鞘磷脂及神经酰胺对人结肠癌细胞(HT-29)的影响

目的　研究鞘磷脂及其代谢产物神经酰胺对人结肠癌细胞的影响。方法　采用噻唑蓝 (MTT)显色法、细胞核分裂指数和3H TdR掺入试验方法 ,所设剂量分别为鞘磷脂 5、10、2 0、40 μg ml和神经酰胺 6 2 5、12 5、2 5、5 0 μmol L ,分别以无水乙醇和二甲基亚砜 (DMSO)为溶剂对照。结果　鞘磷脂对HT 2 9细胞的生长无明显作用 ,神经酰胺对HT 2 9细胞的生长有明显抑制作用。在MTT试验中 ,不同浓度神经酰胺对HT 2 9细胞的 7d抑制率分别为 3 9%、85 %、98%和98% ;在核分裂指数试验中 ,随着神经酰胺剂量的增高 ,其核分裂指数明显降低 ;在3H TdR掺入试验中 ,可见随着神经酰胺剂量的增高 ,3H TdR掺入到HT 2 9细胞中明显的减少 ,与阴性对照组相比 ,差异有显著性 (P <0 0 5 )。结论　鞘磷脂对HT 2 9细胞生长无明显作用 ,神经酰胺对HT 2 9细胞的生长增殖有明显抑制作用 ,这可能是鞘磷脂抑制结肠癌的机制之一。     

Imbalanced DNA synthesis induced by cytosine arabinoside and fludarabine in human leukemia cells

Previous studies have demonstrated that cytosine arabinoside (araC) induces an accumulation of Okazaki fragments, while fludarabine (FaraA) inhibits Okazaki fragment synthesis. We extended these observations in the present study to provide insights into various mechanisms by which these anticancer drugs affect DNA replication and induce genomic instability in human CEM leukemia cells. Neither araC nor FaraA induced a detectable amount of re-replicated DNA in S-phase cells, which indicated that drug-induced alterations in Okazaki fragment synthesis were not accompanied by DNA re-replication. Synthesis on both leading and lagging DNA strands within the c-myc locus was measured in cells incubated with equitoxic concentrations of araC or FaraA. In araC-treated cells, nascent DNA from the lagging strand was enriched about 5-fold compared with the leading strand. In contrast, FaraA did not induce any replication imbalance. AraC- and FaraA induced changes in the frequency of N-(phosphonacetyl)-l-aspartate (PALA) resistance and the extent of CAD gene amplification were monitored as markers of drug-induced genomic instability. At concentrations that reduced cloning efficiency by 50% (IC(50)), araC increased the frequency of PALA resistance about 4-fold, while FaraA did not have a significant effect on the frequency of PALA resistance. Pretreatment with araC also increased the extent of CAD gene amplification. We propose that the imbalanced DNA synthesis induced by araC leads to the accumulation of Okazaki fragments on the lagging arms and single-stranded DNA regions on the leading arms of replication forks. The formation of these abnormal replication structures was associated with the generation of genomic instability.     

氟达拉滨联合环磷酰胺治疗慢性淋巴细胞白血病疗效观察

目的观察氟达拉滨联合环磷酰胺治疗慢性淋巴细胞白血病(CLL)的疗效和安全性。方法将CLL患者24例随机分为治疗组和对照组各12例。治疗组予以氟达拉滨联合环磷酰胺方案治疗,对照组予以CHOP方案化疗。比较2组总有效率及不良反应。结果治疗组总有效率为83.3%高于对照组的66.7%(P<0.05);2组合并感染率、中性粒细胞减少发生率、血小板减少发生率及持续时间比较差异均无统计学意义(P>0.05)。结论氟达拉滨联合环磷酰胺组治疗CLL具有完全缓解率及总有效率高、不良反应轻等优点,值得临床推广应用。     

A Phase III trial of fludarabine,cyclophosphamide, and rituximab vs. pentostatin,cyclophosphamide, and rituximab in B-cell chronic lymphocytic leukemia

