the iBAC system for the delivery of genomic loci whose expression is mediated by complex splicing and promoter usage both for gene therapy applications and functional genomics studies.

Interaction among human leucocyte antigen-peptide-T cell receptor complexes in cow's milk allergy: the significance of human leucocyte antigen and T cell receptor-complementarity determining region 3 loops.

BACKGROUND: Allergic individuals respond to only a few specific antigens, therefore allergic diseases are characterized by antigen specificity. Clarification of the mechanism of antigen specificity will lead to progress in the therapy of allergic diseases. OBJECTIVES: The purpose of this study is to determine the specific association among T cell epitopes, antigen-presenting molecules and T cell receptor (TCR), and to determine the TCR usage in the pathogenesis of allergies using antigen-specific T cell clones (TCCs). The results can clarify the mechanism of the antigen specificity of allergic diseases, and provide new therapeutic possibilities using analogue peptides. METHODS: Short-term T cell clones specific to beta-lactoglobulin (BLG) were established from peripheral blood mononuclear cells (PBMCs) collected from five patients allergic to cow's milk. We then identified the T cell epitopes and antigen-presenting molecules, and examined TCR usage. We also determined the sequence of the TCR-complementarity-determining region 3 (CDR3). RESULTS: Six TCCs established from the five patients recognized three different peptides, and BLGp97-117 was recognized by four of the six TCCs. BLGp101-112 (KYLLFCMENSAE) was the core sequence in the fragment. Sequence analysis of TCR by the RT-PCR method revealed a marked heterogeneity in TCR usage, and similar amino acid sequences were recognized in the CDR3 region. Four of the six TCCs recognized BLG in association with human leucocyte antigen (HLA)-DRB1*0405 as antigen-presenting molecules. CONCLUSION: We proposed the motif of the interaction between the HLA-DRB1*0405 allele and antigen peptide, and suggested that HLA-DRB1*0405 is an immunoregulatory gene product for T cell responses to BLG.

Identification of beta-lactoglobulin-derived peptides and class II HLA molecules recognized by T cells from patients with milk allergy.

BACKGROUND: Cow's milk allergy impairs the health and development of many infants since it deprives them of adequate nutrition. Cow's milk fractions contain many allergens, and beta-lactoglobulin (BLG) is one of the major allergens. OBJECTIVE: The purpose of this study was to determine T cell epitopes, antigen-presenting molecules and cytokine production by T cells in relation to BLG. The results can provide new therapeutic possibilities of using analogue peptides of BLG for infants with cow's milk allergy. METHODS: Using a mixture of a panel of overlapping synthetic peptides that cover the entire BLG molecule, we established polyclonal BLG-specific short-term T cell lines and clones from peripheral blood mononuclear cells of four patients with allergy to cow's milk carrying most of the common human leucocyte antigen (HLA) haplotypes seen in the Japanese population. We then identified the T cell epitopes and antigen-presenting molecules, and measured the production of cytokines interleukin (IL)-4, IL-5 and interferon-gamma in the culture supernatants. RESULTS: The T cell lines established from the four patients responded to seven different peptides. Three of the peptides stimulated the T cells of two donors, regardless of the HLA types. The patterns of inhibition of the proliferative responses of the cell lines by anti-HLA class II antibodies were heterogeneous; three were mainly inhibited by anti-HLA-DR mAbs, and the other was inhibited by anti-HLA-DQ mAbs. High levels of IL-5 were produced by these T cell lines. CONCLUSIONS: Patients' T cells recognized BLG in association with a variety of HLA-DR or -DQ as antigen-presenting molecules. Although some peptides did have a more potent T cell stimulatory activity than others, the T cell receptor ligands formed with the BLG molecule are heterogeneous. Peptides for the desensitization of T cells of the patients with cow's milk allergy need to be designed keeping in mind the different requirements in different ethnic groups.

Antitumor effect of beta2-microglobulin in leukemic cell-bearing mice via apoptosis-inducing activity: activation of caspase-3 and nuclear factor-kappaB.

