重组人粒细胞集落刺激因子在恶性肿瘤化疗后应用的疗效观察

目的:研究国产重组人粒细胞集落刺激因子(rhG-CSF)在恶性肿瘤化疗后所致白细胞(WBC)和中性粒细胞绝对值(ANC)减少的临床效果及不良反应(ADR)。方法:采用自身交叉方法将62例恶性肿瘤病人随机分为AB和BA2组。AB组A周期为治疗周期,即化疗结束48h开始,用国产rhG-CSF治疗,B周期单用化疗,为对照周期;BA组正好相反。隔日查血常规1次,观察WBC和ANC的变化。结果:国产rhG-CSF可以减轻化疗所致的WBC和ANC下降程度,促进化疗患者WBC和ANC的恢复,缩短WBC和ANC减少的持续时间,有助于化疗按期进行。主要ADR有注射部位疼痛、皮疹、发热、肌肉疼痛、乏力。结论:在恶性肿瘤化疗中应用国产rhG-CSF,能有效减轻WBC和ANC降低,缩短WBC和ANC恢复时间,确保化疗顺利进行。     

人粒细胞集落刺激因子基因重组及在E.coli中的表达和鉴定

将RT－PCR扩增的人粒细胞集落刺激因子（hG－CSF）结构基因与表达载体pJGW1重组，转化含有pGP1－2质粒的E.coliDH5α，进行温度诱导表达。经SDS－PAGE分析，rhG－CSF表达量占菌体总蛋白量的30％以上。将其复性，经CM－52FF离子交换层析纯化，纯化后的rhG－CSF（纯度＞98％）经SDS－PAGE测定分子量约为19kD；经胰酶裂解、反相HPLC分析肽谱具有8条特异肽段；等电聚焦测定PI值约为5．8；免疫印迹实验证实其与标准hG－CSF具有相同的抗原反应特异性；NH2-末端氨基酸分析与文献报道一致，采用小鼠白血病细胞株NFS－60测定活性为I×108IU／mg。     

Renal Clearance of a Recombinant Granulocyte Colony-Stimulating Factor,Nartograstim, in Rats

Purpose. To clarify the role of the kidney in the elimination of a recombinant human granulocyte colony-stimulating factor, nartograstim, we have investigated its pharmacokinetics in rats with renal failure. Methods. The steady-state clearance (CL_ss) were determined by the intravenous infusion for 4 hr to unilateral renally-ligated and cisplatin-treated rats, whose renal functions were about 50 and 10 % of controls, respectively. Results. CL_ss of nartograstim (27 ml/hr/kg) in the renally-ligated rats at a high infusion rate was significantly lower (25%) than in control rats (p<0.05). CL_ss in these rats, at a low infusion rate was 95 ml/hr/kg, 14 % lower than in control rats. The saturable CL_ss in these rats, 68 ml/hr/kg, was not significantly different from control rats (75 ml/hr/kg, p>0.05). Also, CL_ss in cisplatin-treated rats with extensive renal failure, at a high infusion rate, decreased to 57 % of controls. Furthermore, the total body clearances (CL_tot) of nartograstim after bolus intravenous administration to renally-ligated and cisplatin-treated rats were reduced to 33–49 % of controls. Conclusions. These results suggest that the kidney may be responsible for 40– 50 % of the nonsaturable clearance of nartograstim. Thus, the kidney should make a major contribution to the elimination of nartograstim when rats are given a high dose of nartograstim, which saturates the receptor-mediated clearance.     

