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Background: The human induced pluripotent stem cells (hiPSCs) can generate all the cells composing the human body, theoretically. Therefore, hiPSCs are thought to be a candidate source of stem cells for regenerative medicine. The major challenge of allogeneic hiPSC-derived cell products is their immunogenicity. The hypoimmunogenic cell strategy is allogenic cell therapy without using immune suppressants. Advances in gene engineering technology now permit the generation of hypoimmunogenic cells to avoid allogeneic immune rejection. In this study, we generated a hypoimmunogenic hiPSC (HyPSC) clone that had diminished expression of human leukocyte antigen (HLA) class Ia and class II and expressed immune checkpoint molecules and a safety switch., Methods: First, we generated HLA class Ia and class II double knockout (HLA class Ia/II DKO) hiPSCs. Then, a HyPSC clone was generated by introducing exogenous β-2-microglobulin (B2M), HLA-G, PD-L1, and PD-L2 genes, and the Rapamycin-activated Caspase 9 (RapaCasp9)-based suicide gene as a safety switch into the HLA class Ia/II DKO hiPSCs. The characteristics and immunogenicity of the HyPSCs and their derivatives were analyzed., Results: We found that the expression of HLA-G on the cell surface can be enhanced by introducing the exogenous HLA-G gene along with B2M gene into HLA class Ia/II DKO hiPSCs. The HyPSCs retained a normal karyotype and had the characteristics of pluripotent stem cells. Moreover, the HyPSCs could differentiate into cells of all three germ layer lineages including CD45 + hematopoietic progenitor cells (HPCs), functional endothelial cells, and hepatocytes. The HyPSCs-derived HPCs exhibited the ability to evade innate and adaptive immunity. Further, we demonstrated that RapaCasp9 could be used as a safety switch in vitro and in vivo., Conclusion: The HLA class Ia/II DKO hiPSCs armed with HLA-G, PD-L1, PD-L2, and RapaCasp9 molecules are a potential source of stem cells for allogeneic transplantation., (© 2024. The Author(s).)

Regenerative medicine is currently the focus of global attention. Countries all around the world are actively working to create new regenerative treatment modalities through pioneering research and novel technologies. This is wonderful news for patients who could not be treated with existing medical options. New venture businesses and companies are being established in regenerative medicine and their rapid industrialization is anticipated. However, to ensure high-quality products, human resources qualified in research and development and the manufacturing of these products are essential. The Forum for Innovative Regenerative Medicine (FIRM) conducted a questionnaire of its industry members to examine the training and hiring of people in research and development, product creation, manufacturing, and more. Regenerative medicine is a brand new field; thus, many different businesses will need to cooperate together. People with a broad range of technical skills, abilities, and knowledge will be in demand, with various levels of expertise, from basic to advanced.

Objectives: The (+)-isomer of indeloxazine AS1069562 has multiple pharmacological actions, such as serotonin (5-HIT) and norepinephrine (NE) reuptake inhibition and analgesic effects in animal models of neuropathic pain. Here, we investigated the analgesic effects of AS1069562 in rat models of inflammatory and noninflammatory nociceptive pain., Methods: Adjuvant-induced arthritis (AIA) and bradykinin-induced knee joint pain were used as rat models of inflammatory pain. The chronic phase of monoiodoacetate-induced arthritis (MIA) was used as a rat model of noninflammatory pain. Analgesic effects were evaluated by weight-bearing deficit in the AIA and MIA models and by pain response in the bradykinin-induced knee joint pain model., Results: In the AIA model and the bradykinin-induced knee joint pain model, AS1069562 significantly ameliorated the pain-related behavior of weight-bearing deficit and the pain response, respectively. AS1069562 also significantly improved the pain-related behavior of weight-bearing deficit in the chronic phase of the MIA model. Further, following monoiodoacetate injection, repeated administration of AS1069562 or duloxetine significantly improved weight-bearing deficit in the MIA model. Interestingly, the analgesic effect of AS1069562 was sustained for 24 hours after the last administration, although the plasma concentration of AS1069562 was reduced to undetectable levels. In contrast, the analgesic effect of duloxetine did not continue after treatment discontinuation., Discussion: AS1069562 exerts analgesic effects on inflammatory and noninflammatory nociceptive pain in rat models of arthritis pain, and repeated administration of AS1069562 exerts a more persistent analgesic effect on arthritis pain than duloxetine. These findings suggest that AS1069562 has an attractive analgesic profile for the treatment of nociceptive pain.

The pharmacological effect of FR177391, isolated from Serratia liquefaciens No. 1821, was studied in normal animals and various types of animal models of hypertriglyceridemia. Treatment of normal mice with FR177391 resulted in an increase in heparin-releasable lipoprotein lipase (LPL) activity in the blood and epididymal fat tissue. FR177391 treatment decreased triglyceride (TG) and increased high-density lipoprotein cholesterol in the blood in normal rats following 7 days treatment, suggesting potent LPL activating properties of FR177391. Both Triton WR1339-induced severe and fructose-induced mild hypertriglyceridemia in rats were attenuated by FR177391 treatment. Severely elevated levels of TG in db/db mice, an insulin resistant diabetic animal model, also significantly decreased from 14 days of treatment with FR177391. FR177391 treatment for 9 days caused a decrease in the elevated levels of TG in mice induced by intraperitoneal inoculation of murine lymphoma EL-4. Overall, this study demonstrated that FR177391 can be possibly a LPL activating agent and that FR177391 treatment improved hypertriglyceridemia in various rat and mouse animal models. These results suggest that FR177391 is a promising candidate compound for the management of hypertriglyceridemia.

