Granulocyte colony-stimulating factor potentiates differentiation induction by all-trans retinoic acid and arsenic trioxide and enhances arsenic uptake in the acute promyelocytic leukemia cell line HT93A

The effects of arsenic trioxide (ATO), all-trans retinoic acid (ATRA) and granulocyte colony-stimulating factor (G-CSF), alone or in combination, were investigated by focusing on differentiation, growth inhibition and arsenic uptake in the acute promyelocytic leukemia (APL) cell line HT93A. ATO induced differentiation at low concentrations (0.125?μM) and apoptosis at high concentrations (1-2?μM). Furthermore, ATRA induced greater differentiation than ATO. No synergistic effect of ATRA and ATO was found on differentiation. G-CSF promoted differentiation-inducing activities of both ATO and ATRA. The combination of ATRA and G-CSF showed maximum differentiation and ATO addition was not beneficial. Addition of 1?μM ATRA and/or 50?ng/ml G-CSF to ATO did not affect apoptosis compared to ATO treatment alone. ATRA induced expression of aquaporin-9 (AQP9), a transmembrane transporter recognized as a major pathway of arsenic uptake, in a time- and dose-dependent manner. However, treatment with 1?μM ATRA decreased arsenic uptake by 43.7% compared to control subject. Although G-CSF addition did not enhance AQP9 expression in the cells, the reduced arsenic uptake was recovered to the same level as that in controls. ATRA decreased cell viability and addition of 50?ng/ml G-CSF to ATRA significantly increased the number of viable cells compared with that in ATRA alone treated cells. G-CSF not only promotes differentiation-inducing activities of both ATRA and ATO, but also makes APL cells vulnerable to increased arsenic uptake. These observations provide new insights into combination therapy using these three agents for the treatment of APL.     

An efficient therapeutic approach to patients with acute promyelocytic leukemia using a combination of arsenic trioxide with low-dose all-trans retinoic acid

The use of arsenic trioxide (As2O3, ATO) combined with all-trans retinoic acid (ATRA) has recently been reported to induce remission in patients with acute promyelocytic leukemia (APL). However, its efficiency remains inconclusive mainly due to the small number of the available cases. In this study, therefore, we present a clinical study using a combination of ATO with low-dose ATRA (LD-ATRA) to treat 108 APL patients (80 newly diagnosed patients, 28 relapsed patients). Therapeutic outcomes using the ATO/LD-ATRA approach were compared with those of APL patients treated either with ATO alone (65 patients) or ATRA alone (51 patients). The results showed that the ATO/LD-ATRA approach provided significantly better therapeutic outcomes as compared to either ATO or ATRA alone, as evidenced by lower mortality, a higher CR rate and a reduced period to CR. In addition, the toxic side-effects have been no worse with the combined ATO/LD-ATRA treatment than with either ATO or ATRO alone and in some cases have been reduced. These data suggest that the ATO/LD-ATRA regimen is superior to either regimen given alone to patients with APL.     

The efficacy and safety of arsenic trioxide with or without all-trans retinoic acid for the treatment of acute promyelocytic leukemia: a meta-analysis

Arsenic trioxide (ATO) and all-trans-retinoic acid (ATRA) could induce apoptosis and differentiation in acute promyelocytic leukemia (APL) cells, respectively, thus the possibility of synergism between them was raised. This meta-analysis assessed the effectiveness and safety of ATO combined with ATRA in the treatment of APL. Compared with ATO alone, induction therapy with ATO/ATRA significantly increased the complete remission (CR) rate (RR: 1.08, 95% CI: 1.00-1.17, P = 0.04), shortened the time to achieve CR (WMD: −6.51, 95% CI: −11.32 to −1.70, P = 0.008), and improved the molecular remission rate after consolidation therapy (RR: 1.74, 95% CI: 1.14-2.66, P = 0.01) and the 1-year disease-free survival rate (RR: 1.22, 95% CI: 1.00-1.50, P = 0.05). There were no statistically significant differences between two treatments in terms of early death and main adverse events. These results suggested that ATO/ATRA could synergistically improve the overall outcome of newly diagnosed and relapsed APL patients, supporting the use of ATO/ATRA as an effective treatment for all APL patients previously untreated with ATO.     