G-CSF (Granulocyte Colony-Stimulating Factor): follow-up and use in a French University hospital

Granulocyte Colony-Stimulating Factor or G-CSF (NEUPOGEN) was approved for use in France in November 1991 for prevention of chemotherapy-induced neutropenia. This retrospective study was conducted at Saint-Louis Hospital, Paris, France, from November 1991 to March 1993 with a more detailed analysis of patient profiles for courses ordered between November 1991 and December 1992. Data were collected on standardized G-CSF-treatment summary forms. The purpose of the study was to define, in clinical terms, the patients treated by G-CSF to determine the average cost per course of therapy and its impact on the hospital pharmacy budget. From November 1991 to December 1992 data from 307 patient profiles were collected and analyzed. The subcutaneous route was the preferred route and only 16.6% of courses were administered intravenously. 45.6% of patients received a single course, 24.3% received two courses, and 30.1% received more than two courses. Each patient completed an average of 2.3 courses at an average cost per course of $2,000.00 (Canadian dollars). During March 1993, 50% of vials dispensed were administered to outpatients. During the 14-month period, an average of 613.8 vials were dispensed per month corresponding to an average monthly expenditure of $104,000.00 (Canadian dollars). In the first 12 months following the commercial availability of G-CSF, G-CSF expenditures accounted for 8% of the pharmacy budget.     

Absorption of Recombinant Human Granulocyte Colony-Stimulating Factor (rhG-CSF) from Rat Nasal Mucosa

Nasal absorption of recombinant human granulocyte colony-stimulating factor (rhG-CSF) was examined in the rat. The relative bioavailability of rhG-CSF for subcutaneous administration was 2%, as evaluated from the immunologically active rhG-CSF concentration in rat plasma and the area under the curve (AUC) of the plasma rhG-CSF concentration versus time for 8 hr. Pharmacological availability relative to subcutaneous administration was determined from the increase in total blood leukocyte numbers. The pharmacological availability was 5–10%, determined from the AUC for the increased ratio of total leukocyte numbers versus time for 48 hr; it was slightly dependent on the pH and the osmotic pressure of the dosing solution. Accordingly, the plasma concentration of rhG-CSF did not always reflect its pharmacological effects. Relative bioavailability and pharmacological availability were increased about 23 times and 3 times, respectively, by polyoxyethylene 9-lauryl ether (Laureth-9), but no increase in availability occurred with sodium glycocholate. The increase in total leukocyte numbers was maintained during multiple rhG-CSF dosing, and the addition of Laureth-9 further increased the pharmacological effects of this agent. This study indicates that nasal administration of rhG-CSF is an effective parenteral administration route.     

Improving the Oral Efficacy of Recombinant Granulocyte Colony-Stimulating Factor and Transferrin Fusion Protein by Spacer Optimization

Purpose To improve the oral efficacy of the recombinant fusion protein containing granulocyte colony-stimulating factor (G-CSF) and transferrin (Tf) by inserting a linker between the two protein domains.Materials and Methods Oligonucleotides encoding flexible and helix-forming peptides were inserted to the recombinant plasmids. The fusion protein without linker insertion was used for comparison. The G-CSF cell-proliferation and Tf receptor-binding activities of the fusion proteins were tested in NFS-60 cells and Caco-2 cells, respectively, and in vivo myelopoietic assay with both subcutaneous and oral administration was performed in BDF1 mice.Results All fusion proteins produced from transfected HEK293 cells were positive in Western-blotting assay with anti-G-CSF and anti-Tf antibodies. Among them, the fusion protein with a long helical (H4-2) linker showed the highest activity in NFS-60 cell proliferation assay, with an EC₅₀ about ten-fold lower than that of the non-linker fusion protein. The fusion protein with H4-2 linker also showed a significantly higher myelopoietic effect when administered either subcutaneously or orally in BDF1 mice.Conclusion The insertion of a linker peptide, such as the helix linker H4-2, between G-CSF and Tf domains in the recombinant fusion protein can improve significantly both in vitro and in vivo myelopoietic activity over the non-linker fusion protein.     