Matrix metalloproteinase (MMP) has been implicated in joint destruction of chronic arthritis diseases, such as rheumatoid arthritis. FR217840 (2R)-1-([5-(4-fluorophenyl)-2-thienyl]sulfonyl)-N-hydroxy-4-(methylsulfonyl)-2-piperazinecarboxamide is a potent, orally active synthetic MMP inhibitor that inhibits human collagenases (MMP-1, MMP-8 and MMP-13), gelatinases (MMP-2 and MMP-9) and membrane type MMP (MT-MMP) (MT1-MMP/MMP-14). FR217840 also inhibits rat collagenase and gelatinase. We studied the effect of FR217840 on a rat adjuvant induced arthritis model. Although oral administration (days 1-21) of FR217840 (3.2, 10, 32 mg/kg) to adjuvant injected Lewis rats did not affect inflammation, as indicated by both hind paw swelling and histological inflammatory infiltration, FR217840 suppressed both bone destruction and serum pyridinoline content in a dose-dependent manner. Also, FR217840 (32 mg/kg) reduced tartrate-resistant acid phosphatase (TRAP) cell number in the ankle joints of rats with arthritis. These results indicate that FR217840 successfully suppressed joint destruction and suggest that FR217840 may have potential as a novel anti-rheumatic drug.

FR255031 (2-[(7S)-7-[5-(4-ethylphenyl)-2-thienyl]-1,1-dioxido-4-(2-pyridinylcarbonyl)hexahydro-1,4-thiazepin-7-yl]-N-hydroxyacetamide) is a novel synthetic matrix metalloproteinase (MMP) inhibitor that inhibits human collagenases (MMP-1, MMP-8 and MMP-13), gelatinases (MMP-2 and MMP-9) and membrane type 1 MMP (MT1-MMP/MMP-14). FR255031 also inhibits rat collagenase and gelatinase. We studied the effect of FR255031 and Trocade, an inhibitor of collagenase and MMP-14, on a rat collagen-induced arthritis (CIA) model. Rat CIA was induced by intradermal injection of type II collagen (IIC) and oral administration of FR255031 or Trocade was performed for 28 days. Body weight loss, hind paw swelling, elevation of serum anti-IIC antibody, and histological and radiographic scores were evaluated. FR255031 markedly inhibited cartilage degradation in a dose-dependent manner in the CIA model, but Trocade failed to prevent the degradation. FR255031 at a dose of 100 mg kg(-1) also had statistically significant effects on bone destruction and pannus formation and on the recovery of body weight loss on day 28. These results indicate that FR255031 is effective for rat CIA, especially on joint cartilage destruction. These data suggest that as well as collagenases or MT-MMP, gelatinases are also involved in joint destruction in arthritis.

Objective: To assess the ability of a marker of collagen type II degradation (CTX-II) to quantify cartilage turnover in vitro in cartilage explants and in vivo in rats with collagen induced arthritis (CIA)., Methods: Bovine articular cartilage explants were cultured in the presence of interleukin 1a, oncostatin M, and plasminogen to induce cartilage degradation. CTX-II, CTX-I (C-telopeptide fragment of collagen type I), glycosaminoglycan, and hydroxyproline contents in culture supernatants were measured. CIA was induced in 12-week-old female Lewis rats by immunization with bovine type II collagen. The incidence and severity of arthritis were monitored by measuring paw swelling, and urinary levels of CTX-II and CTX-I were determined. The knee joints of rats were histopathologically examined after sacrifice. Results. CTX-II but not CTX-I levels correlated well with collagen degradation in bovine articular cartilage in vitro quantified by hydroxyproline release. Urinary CTX-II levels as well as paw volume of CIA rats were significantly higher than normal rats on Days 21, 28, and 42 and were apparently correlated with cartilage destruction, assessed histopathologically. Urinary CTX-I level began to increase on Day 21, but only on Day 42 was it significantly different between CIA and normal rats. The elevation in CTX-I level appeared to occur later than that of CTX-II, in accord with the more delayed onset of bone erosion in the CIA model of rheumatoid arthritis., Conclusion: Urinary CTX-II may be a useful marker for evaluation of dynamics of cartilage destruction in CIA rats.

Objective: To investigate the effects of prophylactic and therapeutic treatments with FK506 (tacrolimus), an immunosuppressive drug that specifically inhibits T cell activation, and methotrexate (MTX) on inflammatory cytokines, tumor necrosis factor (TNF)-a, interleukin (IL)-1beta, and IL-6 levels in rat adjuvant-induced arthritis (AIA)., Methods: AIA was induced in female Lewis rats. Arthritis was assessed by hindpaw swelling. TNF-a, IL-1beta, and IL-6 levels in paw extracts were determined by ELISA. To assess the effects on cytokine levels, rats were treated prophylactically with FK506 (3 mg/kg) or MTX (0.1 mg/kg) from day 1 to day 17, and therapeutically with FK506 (5 mg/kg) or MTX (1 mg/kg) from day 15 to day 17 (3-day treatment) or day 15 to 20 (6-day treatment) by oral administration., Results: TNF-a, IL-1beta, and IL-6 levels in paw tissue were found to significantly increase between day 15 and day 21 after adjuvant injection, when the arthritis was in a developed stage. Prophylactic treatment with FK506 and MTX suppressed arthritis and reduced the levels of those inflammatory cytokines. FK506 caused a marked reduction of TNF-a and IL-1beta levels in paw tissue even in short-term (3-day) therapeutic treatment. It reduced all levels of TNF-a, IL-1beta, and IL-6 in paws in 6-day therapeutic treatment. In contrast, therapeutic treatment with MTX affected neither TNF-a or IL-6 levels in paws. MTX reduced IL-1beta levels only in the 6-day treatment., Conclusion: FK506 is more effective than MTX in reducing elevated levels of inflammatory cytokines TNF-a, IL-1beta, and IL-6 in established stages of AIA. Our findings suggest that inhibition of T cell activation results in a rapid reduction of inflammatory cytokine levels even after the arthritis is established in AIA.

