促红细胞生成素预处理对视网膜神经元缺氧性损伤的保护作用

目的研究缺氧对培养大鼠视网膜神经元的影响及促红细胞生成素(erythropoietin,EPO)预处理的保护作用。方法取体外原代培养3d的大鼠视网膜神经元,利用1mmol.L-1的连二亚硫酸钠消除培养基中的氧合并培养基质缺糖持续6h,导致神经元缺氧性损伤。采用免疫组化染色技术检测了EPO受体在原代培养视网膜神经元的表达,乳酸脱氢酶(LDH)释放率作为评价损伤的指标,TUNEL染色检测视网膜神经元的凋亡,并进行形态学观察。结果原代培养的大鼠视网膜神经元正常下可见EPOR的弱阳性表达,表达部位定位于神经元的胞体和突起,缺氧后EPOR的表达明显增加;神经元LDH释放率和凋亡百分率增加;在损伤前加入浓度2.5～40kU.L-1的rhEPO均可有效抑制LDH释放(P<0.05)。TUNEL结果显示rhEPO预处理能抑制神经元的凋亡,明显改善细胞缺氧性损伤的形态学变化。结论EPO预处理对神经元的缺氧损伤具有保护作用,且具有浓度和时间依赖性。     

促红细胞生成素（EPO）联合化疗治疗癌症相关性贫血

目的 :恶性肿瘤化疗的同时联合应用促红细胞生成素 (EPO) ,观察对癌症相关性贫血 (MTA)临床症状的改善效果。方法 :观察组在化疗前 1周开始给予EPO ,对照组只进行化疗不用EPO ,其他辅助治疗两组相同 (包括营养支持、粒细胞刺激因子应用、出血者止血药物的应用、重要脏器的保护等 )。结果与结论 :化疗联合用EPO能预防和治疗MTA。对靠输血提升血红蛋白后化疗的患者可明显减少输血量 ,甚至可停止输血。EPO对MTA的治疗和与化疗所致贫血的预防有肯定的疗效     

重组人促红素糖指纹图谱分析技术研究

目的:建立重组人促红素N-糖链特征性糖指纹图谱检测分析技术。方法:分别用外切糖苷酶(PNGase F)和神经氨酸酶酶切重组人促红素,超滤离心获得糖的各种组分,使用高pH离子色谱分析N-糖链的结构和组成并确定N-糖链指纹图谱。结果:建立的方法稳定可靠,重复性好,可用于重组人促红素糖结构和组成的研究和质量控制。结论:本研究建立了简便、有效的重组人促红素N-糖链特征性糖指纹图谱检测分析方法。     

Effect of erythropoietin, 5-fluorouracil and SN-38 on the growth of DLD-1 cells

Supplementation of recombinant human erythropoietin (rHuEpo) is one of the methods for the treatment of anemia. The influence of rHuEpo on proliferation or clonogenic growth of cancer cells is not clear and some of the published results are conflicting. The aim of this work was to study the effect of rHuEpo on colon cancer cells when given alone or in combination with cytostatics. Human colon adenocarcinoma cells (DLD-1) were cultured in medium with rHuEpo, 5-fluorouracil (5-FU) and an active metabolite of irinotecan (SN-38). Cell viability was determined using a hematocytometer and 0.4% (w/v) trypan blue dye. Cell proliferation was measured by the MTT assay. Expression of EpoR, Bax, Bcl-2 and Akt1 protein was assessed by Western blot. The results of this study indicate a dose-dependent inhibitory effect of rHuEpo on DLD-1 cell growth and proliferation. Moreover, the combined treatment of rHuEpo and cytotoxic agents such as 5-FU and SN-38 increases the antitumor action, which is indicated by decreases in proliferation in the MTT test, cell numbers and DNA synthesis. We found a significant increase in EpoR, Bcl-2 and Akt1 protein expression in all cells grown in medium containing 3 IU of rHuEpo. We observed that EpoR is constitutively expressed in DLD-1 cells. Our results indicate that rHuEpo acts via EpoR to directly inhibit DLD-1 cell growth and indirectly modulate the cytostatics effects of 5-FU and SN-38.     