亚砷酸治疗急性早幼粒细胞白血病的研究进展

急性早幼粒细胞白血病(acute promyelocytic leukemia,APL)作为急性髓系白血病（acute myeloid leukemia,AML）的独特亚型,以异常的早幼粒细胞增多、危及生命的出血和t(15;17)染色体易位为特征,曾被认为是最凶险的白血病。亚砷酸（application of arsenite acid,ATO）的应用大大提高了该病的治疗效果,显著改善了患者的预后,大量患者长期生存。本文综述相关研究发现:ATO在肝脏中由无机砷（iSA）转化为甲基化砷,通过多种分子机制促进早幼粒细胞分化、凋亡。在任何年龄阶段,无论白细胞是否＞10×109/L,ATO联合其他药物治疗APL,都可以减少化疗药物的剂量,降低患者的累积复发率（CIR）,减少患者的死亡率。单药ATO可高效安全治疗APL。对于复发APL,一部分相关研究支持继续ATO+维甲酸（ATO+retinoic acid,ATRA）或ATO+ATRA+化疗的治疗,一部分研究支持移植前给予ATO治疗可显著改善患者的整体预后和长期生存,观点尚未统一,有待进一步研究。ATO常见的毒副作用主要为分化综合征（differentiation syndrome,DS）、心脏毒性及肝毒性,明显比化疗药物造成的毒副作用轻微。     

Combined treatment with arsenic trioxide and all-trans-retinoic acid in patients with relapsed acute promyelocytic leukemia.

PURPOSE: Arsenic trioxide (ATO) is capable of inducing a high hematologic response rate in patients with relapsed acute promyelocytic leukemia (APL). Preclinical observations have indicated that all-trans-retinoic acid (ATRA) may strongly enhance the response to ATO. PATIENTS AND METHODS: Between 1998 and 2001, we conducted a randomized study of ATO alone versus ATO plus ATRA in 20 patients with relapsed APL, all previously treated with ATRA-containing chemotherapy. The primary objective was to demonstrate a significant reduction in the time necessary to obtain a complete remission (CR) in the ATO/ATRA group compared with the ATO group. Secondary objectives were safety and molecular response. RESULTS: The CR rate after one ATO with or without ATRA induction cycle was 80%. Clinical and pharmacokinetic observations indicated that the main mechanism of action of ATO in vivo was the induction of APL cell differentiation. Hematologic and molecular response, time necessary to reach CR, and outcome were comparable in both treatment groups. Of 16 CR patients, three patients who reached a molecular remission after one induction cycle had all received chemotherapy for a treatment-induced hyperleukocytosis. Three additional patients who received further additional ATO with or without ATRA cycles converted later to molecular negativity. CONCLUSION: ATRA did not seem to significantly improve the response to ATO in patients relapsing from APL. Other potential combinations, including ATO plus chemotherapy, have to be tested.     