We have reported previously that beta2-microglobulin (beta2m) induces apoptosis in leukemic cells in vitro, and that an interaction between beta2m and HLA class I antigen induces apoptosis. Here we examined whether beta2m can induce apoptosis in leukemic cells in vivo and whether it has an antitumor effect in tumor-bearing mice. Daily administration of 50 or 250 microg of beta2m induced apoptosis and an antitumor effect on K562 leukemia cell-bearing mice in the same manner as tumor necrosis factor-alpha. In tumor tissues in beta2m-treated mice, both caspase-3 and nuclear factor-kappaB (NF-kappaB) were stained more strongly than in control mice by anti-caspase-3 and anti-NF-kappaB p65/Rel A polyclonal antibodies. We also observed the in vivo immunological effects of beta2m on lymphoid and hematopoietic organs, such as thymus, bone marrow, Peyer's patches, liver, and spleen in normal mice. Using antibodies against caspase-3 and NF-kappaB, immunohistochemical staining showed that no specific tissues were damaged or stained in normal mice. We conclude that beta2m stimulates caspase-3 and NF-kappaB pathways to induce apoptosis, making it a useful approach to a new therapy for leukemia.

Midaortic syndrome in childhood associated with a ruptured cerebral aneurysm: a case report.

BACKGROUND: We present a patient with a midaortic syndrome who presented with subarachnoid hemorrhage caused by rupture of an anterior communicating artery aneurysm. CASE DESCRIPTION: A 14-year-old boy with midaortic syndrome was admitted to our hospital because of subarachnoid hemorrhage due to rupture of an anterior communicating artery aneurysm. He also developed acute renal failure due to previously controlled hypotension. After blood dialysis, successful clipping of the aneurysm was performed. The postoperative course was complicated by malignant renovascular hypertension due to midaortic syndrome. Medical treatment failed to control his hypertension; left primary nephrectomy improved his condition. CONCLUSION: Although midaortic syndrome is rare, it may be significant as a cause of cerebral hemorrhage in childhood.

Mycobacterium tuberculosis infection within Warthin's tumor: report of two cases.

We report two patients with Warthin's tumor who were also infected with Mycobacterium tuberculosis. Case 1 was a 75-year-old woman with Warthin's tumor and multiple small epithelioid granulomas with caseous necrosis involving the submandibular gland. This patient died of tuberculous meningitis 4 months after biopsy. Case 2 was a 78-year-old man with a 10-year history of a parotid mass which had enlarged rapidly over 2 months. Surgical excision revealed Warthin's tumor and epithelioid granulomas involving the left parotid gland. DNA extracted from paraffin sections was amplified by nested polymerase chain reaction (PCR) with primer sets for the mycobacterial 65-KDa antigen gene. Restriction enzyme digestion of the PCR products could differentiate Mycobacterium tuberculosis from other mycobacteria in both cases. Although the histogenesis of lymphoid components of Warthin's tumor is controversial, the frequent prevalence of inflammation or necrosis and our present findings suggest these components have a similar behavior to regional lymph nodes.

Protection of cultured spinal motor neurons by estradiol.

Estrogens have been reported to exert neuroprotection in the brain, but there have been no reports of such neuroprotection in spinal motor neurons, the neurons selectively involved in amyotrophic lateral sclerosis (ALS). In this study, we demonstrated that 17beta-estradiol and its biologically inactive stereoisomer, 17alpha-estradiol, prevented glutamate- and nitric oxide (NO)-induced selective motor neuronal death observed in primary cultures of the rat spinal cord. The dose of estradiols required for motor neuron protection was greatly reduced by co-administration with glutathione. The results of this study shows that estradiol protects spinal motor neurons from excitotoxic insults in vitro, and may have application as a treatment for ALS.

Long-term study of a female hyper-IgM immunodeficiency.

Hyper-IgM immunodeficiency (HIM) is an immunological disorder characterized by normal or elevated serum IgM levels, and reduced serum IgG and IgA levels, due to the disruption of immunoglobulin class switching in B cells. X-linked hyper-IgM is caused by the defective expression of the CD40 ligand on activated T cells, which induces immunoglobulin class switching along with some cytokines, such as interleukin 4, by the signal transduction of CD40 in B cells. We report on a Japanese girl who initially showed low serum IgM, IgG and IgA levels like patients with common variable immunodeficiency; however, in the course of time, serum IgG levels became reduced and serum IgM levels increased, resulting in the typical immunoglobulin profile of HIM. Neutropenia, one of the features of X-linked HIM, was not observed. In spite of extremely low serum IgG levels, she did not show any predisposition to severe infection, even without gammaglobulin replacement therapy. No mutation of the CD40 ligand or CD40 was detected. Sequencing of the complementarity-determining region of immunoglobulin heavy-chain genes in peripheral B lymphocytes revealed that they were all in frame, and insertion of the N region was detected. These results indicate that the heavy-chain gene rearrangement in the patient's B cells is intact. Non-X-linked HIM has heterogeneous pathogenetic mechanisms, and some groups may show the resistance to infection at the healthy donor level. The underlying defects in non-X-linked HIM might be specifically involved in class switching.