重组人粒细胞集落刺激因子对化疗后中性粒细胞缺乏的有效性和安全性研究

目的：评价重组人粒细胞集落刺激因子（rhG-CSF）对化疗所致中性粒细胞缺乏的恶性血液病患者的临床疗效和安全性。方法：对2012年1~12月我院73例化疗后出现中性粒细胞缺乏的恶性血液病患者进行调查分析,均在粒缺发生后使用rhG-CSF,用SPSS统计软件对信息进行分析。结果：rhG-CSF可以有效缓解化疗后的中性粒细胞缺乏,使白细胞和中性粒细胞数目明显升高,平均恢复时间为（7.77±5.14） d,总有效率为95.9%,研究中根据病人中性粒细胞缺乏发生程度、阶段及持续时间进行个体化给药。结论：rhG-CSF对化疗所致的中性粒细胞缺乏症安全有效。     

Effects of Excipients on the Hydrogen Peroxide-Induced Oxidation of Methionine Residues in Granulocyte Colony-Stimulating Factor

No HeadingPurpose. The objective of this study was to elucidate the different mechanisms of action of different excipients on the oxidation of Met¹, Met¹²², Met¹²⁷, and Met¹³⁸ in granulocyte colony-stimulating factor (G-CSF) by using hydrogen peroxide as the oxidant.Methods. The oxidation of Met¹, Met¹²⁷, and Met¹³⁸ was quantified by peptide mapping analysis. The oxidation of Met¹²² has biphasic oxidation kinetics with a faster second phase. Therefore, the oxidation of Met¹²² was quantified by two different methods: peptide mapping analysis for the first phase of oxidation and direct reverse-phase HPLC for the second phase of oxidation.Results. The current work reveals that the preferential excluding excipients sorbitol, sucrose, and trehalose, in the concentration range 0–30% (w/v), and the preferential binding excipients urea and guanidine hydrochloride, in the concentration range 0–0.8 M, do not affect the oxidation of methionine residues in G-CSF at pH 4.5. The chelating agents citrate and EDTA have different effects on the rates of oxidation of methionine residues in G-CSF. At low concentrations, citrate decreases the rates, while at high concentrations, citrate increases the rates. EDTA decreases the rates of oxidation of methionine residues in G-CSF, such that its effect becomes more and more as its concentration is increased from 0 to 200 mM. The efficacy of EDTA on the rates of oxidation of the four methionine residues in G-CSF follows the order Met¹²² > Met¹²⁷ > Met¹³⁸ > Met¹.Conclusions. Our results indicate that EDTA can protect the methionine residues in G-CSF against oxidation induced by hydrogen peroxide. The more exposed the methionine residue is, the more difficult it is to be protected by EDTA. The mechanism may be due to the specific ion binding of EDTA to proteins.     

注射用重组人粒细胞巨噬细胞刺激因子中内毒素的定量检测

目的建立定量检测注射用重组人粒细胞巨噬细胞刺激因子（rhGM-CSF）的细菌内毒素含量的方法。方法参考《中国药典》2010年版中细菌内毒素的检测方法,通过建立标准曲线和预干扰实验确定样品的稀释倍数,采用终点显色法定量检测rhGM—CSF细菌内毒素含量。结果注射用重组人粒细胞巨噬细胞刺激因子（rhGM-CSF）进行2倍稀释,用细菌内毒素定量法检测无明显干扰作用,内毒素回收率在50％～200％,检测结果与凝胶法结果一致。结论应用终点显色法测定注射用重组人粒细胞巨噬细胞刺激因子（rhGM．CSF）中的细菌内毒素是可行的。     

重组人粒细胞刺激因子用于乳腺癌化疗后辅助治疗专项点评

目的 了解乳腺癌化疗后辅助治疗药物重组人粒细胞刺激因子的使用情况,促进肿瘤化疗患者合理使用此类药物。方法 汇总2016年厦门市妇幼保健院全部乳腺癌化疗病历,抽取150份,建立点评依据,对重组人粒细胞刺激因子用药的适应证、给药时机、用法用量等进行点评。结果 厦门市妇幼保健院重组人粒细胞刺激因子用于乳腺癌化疗后辅助治疗,在预防使用方面主要为给药时机不合理,占总抽查病历的20.22%;而治疗使用方面主要体现为适应证不合理,占总抽查病历的26.23%。结论 临床药师应加强此类药物的监控与评价,寻求、创造更好的循证医学证据,促进临床合理用药。     