全反式维甲酸与亚砷酸联合柔红霉素对NB4细胞CD11b表达的影响

 目的　观察全反式维甲酸（ATRA）与亚砷酸（ATO）单独以及二药联合柔红霉素（DNR）诱导分化治疗过程中对NB4细胞CD11b表达的影响,及其对高白细胞血症、急性早幼粒细胞白血病（APL）分化综合征的作用。方法　采用荧光素标记的CD11b单克隆抗体,流式细胞术动态检测各组药物作用于NB4细胞24、48、72、168 h后细胞CD11b的表达状况。结果　1 μmol／L ATRA作用于NB4细胞后,CD11b表达随作用时间的延长逐渐增加,呈时间依赖性。24、48、72、168 h CD11b表达分别为（33.34±3.15）%、（55.59±5.13）%、（86.08±5.12）%、（90.69±2.69）%,在同一时间点均高于空白对照组（P＜0.01）。1 μmol／L ATO作用于NB4细胞后,随时间延长CD11b表达亦逐渐增加,但各时间点之间CD11b表达差异无统计学意义;在同一时间点CD11b表达低于ATRA组（P＜0.01）,但与空白对照组比较差异无统计学意义（P＞0.05）。1 μmol／L ATRA+1 μmol／L ATO作用于NB4细胞后,24、48 h CD11b表达分别为（16.92±1.05）%、（17.01±0.22）%,与空白对照组比较差异无统计学意义（P＞0.05）,72、168 h CD11b表达高于空白对照组（P＜0.05）,分别为（18.81±1.40）%、（25.61±4.54）%;但在同一时间点CD11b表达均低于ATRA组（P＜0.01）。1 μmol／L ATRA+1 μmol／L ATO+1 μmol／L DNR作用于NB4细胞后,在同一时间点CD11b表达低于ATRA组（P＜0.01）;与空白对照组比较差异无统计学意义（P＞0.05）。结论　ATRA联合ATO或加用DNR诱导治疗APL,可能由于避免了APL细胞诱导分化过程中CD11b表达的增高,从而在一定程度上减少了高白细胞血症、APL分化综合征的发生。     

全反式维甲酸联合亚砷酸治疗儿童急性早幼粒细胞白血病疗效分析

目的:评价全反式维甲酸（all-trans retinoic acid,ATRA）联合亚砷酸(ATO)治疗儿童急性早幼粒细胞性白血病(acute promyelocytic leukemia,APL) 的疗效。方法:2009年8月至2013年4月于我院儿科就诊的APL患儿18例,将亚砷酸注射液（0.1% ATO）按6mg/m2稀释于50g/L的葡萄糖溶液200-400ml中,静脉滴注持续3-5h,1次/d,ATRA 20-35mg/(m·d),口服,3次/d。结果:18例患儿获得完全缓解（CR）率为94.4%;12例初治患儿均获得CR,6例复发患儿中5例获得CR。获得缓解时间（26.2±1.2）天,无明显不良反应发生。结论:ATRA联合亚砷酸治疗儿童APL疗效显著,有效缩短达到CR的时间,毒副作用基本可以耐受,本方案可以有效缩短治疗时间,降低患儿家庭经济负担,对患儿长期治疗具有重要作用,是一种经济有效的治疗方案。     

Apoptosis induction by (+)α-tocopheryl succinate in the absence or presence of all-trans retinoic acid and arsenic trioxide in NB4, NB4-R2 and primary APL cells

We analyzed the effect of (+)α-tocopheryl succinate (α-TOS) alone or associated with arsenic trioxide (ATO) or all-trans retinoid acid (ATRA) in acute promyelocytic leukemia (APL). α-TOS-induced apoptosis in APL clinical samples and in ATRA-sensitive (NB4) and ATRA-resistant (NB4-R2) APL cell lines. The effective dose 50% (ED-50) was calculated to be 71 and 58 μM, for NB4 and NB4-R2, respectively. α-TOS neither induced nor modified ATRA-induced differentiation of APL cells, and did not affect the proliferation and differentiation of normal CD34⁺ hematopoietic progenitors in methylcellulose assays. α-TOS exerted a moderate antagonistic effect to ATO-induced apoptosis when treatment was done simultaneously but when α-TOS was added 24 h after ATO, an additive effect was observed. Our results support the concept of α-TOS as an anti-leukemic compound which spares normal hematopoiesis.     

Contemporary Treatment of APL

Acute promyelocytic leukemia (APL) is characterized by coagulopathy, leukopenic presentation and sensitivity to anthracyclines, all-trans retinoic acid (ATRA) and arsenic trioxide (ATO). For the last 25 years, APL has been treated with a combination of ATRA and chemotherapy for induction followed by consolidation and maintenance therapy. This general treatment approach has resulted in cure rates of 80–90 %. ATO, originally approved in relapsed APL, has been incorporated into contemporary upfront treatment regimens with excellent response rates. Recent studies show that most patients with APL can be cured with ATRA and ATO alone, eliminating cytotoxic chemotherapy and resulting in superior outcomes compared to standard treatment. We will herein review historical treatment of APL, treatment considerations in specific patient populations, and therapeutic updates.     

