Comparative studies of cyclocytidine (NSC-145668) and cytosine arabinoside (NSC-63878) in mice.

The distribution of cyclocytidine and cytosine arabinoside has been studied in normal BDF mice and in mice bearing 6-day solid L1210 lymphocytic leukemia by whole-body radioautography, bioassay, and radiochemical techniques. Radioactivity was widely distributed throughout the tissues between 15 minutes and 12 hours after a single intravenous dose of either cyclocytidine-2-14C or cytosine arabinoside-2-14C. Whole-body radioautograms demonstrated that for most tissues, cytosine arabinoside-derived 14C was uniformly excreted by 48 hours; cyclocytidine-derived 14C, however, was localized in certain tissues as early as 15 minutes after drug administration and was retained in these sites for 48 hours. Depot loci of 14C included salivary and adrenal glands, fat, cardiac muscle, gastrointestinal tract, and L1210 tumor. The distribution and persistence of cyclocytidine-derived radioactivity is consistent with other reports of toxicity induced by the drug in these tissues. Radiochromatography and bioassay data from BDF mice dosed intraperitoneally with cyclocytidine demonstrated that 65%-95% of the 14C-radioactivity in a number of tissues was the parent compound itself. Thus, cyclocytidine contributed in large measur to the generation of the radioautograms. This study demonstrates that the retention of cyclocytidine in body tissues may serve to effect the sustained release of the deaminase-resistant chemotherapeutic drug from these depot sites and thus prolong cytotoxic levels of drug in tumor tissue.     

Bcl-2反义寡核苷酸增强阿糖胞苷诱导原代白血病细胞凋亡的研究

背景与目的:有研究证实,针对bcl-2mRNA翻译起始区和蛋白编码区的2个有效反义作用靶点的反义寡核苷酸能增强HL-60和K562细胞对阿糖胞苷(Ara-C)、柔红霉素、足叶乙甙的敏感性。本研究观察这两个新靶点的反义寡核苷酸对Ara-C诱导原代培养的急性白血病(AL)、慢性淋巴细胞白血病(CLL)细胞凋亡的影响。方法:用细胞记数法观察细胞的生存情况;免疫荧光标记法通过流式细胞仪检测细胞Bcl-2蛋白水平;用姬姆萨染色法观察并用流式细胞仪检测细胞凋亡。结果:靶向bcl-2mRNA翻译起始区与靶向蛋白编码区的两个反义寡核苷酸分别与Ara-C联合作用AL、CLL细胞48h,细胞的活性受到明显的抑制,分别同无关寡核苷酸(NS-ODN)联合Ara-C组、单用Ara-C组进行比较,差异有显著性(P<0.05)。这两个不同靶点的反义寡核苷酸分别与Ara-C联用均能明显下调AL、CLL细胞Bcl-2蛋白的表达,并且联合作用AL、CLL细胞48h的凋亡细胞百分率分别与NS-ODN联合Ara-C组、单用Ara-C组进行比较,差异有显著性(P<0.05)。与靶向bcl-2mRNA翻译起始区的反义寡核苷酸比较,靶向蛋白编码区的反义寡核苷酸提高AL细胞对Ara-C的敏感性作用要强些(P<0.05)。结论:针对bcl-2mRNA翻译起始区和蛋白编码区两个靶点的反义寡核苷酸能增强Ara-C诱导AL、CLL细胞的凋亡。     

