Src family kinase inhibitor PP2 enhances differentiation of acute promyelocytic leukemia cell line induced by combination of all-trans-retinoic acid and arsenic trioxide

An all-trans-retinoic acid (ATRA) and arsenic trioxide (ATO) combination yields high-quality remission and survival in newly-diagnosed acute promyelocytic leukemia (APL). For subsequent similar data, NCCN guidelines indicate that ATRA plus ATO is one of the recommended regimens for the treatment of patients with APL. We demonstrated SFK (Src family kinase) inhibitor PP2-enhanced APL cell differentiation when combined with either ATRA or ATO with difference in activation of RA-induced genes. In this study, we investigated whether SFK inhibitor PP2 could enhance the differentiation of NB4 APL cells when combined with ATRA and ATO and the changes in the expression of intercellular adhesion molecule-1 (ICAM-1) derived from the retinoic acid receptor (RAR) target gene.     

Granulocyte-colony Stimulating Factor and Retinoic Acid Cooperatively Induce Granulocytic Differentiation of Acute Promyelocytic Leukemia Cells in vitro

The interaction of granulocyte-colony stimulating factor (G-CSF) and retinoic acid (RA) in proliferation and differentiation of acute promyelocytic leukemia (APL) cells was examined. G-CSF stimulated proliferation of APL cells at concentrations of 0.1 to 50 ng/ml in a dose dependent manner. More than 10⁻⁸ M RA induced granulocytic differentiation of APL cells. Although G-CSF induced lysozyme activities in APL cells, it alone did not induce terminal differentiation of APL cells. G-CSF significantly enhanced the RA-induced granulocytic differentiation of APL cells in vitro. Enhancement by G-CSF was not due to the prolongation of survival of RA-induced differentiated cells, but the differentiation-inducing effects of G-CSF might be evident only in the presence of RA. Since G-CSF has a potential to induce the granulocytic differentiation of myeloid leukemia cells, G-CSF in combination with RA may be applicable in differentiation induction therapy for some types of myeloid leukemia.     

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.     

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.     

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.     

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.     

Effect of arsenic trioxide or ATRA on primary APL cell or HL-60 cells and their value analysis in hyperleukocytosis

We have done a lots of works to detect the mechanism of hyperleukocytosis in patients with acute promyelocytic leukemia (APL) after treatment with all trans retinoic acid (ATRA). The early results of study showed that some important factors are related to the hyperleukocytosis, for example, the relationship between serum G-CSF and variation of promyelocytes or more matured granulocytes[1,2]. The further study also indicated that effect of G-CSF on leukemia cells was related to the variat…     

All-trans retinoic acid and hematopoietic growth factors regulating the growth and differentiation of blast progenitors in acute promyelocytic leukemia

Although acute leukemia is generally thought to be characterized by maturation arrest, it has been shown that differentiation occurs in blast cells of acute myelogenous leukemia (AML) in vitro as well as in vivo, and that morphologically abnormal mature polymorphonuclear neutrophils (PMNs) often seen in patients with AML are possibly derived from spontaneously differentiating leukemic cells. Acute promyelocytic leukemia (APL) is an unique example in which these features of AML are evident in an almost complete form; administration of all-trans retinoic acid (ATRA) induces differentiation of neoplastic cells into mature neutrophils and successfully induce complete remission in most patients. However, PMNs appearing during ATRA treatment are morphologically abnormal, as indicated not only by the presence of Auer rods but also by neutrophil secondary-granule deficiency that is commonly seen in AML. Moreover, ATRA has heterogeneous effects on the growth of blast progenitors in APL in different patients, being inhibitory, stimulatory or ineffective, which might account in part for the leukemia relapse in patients treated with ATRA alone. Hematopoietic growth factors regulate the growth of blast progenitors in APL. Among them, granulocyte colony-stimulating factor (G-CSF) is unique in that it preferentially stimulates clonal growth, but not self-renewal, in many APL cases, and synergistically enhances the differentiation-inducing effect of ATRA when used in combination. Many other compounds also exert such synergistic effects with ATRA, for which a variety of mechanisms have been suggested. It is crucial to precisely elucidate the functions of these molecules governing the growth/differentiation balance of AML blast progenitors and the mechanisms underlying their deregulated differentiation program in order to achieve effective differentiation therapy for patients with AML, not restricted to APL.     

A novel differentiation-inducing therapy for acute promyelocytic leukemia with a combination of arsenic trioxide and GM-CSF.

Arsenic trioxide (As2O3) effectively induces clinical remission via apoptosis in relapsed acute promyelocytic leukemia (APL). However, because this new anti-leukemic drug is also considered to be a poison, its possible adverse effects are a highly important issue related to its clinical use. We here investigated, both in vitro and in vivo, the effects of a combination of As2O3 and GM-CSF as a novel therapeutic approach for the treatment of APL. Treatment of both retinoic acid (RA)-sensitive and -resistant APL cell lines (NB4 and UF-1 cells, respectively), as well as primary APL cells with a combination of As2O3 and GM-CSF for 4 days resulted in inducing differentiation, but not apoptosis, to mature granulocytes. In addition, a combination of both agents induced degradation of the PML/RARalpha protein. GM-CSF was found to be associated with increased tyrosine phosphorylation of Jak2 kinase in both NB4 and UF-1 cells, and a specific inhibitor of Jak2, AG490, completely blocked the ability of GM-CSF to prevent apoptosis and induce differentiation of As2O3-treated UF-1 cells. In in vivo analysis, As2O3 induced differentiation of APL cells in a RA-resistant APL model of human GM-CSF-producing transgenic SCID mice that had a high level of human GM-CSF in their sera. In contrast, As2O3 alone diminished tumors in UF-1 cells transplanted into NOD/SCID mice via induction of apoptosis. In conclusion, a combination of As2O3 and GM-CSF appears to be a novel differentiation-inducing therapy in patients with APL, including relapsed or RA-resistant cases.     

