Histone deacetylase inhibition restores cisplatin sensitivity of Hodgkin's lymphoma cells

Increasing evidence suggest that epigenetic mechanisms (e.g. histone modification by histone deacetylases) play a major role in the pathogenesis of Hodgkin's lymphoma (HL). We treated HL cell lines with the histone deacetylase inhibitor vorinostat and investigated the gene expression profile of these cells by using DNA microarrays. Vorinostat inhibited cell proliferation and induced chances in the gene expression profile of HL cells, including down regulation of interleukin-26 and CD30. Vorinostat also increased sensitivity for cisplatin. Our data suggest that the combination of vorinostat and chemotherapy might be an interesting option for patients with chemoresistant HL.     

ATP生物荧光肿瘤体外药敏检测技术指导复发合并恶性胸腔积液的非小细胞肺癌的治疗

Objective: This study was aimed to research the feasibility of ATP-bioluminescence assay (ATP-TCA) guiding the treatment on recurrent non-small cell lung cancer (NSCLC) combined with malignant pleural effusion. Methods: We collected 30 pleural fluid samples which were approved to be positive by cytology from recurrent NSCLC patients. These cells were cocultured with chemotherapy medicines, single agent or drugs combination. Five drug concentrations, two parallel holes were examined in vitro for 4 days, the results were measured by adding luciferase-fluorescein working system and luminescence analyzer. We applied chemotherapy medicines according to the results in vitro of ATP-TCA. Results: There were differences among drug sensitivities of individuals. All the samples could be evaluated. Effective single drugs included cisplatinum, mitomycin C, doxorubicin, and pemetrexed disodium; sensitive drugs in the combination therapy were gemcitabine plus cisplatin, vinorelbine plus cisplatin, paclitaxel plus cisplatin, docetaxel plus cisplatin, and mitomycin C, vindesine plus cisplatin, in which gemcitabine + cispiatin (GEM + DDP) in vitro was the most efficient program. Conclusion: ATP-TCA in vitro sensitivity assay is rapid, reliable, and simple to guide the treatment of recurrent NSCLC with malignant pleural effusion, and can help clinicians to make the individual chemotherapy program.     

Growth inhibition and apoptosis induction in ovarian cancer cells

Standard therapy for the treatment of ovarian cancer is radical surgery followed by radiation and/or chemotherapy using cisplatin and paclitaxel. Unfortunately, some patients relapse after this first line chemotherapy and some patients become platinum-refractory. Therefore, we analyzed two different ovarian carcinoma cell lines for their sensitivity for gamma-irradiation and treatment with cisplatin, irinotecan, paclitaxel and gemcitabine. We found that both cell lines were rather resistant against gamma-irradiation and treatment with cisplatin and irinotecan whereas paclitaxel and gemcitabine resulted in a considerable reduction of the viability of the cancer cells. Both paclitaxel and gemcitabine treatment resulted in the induction of apoptosis. This sensitivity profile might be due to a particular subset of p53, which reacted with monoclonal antibodies DO-1 and PAb1801 but not with PAb1620 and PAb421. Gemcitabine and paclitaxel are highly efficient in the induction of apoptosis in ovarian cancer cells, which express a particular subset of the growth suppressor protein p53. Thus, a sensitivity profile for each ovarian carcinoma seems to be highly recommended before starting treatment.     

Clinical evaluation of chemotherapy response predictors developed from breast cancer cell lines

The goal of this study was to develop pharmacogenomic predictors in response to standard chemotherapy drugs in breast cancer cell lines and test their predictive value in patients who received treatment with the same drugs. Nineteen human breast cancer cell lines were tested for sensitivity to paclitaxel (T), 5-fluorouracil (F), doxorubicin (A) and cyclophosphamide (C) in vitro. Baseline gene expression data were obtained for each cell line with Affymetrix U133A gene chips, and multigene predictors of sensitivity were derived for each drug separately. These predictors were applied individually and in combination to human gene expression data generated with the same Affymetrix platform from fine needle aspiration specimens of 133 stage I-III breast cancers. Tumor samples were obtained at baseline, and each patient received 6 months of preoperative TFAC chemotherapy followed by surgery. Cell line-derived prediction results were correlated with the observed pathologic response to chemotherapy. Statistically robust differentially expressed genes between sensitive and resistant cells could only be found for paclitaxel. False discovery rates associated with the informative genes were high for all other drugs. For each drug, the top 100 differentially expressed genes were combined into a drug-specific response predictor. When these cell line-based predictors were applied to patient data, there was no significant correlation between observed response and predicted response either for individual drug predictors or combined predictions. Cell line-derived predictors of response to four commonly used chemotherapy drugs did not predict response accurately in patients.     