A physiological threshold for protection against menadione toxicity by human NAD(P)H:quinone oxidoreductase (NQO1) in Chinese hamster ovary (CHO) cells

NAD(P)H:quinone oxidoreductase 1 (NQO1) has often been suggested to be involved in cancer prevention by means of detoxification of electrophilic quinones. In the present study, a series of Chinese hamster ovary (CHO) cell lines expressing various elevated levels of human NQO1 were generated by stable transfection. The level of NQO1 over-expression ranged from 14 to 29 times the NQO1 activity in the wild-type CHO cells. This panel of cell lines, allowed investigation of the protective role of NQO1 in quinone cytotoxicity. It could be demonstrated that menadione toxicity was significantly reduced in all NQO1-transfected CHO clones compared to the wild-type cells, but the clones did not show differences in their level of protection against menadione. This observation pointed at a critical threshold concentration of NQO1 above which a further increase does not provide further protection against quinone cytotoxicity. Additional studies in which the NQO1 activity was inhibited by dicoumarol showed that only dicoumarol concentrations of about five times the EC(50) for NQO1 inhibition were able to reduce NQO1 levels below the apparent threshold, making the cells more sensitive. The level of this threshold was estimated to be in the range of base line NQO1 activities observed in several tissues and species. Thus, the results of the present study indicate that beneficial effects of NQO1 induction by, for example, cruciferous vegetables might be absent or present depending on the NQO1 activity threshold for optimal protection and the basal level of NQO1 expression in the tissue and species of interest.     

Effect of diisopropanolamine upon choline uptake and phospholipid synthesis in Chinese hamster ovary cells.

Aminoalcohols differ in mammalian toxicity at least in part based upon their ability to alter the metabolism of phospholipids and to cause depletion of the essential nutrient choline in animals. This study examined the incorporation of diisopropanolamine (DIPA) into phospholipids (PLs) and effects of DIPA upon choline uptake and phospholipid synthesis in Chinese hamster ovary (CHO) cells. Results were compared to those of a related secondary alcohol amine, diethanolamine (DEA), whose systemic toxicity is closely associated with its metabolic incorporation into PLs and depletion of choline pools. DIPA caused a dose-related inhibition of (3)H-choline uptake by CHO cells that was approximately 3-4 fold less potent, based upon an IC50, than that reported for DEA. DIPA, in contrast to DEA, did not cause changes in the synthesis rates of (33)P-phosphatidylethanolamine, (33)P-phosphatidylcholine or (33)P-sphingomyelin at either non-toxic or moderately toxic concentrations. Only approximately 0.004%, of administered (14)C-DIPA was metabolically incorporated into PLs, over 30-fold less than the incorporation of (14)C-DEA under similar conditions. Overall, these data and previous pharmacokinetic and toxicity data obtained in vivo suggests that DIPA is distinct from DEA and lacks significant choline and PL metabolism related toxicity in animals.     

Effect of glutathione depletion on apoptosis induced by thiram in Chinese hamster fibroblasts

