Safety and systemic absorption of pulmonary delivered human IFN-beta1a in the nonhuman primate: comparison with subcutaneous dosing.

Safety and bioavailability of pulmonary delivered interferon-beta 1a (IFN-beta1a, AVONEX, Biogen, Inc., Cambridge, MA) was evaluated in the nonhuman primate. Pulmonary bioavailability following intratracheal (i.t.) instillation of 50 microg/kg IFN-beta1a to rhesus macaques was approximately 10%. To evaluate pulmonary safety, IFN-beta1a was administered intrabronchially to rhesus and cynomolgus macaques at a dose of 60 microg/dose one, three, or seven times per week for 4 weeks. At scheduled termination, lungs were evaluated for gross and histomorphologic changes. IFN-beta1a or vehicle (human serum albumin [HSA] in phosphate-buffered saline [PBS]) treatment resulted in minimal to mild subchronic alveolitis, located primarily near the instillation sites. These responses were considered nonspecific and consistent with either instillation of a foreign protein or minor injury associated with the instillation procedure. In one rhesus macaque treated every day for 4 weeks, IFN-beta1a induced mild to moderate eosinophilic alveolitis, considered possibly an isolated type I hypersensitivity response to HSA or IFN-beta1a. Partial resolution of pulmonary lesions was seen in all recovery animals killed 2 weeks after cessation of treatment. In conclusion, this study shows that pulmonary administration of human IFN-beta1a is safe and that the pulmonary route of administration is a possible alternate route for the systemic delivery of IFN-beta1a.     

Pharmacokinetics and pharmacodynamics of IFN-beta 1a in healthy volunteers.

The pharmacokinetics of recombinant human interferon-beta1a (IFN-beta1a) (Rebif, Ares-Serono, Geneva, Switzerland) were investigated in healthy volunteers following intravenous (i.v.) administration of increasing single doses of the drug (22 microg/6 million international units [MIU], 44 microg/12 MIU, and 66 microg/18 MIU); i.v., intramuscular (i.m.), and subcutaneous (s.c.) administration of a 66-microg dose; and repeated s.c. administration of four 66-microg doses at 48-h intervals. The disposition of IFN-beta1a followed triexponential decay after i.v. administration (half-lives 3 min, 42 min, and 22 h, respectively). After s.c. and i. m. administration, absorption was the rate-limiting factor in the terminal phase. The median absolute bioavailabilities were 30% and 27%, respectively. The accumulation ratio after repeated s.c. injections was 2.4, and a terminal half-life of 66 h was observed. Intracellular 2-5A synthetase activity and serum neopterin and beta2-microglobulin concentrations increased after all IFN-beta1a injections and remained elevated following every-other-day administration. The local tolerance was good, and the systemic tolerance was satisfactory.     

Pharmacokinetics and hematological effects of the PEGylated thrombopoietin peptide mimetic GW395058 in rats and monkeys after intravenous or subcutaneous administration

GW395058, a potent PEGylated peptide human thrombopoietin receptor (HuTPOr) agonist in vitro, is being evaluated for the treatment of thrombocytopenia. GW395058 shares no sequence homology with TPO. In this report the pharmacokinetics and hematological effects of GW395058 in rats and monkeys are described. Doses eliciting thrombocytosis in rodents (2 or 10 microg/kg s.c.) produced insufficient plasma concentration data for pharmacokinetic parameter estimate calculations. At higher i.v. doses in rats (500, 1,000 or 2,000 microg/kg) serum t1/2 (half-life) values were >20 h, and the area under the concentration time curve increased proportionally with dose. In cynomolgus monkeys GW395058 plasma t1/2 values ranged from 37 to 68 h after s.c. or i.v. dosing, and similar values were observed in rhesus monkeys following s.c. dosing. Rat platelet counts increased following 2 (1.6-fold) or 10 microg/kg (fourfold) s.c. doses. Cynomolgus and rhesus monkey platelet counts did not change significantly at comparable s.c. doses, but did increase slightly (相似文献   

Induction of tolerance to recombinant therapeutic proteins.