The effects of FK317 (11-acetyl-8-carbamoyloxymethyl-4-formyl-6-methoxy-14- oxa-1,11-diazatraacylo[7.4.1.0(2.7).0(10.2)]-tetradeca-2,4,6-trien-9-yl acetate), a novel anti-cancer agent, and mitomycin C (MMC) on survival time of mice bearing B16BL6 melanoma and Lewis lung carcinoma (LLC), induced by intravenous inoculation of the tumor, were investigated. Treatment with FK317 resulted in a significant prolongation of survival time in both tumor models. Four of ten mice bearing B16BL6 were disease-free following FK317 treatment. In contrast, MMC was not effective in prolonging survival time. Overall, this study demonstrated that FK317 shows more potent survival extension in mice bearing B16BL6 and LLC than MMC, suggesting that FK317 may have therapeutic utility for cancer chemotherapy.

FK506 (Tacrolimus) and cyclosporin A exert their immunosuppressive effects via a common mechanism, calcineurin inhibition, after binding to intracellular proteins termed immunophilins: FK506-binding protein (FKBP) and cyclophilin. In this study, FK506 was found to induce chondrogenic differentiation of ATDC5 cells (clonal mouse embryonal carcinoma cells) in a concentration-dependent manner (0.1-1000 ng/ml). Immunohistochemical staining showed that ATDC5 cells induced to differentiate by FK506 produced proteoglycan and type II collagen, main components of the extracellular matrix of cartilage. Rapamycin, an immunosuppressant that binds to FKBP, antagonized the effect of FK506. Cyclosporin A did not induce chondrogenesis at concentrations up to 1000 ng/ml. Taken together, these results suggest that FK506 induces chondrogenic differentiation of ATDC5 cells via a calcineurin-independent mechanism, after binding to FKBP.

Several lines of evidence have postulated that reduction in the activity of lipoprotein lipase (LPL) is involved in cachexia induction in cancer patients. Recently we have demonstrated that murine melanoma B16 has the ability to reduce the LPL activity and thereby induce cachexia symptoms in mice following intraperitoneal inoculation. In order to further investigate the relationship between LPL activity and cachectic syndrome, cachexia models other than melanoma B16 are required. However, there are few animal cachexia models in which LPL activity is involved in the induction of cachectic symptoms. In this study, cachectic symptoms and plasma LPL activity were investigated in mice bearing EL-4 mouse lymphoma. In EL-4 bearing mice the body weight including tumor weight in the abdominal cavity was rather higher than that of normal mice without tumor, whereas weights of carcass wet and gastrocnemius muscle were significantly decreased in EL-4 bearing mice. Elevated blood levels of triglyceride and non-esterified fatty acid were observed in mice bearing EL-4, associated with the impaired plasma LPL activity. Overall, this study indicated that EL-4 lymphoma in mice results in a severe cachexia which is possibly related to impaired LPL activity and also provided a useful cachexia model for understanding the role of LPL in the development of cancer cachexia.

The aim of this study was to elucidate the in vitro inhibitory potency of FK506 on production of the inflammatory cytokines, tumour necrosis factor (TNF)-alpha and interleukin (IL)-1beta, with a view to assessing this immunosuppressive agent as a potential anti-rheumatic drug. We employed an in vitro model which produces TNF-alpha and IL-1beta through T cell activation. Human peripheral blood mononuclear cells (PBMC) were cultured with immobilized anti-CD3/CD28 monoclonal antibody in this model. FK506 inhibited anti-CD3/CD28 induced TNF-alpha and IL-1beta production at concentrations less than 1 ng ml(-1). Flow cytometric analysis of intracellular TNF-alpha and IL-1beta positive cells showed that FK506 potently suppresses inflammatory cytokine production from CD14+ monocytes as well as from T cells. Cyclosporin A (CsA) and dexamethasone (DEX) also inhibited the anti-CD3/CD28 induced cytokine production, but were less potent than FK506. FK506 and CsA, but not DEX, specifically inhibited anti-CD3/CD28 induced inflammatory cytokine production without affecting the lipopolysaccaride (LPS) induced effect. Methotrexate (MTX) was completely inactive for suppressing cytokine production under either condition. Anti-CD3/CD28 stimulated PBMC culture supernatants were found to enhance the expression of adhesion molecules in human vascular endothelial cells. FK506, CsA and DEX led to the suppression of adhesion molecule expression probably by inhibiting cytokine production from PBMC. The inhibitory potency of agents on TNF-alpha and IL-1beta production was compared with cytotoxicity and FK506 was not cytotoxic at concentrations several orders of magnitude greater than those required for cytokine inhibition. These results strongly suggest that FK506 may be most effective to specifically prevent T cell activation mediated inflammatory cytokine production in a clinical setting.