Background Uncontrolled studies comparing pentostatin (P), cyclophosphamide (C), and rituximab (R) (PCR) to fludarabine plus C+R (FCR) suggest similar efficacy with fewer infectious complications with PCR. We compared FCR and PCR in previously-untreated or minimally-treated B-cell chronic lymphocytic leukemia (CLL). Treatment FCR (F 20 mg/m² Days 1–5, C 600 mg/m² Day 1, R 375 mg/m² Day 1) (28-day cycles) or PCR (P 4 mg/m² Day 1, C 600 mg/m² Day 1, R 375 mg/m² Day 1) (21-day cycles). Dose 1 of R: 100 mg/m² was given on Day 8 Cycle 1 and the remainder on Day 9; in subsequent cycles the entire dose was given on Day 1. Results Ninety-two patients were randomly assigned to each group (N = 184). Groups were balanced; ~20% had received prior chemotherapy. The infection rate (FCR/PCR) was 31%/36%, the infective event rate was 38%/45%; 30 (35%)/37 (44%) patients were hospitalized; total hospitalization days was 271/404. 12 (14%)/6 (7%) patients achieved complete remissions (CR); the overall response rate (ORR) including CR+nodular PR (nPR)+PR was 59%/49%. Grade 3–4 treatment related AEs: neutropenia (69%/57%), leukopenia (34%/17%), thrombocytopenia (13%/6%). Grade 3–4 infections: febrile neutropenia (8%/6%), fever (2%/6%), infection (1%/3%), urinary tract infection (1%/0%), pneumonia (3%/1%), and sepsis (1%/2%); 5 deaths (1 FCR/4 PCR) were treatment-related. Conclusions PCR and FCR have significant activity in CLL and can be given safely in the community setting despite significant toxicity. ORRs were lower than expected; the CR rate was higher (NS) with FCR. This trial did not demonstrate a lower infection rate with PCR.     

氟达拉滨联合环磷酰胺对慢性淋巴细胞白血病的临床疗效研究

目的观察氟达拉滨联合环磷酰胺对慢性淋巴细胞白血病的临床疗效及对外周血Th17和Treg细胞比例的影响。方法 200例慢性淋巴细胞白血病患者随机分为对照组100例和试验组100例。对照组患者第1³天静脉滴注氟达拉滨25 mg·m^-2;试验组第1³天静脉滴注氟达拉滨25 mg·m^-2和环磷酰胺250 mg·m^-2。2组均4周为1个周期,治疗2个周期。比较2组患者的临床疗效、不良反应发生情况以及外周血Th17和Treg细胞比例。结果试验组患者总改善率为98.0%,显著高于对照组的72.0%（P<0.05）。试验组患者总不良反应发生率为8.0%,显著低于对照组的29.0%（P<0.05）。试验组患者Th17细胞和Treg细胞状况均显著优于对照组（P<0.05）。结论氟达拉滨联合环磷酰胺能够有效地治疗慢性淋巴细胞白血病,降低不良反应发生率,并能显著改善外周血Th17和Treg细胞比例。     

Increase in ceramide level after application of various sizes of sphingomyelin liposomes to a cultured human skin model

Sphingomyelin-based liposomes (SPM-L) that were sized (or not) by extrusion through a filter with pores of 100, 200, or 400 nm were applied to a three-dimensional cultured human skin model in order to evaluate which size of SPM-L was most effective at increasing its ceramide level. The diameters of the SPM-L in PBS were 102.7, 181.0, 224.0, and 380.1 nm. The diameters of the liposomes in the culture medium were 117.5, 199.2, 242.1, and 749.8 nm. The diameter of the small liposomes (<200 nm in diameter) did not change much, at least for 7 days. SPM-L in saline or culture medium were applied to the basal layer side or stratum corneum side of the cultured skin model, and ceramide II, III, V, and VI were then extracted from it. The extracted ceramide molecules were separated by HPTLC, and the concentration of each type of ceramide was quantified using a densitometer. When the small SPM-L (110 or 190 nm in diameter) were applied to the basal layer side, the levels of ceramide III and V were increased. When they were applied to the stratum corneum side, the levels of ceramide II, III, V, and VI were significantly increased compared to those of the PBS group, especially after the application of the small SPM-L (110 nm in diameter). Thus, the application of small SPM-L was useful for increasing the ceramide II, III, V, and VI levels of a cultured human skin model.     