Fungicide thiram, which is also known as an inducer of allergic contact dermatitis (ACD), was used as a model compound of thiuram chemicals, and its cellular effects were investigated in cultured Chinese hamster V79 cells. The level of intracellular reduced glutathione (GSH), protein sulfhydryl (PSH) groups, protein carbonyls (PC), membrane lipid peroxidation reflected by enhanced thiobarbituric acid reactive substrates (TBARS) production, as well as apoptotic effect were determined. The apoptosis induction was determined by assessing DNA fragmentation by TUNEL, annexin V binding, and caspases activation assays, using fluorescent microscope or flow cytometry, respectively. The concentrations of thiram required to induce cellular GSH depletion (by 40-50%), protein, and membrane lipid peroxidation (2-fold, and 1.7-fold, respectively), as well as to induce apoptosis in V79 Chinese hamster fibroblasts without causing necrosis through cytotoxic effects were between 50-100 microM. To investigate the role of decreased GSH content in the toxicity of thiram, GSH level was modified prior to exposure. Pretreatment of V79 cells with N-acetyl-L-cysteine (NAC), a GSH biosynthesis precursor, prevented GSH decrease, PC and TBARS production, as well as caspases activation induced by thiram exposure. On the other hand, thiram effects were enhanced by the previous depletion of cellular GSH by L-buthionine-(S,R)-sulfoximine (BSO).     

Effects of selenocystine on lead-exposed Chinese hamster ovary (CHO) and PC-12 cells

Lead is a pervasive environmental toxin that affects multiple organ systems, including the nervous, renal, reproductive, and hematological systems. Even though it is probably the most studied toxic metal, some of the symptoms of lead toxicity still cannot be explained by known molecular mechanisms. Therefore, lead-induced oxidative stress has recently started to gain attention. This in vitro study confirms the existence of oxidative stress due to lead exposure. Administration of lead acetate (PbA) to cultures of Chinese hamster ovary cells (CHO) had a concentration-dependent inhibitory effect on colony formation and cell proliferation. This inhibition was eliminated by 5 microM selenocystine (SeCys). In order to evaluate the nature of SeCys's effect, we measured glutathione (GSH), its oxidized form glutathione disulfide (GSSG), malondialdehyde (MDA), catalase, and GSH peroxidase (GPx) activities in lead-exposed CHO cells both in the presence and absence of SeCys. Increases in MDA, catalase, and GPx activities were observed in cultures that received only PbA, but supplementation with SeCys returned these measures to pretreatment levels. The ratio of GSH to GSSG increased in lead-exposed cells incubated in SeCys-enhanced media but declined in cultures treated with PbA only. In order to determine whether SeCys also reverses lead-induced neurotoxicity, a neuronal cell line, PC-12 cells, was used. Lead's inhibition on neurite formation was significantly eliminated by SeCys in PC-12 cells. Our results suggest that SeCys can confer protection against lead-induced toxicity in CHO cells and neurotoxicity in PC-12 cells.     

Dopamine D(2) receptor-induced COX-2-mediated production of prostaglandin E(2) in D(2)-transfected Chinese hamster ovary cells without simultaneous administration of a Ca(2+)-mobilizing agent

We have earlier demonstrated that dopamine stimulates the liberation of the prostaglandin E(2) (PGE(2)) precursor, arachidonic acid, in Chinese hamster ovary cells transfected with the rat dopamine D(2) receptor (long isoform), also without concomitant administration of a Ca(2+)-releasing agent [Nilsson et al., Br J Pharmacol 1998;124:1651-8]. In the present report, we show that dopamine, under the same conditions, also induces a concentration-dependent increase in the production of PGE(2), with a maximal effect of 235% at approximately 100 microM, and with an EC(50) of 794 nM. The effect was counteracted by the D(2) antagonist eticlopride, pertussis toxin, the inhibitor of intracellular Ca(2+) release TMB-8, incubation in Ca(2+)-free experimental medium, and PKC desensitization obtained by chronic pretreatment with the phorbol ester TPA. It was also antagonized by the non-specific cyclooxygenase (COX) inhibitor, indomethacin, and by the selective COX-2 inhibitor, NS-398, but not by the specific COX-1 inhibitor, valeryl salicylate. Both the non-specific phospholipase A(2) inhibitor, quinacrine, and an inhibitor of cPLA(2) and iPLA(2), AACOF3, counteracted the effect; in contrast, a selective iPLA(2) inhibitor, BEL, and a selective sPLA(2) inhibitor, TAPC, were ineffective. No effects of dopamine were obtained in control cells mock-transfected with the p3C vector only. The results reinforce previous assumptions that dopamine may interact with eicosanoid metabolism by means of D(2) receptor activation, and implicate an involvement of cPLA(2) and COX-2 in this effect. It is suggested that measurement of dopamine-induced PGE(2) production may serve as a convenient way to study D(2) receptor function in vitro.     