The specific IgM and IgG antibody responses to subcutaneous (s.c.) treatment of mice with recombinant human IFN-alpha2a (rHuIFN-alpha2a) or IFN-beta were inhibited in a dose-dependent manner by prior oromucosal (o.m.) administration of rHuIFN-alpha2a or IFN-beta, respectively. Pretreatment of animals once a day for 7 days by the o.m. route with the highest dose of IFN-alpha2a tested (10(7) IU) resulted in complete inhibition of the peak IFN-alpha2a-specific IgG antibody response detected 28 days after subsequent s.c. injection of IFN-alpha2a (p < 0.001). Similarly, prior o.m. administration of 1-10 microg rHuGM-CSF per day for 7 days resulted in a statistically significant (p < 0.001) inhibition of the peak GM-CSF-specific IgG antibody response detected 28 days after s.c. administration of GM-CSF. In contrast, prior o.m. treatment with a quantity of bovine serum albumin (BSA) (100 microg) or human serum albumin (HSA) (10 microg) equivalent, respectively, to the protein content of the highest dose of IFN-alpha2a or GM-CSF administered by the o.m. route, did not affect significantly the IFN-alpha2a-specific or GM-CSF-specific IgG antibody responses detected on subsequent s.c. administration of IFN-alpha2a or GM-CSF. Oromucosal administration of IFN-alpha2a, IFN-beta, or GM-CSF alone did not induce detectable IFN-alpha2a-specific, IFN-beta-specific, or GM-CSF-specific IgM or IgG antibody responses at any of the time points tested. These results suggest that short-term o.m. administration of a recombinant protein is an effective means of inducing peripheral tolerance to subsequent parenteral administration of a therapeutic protein.     

Pulmonary administration of interferon Beta-1a-fc fusion protein in non-human primates using an immunoglobulin transport pathway

Currently, products containing interferon beta (IFNβ) are injected either intramuscularly or subcutaneously. To avoid the necessity of injection, we developed a novel monomeric Fc fusion protein of IFNβ (IFNβFc) that is absorbed via an immunoglobulin transport system present in the upper and central airways upon administration of the drug as an inhaled aerosol. The systemic absorption of IFNβFc through the lung in non-human primates, at deposited doses of 1, 3, and 10?μg/kg, was compared to the absorption of a single 3?μg/kg dose of IFNβ-1a (Avonex?) subcutaneously administered. IFNβFc was well absorbed through the lung, displaying dose proportional increases in serum concentrations, and was biologically active, as shown by increases in plasma neopterin levels. The circulating half-life of IFNβFc was ～3 times longer (～30?h) than that of IFNβ-1a, (8-9?h). At approximately equimolar doses of IFNβFc (10?μg/kg) and IFNβ-1a (3?μg/kg), the stimulation of neopterin over background levels was approximately equivalent, demonstrating that the longer half-life of IFNβFc compensated for the lower relative specific antiviral activity of IFNβFc measured in vitro. In conclusion, IFNβFc was efficiently absorbed after pulmonary delivery in non-human primates, retained its biological activity, and may offer a convenient alternative to injectable IFNβ.     

Detection and plasma pharmacokinetics of an anti-vascular endothelial growth factor oligonucleotide-aptamer (NX1838) in rhesus monkeys.

Aptamers are oligonucleotide ligands selected, in vitro, to bind a specified target protein. The first aptamer to reach human clinical testing is NX1838, a polyethylene glycol conjugated aptamer that inhibits vascular endothelial growth factor. This paper describes the validation of a high-performance liquid chromatographic anion-exchange method for the determination of NX1838 in plasma. Measurements of intact NX1838 had a coefficient of variation of less than 8% and an accuracy between 107% and 115%. The assay was utilized to determine NX1838 plasma pharmacokinetics in rhesus monkeys following a single 1 mg/kg intravenous or subcutaneous dose. Following intravenous administration, the maximum achieved plasma concentration was 25.5 microg/ml with a terminal half-life of 9.3 h and clearance rate of 6.2 ml/h. After subcutaneous administration, the fraction of the dose absorbed into the plasma compartment was 0.78 with a time to peak concentration (4.9 microg/ml) of 8 to 12 h.     