Inotuzumab ozogamicin in the treatment of B-cell acute lymphoblastic leukemia

INTRODUCTION: Inotuzumab ozogamicin (CMC-544) is a humanized anti-CD22 monoclonal antibody conjugated with calicheamicin. Preclinical data indicate activity against B-cell tumors and early results from clinical trials indicate activity against B-cell lineage acute lymphoblastic leukemia (ALL). AREAS COVERED: This paper reviews the design, pharmacokinetic and pharmacodynamic characteristics, and preclinical and clinical experience of inotuzumab ozogamicin in adult ALL. EXPERT OPINION: Inotuzumab ozogamicin appears as an effective salvage therapy in patients with advanced ALL, allowing more patients to receive stem cell transplant (SCT) with encouraging response rates. This agent should provide a unique opportunity to treat selected ALL patient subpopulations.     

Changes in drug sensitivity following acute and chronic exercise

A number of factors are known to influence drug sensitivity. These include biological variables such as genetics, age, endocrine status and gender, as well as environmental variables such as operant schedules, ambient temperature and sleep deprivation. Additional factors function as either biological or environmental variables in different situations. For example, chronic drug administration can produce tolerance and cross tolerance and function as a biological variable. Acute administration of the same compound can function as an environmental variable. The present study examined exercise as both a biological and an environmental variable influencing drug sensitivity. Chronic exercise leads to relatively long term changes in physical fitness level, and functions as a biological variable. Fitness level did not influence drug sensitivity when physically conditioned animals and non-exercised control subjects were compared under rested conditions. Mild acute exercise, an environmental variable, increased sensitivity to muscarinic antagonists in the control subjects but not in the exercise trained animals. These results indicate that exercise state should be considered as an environmental variable capable of influencing drug response and that biological fitness level modifies this effect.     

Anti-CD44 mAb for the treatment of B-cell chronic lymphocytic leukemia and other hematological malignancies: evaluation of WO2013063498

Introduction: The CD44 transmembrane glycoprotein family mediates cellular responses to the microenvironment through binding of hyaluronic acid (HA) and other proteins of the extracellular matrix. These interactions start intracellular signaling cascades that foster tumor growth, survival and spread.

Areas covered: The patent concerns the use of a humanized anti-CD44 mAb (RG7356) to treat patients with aggressive forms of chronic lymphocytic leukemia (CLL). The RG7356 humanized antibody was designed to bind the constant region of CD44, preventing binding with HA. The interruption of this circuit in vitro is followed by the induction of caspase-dependent apoptosis in leukemic cells. In agreement with a functional association between CD44 and zeta-associated protein of 70 kDa (ZAP-70) in activating intracellular signaling, the strongest effects were observed in ZAP-70⁺ CLL cells, generally associated to a poor prognosis. Furthermore, these effects were confirmed using in vivo models, where CLL cells were xenografted in immunocompromised mice.

Expert opinion: In summary, these studies suggest that this new humanized mAb may provide additional clinical benefit to CLL patients receiving current standard treatments, specifically to those with a more aggressive disease.     

Plasmapheresis for refractory urticarial vasculitis in a patient with B-cell chronic lymphocytic leukemia

BACKGROUND: Urticarial vasculitis is a form of cutaneous leukocytoclastic vasculitis clinically characterized by persistent and often painful urticarial lesions. Numerous systemic diseases have been associated with urticarial vasculitis, including certain hematologic disorders. This distinctive form of cutaneous necrotizing vasculitis can be resistant to standard therapeutic modalities, necessitating more aggressive intervention. METHODS: We report a case of refractory urticarial vasculitis developing in association with B-cell chronic lymphocytic leukemia in a 46-year-old man. We also reviewed the literature to identify other cases of urticarial vasculitis managed with this therapeutic modality. RESULTS: The disease progressively improved during 6 treatments with plasmapheresis (plasma exchange). In the additional cases identified in the literature, plasmapheresis was generally effective and well tolerated. CONCLUSION: On the basis of these findings, we propose that plasmapheresis be considered a treatment option for refractory urticarial vasculitis.     

Lenalidomide in the treatment of chronic lymphocytic leukemia

Introduction: Lenalidomide is an immunomodulatory drug (IMiD) with a unique mode of action (MOA) that may vary across disease-type. It is currently approved in multiple myeloma (MM), myelodysplastic syndrome (MDS) and mantle cell lymphoma (MCL), yet is also clinically active in a host of lymphoproliferative diseases, including chronic lymphocytic leukemia (CLL). Due to its protean effects on the immune system, lenalidomide may be particularly appealing in CLL, which is distinct in its ability to evade immune recognition and cause immunosuppression.