In vitro cytotoxicity of polychlorinated biphenyls (aroclors 1016, 1242, 1254 and 1260) and their effect on phospholipid and neutral lipid composition of Chinese hamster ovary (CHO-K1) cells

Cytotoxicity of 4 Aroclors (1016, 1242, 1254 and 1260) was compared in Chinese hamster ovary (CHO-K1) cells in Ham's F-12 medium. When parameters of toxicity were cell numbers or tissue protein, 50% lethality occurred at Aroclor concentrations between 30 and 45 ppm. An in vitro clonal assay with CHO-K1 cells was a sensitive indicator of cytotoxicity of the polychlorinated biphenyls (PCBs). From EC₅₀ values (concentration that allowed 50% survival of formed colonies), cytotoxicity was lower with Aroclor 1016 (32 ppm) and higher with Aroclors 1254 (27 ppm) and 1260 (28 ppm). In cells exposed 24 h to a marginally cytotoxic dose (20 ppm) of each Aroclor, phospholipid (PL) thin-layer chromatography (TLC) showed an increase in phosphatidylcholine (PC) and a decrease in phosphatidylethanolamine (PE) and diphosphatidylglycerol (DPG). Neutral lipid (NL) TLC of cells given Aroclors 1242, 1254 or 1260 showed a 3–4-fold increase in triglyceride (TG) and a similar reduction in cholesteryl esters (CE); in contrast to Aroclor 1016 which produced no change in TG and a smaller (2-fold) reduction in CE. Cholesterol and free fatty acid fractions were unaffected by any of the Aroclors. The TG:PL ratio remained unchanged in cells given Aroclor 1016, but increased 3–4-fold with Aroclors 1242, 1254, or 1260. Compared to total values in the untreated controls, CHO-K1 cells contained less neutral lipid and more phospholipid only with Aroclor 1016.These results support the concept that differences in the behavior of Aroclor 1016 are related to its PCB composition. Changes in membrane PL and NL components, observed at marginally cytotoxic levels of each Aroclor, provided further evidence that the PCBs may affect membrane integrity and associated metabolic functions.     

三-(2-甲基-1-氮丙啶)磷化氧对中国仓鼠卵巢细胞的体外诱变性

以中国仓鼠卵巢(CHO)细胞为材料,检测了三-(2-甲基-1氮丙啶)磷化氧(MAPO)的诱变作用,结果表明,在加S9活化与不加S9活化条件下,MAPO在0.1～50.0μg·ml~1浓度范围内,可诱发CHO细胞姐妹染色单体互换率及染色体畸变率升高而在1.0～100.0μg·ml~1浓度范围内,可引起微核率明显升高,并诱发次黄嘌呤鸟嘌呤磷酸核糖基转移酶位点发生正向突变,所有结果均具有较好的剂量反应关系,表明MAPO具有很强的诱变作用。     

Independent coupling of the human tachykinin NK2 receptor to phospholipases C and A2 in transfected Chinese hamster ovary cells