Drug-resistance enables selective killing of resistant leukemia cells: exploiting of drug resistance instead of reversal.

Drug resistance is a well recognized problem in cancer therapy. Despite the current dogma that drug resistance is always an obstacle for treatment, here I show that it provides opportunities for selective protection of non-resistant cells with killing of drug-resistant cancer cells. According to the proposed 'two-drug' strategy, the first drug should be ineffective against a target drug-resistant cell (ie the drug is a substrate of MRP or Pgp pumps). In addition, it must be cytostatic but not cytotoxic. The second drug, which is applied in sequence, must be a cycle-dependent apoptotic drug to which the target cell is not cross-resistant. Thus, low doses of adriamycin, etoposide and actinomycin D, used as the first drugs, were cytostatic to parental HL60 cells. Therefore, these drugs precluded Bcl-2/Raf-1 phosphorylation, PARP cleavage and cell death which are otherwise induced by paclitaxel, a mitosis-selective apoptotic drug for HL60 cells. In contrast, HL60/ADR cells which express MRP, a transporter which pumps out the first drugs from a cell, were insensitive to the first drugs and therefore readily underwent apoptosis following the second drug. This strategy also allowed a selective killing of HL60/TX cells which express MDR-1, with the only difference being that the second drug, paclitaxel, was substituted for epothilones, non-Pgp substrates. Lack of protection by the first drug, a Pgp substrate, resulted in HL60/TX killing by the second drug, whereas parental HL-60 cells were fully protected. Therefore, drug resistant cells can be selectively killed by a combination of drugs not killing sensitive cells. Lack of toxicity against normal cells will be clinically translated in reduction of adverse side-effects of chemotherapy against drug-resistant malignancies.     

Flavopiridol sensitivity of cancer cells isolated from ascites and pleural fluids.

PURPOSE: We examined the efficacy of flavopiridol, a cyclin-dependent kinase inhibitor that is undergoing clinical trials, on primary cancer cells isolated from the ascites or pleural fluids of patients with metastatic cancers. EXPERIMENTAL DESIGN: Metastasized cancer cells were isolated from the pleural fluids (n = 20) or ascites (n = 15) of patients, most of whom were refractory to chemotherapy. These primary cancer cells were used within 2 weeks of isolation without selecting for proliferative capacities. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide viability assay was used to characterize the response of these cancer cells to commonly used chemotherapeutic agents, and their response to flavopiridol was compared with rapidly dividing cultured cell lines. RESULTS: The primary cancer cells displayed phenotypes that were different from established cell lines; they had very low replication rates, dividing every 1 to 2 weeks, and underwent replicative senescence within five passages. These primary tumor cells retained their resistance to chemotherapeutic drugs exhibited by the respective patients but did not show cross-resistance to other agents. However, these cancer cells showed sensitivity to flavopiridol with an average LD50 of 50 nmol/L (range, 21.5-69 nmol/L), similar to the LD50 in established cell lines. Because senescent cells also showed similar sensitivity to flavopiridol, it suggests that the mechanism of action is not dependent on the activity of cyclin-dependent kinases that regulate the progression of the cell cycle. CONCLUSION: Using cancer cells isolated from the ascites or pleural fluids, this study shows the potential of flavopiridol against cancer cells that have developed resistance to conventional chemotherapeutic agents.     

Rituximab-mediated sensitization of B-non-Hodgkin's lymphoma (NHL) to cytotoxicity induced by paclitaxel,gemcitabine, and vinorelbine

Non-Hodgkin's lymphoma (NHL) responds initially to chemotherapy, however, the disease often relapses and acquires a chemoresistant phenotype. Third line treatment options such as paclitaxel, gemcitabine, and vinorelbine have been tried in patients with refractory or relapsed NHL with antilymphoma activity. Currently, rituximab (anti-CD20) has been approved for treatment of indolent NHL, with significant activity in a wide spectrum of B-cell malignancies, though a percentage of patients do not respond to rituximab treatment. Previous findings from our laboratory have demonstrated that rituximab can sensitize drug-resistant NHL B-lymphoma in vitro to some chemotherapeutic drugs. Hence, in this study, we examined the effect of combination treatment of rituximab and the drugs paclitaxel, gemcitabine, and vinorelbine on various NHL cell lines. Our findings indicate that pretreatment of NHL tumor cells with rituximab sensitizes drug-resistant NHL tumor cells to drug-mediated cytotoxicity. These findings suggest the potential clinical application of combination treatments of rituximab and paclitaxel, gemcitabine or vinorelbine in the treatment of drug/rituximab refractory non-Hodgkin's lymphoma.     