全反式维甲酸对阿糖胞苷诱导HL-60细胞凋亡的影响

目的 :探讨 HL- 6 0细胞经全反式维甲酸 (ATRA)作用后对化疗药物阿糖胞苷 (Ara- C)诱导凋亡敏感性的变化。方法 :应用光镜检查凋亡细胞形态 ,DNA电泳检查梯状条带及流式细胞仪检测细胞周期分布、凋亡细胞率和 bcl- 2蛋白表达的阳性细胞率和相对荧光强度 (MFI)。结果 :ATRA0 .3mg/ L 作用 HL- 6 0细胞 72 h后 ,S期细胞显著减少至 32 .9% (P<0 .0 5 ) ,G0 / G1 期细胞明显增加达 5 8.5 % (P<0 .0 5 ) ,bcl- 2阳性细胞率和 MFI分别下降至 18%和 0 .6 3(P<0 .0 5 ) ;Ara- C1.5 m g/ L 作用 HL- 6 0细胞 4h,凋亡细胞率为 5 5 .1% ,DNA电泳见明显的梯状条带。当 HL- 6 0细胞经 ATRA0 .3m g/ L 作用 72 h后再加 Ara- C1.5 m g/ L 继续培养 4h,细胞凋亡率明显减少至 34 .4% (P<0 .0 5 ) ,DNA电泳见梯状条带亮度减弱。结论 :ATRA降低 HL- 6 0细胞对 Ara- C诱导凋亡敏感性 ,其机制可能与 ATRA阻滞 G0 / G1 期细胞进入 S期有关     

rhG-CSF对Ara-C杀伤急性髓性白血病细胞影响的实验研究

目的探讨ｒｈＧ－ＣＳＦ对２１例急性髓性白血病（ＡＭＬ）细胞体外对阿糖胞苷（Ａｒａ－Ｃ）敏感性的影响。方法采用ＭＴＴ法检测Ａｒａ－Ｃ的细胞毒作用。结果ｒｈＧ－ＣＳＦ可显著增强Ａｒａ－Ｃ对耐药ＡＭＬ细胞的毒性,与对照组相比Ｐ＜０．０５;结论ｒｈＧ－ＣＳＦ可能通过促进耐药ＡＭＬ细胞进入增殖周期,而增强Ａｒａ－Ｃ对耐药ＡＭＬ细胞的毒性,从而具有耐药的逆转效应,预示着ｒｈＧ－ＣＳＦ与Ａｒａ－Ｃ联合运用治疗耐药ＡＭＬ的潜在价值     

吡喃阿霉素为主方案治疗难治和复发性急性白血病

 目的:探讨吡喃阿霉素为主方案治疗难治和复发性急性白血病的疗效。 方法:TA、THA和TOAP方案。 结果:8例病人中5例达完全缓解，且例部分缓解。 结论:吡喃阿霉素为主方案治疗难治和复发性急性白血病有一定疗效, 其副作用可以耐受。     

Successful reinduction therapy with amsacrine and cyclocytidine in acute nonlymphoblastic leukemia in children. A report from the Childrens Cancer Study Group.

Amsacrine (AMSA) and cyclocytidine were studied as retrieval therapy in 122 pediatric patients with acute nonlymphoblastic leukemia (ANLL). Patients either failed to achieve sustained initial remissions or were in relapse. Induction therapy consisted of intravenous (IV) AMSA (75 mg/m2) from days 1 to 5 and subcutaneous cyclocytidine (600 mg/m2) from days 1 to 7. Maintenance therapy consisted of IV etoposide (VP-16) (100 mg/m2) for 5 days and IV AMSA (100 mg/m2) on day 1. Of 122 patients, 109 were evaluable. There were 13 early deaths. Ninety-six patients received adequate therapy defined as completion of two courses of therapy. Of these 96 patients, 52 achieved complete remission. Fifteen of 33 patients who failed initial induction achieved complete remission. Eighteen of 39 patients who were resistant to anthracyclines had complete responses. There was no direct evidence of AMSA-induced cardiotoxicity. Remission duration was 28 days to 3 or more years (median, 98 days). AMSA and cyclocytidine were effective retrieval therapy for patients who were in relapse or unresponsive to frontline therapy. Duration of remission was short (median, 98 days).     