Combination therapy with arsenic trioxide, all-trans retinoic acid, and gemtuzumab ozogamicin in recurrent acute promyelocytic leukemia

BACKGROUND: From 20% to 30% of patients with acute promyelocytic leukemia (APL) who are treated with all-trans retinoic acid (ATRA) develop recurrent disease. Arsenic trioxide (ATO) is an effective agent for the salvage of patients with recurrent APL, and gemtuzumab ozogamicin (GO) has shown activity in patients with APL. METHODS: The authors investigated the efficacy of a combination of ATO, ATRA, and GO in 8 patients with APL in first recurrence (7 patients with hematologic recurrences and 1 patient with a molecular recurrence). All patients had received previous treatment with ATRA either alone or in combination with other agents. Patients received ATO 0.15 mg/kg intravenously until they achieved a bone marrow complete remission (CR). Once in CR, patients received consolidation with ATO, ATRA, and GO for 10 months. Patients then received maintenance with idarubicin, ATRA, 6-marcaptopurine, and oral methotrexate for 11 months. RESULTS: All 7 patients who had hematologic recurrences achieved CR after a median of 39 days (range, 21-56 days). After a median follow-up of >/=36 months (range, 4-55 months), 6 patients remained alive in CR, and 2 patients died in CR. Six of 8 patients remained in second CR that was longer than their first CR. All 7 evaluable patients achieved molecular remission. There were no grade 3 or 4 extramedullary toxicities. Two patients died, 1 secondary to a complication of metastatic lung adenocarcinoma, and the other of sepsis. CONCLUSIONS: The combination of ATO, ATRA, and GO was effective and may achieve durable remissions in patients with APL in first recurrence. It should be considered as an effective alternative to allogeneic or autologous transplantation.     

Src inhibitors, PP2 and dasatinib, increase retinoic acid-induced association of Lyn and c-Raf (S259) and enhance MAPK-dependent differentiation of myeloid leukemia cells

All-trans-retinoic-acid (ATRA)-induced differentiation of human myeloid leukemia cells is characterized by persistent mitogen-activated protein kinase (MAPK) signaling. Fragmentary data suggests Src family kinase (SFK) inhibitors enhance differentiation, and thus have potential therapeutic value. The present study shows that SFK inhibitors PP2 and dasatinib enhance aspects of MAPK signaling and regulate a panel of differentiation markers, including CD11b and p47(phox). HL-60 and NB4 myeloid leukemia cells show accelerated ATRA-induced G1/0 arrest/differentiation with inhibitor co-treatment. We also identified components of a Lyn- and c-Raf-containing MAPK signaling complex augmented by the inhibitors. PP2 and dasatinib increased the ATRA-induced expression of Lyn and c-Raf (total and c-RafpS259) and their interaction. The Lyn-associated serine/threonine kinase, casein kinase II (CK2), also complexed with c-Raf and c-RafpS259, and the kinase suppressor of Ras 1 (KSR1) scaffold protein bound c-Raf, Lyn and extracellular signal-regulated kinase (ERK). c-Raf/ERK association was increased by the inhibitors, which is significant as ERK may cause c-Raf C-terminal domain (CTD) phosphorylation in a putative feedback mechanism. Consistent with this, inhibitor treatment caused more CTD phosphorylation. Lyn knockdown decreased c-Raf CTD and S259 phosphorylation. This is the first evidence suggesting SFK inhibitors enhance ATRA-induced differentiation through a possible feedback loop involving KSR1-scaffolded c-Raf and ERK complexed with Lyn and CK2.     