Pulmonary Delivery of Powders and Solutions Containing Recombinant Human Granulocyte Colony-Stimulating Factor (rhG-CSF) to the Rabbit

Two powder formulations (MMAD <4 µm) containing rhG-CSF were insufflated (IF) via an endotracheal tube at doses of 5, 75 or 500 µg/kg to New Zealand white rabbits. Doses of 5 and 500 µg/kg of solutions were administered by intratracheal instillation (IT), subcutaneous (SC) injection in the thigh and intravenous injection (IV) via the marginal ear vein. Blood samples were removed at regular intervals from an indwelling jugular catheter. Blood was analyzed directly for total white blood cell counts (WBC). Plasma was assayed for rhG-CSF by a specific ELISA. The distribution of radioactive dose in lung tissue was found after administering Tc99m HSA in solution or when incorporated into powders. The pharmacokinetics and pharmacodynamics were determined for all routes of administration. High dose IV concentration vs. time profiles declined biexponentially (t_1/2 = 0.6 ± 0.2 hrs, t_1/2 = 4.6 ± 0.2 hrs, n = 8). Clearance was dose dependent (11.6 ± 2.6 [500 µg/kg, n = 8] vs. 21.8 ± 3.3 ml/hr/kg [5 µg/kg, n = 5]). A normal systemic response was obtained after IF, indicating that rhG-CSF retains activity in the solid state. Dissolution and absorption of rhG-CSF from the powders were not rate limiting. The plasma concentration vs. time profiles peaked at similar times to those after IT (T_max 1 -2 hrs) but were earlier than obtained after SC (T_max 6-10 hrs). Powders were less efficiently dosed to the lung lobes after insufflation compared with instillates (14.7 ± 10.5 vs. 60.1 ± 10.6%), resulting in bioavailabilities ranging from 5 to 33%. Bioavailability after SC was 11.0 ± 7.0% and 95.3 ± 7.9% (n = 6) for the low and high doses, respectively.     

重组人粒细胞集落刺激因子（rhG-CSF）的安全性与临床评价

目的：评价重组人粒细胞集落刺激因子在临床应用的作用机制、药代动力学、适应证、用法、用量及其禁忌证。方法：采用国内外文献综述方法。结果及结论：rhG-CSF在治疗粒细胞减少症、严重感染等方面有良好的前景,但选择适应证、个体化用药很重要。     

Effects of Antioxidants on the Hydrogen Peroxide-Mediated Oxidation of Methionine Residues in Granulocyte Colony-Stimulating Factor and Human Parathyroid Hormone Fragment 13-34