Cross-Resistance Studies of Isogenic Drug-Resistant Breast Tumor Cell Lines Support Recent Clinical Evidence Suggesting that Sensitivity to Paclitaxel may be Strongly Compromised by Prior Doxorubicin Exposure

Less than half of breast cancer patients respond to second-line chemotherapy with paclitaxel after failing treatment with anthracyclines such as doxorubicin. A recent clinical trial by Paridaens et al. [J. Clin. Oncol. 18 : 724-733, 2000] examined whether patients may derive a better clinical benefit if paclitaxel was administered before doxorubicin. While overall survival was similar regardless of the order of drug administration, a >4-fold reduction in the response rate to paclitaxel was observed after late crossover from doxorubicin, compared to the response rate to doxorubicin after late crossover from paclitaxel. This may be related to differences in the ability of the drugs to induce cross-resistance to each other. To test this hypothesis, we examined whether isogenic breast tumor cells selected for resistance to doxorubicin exhibit greater cross-resistance to paclitaxel and other drugs than identical cells selected for resistance to paclitaxel. We found that cells selected for resistance to paclitaxel showed strong resistance (>/=40-fold) to paclitaxel and docetaxel, with little cross-resistance (4-fold) to doxorubicin. In contrast, cells selected for resistance to doxorubicin exhibited 50-fold resistance to doxorubicin and a dramatic 4700-fold and 14,600-fold cross-resistance to paclitaxel and docetaxel, respectively. Doxorubicin-resistant cells exhibited higher P-glycoprotein and breast cancer resistance protein (BCRP) levels than paclitaxel-resistant cells. In addition, procaspase-9 was strongly downregulated in doxorubicin-resistant cells but not in paclitaxel-resistant cells. These differences may account for the contrasting cross-resistance profiles observed for the two cell lines and may help to explain why treatment of breast cancer patients with paclitaxel appears to be compromized by prior doxorubicin exposure.     

霍奇金淋巴瘤治疗进展:第54届美国血液学会年会深度报道

 【摘要】　目前,早期霍奇金淋巴瘤（HL）可通过化疗联合受累野放疗或单纯化疗的方式被治愈,但对放疗在早期HL治疗中的必要性仍有争议。而复发、难治HL的治疗仍存在着巨大挑战,除了大剂量化疗序贯自体造血干细胞移植外,尚缺乏有效的治疗手段。安全有效的新型药物如brentuximab vedotin的应用将有助于改善这类患者的生存预后。中期PET（PET-i）检查对早期和晚期HL患者都具有重要预后价值,尤其是在化疗2个周期后行PET检查,但依据PET-i结果而改变治疗方法除临床试验外并不被推荐。     

Effective cytotoxicity against human leukemias and chemotherapy-resistant leukemia cell lines by N-N-dimethylsphingosine.

We evaluated the cytotoxicity of dimethylsphingosine (DMS) against four human leukemia cell lines: two acute (HL60 and a multi-drug resistance MDR-positive derivative HL60-dox) and two blast crisis chronic myelogenous leukemias (JFP1, from a treatment refractory patient and K562), and against blasts isolated from 11 leukemia patients. Cell line viability decreased proportionally to DMS concentration and treatment time (P<0.001). HL60-dox and JFP1 were the most sensitive, indicating DMS efficacy against human leukemia MDR. Importantly, leukemia samples showed a similar sensitivity to DMS as that of the cell lines, firstly demonstrating PKC-independent sphingolipid activity against fresh human tumor specimens. DMS-based chemotherapy may improve leukemia treatment.     