重组人血清白蛋白/粒细胞刺激因子融合蛋白的药效学、药理学和毒理学研究

目的对重组人血清白蛋白/粒细胞刺激因子融合蛋白(rHSA/GCSF)开展药效学、药理学和毒理学动物评价研究,以确认其安全性和有效性。方法①对rHSA/GCSF开展的药效学研究内容主要有:以恒河猴骨髓细胞为研究系统,在体外条件下培养骨髓粒细胞-巨噬细胞集落(CFU-GM),计数不同浓度的rHSA/GCSF对CFU-GM数的影响的体外药效学评价;以小鼠~(60)Coγ照射、氟尿嘧啶注射液所致的鼠白细胞低下和对~(60)Coγ射线照射致食蟹猴白细胞低下的治疗作用。②药理学研究观察了rHSA/GCSF对小鼠中枢神经系统和狗呼吸及心血管系统功能的影响。③毒理学研究考察了rHSA/GCSF静脉和皮下给予小鼠、食蟹猴的急性毒性反应,以及长期和反复给药对大鼠和食蟹猴的安全性做出评价。④rHSA/GCSF的药代/毒代试验则是对~(125)I-rHSA/GCSF不同剂量单次皮下注射给予小鼠、大鼠后,rHSA/GCSF在各器官的分布、总放射性和TCA沉淀放射性的动力学参数测定,以及对不同剂量rHSA/GCSF连续给药食蟹猴后的血浆药物浓度及代谢动力学参数开展了考察。结果①rHSA/GCSF可以使骨髓的粒白细胞系(粒系)增生活跃,骨髓粒系造血细胞及成熟中性粒细胞均明显增多;对氟尿嘧啶所致小鼠白细胞减少症有明显的治疗作用,在使用化疗药早期,可以减缓白细胞的降低,特别是可以使粒白细胞提前恢复到正常水平;rHSA/GCSF可显著缩短食蟹猴白细胞减少症放疗模型产生的白细胞低下持续时间,使外周血白细胞加速恢复,尤其以中性粒细胞的增加为主,同时对红细胞和血小板的影响不大。②从获得的PD/PK数据t_(1/2)来看:rHSA/GCSF半衰期平均值约为38.6 h;单次皮下注射rHSA/GCSF,在500、1500、3000μg/kg剂量范围内对小鼠中枢神经系统无影响,与戊巴比妥钠无协同作用;单次皮下注射rHSA/GCSF在50、200μg/kg剂量范围内对狗呼吸和心血管系统无明显影响。实验表明rHSA/GCSF单次皮下和静脉注射给予小鼠的最大耐受量(MTD)≥37.5 mg/kg,单次皮下注射给予食蟹猴的最大耐受剂量为11.6 mg/kg:长期反复给药对大鼠的基本安全剂量为300μg/kg,对食蟹猴则是≥150μg/kg。结论 rHSA/GCSF融合蛋白每4天给药1次,对放、化疗所致的动物外周血白细胞减少症具有治疗作用,与市售常规rhGCsF每天注射1次相比具有长效作用。所获得的大量药效学、药代动力学、毒理学、毒代动力学的试验数据可供正在开展的临床试验研究参考,并具有很好的指导意义。     

Pharmacodynamic comparison of single doses of IFN-beta1a and IFN-beta1b in healthy volunteers.

In a study in 75 volunteers, preparations of interferon-beta1b (IFN-beta1b) and IFN-beta1a were compared in terms of the resulting serum concentrations of three biologic markers, neopterin, human Mx protein, and 2',5' oligoadenylate synthetase. Each preparation was tested at five dose levels, the middle dose being that recommended for use in patients with multiple sclerosis on the basis of large clinical trials. Five randomly chosen volunteers each received a single subcutaneous dose of one of the IFN or of IFN-beta1a given intramuscularly. The amounts of each marker induced were dose related. There were no major differences between the results with the two IFN or in the duration of the changes in the markers after the two routes of injection. The data indicated that 8 million international units (MIU) of IFN-beta1b and 6 MIU of IFN-beta1a had very similar effects. Even after the highest single dose tested, the increase in the biologic markers were not sustained for a full week.     

Pharmacokinetics of theophylline and caffeine after intravenous administration of aminophylline to premature neonates in Korea