The human tachykinin NK₂ receptor stably expressed in Chinese hamster ovary cells (CHO-hNK₂R cells) was characterized by studying the effect of neurokinin A (NKA), the preferred natural ligand, and that of other agonists and antagonists in both binding experiments and functional assays. Competition experiments using [¹²⁵I]NKA showed that CHO-hNK₂R cells express binding sites which have high affinity for NKA (K _i=3.4±0.9 nM), GR 64349 (K _i=12±3 nM) and [βAla⁸]NKA(4–10) (K _i=21±8 nM) and for the antagonists MEN 10627 (K _i=0.55±0.2 nM), and MEN 11420 (K _i=2.4±0.8 nM). In contrast, the tachykinin NK₁ and NK₃ receptor agonists [Sar⁹,Met(O₂)¹¹]SP and senktide, respectively, were recognized with low affinity (K _i>10 μM). NKA (EC₅₀=68±18 nM) induced a rapid and concentration-dependent increase in the intracellular level of inositoltrisphosphate (IP₃). The concentration-response curve to GR 64349 (EC₅₀=155±14 nM) was close to that of NKA, whereas [βAla⁸]NKA(4–10) (EC₅₀=445±78 nM) and SP (EC₅₀=3197±669 nM) were 7- and 50-fold less potent, respectively. In addition, NKA stimulated the release of arachidonic acid and the production of prostaglandin E₂ (PGE₂) in a concentration-dependent manner. Also in this assay, NKA was found to be more potent than the other agonists tested (the EC₅₀ values were 3±0.3, 9±3, 7.8±0.9 and 217±37 nM for NKA, GR 64349, [βAla⁸]NKA(4–10) and SP, respectively). MEN 10627 and MEN 11420 were potent and competitive antagonists in blocking NKA-induced IP₃ formation and PGE₂ release: MEN 10627 and MEN 11420 displayed comparable potencies in blocking the two functional responses initiated by occupancy of the NK₂ receptor by NKA. Pretreatment of the cells with pertussis toxin (500 ng/ml for 18 h) did not significantly modify the basal or stimulated phosphatidylinositol turnover but reduced the basal and NKA-induced PGE₂ release by about 35%. The phospholipase C inhibitor U-73122 (10 μM) prevented the NKA-induced formation of IP₃ but did not affect PGE₂ release. Conversely, the phospholipase A₂ inhibitor quinacrine (100 μM) blocked the release of arachidonic acid and PGE₂ without affecting the NKA-stimulated formation of IP₃. Chelation of extracellular calcium with 3 mM EGTA inhibited the NKA-induced PGE₂ release by 81% but was without effect on basal and NKA-stimulated IP₃ production. The calcium channel blockers verapamil (10 μM) and ω-conotoxin GVIA (0.1 μM) did not modify the basal PGE₂ production and had no significant effect on the response to tachykinins while the blocker of non-selective cation channels, SKF-96365 (10 μM), inhibited the response to NKA by about 74%. SKF-96365 did not affect the basal or the NKA-induced IP₃ formation. In conclusion, our data demonstrate that the human tachykinin NK₂ receptor expressed in CHO cells displays binding affinity and functional properties which are those of a native NK₂ receptor. No pharmacological evidence for heterogeneity of the human NK₂ receptor was obtained in this study. Our findings indicate that the human tachykinin NK₂ receptor is independently coupled to both PLC and PLA₂ signaling pathways. Activation of the PLA₂ pathway may be linked to the opening of a voltage-independent cation channel which activates a Ca²⁺-dependent PLA₂. Received: 7 April 1998 / Accepted: 17 July 1998     

Attenuation of cytogenetic effects by erythropoietin in human lymphocytes in vitro and P388 ascites tumor cells in vivo treated with irinotecan (CPT-11)

Erythropoietin (EPO) is a protein widely used against drug induced anemia at cancer patients. Irinotecan (CPT-11) is a genotoxic topoisomerase I inhibitor. We investigated the genotoxic, cytostatic and cytotoxic effects of EPO in the presence and in the absence of CPT-11 in human lymphocytes in vitro and in ascites cells of P388 leukemia in vivo. The levels of genotoxicity, cytostaticity and cytotoxicity were evaluated in human lymphocytes in vitro, and in P388 ascites tumor cells in vivo. The results show that EPO is not genotoxic. Unlikely to EPO, CPT-11 caused severe genotoxic, cytostatic and cytotoxic effects by significantly increasing SCE levels and decreasing PRI and MI values in peripheral lymphocytes in vitro and in P388 ascites tumor cells in vivo. Adding EPO in human lymphocyte cultures in vitro and in P388 leukemia bearing mice in vivo in the presence of CPT-11 decreased SCEs levels and increased PRIs and MIs were observed compared with cells treated either in vitro or in vivo with CPT-11 alone, which shows that EPO protected cells from the toxic action of CPT-11. EPO’s protective action on human peripheral lymphocytes in vitro and P388 cells in vivo from the topoisomerase I inhibitor CPT-11, lead us to propose it as a geno- and cytoprotective agent.     