No HeadingPurpose. The effects and mechanisms of different antioxidants, methionine, glutathione, acetylcysteine, and ascorbic acid (AscH₂), on the oxidation of methionine residues in granulocyte colony-stimulating factor (G-CSF) and human parathyroid hormone fragment 13-34 (hPTH 13-34) by hydrogen peroxide (H₂O₂) were quantified and analyzed.Methods. The rates of oxidation of methionine residues in G-CSF were determined by peptide mapping analyses, and the oxidation of methionine residue in hPTH 13-34 was quantified by reverse-phase HPLC.Results. At pH 4.5, free methionine reduces, glutathione and acetylcysteine have no obvious effect on, and AscH₂ promotes the rates of oxidation of methionine residues in G-CSF. The H₂O₂-induced oxidation rate constants for free methionine, acetylcysteine, and glutathione at pH 4.5 were measured to be 32.07, 1.00, and 1.63 M^-1h^-1, respectively, while the oxidation rate constant for Met¹, the most readily oxidizable methionine residue in G-CSF, is 13.95 M^–1h^–1. Therefore, the different effects of free methionine, acetylcysteine, and glutathione on the rates of oxidation of methionine residues in G-CSF are consistent with their different reactivity toward oxidation by H₂O₂. By using hPTH 13-34, the effect of AscH₂ on the H₂O₂-induced oxidation of methionine residue was quantified, and the mechanisms involved were proposed. Because of the presence of trace transition metal ions in solution, at low concentrations, AscH₂ is prone to be a prooxidant, increasing the hydroxyl radical (OH) production rate via Fenton-type reactions. In addition to peroxide oxidation, these radicals lead to the degradation of hPTH 13-34 to smaller peptide fragments. At high concentrations, AscH₂ tends to act as an OH scavenger. EDTA inhibits OH production and thus eliminates the degradation of hPTH 13-34 by forming complexes with transition metal ions. However, the rate of oxidation of the methionine residue in hPTH 13-34 increases as the concentration of AscH₂ is increased from 0 to 200 mM, and the reason for this is still not clear.Conclusions. Our results demonstrate that free methionine is an effective antioxidant to protect G-CSF against methionine oxidation at pH 4.5. Acetylcysteine and glutathione are not effective antioxidants at pH 4.5. Their oxidation rates at different pH values imply that they would be much more effective antioxidants than free methionine at alkaline conditions. AscH₂ is a powerful electron donor. It acts as a prooxidant in the conditions in this study and is unlikely to prevent oxidation by H₂O₂ in protein formulation, whether or not EDTA is present.     

乳腺癌化疗对人表皮生长因子受体表达变化及其临床意义

目的探讨乳腺癌患者化疗前后人表皮生长因子受体(HER)表达的变化及其临床意义。方法选择乳腺癌化疗患者42例,采用免疫组织化学法检测化疗前后乳腺癌组织HER1和HER2蛋白表达的变化,并分析其与化疗疗效和淋巴结转移之间的关系。结果与化疗前相比较,化疗后乳腺癌组织HER1和HER2蛋白表达阳性率均显著下降;HER1和HER2阴性患者化疗有效率较阳性患者显著增加;HER2阳性患者淋巴结转移发生率较阴性患者显著增加。结论术前化疗有助于抑制乳腺癌的增殖,其机制与下调HER的表达有关,HER2可以作为预测乳腺癌化疗疗效和淋巴结转移的指标。     

回哞与展望——细胞集落刺激因子的临床评价

目的:回顾细胞集落刺激因子进展,借以评价其疗效及安全性.方法:采用近年来国内、外文献进行综述.结果与结论:于20世纪90年代上市的细胞集落刺激因子,作为生物工程药品中多肽类的代表,进展十分迅猛,临床应用也同步增多,近期国内、外专业文献的有关报道亦浩如烟海.大量文献证实,应用细胞集落刺激因子可克服肿瘤化疗和放疗引起的骨髓毒性,有利于大量强化治疗,缩短应用肿瘤化疗周期,并减少感染的并发症.     

重组人粒细胞集落刺激因子预防放射性口腔黏膜炎的临床观察

目的:观察重组人粒细胞集落刺激因子(rhG-CSF)预防放射性口腔黏膜炎的临床疗效。方法:将86例接受放射治疗的头颈部恶性肿瘤患者随机分为治疗组和对照组。治疗组44例,采用复方氯己定含漱液清洗口腔后,rhG-CSF漱口液含漱;对照组42例,单纯采用复方氯己定含漱液清洗口腔。结果:治疗组Ⅲ、Ⅳ级放射性口腔黏膜炎的发生率明显低于对照组,分别为38.6%和71.4%,2组比较有显著性差异(P<0.05)。治疗组发生放射性口腔黏膜炎的时间较晚,放射治疗20Gy时,治疗组口腔黏膜炎的发生率为68.2%,对照组为92.9%,2组比较差异有统计学意义(P<0.05)。结论:rhG-CSF局部预防用药能推迟放射性口腔黏膜炎的发生,并可降低Ⅲ、Ⅳ级放射性口腔黏膜炎的发生率。     