Induction of differentiation of human myeloid leukemic k562 cells by 3'-deoxy-3'-fluorothymidine (comparison with arabinosylcytosine)

3'-Deoxy-3'-fluorothymidine (FT), at a concentration of 30 muM, is capable of inducing about 50% of human myeloid K562 cells to differentiate while reducing the growth potential (clonogenicity, growth rate) approximately by one half. For comparison, arabinosylcytosine (Ara C), at 0.1 muM, causes the same percentage of differentiation, but cell proliferation is abolished by >80%. Induction of differentiation is more correlated with the inducer concentration than with incubation periods for both agents. After removal of FT after 5 days of treatment, the cells resume normal growth for the next 5 days. The absolute number of differentiated cells increases within this period. In accordance with their effect on the growth potential, the shift of the colony/cluster ratio towards higher numbers of clusters is greater for Ara C than for FT. Cells treated with either compound are larger than control cells. While in control colonies only very few differentiated cells are found by microscopic inspection, up to 50% of FT treated colony cells are estimated as differentiated. In many clusters and small colonies of Ara C treated cells more than 80% of the cells are benzidine-positive. Loss of clonogenicity of CFU-GM from mouse bone marrow is stronger with Ara C treatment in liquid cell culture for 2 days than with FT treatment in the same concentration range.     

Clinical Study of Cyclocytidine in Children With Acute Leukemia

One hundred children with various types of leukemia were given113 treatments with cyclocytidine. Eighty-four of these patientshad been treated previously. Cyclocytidine was daily given intravenouslyfor four to 11 consecutive days in amounts of 20mg/kg or 500mg/m².The treatment course . was repeated, when necessary. Complete remission was obtained in one case of monocytic leukemiaamong six treated by cyclocytidine alone. Combined therapy withsteroid hormone gave 14 complete and eight partial remissionsin 38 cases, the remission rate standing at 58%. There were30 complete and 16 partial remissions in 69 cases treated withcombination therapy with two or more agents, and the remissionrate was 67%. Transient pains in the region of the parotid gland were themost characteristic toxicity. GI tract symptoms were less commonand milder than those of cytosine arabinoside. Dizziness wasnoticed in three cases. Nine children with acute leukemia who were given cyclocytidineorally showed no remarkable response. Cyclocytidine, given parenterally, seems beneficial for varioustypes of leukemia in children.     

The action of DNA antagonists on Epstein-Barr virus (EBV)-associated early antigen (EA) in Burkitt lymphoma lines

The effect of two DNA antagonists (IUDR and Ara C) on EBV-associated antigens was studied in two BL-derived carrier culture lines (P3HR-1 and Onesmas). Both drugs led to an accumulation of the early antigen (EA) from 2% in the untreated to 5–40% in the treated cultures. Ara C blocked the production of viral capsid antigen (VCA) whereas a small number of VCA + cells were still present after IUDR treatment. Reversion of Ara C-induced DNA inhibition led to the appearance of VCA + cells, reaching a higher level than in the untreated control samples. Combined immunofluorescence and autoradiography showed that the majority of VCA + cells incorporated H³-thymidine. These facts are in line with the hypothesis that EA can be made in the absence of cellular DNA synthesis, whereas VCA production is dependent on the DNA synthesis. The relationship between EA and two other EBV-associated antigens, MA (membrane antigen) and VCA (viral capsid antigen) was studied by the two-color immuno-fluorescence technique. VCA + cells were EA+ and MA+. EA + VCA - cells were partly MA + and partly MA-. This is in good agreement with the corresponding findings on the EBV-infected Raji cell system (Gergely et al., 1970a).     