A new apoptotic pathway for the complement factor B-derived fragment Bb.

Apoptosis is involved in both the cellular and humoral immune system destroying tumors. An apoptosis-inducing factor from HL-60 myeloid leukemia cells was obtained, purified, and sequenced. The protein found has been identified as a human complement factor B-derived fragment Bb, although it is known that factor B is able to induce apoptosis in several leukemia cell lines. Monoclonal antibodies against fragment Ba and Bb inhibited the apoptotic activity of factor B. When the purified fragment Bb was used for apoptosis induction, only the anti-Bb antibody inhibited Bb-induced apoptosis, and not the anti-Ba antibody. The apoptosis-inducing activity was found to be enhanced under conditions facilitating the formation of Bb. Blocking TNF/TNFR or FasL/Fas interactions did not interfere with the factor B-induced apoptosis. CD11c (iC3bR) acts as the main subunit of a heterodimer binding to fragment Bb in the apoptosis pathway, and the factor B-derived fragment Bb was found to possess the previously unknown function of inducing apoptosis in leukemic cells through a suicide mechanism of myeloid lineage cells during the differentiation stage.

Attempted gene therapy for intractable pain: dexamethasone-mediated exogenous control of beta-endorphin secretion in genetically modified cells and intrathecal transplantation.

For optimal neural transplantation using gene engineering, it might be important to control the expression of the transfected gene extrinsically as required. This strategy could be very useful for the treatment of intractable pain that responds to opioids. For this purpose, we established a genetically modified embryonal carcinoma cell line (P19) in which the expression of beta-endorphin (beta-EP) could be controlled by the addition of dexamethasone. To obtain extrinsic control, we transfected the cells with pMAMneo containing mouse MMTV-LTR as a promoter and cDNA of the artificial beta-EP. The upregulation of beta-EP, through the activation of MMTV by the administration of dexamethasone, was confirmed in vitro. Then we transplanted these cells into the subarachonoid space in rats and evaluated the analgesic potential of these cells in vivo by hot plate test and formalin test. In the rats that received beta-EP-producing cells, we observed prominent analgesic effects after the transplantation for a month. The administration of naloxone blocked these effects. Intraperitoneal injection of 100 mg/kg dexamethasone further enhanced these effects by up to two times. These data indicate obvious analgesic effects of the cells after the transplantation and the possible exogenous upregulation of transfected beta-EP gene expression in vivo. The application of this technique might provide a new therapeutic approach to various neurological diseases.

Intracellular ATP slows time-dependent decline of muscarinic cation current in guinea pig ileal smooth muscle.

The effects of intracellular nucleotide triphosphates on time-dependent changes in muscarinic receptor cation currents (I(cat)) were investigated using the whole cell patch-clamp technique in guinea pig ileal muscle. In the absence of nucleotide phosphates in the patch pipette, I(cat) evoked every 10 min decayed progressively. This decay was slowed dose dependently by inclusion of millimolar concentrations of ATP in the pipette. This required a comparable concentration of Mg(2+), was mimicked by UTP and CTP, and was attenuated by simultaneous application of alkaline phosphatase or inhibitors of tyrosine kinase. In contrast, a sudden photolytic release of millimolar ATP (probably in the free form) caused a marked suppression of I(cat). Submillimolar concentrations of GTP dose dependently increased the amplitude of I(cat) as long as ATP and Mg(2+) were in the pipette, but, in their absence, GTP was ineffective at preventing I(cat) decay. The decay of I(cat) was paralleled by altered voltage-dependent gating, i.e., a positive shift in the activation curve and reduction in the maximal conductance. It is thus likely that ATP exerts two reciprocal actions on I(cat), through Mg(2+)-dependent and -independent mechanisms, and that the enhancing effect of GTP on I(cat) is essentially different from that of ATP.