Our recent study has demonstrated that ponalrestat, an aldose reductase inhibitor, activates lipoprotein lipase (LPL) activity in the adipose tissue and alleviates the cachectic symptoms induced by B16 melanoma in mice. In this study, the effect of ponalrestat on cachexia symptoms in nude mice bearing human melanomas G361 and SEKI was investigated because it has been suggested that the suppression of LPL has an important role in cachexia induction by these two melanomas in nude mice. Mice bearing G361 subcutaneously did not gain weight and became cachectic, associated with the tumor growth. Tumor growth was not affected by ponalrestat, nevertheless treatment with ponalrestat resulted in an amelioration of the reduction in the weight of body mass, epididymal fat, gastrocnemius muscle, carcass and whole body lipid induced by the presence of G361. A severe weight loss observed in nude mice bearing SEKI was also partially attenuated by ponalrestat treatment. Overall, this study showed that ponalrestat is effective in the attenuation of the cachectic symptoms induced by human melanomas G361 and SEKI in nude mice, suggesting that ponalrestat has a potential usefulness for the treatment of cancer cachexia.

Our recent study has demonstrated that ponalrestat, an aldose reductase inhibitor, activates lipoprotein lipase activity and alleviates B16 melanoma-induced cachexia in mice. In this study, the effect of ponalrestat on murine adenocarcinoma colon26-induced cachexia was investigated in mice. Mice bearing colon26 subcutaneously lost weight and became cachectic, associated with the tumor growth. Although tumor growth was slightly stimulated when tumor bearing mice were treated with ponalrestat: nevertheless, the drug attenuated the reduction in the weight of body mass, epididymal fat, gastrocnemius muscle and carcass induced by colon26, as well as significantly prolonged the survival of the colon26 bearing mice. Ponalrestat inhibited the production of interleukin-1 (IL-1) from human monocytes stimulated by Lipopolysaccharide (LPS) in vitro, and also suppressed LPS-induced increase of IL-1 in the blood in mice. Overall, this study showed that ponalrestat suppresses IL-1 production both in vitro and in vivo, and inhibits the cachectic symptoms induced by colon26 adenocarcinoma in mice, suggesting that ponalrestat has a therapeutic potential for the treatment of cancer cachexia.

Dapsone (4,4'-diaminodiphenyl sulphone), an antileprotic and antimalarial drug, has been reported to be of therapeutic benefit in idiopathic thrombocytopenic purpura in the clinic. However, adverse reactions such as haemolytic anaemia have often been observed. In this study, we found that dapsone increased the number of platelets and decreased the number of red blood cells in male (NZWxBXSB)F1 (W/BF1) mice, an animal model of idiopathic thrombocytopenic purpura. In studies to prepare derivatives of dapsone with weaker side effects than the parent compound, FR115092 (2-[5-(2-pyridylsulphonyl)thiazolyl]amine) was discovered. The effect of FR115092 on the number of blood cells was studied and compared with dapsone in mice. FR 115092 increased the number of platelets without reducing the number of red blood cells in W/BF1 mice. This drug significantly suppressed the increase in circulating autoantibodies against platelets and increased the number of megakaryocytes. Furthermore, FR115092 inhibited the reduction of the number of platelets in mitomycin C-induced thrombocytopenic mice, as a consequence of its enhancement of growth and maturation of megakaryocytes. These findings suggest that FR115092 may be effective against various thrombocytopenias, without inducing haemolytic anaemia.

The antitumor effects of FK317, a novel substituted dihydrobenzoxazine, were evaluated using human tumor xenografts (small cell lung cancer, non-small cell lung cancer, stomach cancer, colon cancer, pancreatic cancer, breast cancer, cervical cancer and ovarian cancer). Tumor growth-inhibitory effects and the effective dose-range of FK317 were much stronger and broader, respectively, than those of reference drugs such as mitomycin C, adriamycin, cisplatin, taxol and irinotecan. Furthermore, the body weight decrease and myelosuppression in FK317-treated mice were less than in the animals given any of the reference drugs. To explain this tumor selectivity, the distribution of FK317 was investigated after dosing tumor-bearing mice with the 14C-labelled compound. The concentration of FK317 in tumor tissues was relatively low, and long tumor retention was not observed. However, thin-layer chromatographic separation revealed that the radioactivity in the tumor resided mainly in strongly cytotoxic metabolites, while that in other tissues resided mainly in non-cytotoxic metabolites. These results suggest that FK317 shows strong antitumor activity without side effects, and one reason for this is its specific metabolite pattern. FK317 is now undergoing phase I clinical trials.

The effects of FK317 (11-acetyl-8-carbamoyloxymethyl-4-formyl-6- methoxy-14-oxa-1,11-diazatetracyclo[7.4.1.0(2, 7). 0(10, 2] tetradeca-2,4,6-trien-9-yl acetate), a novel anti-cancer agent, on murine adenocarcinoma colon26- and human lung carcinoma LX-1-induced cachexia were investigated in mice. Mice bearing colon26 or LX-1 s.c. lost weight and became cachectic, associated with tumor growth. FK317 and mitomycin C (MMC) inhibited the growth of both tumors. FK317 ameliorated the weight loss induced by the presence of colon26 or LX-1, while MMC enhanced it. An attenuation of the reduction in the weights of epididymal fat, gastrocnemius muscle and carcass was observed in FK317-treated tumor-bearing mice in both cachexia models, but not in MMC-treated mice. The decreases in the circulating levels of triglyceride, glucose and non-esterified fatty acid, which were induced by the presence of colon26, was partially inhibited by treatment with FK317. Overall, this study revealed that FK317 is a potent anti-cancer drug with anti-cachectic activity, suggesting that FK317 has potential utility for the treatment of cancer.