Areas covered: This review recaps the biological mechanisms of lenalidomide specific for CLL, and summarizes the clinical data in previously untreated and relapsed/refractory (R/R) CLL patients, with emphasis on toxicity. Moreover, lenalidomide treatment is put into the context of the highly effective targeted agents that are drastically changing the therapeutic approach in CLL.

Expert opinion: Lenalidomide is a potent drug in CLL, both in first line and relapse. However, in comparison to other newly available agents, lenalidomide has slow onset of efficacy and notable toxicity profile that limits both its single agent use and combinations with chemotherapy. Future trials will hopefully direct our ability to harness lenalidomide MOA to best incorporate it in the rapidly evolving landscape of CLL treatment.     

Lenalidomide in the treatment of chronic lymphocytic leukemia

INTRODUCTION: insights into the role of the tumor microenvironment and of immune dysfunction in chronic lymphocytic leukemia (CLL) have opened the way for further augmenting the therapeutic armamentarium for CLL patients. In this respect, lenalidomide represents an exciting drug since it is able to eliminate CLL cells without immunosuppression. AREAS COVERED: mechanism of action and clinical trials of lenalidomide in CLL, and suggestions for its future utilization are reviewed. The most relevant papers and the meeting abstracts published up to July 2010 were used as sources for this review. This review will help readers understand the mechanism of action of lenalidomide and will provide a comprehensive summary regarding efficacy and safety of this drug in CLL patients. EXPERT OPINION: lenalidomide shows good activity against CLL. However, the toxicity profile is significant and can result in serious and potentially life-threatening side effects. Definitive data from ongoing trials will aid better definition of its status in CLL therapy. Moreover, clarification of the exact mechanism(s) of action in CLL will allow more precise use of lenalidomide and design of more efficacious combination therapies.     

复发难治B系淋巴瘤CAR-T治疗后细胞因子变化

目的分析于我中心行CAR-T治疗患者治疗前后细胞因子变化,评估相关细胞因子在免疫治疗中的作用及相关机制。方法分别在复发难治B系淋巴瘤患者CAR-T细胞回输前及回输后4、7、14 d采集外周血,利用流式细胞术监测患者相关细胞因子(Th1、Th17、Foxp3、TNF-α、TGF-β、IL-10、IFN-γ、T抑制细胞、NK细胞及CRP)水平,分析变化趋势。结果 4次CAR-T细胞回输数目分别为:8×10~7、10×10~7、9.6×10~7、10×10~7。CAR-T细胞回输前及回输后4、7、14 d,Th1、Th17及CRP变化趋势与临床疗效相关,NK细胞、CD8~+细胞未见明显相关性,而Tregs细胞作用尚未明确。结论应用CAR-T细胞回输后,Th1、Th17相关效应性T细胞细胞因子上升趋势与临床疗效密切相关,NK细胞、CD8+T细胞未见明显相关性,而Tregs细胞作用尚未明确。关于CAR-T治疗前后序贯治疗、移植与CAR-T治疗的关系,是否可以应用细胞因子制剂促进CAR-T细胞的扩增与效应,仍有待进行相关大样本临床研究。     

神经酰胺诱导人结肠癌细胞凋亡作用

目的探讨外源性神经酰胺诱导人结肠癌HT-29细胞的凋亡作用。方法采用光镜、电镜、荧光显微镜和琼脂糖凝胶电泳方法检测C2-神经酰胺诱导HT-29细胞凋亡。MTT法检测C2-神经酰胺对HT-29细胞线粒体功能的影响。结果C2-神经酰胺使HT-29细胞发生核染色质断裂、DNA Ladder、凋亡小体等典型凋亡表现,12和24 h凋亡细胞率分别为64.1%和81.3%,呈现时间-剂量依赖关系。同时C2-神经酰胺处理细胞6 h后,细胞线粒体功能即出现损伤。结论外源性神经酰胺能诱导人结肠癌HT-29细胞凋亡,参与其凋亡调控。     