生血宁片联合促红细胞生成素和左卡尼汀治疗肾性贫血的临床研究

目的探讨生血宁片联合促红细胞生成素和左卡尼汀治疗肾性贫血的临床疗效。方法选取2015年2月—2015年10月成都新华医院肾内科收治的肾性贫血患者94例,随机分为对照组和治疗组,每组各47例。对照组sc重组人促红素注射液(CHO细胞),根据促红细胞生成素使用指南来调整药量;并在透析结束后iv注射用左卡尼汀,1 g加入到生理盐水20m L中。治疗组在对照组基础上口服生血宁片,2片/次,3次/d。两组患者均连续治疗3个月。观察两组患者临床疗效,比较贫血和铁代谢指标及氧化应激水平变化。结果治疗后,对照组和治疗组的总有效率分别为87.23%、97.87%,两组比较差异有统计学意义(P0.05)。治疗后,两组患者的血红蛋白(Hb)、红细胞压积(HCT)、转铁蛋白饱和度(TSAT)及铁蛋白(SF)均明显升高,同组治疗前后差异具有统计学意义(P0.05);且治疗组的这些观察指标改善程度明显优于对照组,两组比较差异具有统计学意义(P0.05)。治疗后,两组患者晚期氧化蛋白产物(AOPP)、丙二醛(MDA)、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)水平均明显升高,同组治疗前后差异具有统计学意义(P0.05)。治疗后,治疗组AOPP和MDA水平升高幅度低于较对照组,但SOD和GSH-Px水平改善幅度高于对照组,两组比较差异具有统计学意义(P0.05)。结论生血宁片联合促红细胞生成素和左卡尼汀治疗肾性贫血的临床疗效较好,能明显改善贫血和铁代谢,缓解氧化应激反应,具有一定的临床推广应用价值。     

Effect of trivalent and pentavalent arsenic in causing chromosome alterations in cultured Chinese hamster ovary (CHO) cells

Sodium salts of trivalent and pentavalent arsenic were tested for their effect in inducing chromosome aberrations and sister-chromatid exchange (SCE) in cultured Chinese hamster ovary (CHO) cells. It was discovered that arsenite (As 3) produced excessive endoreduplication of the chromosomes at higher levels. No endoreduplication was observed with arsenate (As 5) treatment. These agents also elevated the frequencies of SCE, but less so compared to aberrations. The results obtained indicate that arsenic may be carcinogenic in animal system.     

Genotoxic and cytotoxic evaluation of the herbicide flurochloridone on Chinese hamster ovary (CHO-K1) cells

The in vitro effects of flurochloridone (FLC) and its formulations Twin Gold Pack® (25% a.i.) and Rainbow® (25% a.i.) were evaluated on Chinese hamster ovary (CHO-K1) cells by genotoxicity [sister chromatid exchange (SCE)] and cytotoxicity [cell-cycle progression, proliferative rate index (PRI), mitotic index (MI), MTT, and neutral red] end points. Cells were treated for 24 h within the 0.25–15 μg/ml concentration range. FLC and Twin Pack Gold® induced a significant and equivalent increase in SCEs regardless of the concentration. Rainbow®-induced SCEs at concentrations higher than 2.5 μg/ml; however, the increases were always lower than those induced by FLC and Twin Pack Gold®. For all compounds, the PRI decreased as a function of the concentration titrated into cultures. Whereas only the highest FLC and Twin Pack Gold® concentrations induced a significant reduction of the MI, all tested Rainbow® concentrations induced MI inhibition. Overall, the results demonstrated that although all compounds were not able to reduce the lysosomal activity, the mitochondrial activity was diminished when the highest concentrations were employed. These observations represent the first study analyzing the genotoxic and cytotoxic effects exerted by FLC and two formulated products on mammalian cells in vitro, at least on CHO-K1 cells.     