FK317, a novel substituted dihydrobenzoxazine, was examined for antitumor effects on multidrug-resistant (MDR) tumor cells in vitro and in vivo. In nude mice, FK317 markedly inhibited the growth of s.c. implanted KB-V1 vinblastine (VLB)-resistant human epidermal carcinoma KB cells, as well as the parent cells (KB-3-1). However, KB-V1 showed much greater resistance to FK317 than to VLB and adriamycin (ADM) in the in vitro study. This resistance was reversed by the addition of verapamil, whereby intracellular accumulation of FK317 in the KB-V1 cells was also decreased. After incubation of FK317 in human and mouse blood, it was shown to be rapidly metabolized to a monodeacetylated form, and slowly metabolized further to a dideacetylated form. With the removal of the acetyl groups from FK317, resistance indexes in KB-V1 and SBC-3/ADM, ADM-resistant human lung carcinoma, decreased. In addition, photolabeling of P-glycoprotein with [3H]azidopine in KB-V1 plasma membrane was completely inhibited by FK317, but not by the deacetylated metabolites. These results indicate that FK317 is metabolized to deacetylated forms, which do not bind to P-glycoprotein and are incorporated into MDR cells, causing cytotoxic effects.

FK317 is a member of a new class of bioreductive agents that exhibit strong cytotoxicity against various human cancer cells. The effect of FK317 was found to be stronger than that of mitomycin C (MMC), adriamycin (ADR) or cisplatin (CDDP). Alkaline elution analysis indicated that FK317 formed interstrand DNA-DNA and DNA-protein cross-links in cells. On the other hand, no DNA single-strand breaks were observed in the cells treated with FK317. In a cell-free system the deacetylated metabolites produced cross-linked DNA under reductive conditions, though FK317 itself did not form DNA-DNA cross-links. In order to elucidate the metabolic activation mechanisms, we established an FK317-resistant subline from human non-small cell lung cancer cells (Lu99) by stepwise and brief exposure (1 h) to FK317. The resistant subline (Lu99/317) showed cross-resistance to MMC and carboquone (CQ), but not to ADR or CDDP. DT-diaphorase, which is one of the activation enzymes of MMC and CQ, was deficient in Lu99/317 cells as determined by enzyme activity assay. However, the levels of NADPH:cytochrome P450 reductase, which is another activation enzyme for MMC and CQ, were comparable in resistant and parent cell lines. Treatment of the cells with dicumarol, an inhibitor of DT-diaphorase, reduced the cytotoxicity of FK317 to Lu99 cells, but not to Lu99/317 cells. These results indicate that deacetylation of FK317 is necessary for its reductive activation, and deacetylated FK317 is reduced by DT-diaphorase to form an active metabolite, which produces DNA-DNA interstrand and DNA-protein cross-links that lead to cell death.

A series of dapsone-related 4-aminopheynl and 2-aminothiazolyl derivatives was prepared, and their antinephritic activity and blood toxicity were evaluated. 5-(2-Pyridylsulfonyl)-2-thiazolamine (FR115092, 26) was effective against two nephritis models, namely graft-versus-host disease (GVHD) and autoimmune W/BF1 mice, and showed none of the blood toxicity observed with dapsone.

Cancer cachexia, characterized by weight loss and progressive tissue wasting, has been postulated to be mediated by cytokines. In this study the effect of FR143430, (2-(4-fluorophenyl)-4, 5, 6, 7-tetrahydro-3-(4-pyridyl)pyrazolo[1, 5-a]pyrimidine monohydrochloride), an inhibitor of Interleukin-1 and Tumor necrosis factor-a (TNF- a), on adenocarcinoma colon26-induced cachexia was investigated in mice. Tumor growth was not affected. Nevertheless, treatment with FR143430 (0.1 to lmg) into the tumor resulted in the attenuation of the reduction in body weight, food intake, epididymal fat and carcass weight, the decrease in the circulating levels of triglyceride and glucose, and the increase in the circulating levels of total cholesterol, non esterified free fatty acid (NEFA) and total protein, which were induced by the presence of the tumor. However, oral treatment with FR143430 failed to show an inhibitory effect on cachexia induction. Overall, this study demonstrated that the cachexia induced by colon26 was alleviated by the injection of FR143430 into the tumor in sufficient quantity, without any effect on tumor growth, suggesting the potential utility of cytokine suppressive agents e for the treatment of cancer cachexia.