Bosutinib for the treatment of chronic myeloid leukemia in chronic phase

The clinical outcome for patients with chronic myeloid leukemia in chronic phase (CML-CP) is currently very favorable due to the availability of tyrosine kinase inhibitors (TKIs) that are well tolerated and effectively suppress the constitutively activated BCR-ABL1 kinase that underlies the pathogenesis of this malignancy. Three TKIs -imatinib, nilotinib and dasatinib- have been approved as frontline therapy in CML-CP. Another TKI, bosutinib, inhibits with high potency numerous tyrosine kinases, including BCR-ABL1, Src family of kinases and MAPK, among others. Like nilotinib and dasatinib, bosutinib is a second-generation TKI that inhibits the majority of mutations associated with imatinib resistance, with the exception of T315I. In patients with CML-CP with prior intolerance or resistance to imatinib therapy, bosutinib rendered response rates similar to those observed in the same patient population treated with nilotinib or dasatinib. Preliminary results from the ongoing phase III BELA study in which bosutinib is compared in a randomized fashion to imatinib for patients with newly diagnosed CML-CP have been recently reported. We herein summarize the preclinical and clinical experience of bosutinib in CML.     

慢性粒细胞白血病不同药物治疗后树突细胞的变化

目的探讨干扰素α(IFN-α)、伊马替尼、羟基脲3种药物治疗慢性粒细胞白血病后树突细胞(Dendritic cells,DC)的含量变化。方法采用流式细胞仪检测不同药物治疗后26例慢性粒细胞白血病患者及9例正常人外周血树突细胞的含量。结果羟基脲组外周血树突细胞含量较正常人明显下降(P<0.05)。伊马替尼组和TNF-α组外周血树突细胞含量比羟基脲组明显增加(P<0.05),但和正常人相似(P>0.05)。伊马替尼和TNF-α两组之间树突细胞含量比较差异无统计学意义(P>0.05)。结论慢性粒细胞白血病患者体内DC含量明显减少,抗肿瘤作用下降。而伊马替尼和IFN-α可促使体内DC的增殖,从而达到抗肿瘤的作用,两者间促使DC增殖的作用无显著差异。     

Dequalinium induces apoptosis in peripheral blood mononuclear cells isolated from human chronic lymphocytic leukemia

B-cell chronic lymphocytic leukemia (B-CLL) is an abnormal neoplasic proliferation of B cells, which accumulate mainly in the bone marrow and blood preventing both B cells development in the lymph nodes and the ability to fight against infection. The antitumor agents used in chemotherapy are aimed at inducing malignant cell death, thus limiting the growth and spreading of these cells. However, the lack of specificity for tumor cells exhibited by these agents causes undesirable side effects that have led to the investigation of new therapeutic strategies designed to specifically target malignant cells and thus trigger selective cell destruction. Dequalinium (DQA) is an antitumoral agent that selectively accumulates in the mitochondria and has been shown to display anticancer activity in cells from different malignancies. In the present study, the DQA-induced cytotoxicity in B-CLL cells was analyzed by measuring cell viability and cell death, either by necrosis or apoptosis. Our results support the importance of DQA as a selective and potential antileukemic drug with a higher cytotoxic effect on peripheral blood mononuclear cells from B-CLL patients than in those from healthy donors and encourage the performance of further studies in combination with other agents.     

Bendamustine for treatment of chronic lymphocytic leukemia

INTRODUCTION: Chronic lymphocytic leukemia (CLL) often has an extended disease course. With a median age at diagnosis of 72 years, newer treatment options with less toxicity than standard nucleoside analogue-based regimens are needed. Historically, few therapy options are available once CLL has become refractory to nucleoside analogues. Bendamustine has emerged as a feasible therapy for older and less fit CLL patients, with clinical efficacy in previously untreated and refractory CLL. AREAS COVERED: This paper reviews several of the pivotal clinical trials that established the clinical activity of bendamustine in previously untreated and relapsed/refractory CLL. The toxicity profile of bendamustine, primarily myelosuppression and infections, is reviewed and compared across different CLL populations. A review of the clinical data focuses on potential explanations for differences in response rates and duration of remission reported across studies and how this may impact the development of therapies for CLL. EXPERT OPINION: Bendamustine is a valuable new agent for the management of CLL. Ongoing clinical trials are comparing bendamustine with standard CLL regimens in untreated disease, and investigating bendamustine combinations with novel targeted therapies and monoclonal antibodies. These studies will help to define the optimal role for bendamustine in CLL management.