Methotrexate-induced resistance to dactinomycin in choriocarcinoma

NaUCC-2, a choriocarcinoma cell line, was derived from a patient who had a very poor clinical response to combination chemotherapy. Methotrexate (MTX) might have inhibited the antitumor effect of dactinomycin. To investigate this point, in vitro studies were performed to determine the sensitivity and uptake of MTX and dactinomycin (administered individually and in combination) to NaUCC-2 and three other choriocarcinoma cell lines. Dihydrofolate reductase (DHFR) concentrations were studied as well. Although NaUCC-2 showed sensitivity to MTX and dactinomycin, which were comparable to the other cell lines when they were given separately, NaUCC-2 was unique in that the combination of MTX and dactinomycin was less lethal than dactinomycin given by itself. The uptake of MTX in NaUCC-2 was significantly higher than that in the other cell lines, and MTX also induced an increase in dactinomycin uptake in NaUCC-2. There was no significant difference in DHFR activity. Although additional studies are necessary to determine the mechanism responsible for this effect, these findings suggest that a mechanism other than drug uptake or DHFR activity must play a role in the drug resistance for choriocarcinoma. These findings also suggest that the most commonly used combination chemotherapy for choriocarcinoma, dactinomycin and MTX, may not always be the best method.     

Cytotoxicity of 2-chlorodeoxadenosine (cladribine, 2-cdA) in combination with other chemotherapy drugs against two lymphoma cell lines

Cladribine is a purine analog with impressive activity in patients with low-grade lymphoproliferative disorders. We studied the combination of cladribine with other antineoplastic drugs against two human-derived B-cell lymphoma cell lines in vitro. Cladribine was combined with cisplatin, daunorubicin, chlorambucil, paclitaxel or etoposide. Under the experimental conditions studied, only the combination of cladribine and cisplatin showed significantly increased cytotoxicity compared to the effect of either drug alone.     

MTT肿瘤药物敏感试验的方法学研究

目的：对MTT法肿瘤药物敏感试验的方法进行选择,旨在建立一种MTT肿瘤药敏试验的规范化操作流程。方法：MTT法测定K562细胞对抗癌药卡铂与紫杉醇的敏感性实验及其影响因素。结果：在每孔K562细胞数为5×10^3（浓度为5×10^4个/ml）、药物作用时间48h、MTT20μl作用6h、单波长570nm处测OD值时,药物对肿瘤细胞的抑制率较高,实验效果较好。结论：规范化的MTT肿瘤药敏试验可以为临床医师开展和实现化疗个体化提供较为可靠的实验依据。     

Favorable response of heavily treated wilms' tumor to Paclitaxel and Carboplatin

Background: Heavily treated Wilms' tumor responding to the combination of paclitaxel and carboplatin has not yet been reported. Case Report: A 17-year-old man presented with hematuria. He received a diagnosis of Wilms' tumor with multiple lung metastases and was treated with preoperative chemotherapy including vincristine, dactinomycin, and doxorubicin, a right nephrectomy, and adjuvant chemotherapy, followed by pulmonary metastasectomy. During the next 8 years, he suffered from 4 relapses and has been treated with multiple anticancer agents including high-dose chemotherapy with autologous peripheral blood stem cell transplantation. Finally, the disease progressed due to peritoneal and pleural metastases. With opioid administration for left shoulder pain due to pleural metastasis, he received combination chemotherapy with carboplatin (area under the curve = 4) and paclitaxel (175 mg/m(2)) on day 1. After 2 cycles, he achieved a partial response with mild toxicity. He received 7 cycles of the chemotherapy and the time to progression was 200 days. Conclusion: In a refractory case after intensive treatments, we succeeded to control the disease for a while.     

霍奇金淋巴瘤的治疗共识及临床研究进展

霍奇金淋巴瘤（HL）起源于淋巴造血组织,是治疗效果较好、治愈率较高的恶性肿瘤之一.目前,早期HL可通过化疗联合受累野放疗或单纯化疗的方式治愈,但复发、难治性HL患者仍缺乏有效治疗手段.近年来随着个体化分层治疗、靶向药物brentuximab vedotin、PET-CT检查等新型策略的应用,HL的预后评价及治疗研究均取得了新进展.     