Purpose: FK973, a substituted dihydrobenzoxazine, is an antitumor antibiotic which has shown high therapeutic efficacy in a phase I study, but its development has been abandoned because of the side effect of vascular leak syndrome (VLS) in the clinical study. This study was performed to investigate whether or not FK317, a new benzmethoxy derivative of FK973, retains the antitumor activity of FK973 without the side effect of VLS., Methods: VLS was evaluated by the volume of pleural effusion in rats. Cytotoxic activities were determined by a tetrazolium-based colorimetric assay (MTT assay) against murine (B16, P388) and human (HeLa S3, KB) tumor cell lines. Antitumor activities against murine ascitic leukemia (P388, L1210), murine solid tumors (reticulum cell sarcoma M5076, Colon 38 carcinoma) and human xenografts (mammary carcinoma MX-1, lung carcinoma LX-1) were examined., Results: FK973 (1.8 mg/kg) given i.v. to rats induced pleural effusion, one of the elements of VLS, 36 days after the first dosing, but did not 28 days after dosing. This model reflects clinical VLS delayed-type effusion with high protein concentrations. In contrast, FK317 (1.0-3.2 mg/kg) did not induce pleural effusion at all. FK317 had stronger cytotoxic effects against in vitro cultured B16, P388, HeLa S3 and KB tumor cell lines, and in in vivo experiments, FK317 showed equivalent antitumor activity against P388, M5076 and MX-1, and more potent antitumor activity against L1210, Colon 38 and LX-1 compared with FK973., Conclusion: These results suggest that FK317 retains the antitumor activity of FK973 and does not induce VLS, and FK317 is a drug with high clinical potential for treating tumors in humans.

Home medical care is becoming a greater matter of concern along with the increasing population of elderly persons. The various difficulties in home care are also found in regional care, and it could be said that in Japan regional care serves as an "advanced model" for home medical care. Ohya Town is located in the mountains of northern Hyogo Prefecture. It has a population of about 5,000 of which 32% is elderly people. There is no railroad, and it takes around 30 minutes to travel to the central hospital from the area. The three town clinics are in contact with each other and with the central hospital concerning medical information. The characteristics of Home Parenteral Nutrition in this underpopulated district are discussed and reported in reference mainly to cases in which the 3 way valved peripherally inserted central venous catheter (Groshong Catheter) is used, in comparison with HPN cases in the city university hospital.

The pharmacological and antitumor effects of FR901537, a new aromatase inhibitor, isolated from Bacillus sp. No. 3072, were studied. Treatment for four consecutive days with FR901537 inhibited the androstenedione-induced increase in the uterus weight in immature rats. FR901537 had no effect on the uterus, adrenal glands, ovary or pituitary weights in mature rats following 14 days of treatment. The antitumor activity of FR901537 on 7,12-dimethylbenz(a)anthracene-induced mammary tumors was studied in ovariectomized, testosterone propionate (TP)-treated rats as a postmenopausal tumor model. Ovariectomy caused the regression of the mammary tumors and the growth of tumors was remarkably stimulated following TP treatment. Further, in the rats treated with FR901537 and TP, the TP-induced tumor growth was significantly inhibited by FR901537. These results suggest that FR901537 is a promising drug in the treatment of estrogen-dependent mammary tumors in postmenopausal women.

Recombinant tumor necrosis factor-alpha (rTNF-alpha) inhibited tumor growth of Meth A fibrosarcoma (Meth A) solid tumor in mice, and the antitumor effect of rTNF-alpha was significantly decreased by pretreatment with small doses or rTNF-alpha in mice. In in vitro experiments, incubation of human umbilical vein endothelial cells with rTNF-alpha enhanced procoagulant activity (PCA), which was drastically augmented after an addition of the conditioned medium of Meth A tumor cells. Furthermore, rTNF-alpha-induced PCA was decreased by pretreatment with rTNF-alpha in endothelial cells. This PCA was completely blocked after the addition of anti-human tissue factor (TF) murine monoclonal antibody. These results imply that in vivo antitumor effects of rTNF-alpha are mediated by expression of TF in endothelial cells, which is augmented by tumor released factor(s).

Tumor necrosis factor-alpha (TNF-alpha) was reported to be important in the induction of septic shock. After i.v. injection of recombinant TNF-alpha (rTNF-alpha), BALB/c mice bearing Meth A fibrosarcoma (Meth A), but not normal mice, died of shock. Tumor cells are known to release many biological components. In this study, we examined the role of the tumor in the toxicity of rTNF-alpha in mice. Meth A cells maintained i.p. in mice were cultured for 24 hr in vitro. Conditioned medium (CM) obtained from the Meth A cells was given i.v. to mice, and 2 to 7 days later, i.v. injection of rTNF-alpha induced death in the animals. rTNF-alpha treatment 4 days after Meth A CM gave the maximum effect. rTNF-alpha did not induce death in mice treated with CM from spleen cells. However, after the Meth A cells were passaged 2 or 3 times in in vitro culture, the CM did not potentiate the toxicity of rTNF-alpha in mice. rTNF-alpha induced symptoms of disseminated intravascular coagulation (DIC) on coagulation parameters in the blood, and high plasma tissue factor (TF) activity in Meth A CM-treated mice and Meth A tumor-bearing mice. These results suggest that factor(s) are released from tumor cells activated by interaction with host cells, and injection of rTNF-alpha and the factor(s) results in the induction of DIC syndrome leading to host death.

The binding sites, other than the estrogen receptor (ER), of the antiestrogens droloxifene (DROL, (E)-a-[p-[2-(dimethylamino)ethoxy]-phenyl]-a'-ethyl-3-stilbenol) and tamoxifen (TAM), and estradiol-17 beta (E2) in the cytosol of 7,12-dimethylbenz[a]anthracene-induced rat mammary ER-positive tumor cells were studied using a high-performance liquid chromatography (HPLC) gel filtration assay. The cytosol was incubated with 3H-labeled drug with or without unlabeled drug, and separated by HPLC gel filtration. 3H-E2 produced two major peaks of radioactivity at fractions No. 40 and No. 70. The peak at fraction No. 70 was identified as the ER in an ER-enzyme-immuno assay. This peak was dose-dependently inhibited by unlabeled DROL or TAM, DROL being a more potent inhibitor than TAM. The peak at fraction No. 40 was also inhibited by co-incubation with unlabeled DROL or TAM. 3H-DROL or 3H-TAM provided only one peak at fraction No. 43. This peak was thought to be an antiestrogen binding site (AEBS), because it was inhibited by unlabeled antiestrogen but not by E2. The results suggest that the antiestrogens DROL and TAM have a higher affinity for the AEBS than for the ER in the absence of E2, while in the presence of E2 both have an affinity for the ER and inhibit E2 binding to the ER.