Theophylline has been widely used to treat apnea of premature neonates. The purpose of this study was to compare the pharmacokinetic parameters of theophylline and caffeine after intravenous administration of aminophylline to seven Korean low-birthweight neonates with apnea to those in other countries. The serum concentrations of theophylline and caffeine were measured simultaneously by high-performance liquid-chromatography (HPLC). The mean (+/- S.E.M.) birth weight and gestational period were 1190 +/- 253 g and 31.5 +/- 1.99 weeks, respectively. The mean (+/- S.E.M.) theophylline maintenance dosage was 1.28 +/- 0.15 mg/kg (given as equivalent aminophylline solution) every six hours. The mean (+/- S.E.M.) volume of distribution, 0.937 +/- 0.232 l/kg, elimination rate constant, 0.0249 +/- 0.0095/h, elimination half-life, 32.1 +/- 12.1 h, and total body clearance, 21.7 +/- 6.18 ml/h/kg, of theophylline in Korean premature neonates were comparable to the values of neonates in other countries. For caffeine, the mean (+/- S.E.M.) elimination half-life was 95.1 +/- 25.4 h and the elimination rate constant was 0.0079 +/- 0.0024/h. The mean (+/- S.E.M.) serum concentrations of theophylline and caffeine on the sixth day after aminophylline infusion were 10.4 +/- 2.28 microg/ml (range, 6.38-13.4 microg/ml) and 2.94 +/- 0.98 microg/ml (range, 1.80-4.44 microg/ml), respectively. The mean (+/- S.E.M.) caffeine to theophylline concentration ratio on the day after discontinuation of aminophylline infusion was 0.71 +/- 0.23 (range, 0.39-1.03).     

Early clearance of circulating hepatitis C virus enhanced by induction therapy with twice-a-day intravenous injection of IFN-beta.

To improve the long-term efficacy of interferon (IFN) for treatment of chronic hepatitis C virus (HCV) infection, we proposed induction therapy with twice-a-day IFN-beta injection. This study was intended to clarify the antiviral mechanism. Thirty patients were randomly assigned to two groups: group A (twice-a-day therapy) received 3 MU IFN-beta intravenously (i.v.) twice a day for 2 weeks; group B (once-a-day therapy) received 6 MU of IFN-beta daily. HCV RNA, IFN-beta, alanine aminotransferase (ALT), 2'5'-oligoadenylate synthetase (2'5'-AS) activity, and beta2-microglobulin in serum were compared between the two groups during the first 2 weeks of IFN therapy. The clearance rate of serum HCV RNA in group A (86.7%) was significantly higher than that in group B (13.3%) at day 3 (p = 0.0006). No accumulation of IFN-beta was shown in serum throughout the therapy. The ratio (day 3/day 1) of 2'5'-AS activity was significantly higher in group A. Multivariate analysis indicated twice-a-day IFN-beta injection therapy led to significantly early clearance of circulating HCV. Twice-a-day IFN-beta injection therapy could induce biologically enhanced antiviral activities and be an efficient induction therapy for eradication of HCV.     

Extended activity in cynomolgus monkeys of a granulocyte colony-stimulating factor mutein conjugated with high molecular weight polyethylene glycol

The activity of a granulocyte colony-stimulating factor (G-CSF) mutein (nartograstim; [NTG]) conjugated with an average of two polyethylene glycol (PEG) chains per protein molecule was examined in cynomolgus monkeys following a single s.c. injection. Groups of monkeys were given 10 microg/kg, 30 microg/kg, or 100 microg/kg. For comparison, one group of monkeys was given 5 microg/kg of recombinant human G-CSF (rHuG-CSF) daily for six days. In monkeys given 100 microg/kg of PEG-NTG, neutrophil levels reached a peak one day after injection approximately 20-fold higher than baseline levels. Neutrophil numbers in these animals were still significantly elevated six days after injection. In contrast, peak neutrophil levels in monkeys given six injections of rHuG-CSF reached a peak only on day 6 and were approximately the same as that in monkeys given a single dose of PEG-NTG six days before. Pharmacokinetics of PEG-NTG in these monkeys indicated that the area under the plasma concentration time curve (AUC) increased with increasing the dose from 497 ng x h/ml at 10 microg/kg, 6,140 ng x h/ml at 30 microg/kg to 27,900 ng x h/ml at 100 microg/kg. In a separate study, the effects of single doses of 100 microg/kg of PEG-NTG, rHuG-CSF, and unmodified NTG were compared. In this experiment, peak numbers of neutrophils were reached two days after injection in animals receiving PEG-NTG and one day after in animals given unmodified proteins. The pharmacokinetic parameters demonstrated increased exposure for PEG-NTG relative to the unmodified proteins with an AUC0. of 21,012 ng x h/ml compared with 5,492 ng x h/ml for rHuG-CSF and 5,153 ng x h/ml for NTG. These results demonstrate that conjugation of a G-CSF mutein with high molecular weight PEG results in a preparation that can induce prolonged elevation of neutrophils in normal nonhuman primates following a single injection.     