Treatment of acute promyelocytic leukemia in the very elderly: case report and review of the literature

Acute promyelocytic leukemia (APL) is the most curable subtype of AML yet it is not known to what extent newer therapies will succeed in the very elderly. Conventional chemotherapeutic induction regimens are usually too toxic for older patients, however, all trans-retinoic acid (ATRA) and arsenic trioxide (ATO) may be useful therapeutic options if used judiciously. This case series describes three octogenarians with APL all treated with ATRA and achieved complete remissions. The last patient received ATO at the time of first relapse and achieved a second remission. To our knowledge, this is the first report of successful use of ATO for induction in an octogenarian with APL.     

三氧化二砷治疗复发及难治性血液病的疗效及作用机制

三氧化二砷（ATO）被认为是不同类型癌症治疗中的有效药物。它是目前治疗急性早幼粒细胞白血病（APL）最有效的药物,特别是对全反式维甲酸（ATRA）及常规化疗耐药的APL。此外,实验室数据表明ATO在多种血液病及实体肿瘤细胞系中也具有活性。但是,作用机制尚不完全清楚。高剂量的ATO引发细胞凋亡,而在较低浓度时,它诱导部分分化。ATO作用机制涉及对导致细胞凋亡的线粒体跨膜电位的影响。它还作用于半胱天冬酶,NF-κB核因子或促凋亡蛋白和抗凋亡蛋白的活性等。本文主要综述了ATO在治疗复发及难治性APL、FLT3-ITD突变的AML、慢性粒细胞白血病急变期、重型再生障碍性贫血以及复发或难治性淋巴瘤的疗效及作用机制。这将为以后的研究者开展相关临床实验提供借鉴。     

白血病多靶点联合治疗的基础和临床研究

作为靶向诱导肿瘤细胞分化和凋亡的代表性模式,全反式维甲酸（ATRA）和三氧化二砷（ATO）治疗急性早幼粒细胞白血病（APL）取得了巨大的成功。ATRA与ATO均作用于异常转录因子PML—RARα,通过不同的途径降解PML—RARα致病蛋白,导致APL细胞分化和凋亡。临床试验证实了两药的协同作用,两药联合治疗初发APL获得了迄今急性白血病治疗的最好疗效。体外实验证实ANLL—M2b型白血病存在c—kif突变和过度表达,提示异常c—kif可以作为靶向治疗该类型白血病的候选靶点。冬凌草甲素（Oridonin）可以特异性地降解AML1-ETO,有可能成为治疗该类白血病的候选药物。酪氨酸激酶抑制剂在慢性粒细胞白血病（CML）治疗中获得了成功,该药联合砷剂治疗CML已在体外实验中初步证实其有效性。     

Upfront maintenance therapy with arsenic trioxide in acute promyelocytic leukemia provides no benefit for non‐t(15;17) subtype

Aims: The optimal maintenance therapy for patients with acute promyelocytic leukemia (APL) who achieved complete remission (CR) and complete consolidation chemotherapy is still controversial. Whether the use of arsenic trioxide (ATO) alone or along with all‐trans retinoic acid (ATRA) improves overall survival (OS) or disease‐free survival (DFS) is still debated. Methods: A retrospective reivew was conducted of 20 patients diagnosed with APL according to the French – American – British system. After achieving CR and receiving consolidation chemotherapy, nine patients were given maintenance therapy for 1 year (ATRA 45 mg/m²/day p.o., mercaptopurine 60 mg/m²/day p.o. and ATO 0.15 mg/kg/day × 5 days/week for six cycles in five patients; ATRA 45 mg/m²/d p.o. alternating with ATO 0.15 mg/kg/day × 5 days/week in 1 patient; ATRA only in three patients). Results: In all patients the rates of CR, 3‐year OS and 5‐year OS were 75, 71 and 57%, respectively. For patients treated with ATO maintenance, the rates were 100% for both 5‐year OS and 5‐year DFS. Four of six patients on ATO maintenance had grade 1 or grade 2 adverse events. Excluding the two patients who died from intracerebral hemorrhage within 4 days after diagnosis, these rates were 85, 82 and 78%, respectively. Conclusion: Upfront ATO maintenance therapy for one year is safe and appears to be effective, with the benefits restricted to patients with APL with t(15;17) translocation. Larger studies will be required to confirm this observation.     