The antitumor activities of FR901228, (E)-(1S,4S,10S,21R)-7-[(Z)- ethylidene]-4,21-diisopropyl-2-oxa-12,13-dithia-5,8,20,23- tetraazabicyclo[8,7,6]-tricos-16-ene-3,6,9,19,22-pentanone, isolated from Chromobacterium violaceum No. 968, were studied in animals. FR901228 (ip) prolonged the life of mice bearing such murine ascitic tumors as P388 and L1210 leukemias and B16 melanoma, and inhibited (iv) the growth of murine solid tumors (Colon 38 carcinoma, M5076 reticulum cell sarcoma and Meth A fibrosarcoma) and human solid tumors (Lu-65 and LC-6 lung carcinomas, and SC-6 stomach adenocarcinoma) implanted in normal and nude mice, respectively. Its antitumor activity was especially potent against murine Meth A fibrosarcoma and human SC-6 stomach adenocarcinoma which were refractory to mitomycin C or cisplatin. FR901228 also was more effective against mitomycin C-, cyclophosphamide-, vincristine- and 5-fluorouracil-resistant P388 leukemias than against non-resistant P388 in mice. These results suggest that FR901228 will be a new type of drug for the treatment of cancer.

Insulin-like growth factor-I (IGF-I) is an important mitogen in breast cancer. We studied here the effects of a new antiestrogen drug, droloxifene (DROL, (E)-alpha-[p-[2-(dimethylamino) ethoxy]-phenyl]-alpha'-ethyl-3-stilbenol) and tamoxifen (TAM) on the IGF-I-stimulated growth of estrogen receptor (ER) positive breast cancer cells, MCF-7 and their mechanism of action. IGF-I secretion from MCF-7 was increased by the addition of estrogen. Externally added IGF-I stimulated the growth of MCF-7 but not ER negative breast cancer cells, MDA-MB-231. DROL and TAM inhibited the IGF-I-stimulated growth of MCF-7. A 2 hr treatment with both drugs did not block IGF-I binding to the receptors in MCF-7. However, a 4 day treatment with DROL decreased the number of IGF-I receptors without altering the binding affinity in MCF-7. These results suggest that DROL can exert its antitumor activity against ER positive breast cancer not only by blocking the E2 binding to the ER, but also by counteracting the mitogenic effect of IGF-I.

The induction of macrophage colony-stimulating factor (M-CSF) in monkey plasma following administration of FK565 was observed within 2 h of injection peaked at 4 h, and remained high after 24 h. Interleukin-6 (IL-6) and M-CSF levels increased in monkeys treated with FK565, even at doses as low as 0.01 mg/kg. Granulocyte CSF (G-CSF) levels increased slightly following a dose of 1 mg/kg, but granulocyte macrophage CSF (GM-CSF) was not detected at any doses of FK565 studied. To examine the thrombopoietic activity of FK565 in vivo, single doses of drug (0.01, 0.1 or 1.0 mg/kg) were administered i.v. to cynomolgus monkeys or normal mice on day 0. The promotes platelet (PLT) count after FK565 injection decreased transiently on days 1 and 2, and then increased in a dose-dependent manner on day 5 and was still high on day 14. The experiment using anti-PLT antibody showed that the increased PLT count was not simply due to a rebound phenomenon after the transient decrease in PLT. The effect of i.v. FK565 was studied in mice myelosuppressed with a single dose of mitomycin C (MMC) (5.6 mg/kg). The fall in PLT count was suppressed on day 7 by 0.1 and 1.0 mg/kg FK565. Although intact cells or tissues are necessary for an increase in PLT following FK565 treatment, FK565 suppressed the impaired hematopoietic function seen after chemotherapy. FK565 is proposed as a drug to restore reduced neutrophil and platelet counts found in AIDS or cancer therapy.

Pretransplant tests necessary for kidney transplantation are HLA typing, mixed lymphocyte culture response (MLR), and direct crossmatch. HLA typing and MLR are closely related to graft survival rates. The significance of HLA matching is generally known. In our analysis of 25 living related kidney grafts, graft survival rate of two haplo identical donor transplants was 4/4 (100%), while that of one haplo identical donor transplant was 18/21 (85.7%). Acute rejection rate in the MLR low response group (S.I. less than or equal to 5) was 2/7 (28.6%), while that in the high response group (S.I. greater than 5) was 11/19 (57.9%). HLA typing and MLR are useful in selecting the most suitable recipient. In order to reduce the risk of hyperacute or accelerated graft rejection, T warm direct crossmatch is performed. Anti-T warm antibodies are mainly produced by blood transfusion. In our study of 239 hemodialysis patients, there were 31 patients with positive T warm (13.0%), the positive rate became higher in proportion to increases in blood transfusion. Recently, there have been reports of kidney transplants successfully performed across T warm-positive crossmatches due to IgM antibodies. We also investigated the immunoglobulin class. Not only pretransplant crossmatch but also posttransplant crossmatch is necessary. In the case of accelerated, acute or chronic rejection, anti-donor HLA antibodies are produced in patients' peripheral blood. Thus, the test of posttransplant anti-donor antibodies is useful for the early detection of rejection, its diagnosis, and index of the prognosis. The sensitivity of flow cytometry crossmatches was compared to standard cytotoxicity crossmatch. Titration studies indicate a 32-64 fold range of greater sensitivity than the cytotoxicity test.(ABSTRACT TRUNCATED AT 250 WORDS)