Ara-C INDUCED APOPTOSIS IN HUMAN MYELOID LEUKEMIA CELL LINE HL-60: INDUCING APOPTOSIS IS THE PRIMARY MECHANISM OF CHEMOTHERAPY

ＡｒａＣＩＮＤＵＣＥＤＡＰＯＰＴＯＳＩＳＩＮＨＵＭＡＮＭＹＥＬＯＩＤＬＥＵＫＥＭＩＡＣＥＬＬＬＩＮＥＨＬ６０：ＩＮＤＵＣＩＮＧＡＰＯＰＴＯＳＩＳＩＳＴＨＥＰＲＩＭＡＲＹＭＥＣＨＡＮＩＳＭＯＦＣＨＥＭＯＴＨＥＲＡＰＹＺｈｏｕＪｉａｎｆｅｎｇ周剑锋...     

短程大剂量盐酸米托蒽醌与阿糖胞苷治疗难治复发急性白血病的临床研究

目的 :评价大剂量盐酸米托蒽醌 40 mg/ m2单剂与阿糖胞苷 (1～ 1.5 ) g/ m2连用 5 d,治疗难治复发急性白血病的疗效及推广应用价值。方法 :以该方案对 2 0例难治复发急性白血病进行 2 2例次治疗。结果 :2 2例次治疗中的 2 0例次获得完全缓解 (CR90 .9% ) ,1例次取得部分缓解 (PR4.5 4% ) ,其中 14例急淋的 16例次治疗 ,14例次取得完全缓得 ,1例次部分缓解 ,1例无效 ,6例急性粒细胞白血病治疗均取得完全缓解 ,无治疗相关死亡。结论 :短程大剂量米托蒽醌联合阿糖胞苷对难治复发急性白血病有较高的疗效 ,可提高缓解率 ,降低死亡率。     

化疗药物诱导急性白血病原代细胞凋亡的研究

目的：探讨化疗药物诱导急性白血病（AL）原代细胞凋亡的时间,剂量,效应,关系,方法：以成人AL原代细胞为研究,采用高,中、低3种浓度梯度,模拟人体应用HHar,Ara-C和ACR后的最大血液浓度,应用一系列经典方法检测细胞凋亡,并分析得出相应的时间,剂量,效应关系。结果：HHar,Ara-C具有时间－效应及剂量－效应线性趋势,而CR低剂量组诱导细胞凋亡的能力大于高剂量组,并且有时间－效应线趋势;高剂量组HHar,Ara-C诱导细胞凋亡的能力大于高剂量组ACR,而低剂量组ACR诱导细胞凋亡的能力大于低剂量组HHar及Aar-C,3种花物对AML原代细胞诱导凋亡的效果优于对ALL原代细胞,结论：AL原代细胞对细胞凋亡的敏感性也是决定化疗效果的关键因素。     

小剂量阿糖胞苷和维甲酸治疗高危MDS25例疗效观察

目的：研究高危ＭＤＳ的治疗方法,观察小剂量阿糖胞苷（ＬＤ－Ａｒａ－ｃ）和全反式维甲酸（ＡＴＲＡ）诱导治疗高危ＭＤＳ的疗效。方法：高危ＭＤＳ２５例,应用ＬＤ－Ａｒａ－ｃ１０－１５ｍｇ／ｍ＾２,每１２小时１次皮下注射,２１天一疗程,间隔１０天～１５天重复;ＡＴＲＡ３０～６０ｍｇ／ｄ,分３次口服。结果：完全缓解９例（３６％）,部分缓解４例（１６％）,４例进步,８例无效,总有效率５２％;１０例（４０％）在     

Biochemical and cytochemical effects of cytosine arabinoside (ARA C) and hydrocortisone on HeLa cells.

The biochemical and cytochemical effects of cytosine arabinoside (Ara C) and of hydrocortisone on HeLa cell in vitro have been studied. In cultures treated with Ara C, a relationship exists between the cytocidal effects of the drug and an increase in alkaline phosphatase. Although hydrocortisone had no influence on the proliferative rhythm, it induced an increase in alkaline phosphatase. Results obtained with the cytochemical technique were evaluated, particularly in relation to enzyme localization.     

A randomized trial of amsacrine and rubidazone in 39 patients with acute promyelocytic leukemia.