Combination therapy with arsenic trioxide, all-trans retinoic acid, and chemotherapy in acute promyelocytic leukemia patients with various relapse risks

To improve the recovery rate of high-risk patients with acute promyelocytic leukemia (APL), we used all-trans retinoic acid (ATRA)/arsenic trioxide (ATO)/daunorubicin combination in remission induction, daunorubicin and cytarabine in consolidation, and ATRA/ATO/methotrexate ± 6-mercaptopurine in maintenance treatment of APL patients with various risks for relapse. Our results showed a high complete remission rate of 95.3%. Excluding the cases of early-death, no significant differences in event-free survival were observed between the intermediate-risk and high-risk group (p = 0.393) and the low-risk and high-risk group (p = 0.162). In addition, there were no significant differences between the groups in cumulative incidence of central nervous system relapse. In conclusion, our results suggest that APL patients benefit from combination ATO/ATRA/chemotherapy, and that this regimen is especially beneficial for patients with high-risk prognostic factors.     

三氧化二砷联合全反式维甲酸治疗初发急性早幼粒细胞白血病50例疗效观察

 目的　观察三氧化二砷（ATO）联合全反式维甲酸（ATRA）治疗初发急性早幼粒细胞白血病（APL）的疗效。方法　98例初发APL患者分为对照组和治疗组,对照组48例,治疗组50例。对照组采用常规ATRA+DA双诱导方案治疗;治疗组采用ATRA每天25 mg／m2,ATO每天0.15 mg／kg（ATRA后第10天开始）联合治疗,直至完全缓解（CR）,CR后接受ATO和ATRA联合巩固治疗。比较两组CR率、PML-RARα融合基因转阴时间及5年无病生存率。结果　对照组和治疗组CR率分别为89.5 %（43／48）和90.0 %（45／50）,获得CR时间分别为（30.0±5.1）d和（28.1±4.4）d,两组CR率（χ2＝-0.068,P＝0.946）及获得CR时间（t＝1.757,P＝0.083）相比差异均无统计学意义。在所有获得CR的患者中,3例分别在CR后第276、385和394天复发。所有患者发病时PML-RARα融合基因均阳性,对照组和治疗组CR时分别有25.0 %（5／20）和29.4 %（5／17）转阴,巩固后分别有92.5 %（37／40）和97.6 %（41／42）转阴。对照组和治疗组5年无病生存率分别为（85.3±5.9）%和（87.6±5.6）%,差异无统计学意义（χ2＝0.232,P＝0.630）。结论　ATO联合ATRA能有效治疗初发APL患者,可以作为常规化疗方案外的另一选择。     

Outcome of therapy‐related acute promyelocytic leukemia with or without arsenic trioxide as a component of frontline therapy

BACKGROUND:

Patients with therapy‐related acute promyelocytic leukemia (t‐APL) have been commonly exposed to topoisomerase inhibitors and may potentially benefit from induction regimens omitting anthracyclines.

METHODS:

Retrospective analysis of the outcomes of 29 patients with t‐APL who were either treated with arsenic trioxide (ATO) and all‐trans‐retinoic acid (ATRA) or with standard ATRA plus anthracycline‐based chemotherapy was performed.

RESULTS:

Prior therapy included chemotherapy alone, radiation alone, or a combination of the 2 in 19%, 33%, and 47% of patients, respectively. The combination of ATO and ATRA (n = 19) for induction resulted in a similar remission rate compared with ATRA plus chemotherapy (n = 10) (89% vs 70%; P = .35). The median overall survival for the patients treated with ATRA plus ATO was not reached compared with that for patients treated with ATRA plus chemotherapy (161 weeks; P = .79).

CONCLUSIONS:

In this cohort of t‐APL patients, outcomes with ATO and ATRA appeared to be comparable to anthracycline‐containing induction regimens. This combination may be preferable in t‐APL patients to avoid any risk of anthracycline‐induced toxicities. Cancer 2011. © 2010 American Cancer Society.