The antitumor effects of recombinant human tumor necrosis factor-alpha (rTNF-alpha) and 5-fluorouracil (5-FU) in combination treatment were examined on Meth A fibrosarcoma implanted intradermally in mice. Growth of the tumor was inhibited when rTNF-alpha was given i.v. on day 7 or 11 after implantation, but the effect was countered when 5-FU was additionally given i.p. once a day on days 1-4 after implantation. Conversely, 5-FU given on days 5-8 after implantation augmented the antitumor effects of rTNF-alpha. Injection of carbon particles showed that fine capillaries did not develop in the tumors of mice treated with 5-FU on days 1-4 after implantation, but that a delicate network of capillaries developed in the tumors of both the mice treated with 5-FU on days 5-8 after implantation and the controls given saline. The results show that the timing of 5-FU treatment is important when attempting to enhance the antitumor effects of rTNF-alpha, and suggest that these effects are directly associated with newly formed fine capillaries in the tumor.

The possible involvement of chemical mediator(s) in the induction of the antitumor effects of recombinant human tumor necrosis factor-alpha (rTNF-alpha) on Meth A fibrosarcoma (Meth A) in mice was studied. On day 7 after intradermal implantation of Meth A in mice, rTNF-alpha caused tumor necrosis and inhibited the tumor growth. Ketanserin, cyproheptadine, and spiperone [serotonin (5-HT) receptor blockers] inhibited or attenuated the antitumor effects of rTNF-alpha, but the other types of receptor blockers tested (histamine H1 and H2, adrenaline alpha and beta, dopamine, and acetylcholine receptor blockers) did not. The large i.v. doses of 5-HT caused tumor necrosis and inhibited tumor growth in mice when given i.v. on day 7 but not when given on day 3 after Meth A implantation, which effects closely resemble those of rTNF-alpha. Its anti-tumor effects were completely inhibited by the 5-HT receptor blockers. 5-HT, like rTNF-alpha, showed no cytotoxicity against in vitro cultured Meth A cells. The results suggest that 5-HT is, at least in part, important for the induction of antitumor effects of rTNF-alpha on Meth A in mice.

The antitumor activity of recombinant human tumor necrosis factor (rTNF-alpha) was examined on murine tumors in mice and in cultured cells in vitro. Mice were implanted intradermally with Meth A fibrosarcoma (Meth A) on day 0. rTNF-alpha caused tumor necrosis and inhibited the tumor growth when given i.v. on day 7 or 10, but not when given on day 3. When rTNF-alpha was given i.v. in doses of 0.1-3.2 micrograms/mouse twice a week for 3 weeks beginning on day 7 or 11, the growth of solid Meth A, Colon 26 adenocarcinoma, Colon 38 carcinoma, Sarcoma-180, and M5076 reticulum cell sarcoma tumors implanted s.c. or intradermally was markedly inhibited, and the life of the mice bearing these tumors, except M5076 reticulum cell sarcoma, was prolonged. The growth of Meth A implanted i.m. was also markedly inhibited by rTNF-alpha given i.v. However, the life of mice bearing i.p. Colon 26 adenocarcinoma, MH134 hepatoma, Sarcoma-180, and Ehrlich carcinoma was not prolonged by rTNF-alpha given i.p. nine times (days 1-9) in doses up to 1.0 or 3.2 micrograms/mouse. Only in the case of mice bearing i.p. Meth A, the life was slightly prolonged by i.p. treatment with rTNF-alpha but not by i.v. treatment. In experiments against in vitro cultured cells, rTNF-alpha did not show any direct cytotoxicity against mouse tumor cells: Meth A, Colon 26 adenocarcinoma, Colon 38 carcinoma, and Sarcoma-180, but had a cytotoxic effect against L929 mouse fibroblast. The results suggest that rTNF-alpha is a unique antitumor drug with potent necrotizing activity against solid tumors in mice, and that this activity may derive from indirect mechanisms related to the growth of tumors and not to the direct cytotoxicity of the drug.

It is important to examine the presence of preformed antibodies in hemodialysis patients before renal transplantation, because it causes the graft rejection in organ transplantation. In the 239 hemodialysis patients we analysed 3 preformed antibodies divided TW, BW and BC. Sensitized rate (TW+, BW+ and BC+) was 0% at no blood transfusions (BT), 9.3% at 1-5 BT and 28.9% at 6 more BT. The number of blood transfusions increased the sensitized rate (TW). Sensitized rate (BW+, BC+) was 6.2% at no BT, 9.3% at 1-5BT and 12.0% at 6 more BT, while sensitized rate (BC+) was 33.3% at no BT, 26.7% at 1-5BT and 14.5% at 6 more BT. There was no correlation between the number of blood transfusions and the sensitized rate (BW+, BC+). At one year after the first check, 69.2% of TW positive patients was converted to TW negative and 53.8% of BW positive patients was converted to BW negative each. Further studies of the preformed antibodies are required to assess the clinical course of long surviving patients.