Thirty-nine patients with untreated acute promyelocytic leukemia (APL) were randomly allocated to receive rubidazone (zorubicin) 200 mg/m2/d, days 1 to 4 plus cytarabine (Ara C) 200 mg/m2/d, days 1 to 7 (arm A, 21 patients), or amsacrine (Amsa) 150 mg/m2/d, days 1 to 4 plus Ara C 200 mg/m2/d, days 1 to 7 (arm B, 18 patients). Prophylaxis of disseminated intravascular coagulation was made by platelet transfusions and heparin. In case of leukemic resistance, patients received a second course with 2 days of rubidazone (arm A) or Amsa (arm B) and 3 days of Ara C. Patients who achieved complete remission (CR) received three consolidation courses with the two drugs used for induction and maintenance therapy for 3 years. Two patients in arm A and one in arm B were allografted in first CR. Initial characteristics were similar in both arms. In arm A, 18 patients (86%) reached CR, two had hypoplastic death, and one had leukemic resistance after two courses. In arm B, 12 patients (66%) achieved CR, two had early death (CNS bleeding, one case; ventricular fibrillation, one case), and four had resistant leukemia after two courses. The difference in CR rate between the two arms was not significant. In arm A, disease-free survival (DFS) showed a plateau at 54.3% after 34 months (95% confidence interval [CI], 32.1% to 74.9%), with eight CRs longer than 34 months. In arm B, DFS was significantly shorter (P less than .03), showing a plateau at 16.7% after 38 months (95% confidence interval, 4.7% to 44.6%), and only two prolonged CRs were seen. The difference in DFS remained significant after censoring allografted patients and patients who died in CR (one in arm A, two in arm B). Our results suggest that Amsa-Ara C combinations may be inferior to anthracycline-Ara C combinations in the treatment of APL, because they seem to provide shorter DFS and, possibly, a higher incidence of initial leukemic resistance. However, studies with larger numbers of patients are required.     

Phase II-Trial of Double-Consolidation Following Intensive Induction Treatment for Improvement of Survival in Elderly Patients with Acute Myeloid Leukemia

Thirty-two consecutive, unselected acute myeloid leukemia (AML) patients (pts) of all FAB-subtypes with a median age of 68 years were treated with intensive induction chemotherapy consisting of one or two cycles of daunorubicin 30 mg/m² day 1-3 and Ara C 100 mg/m² as continuous infusion day 1-7. The overall CR rate was 50%, 14/24 (58%) in de novo AML, and 2/8 (25%) with preceding MDS. One patient achieved a PR of 21 months duration, 3 pts died within 7 days of the induction treatment (ED), 6 died during hypoplasia (HD), and 6 remained refractory to 2 cycles of induction. Four pts died after achieving CR. Of the remaining 12 responders, 11 pts received 2 cycles of consolidation consisting of daunorubicin 30 mg/m² day 1, and Ara C 100 mg/m² continuous IV infusion day 1-7. No deaths were observed during consolidation. DFS and survival of responders were 7 and 13 months respectively, survival of all pts, responders and non-responders, was 7 months. Large cooperative trials are necessary to identify those elderly pts who may benefit from intensified consolidation treatment.     

Detection of early antigens in nuclei of cells infected with cytomegalovirus or herpes simplex virus type 1 and 2 by anti-complement immunofluorescence, and use of a blocking assay to demonstrate their specificity.

Skin fibroblasts exposed to cytosine arabinoside (Ara C) were infected with either cytomegalovirus (CMV) or herpes simplex virus (HSV) type 1 and 2. Herpesvirus-determined early antigens (HV-EA), detected by anti-complement immunofluorescence (ACIF), occurred primarily in the nucleic, and the specificity of these results was established by an ACIF blocking reaction using F(ab')2 fragments of human and hyperimmune reference sera. Direct tests with selected sera and cross-blocking experiments between early antigenic systems of CMV, HSV-1 and the Epstein-Barr virus (EBV) did not demonstrate common HV-EA.     