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.     

The impact of arsenic trioxide and all-trans retinoic acid on p53 R273H-codon mutant glioblastoma

Glioblastoma (GBM) is the most common primary brain tumor in adults and demonstrates a 1-year median survival time. Codon-specific hotspot mutations of p53 result in constitutively active mutant p53, which promotes aberrant proliferation, anti-apoptosis, and cell cycle checkpoint failure in GBM. Recently identified CD133⁺ cancer stem cell populations (CSC) within GBM also confer therapeutic resistance. We studied targeted therapy in a codon-specific p53 mutant (R273H) created by site-directed mutagenesis in U87MG. The effects of arsenic trioxide (ATO, 1 μM) and all-trans retinoic acid (ATRA, 10 μM), possible targeted treatments of CSCs, were investigated in U87MG neurospheres. The results showed that U87-p53^R273H cells generated more rapid neurosphere growth than U87-p53^wt but inhibition of neurosphere proliferation was seen with both ATO and ATRA. U87-p53^R273H neurospheres showed resistance to differentiation into glial cells and neuronal cells with ATO and ATRA exposure. ATO was able to generate apoptosis at high doses and proliferation of U87-p53^wt and U87-p53^R273H cells was reduced with ATO and ATRA in a dose-dependent manner. Elevated pERK1/2 and p53 expression was seen in U87-p53^R273H neurospheres, which could be reduced with ATO and ATRA treatment. Additionally, differential responses in pERK1/2 were seen with ATO treatment in neurospheres and non-neurosphere cells. In conclusion, codon-specific mutant p53 conferred a more aggressive phenotype to our CSC model. However, ATO and ATRA could potently suppress CSC properties in vitro and may support further clinical investigation of these agents.     

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。     

Treatment of acute promyelocytic leukemia with arsenic trioxide: clinical results and open questions

Acute promyelocytic leukemia (APL) is a rare form of acute myeloid leukemia. The specific translocation t(15;17), which results in the fusion gene PML–RARA is the diagnostic and pathomechanistic hallmark of APL. By combination, treatment consisting of the differentiating agent all-trans retinoic acid (ATRA), which targets this molecular lesion, and cytotoxic chemotherapy, cure can be achieved in over 70% of patients. Recently, arsenic trioxide (ATO) has emerged to be the most active single agent in the treatment of APL. Previous studies employing ATO in relapse settings reported average complete remission rates of 85% and a mean overall survival of over 60%. In recent approaches installing ATO in first-line treatment, ATO-induced response rates comparable to previous combination regimen. The results of these newer studies indicate that the backbone of chemotherapy can be dramatically reduced or completely replaced by ATO and ATRA with similar or even better outcome.     

Differentiating agents in pediatric malignancies: All-trans-retinoic acid and arsenic in acute promyelocytic leukemia

Acute promyelocytic leukemia (APL) is the most potentially curable type of acute myeloid leukemia. It is characterized by the chromosomal translocation t(15;17), which results in the fusion gene PML-RAR-α. The introduction of all-trans- retinoic acid (ATRA) was a major advance in treatment of this disease. This agent induces terminal differentiation of malignant myeloid cells to mature neutrophils, and its side effects are usually well tolerated in children. ATRA does not eradicate the malignant myeloid clone in APL and, eventually, resistance develops. Arsenic trioxide induces nonterminal differentiation of malignant promyelocytes and promotes apoptosis. APL patients treated with ATRA or arsenic trioxide have rapid resolution of their coagulopathy. Because both of these drugs are well tolerated in children and their synergy has been shown in animal models, the possibility of combining ATRA and arsenic trioxide in frontline therapy for children with APL is being considered.     

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.     

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。     

急性早幼粒细胞白血病对全反式维甲酸耐药的机制及其逆转

目的　揭示急性早幼粒细胞白血病 (APL)产生维甲酸耐药的机制 ,探讨耐药性逆转的方法。方法　MTT法测定细胞增殖 ,极限稀释法诱导APL细胞耐药性。RT PCR及流式细胞仪测定MDR1、消炎痛及PGE1增殖影响实验分别间接测定GST酶活性或cAMP作用。干扰素、高三尖杉酯碱(HHT)及三氧化二砷 (As2 O3 )进行体外耐药细胞逆转实验。结果　对全反式维甲酸 (ATRA)耐药的HL6 0细胞MDR1阴性 ,初发时MDR1为阴性的APL患者复发后仍为阴性。消炎痛对耐药的HL6 0增殖分化无影响 ,前列腺素E(PGE)可部分恢复ATRA对耐药HL6 0的作用。干扰素可明显逆转HL6 0的耐药性。As2 O3 与HHT对ATRA耐药的HL6 0细胞有明显作用。结论　APL细胞耐药并非多药耐药 ,消炎痛或干扰素明显逆转其耐药性 ,ATRA与As2 O3 化疗药无交叉耐药性。