Acute and chronic administration of immunomodulators induces anorexia in Zucker rats

To investigate the possible involvement of leptin signaling in lipopolysaccharide (LPS) anorexia, we compared the anorectic effect of LPS in genetically obese (fa/fa) Zucker rats and in their lean (Fa/?) counterparts. The effects of interleukin-1beta (IL-1beta) and muramyl dipeptide (MDP) were also tested. LPS [100 microg/kg body weight (BW)], IL-1beta (2 microg/kg BW) and MDP (2.2 mg/kg BW) injected intraperitoneally (i.p.) at lights out reduced food intake similarly in obese and lean rats. LPS injection at 500 or 1000 microg/kg BW (i.p.) also reduced food intake and BW similarly in obese and lean rats, but obese regained BW faster than lean rats. LPS (2.45 microg or 9.8 microg/h/rat) administered chronically with i.p. implanted osmotic pumps reduced food intake similarly on experimental day 1, regardless of the genotype. After day 3, the lean rats' anorectic response and recovery were dose-dependent, whereas the anorectic response in obese rats was minimally affected by dose (significant dose effect only on day 3). Again, obese rats regained lost BW faster than lean rats. These results do not support a role for leptin as the sole mediator of anorexia induced by bacterial products (LPS and MDP) and IL-1beta.     

Novel controlled-release Lemna-derived IFN-alpha2b (Locteron): pharmacokinetics, pharmacodynamics, and tolerability in a phase I clinical trial.

Locteron, a newly developed controlled-release formulation of Lemna-derived free (unpegylated) recombinant interferon-alpha2b (IFN-alpha2b, Biolex Therapeutics, Pittsboro, NC) in poly(ether-ester) microspheres (PolyActive, OctoPlus N.V., Leiden, the Netherlands), was evaluated in 27 volunteers injected with either 20, 80, or 320 microg Locteron (equivalent to 6.25, 25, or 100 x 10(6) IU, respectively), 80 microg pegylated IFN-alpha2b (PEG-IFN-alpha2b), microspheres not containing IFN-alpha2b, or placebo. Serum free or PEG-IFN-alpha2b and two biomarkers of IFN activity, neopterin and 2',5'-oligoadenylate synthetase (2',5'-OAS), were measured. After injection of 320 microg Locteron, serum IFN-alpha2b remained elevated through 14 days. The elimination half-life of Locteron was more than 2-fold that of PEG-IFN-alpha2b. The effects of 80 microg Locteron and 80 microg PEG-IFN-alpha2b on both neopterin and 2',5'-OAS were in a comparable range. Serum persistence of both these biomarkers was similar at 14 days after 320 microg Locteron compared with 7 days after 80 microg PEG-IFN-alpha2b. Mild, moderate, or severe influenza-like symptoms developed in all 6 subjects receiving 80 microg PEG-IFN-alpha2b. No such symptoms occurred after 20 or 80 microg Locteron doses. Among the 4 recipients of 320 microg Locteron, 1 experienced mild and 2 experienced moderate influenza-like symptoms. Locteron merits further clinical investigation as a hepatitis C therapy suitable for dosing once per 2 weeks.     

Interferon-beta1a administration results in a transient increase of serum amyloid A protein and C-reactive protein: comparison with other markers of inflammation

Putative markers of inflammation such as serum beta2-microglobulin and neopterin have been shown to be transiently upregulated following interferon-beta (IFN-beta) administration to multiple sclerosis (MS) patients. However, to date the role of the important inflammatory mediators serum amyloid A protein (SAA) and C-reactive protein (CRP) have not been described. Here we show that SAA but not CRP is elevated in relapsing-remitting MS patients compared to normal healthy individuals, and furthermore that both are transiently upregulated following intramuscular injection with IFN-beta1a (Avonex). This pattern of expression was found to parallel that of beta2-microglobulin and neopterin following injection and was mirrored by a selective activation of peripheral monocytes with respect to upregulation of receptors known to be involved in the inflammatory response (HLA-DR, CD16 and CD86). Injection of saline solution intramuscularly to six healthy control individuals did not produce a similar upregulation of any of the inflammatory markers investigated. Following IFN-beta1a injection, all inflammatory responses were attenuated at week 12 of therapy in comparison to those following the initial injection in a group of follow-up patients. In addition, IFN-beta1a injected on a weekly basis did not produce a sustained modulation of any of the markers investigated in patients treated for 32 weeks.     