碱性成纤维细胞生长因子对非贴壁骨髓基质前体细胞增殖及分化作用的研究

目的　探讨碱性成纤维细胞生长因子 (bFGF)对非贴壁骨髓前体细胞的增殖及分化作用。方法　分离骨髓前体细胞体外培养 ,用碱性磷酸酶化学染色及图像分析仪测定细胞克隆数 ;采用免疫组化 (ABC法 )检测PCNA、Ⅰ型胶原、钙及骨钙素。研究bFGF对非贴壁前体细胞的促增殖及分化作用。结果　骨髓细胞中许多基质前体细胞以非贴壁前体细胞 (NASP)形式存在 ,bFGF不仅能促进其增殖 ,而且能诱导其向成骨细胞分化。结论　bFGF主要作用于非贴壁生长的基质前体细胞 :能增加骨细胞的数量 ,促进骨样组织形成 ,为临床用bFGF治疗骨折、骨不连提供理论依据。     

Morphological cell transformation of Syrian hamster embryo (SHE) cells by the cyanotoxin, cylindrospermopsin

Cylindrospermopsin (CYN) is a cyanotoxin which has been implicated in human intoxication and animal mortality. Genotoxic activity of this hepatotoxin is known but its carcinogenic activity remains to be elucidated. In this work, CYN was assessed for its cell-transforming activity using the Syrian hamster embryo (SHE) cell transformation assay. This in vitro assay is used to evaluate the carcinogenic potential of chemical, physical and biological agents in SHE cells, which are primary, normal, diploid, genetically stable and capable of metabolic activation. We demonstrated that CYN induced a significant increase in morphological cell transformation in SHE cells following a 7-day continuous treatment in the range of non-cytotoxic concentrations 1 × 10⁻⁷-1 × 10⁻² ng/mL.     

The cardioprotective and DNA topoisomerase II inhibitory agent dexrazoxane (ICRF-187) antagonizes camptothecin-mediated growth inhibition of Chinese hamster ovary cells by inhibition of DNA synthesis

Dexrazoxane (ICRF-187), which is clinically used to reduce doxorubicin-induced cardiotoxicity, has cell growth inhibitory properties through its ability to inhibit the catalytic activity of DNA topoisomerase II. A study was undertaken to investigate whether preincubating Chinese hamster ovary cells (CHO) with dexrazoxane prior to camptothecin treatment resulted in potentiation. Camptothecin is a DNA topoisomerase I poison. It was found that pretreating CHO cells with concentrations of dexrazoxane sufficient to strongly inhibit topoisomerase II for periods from 18 to 96 h resulted in significant antagonism of camptothecin-mediated growth inhibition. Lower concentrations that were sufficient to cause partial inhibition of topoisomerase II and partial dexrazoxane-mediated cell growth inhibition had little effect on camptothecin-mediated growth inhibition. Neither topoisomerase I protein levels nor camptothecin-induced topoisomerase I-DNA covalent complexes were affected by dexrazoxane concentrations that were sufficient to cause antagonism of camptothecin-induced growth inhibition. However, under these experimental conditions, dexrazoxane caused a decrease in DNA synthesis. Therefore, results presented here confirm the importance of the DNA synthesis-dependent replication fork interaction with topoisomerase I-DNA covalent complexes for the expression of camptothecin activity. It is concluded that dexrazoxane and camptothecin analogs should be used with caution in combination chemotherapy.