Non-cytotoxic and sublethal paclitaxel treatment potentiates the sensitivity of cultured ovarian tumor SKOV-3 cells to lysis by lymphokine-activated killer cells

BACKGROUND: The standard treatment of epithelial ovarian cancer is tumor debulking by surgery, followed by six cycles of chemotherapy consisting of cisplatinum and paclitaxel. However, this therapy protocol is not satisfactory, since about 50% of the treated patients eventually experience recurrence within a few years of follow-up. Thus, a more innovative treatment modality is urgently needed for patients with this malignancy. We hypothesized that pretreatment of ovarian cancer SKOV-3 cells at a non-cytotoxic to sublethal dose range of paclitaxel would result in increased sensitivity to LAK-mediated killing. MATERIALS AND METHODS: MTT and trypan blue dye exclusion were used to determine the non-cytotoxic to sublethal range of paclitaxel against SKOV-3 cells. A 4-h 51Cr release cytotoxicity assay was used to evaluate the sensitivity of paclitaxel-treated and untreated SKOV-3 cells. Immunofluorescence/flow cytometric analysis was used for phenotypic changes of cells with or without paclitaxel treatment. RESULTS: Our results with trypan blue dye exclusion and MTT assays showed that the non-cytotoxic to sublethal range was between 0.001 microM and 0.01 microM. Pretreatment of SKOV-3 cells with paclitaxel at these doses for 72 h revealed significantly enhanced LAK-mediated killing against SKOV-3 cells with the highest sensitivity achieved with cells treated with 0.001 microM paclitaxel, as compared with the baseline killing of untreated cells using LAK cell alone (p < 0.05). The enhanced sensitivity of LAK-mediated killing appeared to be in part due to paclitaxel-induced expression of ICAM-1 on SKOV-3 cells. CONCLUSION: This treatment approach may be useful for further development of an effective therapeutic mode for patients with ovarian cancer.     

MTT法体外药敏实验预测宫颈癌细胞药物敏感性的初步探讨

背景与目的:近年来,宫颈癌新辅助化疗逐渐受到重视,应用广泛,但不同个体对不同化疗药物敏感性不同.本研究通过宫颈癌体外药敏实验来预测肿瘤细胞对化疗药物的敏感性,为临床筛选对宫颈癌相对敏感的化疗药物提供参考依据.方法:采用MTT法测定9种药物对32例宫颈癌患者的原代培养癌细胞的抑制水平.结果:①宫颈癌细胞对9种药物的敏感性依次为:力扑素＞泰素＞卡铂＞异环磷酰胺＞鬼臼乙叉甙＞氟尿嘧啶＞顺铂＞博来霉素＞环磷酰胺,力扑素、泰素和卡铂对肿瘤细胞的平均抑制率(IR)较高,分别为56.56%、55.66%和46.81%,与其它药物相比,差异有显著性(P＜0.05).②对Ⅰ b1期宫颈癌细胞,泰素、力扑素和卡铂IR值较其它药物高,分别为58.71%、53.00%和49.25%;对Ⅰ b2期宫颈癌细胞,力扑素和泰素的IR值较其它药物高,分别为65.26%和50.06%.③对于宫颈中分化鳞癌细胞,泰素、力扑素和卡铂IR值较其它药物高,分别为52.01%、49.21%和40.24%;对低分化鳞癌细胞,力扑素、泰素和卡铂的IR值较其它药物高,分别为61.16%、60.62%和48.75%.结论:①MTT法作为一种应用广泛的药敏实验方法,对宫颈癌新辅助化疗筛选敏感药物、选择临床化疗方案有一定的参考价值.②力扑素、泰素和卡铂对宫颈癌细胞较其它药物敏感.     

Cell type variation in responses to antimitotic drugs that target microtubules and kinesin-5

To improve cancer chemotherapy, we need to understand the mechanisms that determine drug sensitivity in cancer and normal cells. Here, we investigate this question across a panel of 11 cell lines at a phenotypic and molecular level for three antimitotic drugs: paclitaxel, nocodazole, and an inhibitor of kinesin-5 (also known as KSP, Eg5, Kif11). Using automated microscopy with markers for mitosis and apoptosis (high content screening), we find that the mitotic arrest response shows relatively little variation between cell types, whereas the tendency to undergo apoptosis shows large variation. We found no correlation between levels of mitotic arrest and apoptosis. Apoptosis depended on entry into mitosis and occurred both from within mitosis and after exit. Response to the three drugs strongly correlated, although paclitaxel caused more apoptosis in some cell lines at similar levels of mitotic arrest. Molecular investigations showed that sensitivity to apoptosis correlated with loss of an antiapoptotic protein, XIAP, during the drug response, but not its preresponse levels, and to some extent also correlated with activation of the p38 and c-Jun NH(2) kinase pathways. We conclude that variation in sensitivity to antimitotic drugs in drug-naive cell lines is governed more by differences in apoptotic signaling than by differences in mitotic spindle or spindle assembly checkpoint proteins and that antimitotics with different mechanisms trigger very similar, but not identical, responses.