长、短效粒细胞刺激因子预防和治疗肺部恶性肿瘤化疗所致骨髓抑制的成本-效果分析

目的:对比长、短效粒细胞刺激因子预防和治疗肺部恶性肿瘤化疗所致骨髓抑制的成本-效果,为临床合理用药提供参考。方法:回顾性分析郑州大学附属肿瘤医院2017年1月-2018年6月期间使用长、短效粒细胞刺激因子预防和治疗肺部恶性肿瘤化疗所致骨髓抑制的132例病例,其中使用重组人粒细胞刺激因子注射液(短效,A组)60例,使用聚乙二醇化重组人粒细胞刺激因子注射液(长效,B组)72例,比较两组患者的临床疗效、骨髓抑制情况、不良反应发生情况,计算两组治疗方案的成本,进行成本-效果分析,并以下调药品价格20%进行敏感性分析。结果:A、B组患者的总有效率分别为71.7%、75.0%,差异无统计学意义(P>0.05),不同程度骨髓抑制情况的发生率和不良反应发生率差异也均无统计学意义(P>0.05);平均成本分别为(335.91±180.34)、(1 982.75±603.15)元,A组成本明显低于B组(P<0.05),成本-效果比分别为4.69、26.44。以A组为参照,B组的增量成本-效果比为494.55。敏感性分析结果与成本-效果分析结果无差异。结论:重组人粒细胞刺激因子注射液和聚乙二醇化重组人粒细胞刺激因子注射液预防和治疗肺部恶性肿瘤化疗所致骨髓抑制的有效性相当,但前者的成本-效果比低于后者。     

聚乙二醇化重组人粒细胞刺激因子专项处方点评与分析

目的:通过实施处方专项点评,了解肿瘤患者PEG-rhG-CSF临床用药情况,发现不适宜用药问题,为指导合理用药提供依据.方法:随机抽取2020年从本院出院并使用PEG-rhG-CSF的病例200例,参照相关指南及药品说明书制定点评标准,并依据此标准评价PEG-rhG-CSF的用药合理性.结果:在200例中,不合理用药达160例(占80.00％),其中无适应症用药最多(144例,占90.00％),其次为用药剂量偏低(64例,占40.00％)、用药途径不适宜(6例,占3.75％)等.无适应症用药在低FN风险患者中最为常见(135例,占93.75％).结论:PEG-rhG-CSF临床应用中不合理用药现象较为普遍,尤其对于低FN风险患者中的预防用药缺乏依据,需引起临床重视.     

微波灼伤创面应用生长因子后的DNA含量分析

目的:探讨重组人酸性成纤维细胞生长因子对微波灼伤创面组织细胞的促增殖作用。方法:运用NS-FⅡ型多功能光谱治疗仪器建立微波灼伤动物模型,分为5组,每个创面按分组分别滴加0．9％氯化钠注射液(NS)、重组人表皮生长因子(rh-EGF,5 mg·L-1)、重组人酸性成纤维细胞生长因子(rh-aFGF,5mg·L-1)、重组人碱性成纤维细胞生长因子(rh-bFGF,5 mg·L-1)和牛脑提取液(bBE,1O mg·L-1)各0．2 ml,直到伤后第5d,取创面组织经流式细胞仪检测全层皮肤组织细胞周期及DNA含量。结果:应用生长因子各组的SPF、PI及DNA含量均明显高于NS组(P<0．05)。结论:重组人酸性成纤维细胞生长因子具有促进微波灼伤创面组织细胞增殖作用,促进创面的修复,缩短创面愈合时间。