A Comparative Study of DCMP versus DCcMP Combination Chemotherapy for Adult Acute Leukemia

An experimental study using L-1210 mouse leukemia indicatedthat daunomycin, cyclocytidine, and 6-mercaptopurine (DCcM)combination chemotherapy was more effective than daunomycin,cytosine arabinoside, and 6-mercaptopurine (DCM). Based on theseresults, a clinical study comparing daunomycin, cytosine arabinoside,6-mercaptopurine, and predni-solone (DCMP) combination chemotherapywith daunomycin, cyclocytidine, 6-mercaptopurine, and prednisolone(DCcMP) in the treatment of adult acute nonlymphocytic leukemiawas conducted. Patients were randomly allocated to receive eitherregimen. Ten of 15 patients treated with DCMP had complete remissionfor a median duration of 43 weeks and one had partial remission.Seven of 15 patients treated with DCcMP had complete remissionfor a median duration of 18 weeks and two had partial remission.The patient characteristics before chemotherapy, the toxicityof the chemotherapeutic regimens and the causes for differencesin antitumor activity between the experimental study and theclinical study are discussed.     

Differential effects of the MDR1 (multidrug resistance) gene-activating agents on protein kinase C: evidence for redundancy of mechanisms of acquired MDR in leukemia cells.

Human leukemia cells may acquire MDR1/P-glycoprotein-mediated multidrug resistance (MDR) in the course of short-term (within hours) exposure to many stress stimuli. This effect is thought to be associated with the activity of protein kinase C (PKC) (Chaudhary, Roninson, 1992. 1993). However, we show here that cytosine beta-D-arabinofuranoside (Ara C) and 12-O-tetradecanoylphorbol 13-acetate (TPA), agents that activated the MDR1 gene in the H9 T-cell leukemia line, caused different effects on PKC. Namely, TPA activated PKC whereas Ara C was without the effect. Furthermore, cell permeable ceramide, a lipid messenger known to mediate cellular effects of chemotherapeutic drugs and TPA, activated the MDR1 gene and down-regulated PKC. These results suggest that the MDR1 gene can be activated via the pathway(s) that requires PKC activity as well as via bypass of PKC. The redundancy of signaling pathways that regulate the acquisition of MDR should be taken into consideration for prevention of secondary drug resistance in hematological malignancies.     

Myeloid and monocytoid leukemia cells have different sensitivity to differentiation-inducing activity of deoxyadenosine analogs

The differentiation-inducing effect of clinically applicable analogs of deoxyadenosine on myelomonocytic leukemia cells was examined. Monocytoid leukemia cells were more sensitive to the analogs than were erythroid or myeloid leukemia cells based on the inhibition of cell growth and induction of cell differentiation. Monocytoid leukemia cells were highly sensitive to combined treatment with 2'-deoxycoformycin (dCF) and 9-beta-D-arabinofuranosyladenine (Ara A) for inducing cell differentiation. Ara A induced the differentiation of monocytoid leukemia U937 and THP-1 cells at concentrations which were 1/1000-10000 of that at which it induced the differentiation of other cell lines in the presence of dCF. In combination with a low concentration of 1alpha,25-dihydroxyvitamin D3 (VD3), the induction of the monocytic differentiation was greater in monoblastic U937 cells. Adenosine deaminase-resistant analogs such as fludarabine (FLU) and cladribine (CdA) also induced the differentiation of human myelomonocytic leukemia cells, and these analogs synergistically enhanced the differentiation induced by all-trans retinoic acid (ATRA) or VD3. CdA was the most potent analog for inducing the differentiation of myeloid leukemia NB4 and HL-60 cells in the presence or absence of ATRA. These findings indicate that dCF + Ara A and CdA may be effective for the therapy of acute monocytoid and myeloid leukemia, respectively.