Randomized multicenter phase II trial of subcutaneous recombinant human interleukin-12 versus interferon-alpha 2a for patients with advanced renal cell carcinoma.

Recombinant human interleukin-12 (rHuIL-12) is a pleiotropic cytokine with anticancer activity against renal cell carcinoma (RCC) in preclinical models and in a phase I trial. A randomized phase II study of rHuIL-12 compared with interferon-alpha (IFN-alpha) evaluated clinical response for patients with previously untreated, advanced RCC. Patients were randomly assigned 2:1 to receive either rHuIL-12 or IFN-alpha2a. rHuIL-12 was administered by subcutaneous (s.c.) injection on days 1, 8, and 15 of each 28-day cycle. The dose of IL-12 was escalated during cycle 1 to a maintenance dose of 1.25 microg/kg. IFN was administered at 9 million units by s.c. injection three times per week. Serum concentrations of IL-12, IFN-gamma, IL-10, and neopterin were obtained in 10 patients treated with rHuIL-12 after the first full dose of 1.25 microg/kg given on day 15 (dose 3) of cycle 1 and again after multiple doses on day 15 (dose 6) of cycle 2. Thirty patients were treated with rHuIL-12, and 16 patients were treated with IFN-alpha. Two (7%) of 30 patients treated with rHuIL-12 achieved a partial response, and the trial was closed to accrual based on the low response proportion. IL-12 was absorbed rapidly after s.c. drug administration, with the peak serum concentration appearing at approximately 12 h in both cycles. Serum IL-12 concentrations remained stable on multiple dosing. Levels of IFN-gamma, IL-10, and neopterin increased with rHuIL-12 and were maintained in cycle 2. rHuIL-12 is a novel cytokine with unique pharmacologic and pharmacodynamic features under study for the treatment of malignancy and other medical conditions. The low response proportion associated with rHuIL-12 single-agent therapy against metastatic RCC was disappointing, given the preclinical data. Further study of rHuIL-12 for other medical conditions is underway. For RCC, the study of new cytokines is of the highest priority.     

Identification and characterization of two novel staphylococcal enterotoxins, types S and T

In addition to two known staphylococcal enterotoxin-like genes (selj and selr), two novel genes coding for two superantigens, staphylococcal enterotoxins S and T (SES and SET), were identified in plasmid pF5, which is harbored by food poisoning-related Staphylococcus aureus strain Fukuoka 5. This strain was implicated in a food poisoning incident in Fukuoka City, Japan, in 1997. Recombinant SES (rSES) specifically stimulated human T cells in a T-cell receptor Vbeta9- and Vbeta16-specific manner in the presence of major histocompatibility complex (MHC) class II(+) antigen-presenting cells (APC). rSET also stimulated T cells in the presence of MHC class II(+) APC, although its Vbeta skewing was not found in reactive T cells. Subsequently, we examined the emetic activity of SES and SET. We also studied SElR to determine emetic activity in primates. This toxin was identified in previous studies but was not examined in terms of possession of emetic activity for primates. rSES induced emetic reactions in two of four monkeys at a dose of 100 microg/kg within 5 h of intragastric administration. In one monkey, rSET induced a delayed reaction (24 h postadministration) at a dose of 100 microg/kg, and in the other one, the reaction occurred 5 days postadministration. rSElR induced a reaction in two of six animals within 5 h at 100 microg/kg. On this basis, we speculate that the causative toxins of vomiting in the Fukuoka case are SES and SER. Additionally, SES, SER, and SET also induced emesis in house musk shrews as in the monkeys.     

人血清白蛋白对近端肾小管上皮细胞骨调素和CD44表达的影响

目的：研究人血清白蛋白能否刺激人近端肾小管上皮细胞产生骨调素（OPN）和CD44。方法： 用人血清白蛋白刺激人近端肾小管上皮细胞系（HK-2细胞），分不同时间（0-48 h）、不同浓度（0.1-10 g/L）两组，以RT-PCR方法分别检测两组细胞表达的OPN mRNA，以Western blotting分别检测两组细胞表达的OPN。用免疫荧光法、激光共聚焦显微镜观察1 g/L的人血清白蛋白刺激HK-2细胞24 h、48 h后OPN、CD44的表达。结果： 人血清白蛋白刺激HK-2细胞上调表达OPN mRNA和蛋白，呈时间和剂量相关性。CD44的表达与OPN同步。 结论： 人血清白蛋白可刺激HK-2细胞表达OPN与CD44。     

Leridistim, a chimeric dual G-CSF and IL-3 receptor agonist, enhances multilineage hematopoietic recovery in a nonhuman primate model of radiation-induced myelosuppression: effect of schedule, dose, and route of administration.

Leridistim is from the myelopoietin family of proteins, which are dual receptor agonists of the human interleukin-3 and G-CSF receptor complexes. This study investigated the effect of dosage, administration route, and schedule of leridistim to stimulate multilineage hematopoietic recovery in total body irradiated rhesus monkeys. Animals were x-irradiated on day 0 (600 cGy, 250 kVp) and then received, on day 1, leridistim s.c. in an abbreviated, every-other-day schedule at 200 microg/kg, or daily at 50 microg/kg, or i.v. daily or every-other-day schedules at 200 microg/kg dose. Other cohorts received G-CSF (Neupogen((R)) [Filgrastim]) in an every-other-day schedule at 100 microg/kg/day, or autologous serum (0.1%) s.c. daily. Hematopoietic recovery was assessed by bone marrow clonogenic activity, peripheral blood cell nadirs, duration of cytopenias, time to recovery to cellular thresholds, and requirements for clinical support. Leridistim, administered s.c. every other day, or i.v. daily, significantly improved neutrophil, platelet, and lymphocyte nadirs, shortened the respective durations of cytopenia, hastened trilineage hematopoietic recovery, and reduced antibiotic and transfusion requirements. A lower dose of leridistim administered daily s.c. enhanced recovery of neutrophil and platelet parameters but did not affect lymphocyte recovery relative to controls. Leridistim, a novel engineered hematopoietic growth factor administered at the appropriate dose, route and schedule, stimulates multilineage hematopoietic reconstitution in radiation-myelosuppressed nonhuman primates.     

Monocyte-derived dendritic cells release neopterin

Increased neopterin concentrations in body fluids are found in diseases associated with activated, cell-mediated immunity including infections, autoimmune diseases, and certain malignancies. Monocytes/macrophages are known to secrete large amounts of neopterin upon stimulation with interferon-gamma (IFN-gamma). Ontogenetically, the major part of dendritic cells (DC) belongs to the myeloid lineage. Therefore, we investigated whether cultured monocyte-derived DC can elaborate neopterin. Cells were treated with cytokines in the presence or absence of monocyte-conditioned medium as a maturation stimulus. DC secreted an average 3.5 nmol/l neopterin. In response to IFN-gamma, cells significantly increased their output of neopterin. In distinction to monocytes/macrophages, neopterin production in DC was highly sensitive to IFN-alpha and IFN-beta. Further, lipopolysaccharides (LPS) enhanced neopterin synthesis, whereas tumor necrosis factor alpha, interleukin (IL)-1beta, IL-2, IL-10, and IL-18 were ineffective. Simultaneously, tryptophan degradation by induction of indoleamine (2,3)-dioxygenase (IDO) was tested in stimulated cells. Our results showed that IFN-gamma as well as LPS are inducers of IDO in DC.     

A single dose of pegylated leridistim significantly improves neutrophil recovery in sublethally irradiated rhesus macaques.

Leridistim, a member of the myelopoietin family of dual receptor agonists that binds interleukin-3 and G-CSF receptors, has been shown to enhance hematopoietic activity in rhesus monkeys above that observed with either cytokine alone or in combination. This study demonstrated the ability of a pegylated form of leridistim (peg-leridistim), administered s.c., as a single- or two-dose regimen separated by 4 or 7 days, to significantly improve neutrophil recovery following radiation-induced myelosuppression. Rhesus macaques were total body x-irradiated (250 kVp, TBI) to 600 cGy. Following TBI, two groups received peg-leridistim (n = 5) or leridistim (n = 4) at a dose of 600 microg/kg on day 1, while two other groups (both n = 4) received peg-leridistim at 200 microg/kg on day 1 and day 4, or day 1 and day 7. The irradiation controls (n = 7) received 0.1% autologous serum for 18 days. All peg-leridistim treatment schedules significantly improved all neutrophil-related parameters following TBI as compared with nontreated controls and were equivalent in effect when compared among themselves. Administration of a single high dose or two separate lower doses of peg-leridistim significantly improved neutrophil regeneration, in a manner equal to that of conventional daily or abbreviated every-other-day administration of leridistim in this nonhuman primate model of severe myelosuppression.