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1.
BMC Cancer ; 14: 709, 2014 Sep 24.
Artículo en Inglés | MEDLINE | ID: mdl-25253633

RESUMEN

BACKGROUND: Patients with malignant mesothelioma have a poor prognosis and only 40% respond to first line treatment; a combination of pemetrexed and cisplatin or carboplatin. We used primary malignant mesothelioma cells and an ex vivo chemosensitivity assay with future purpose to predict best choice of treatment. The clinical outcome of these patients might be predicted by measuring drug sensitivity. METHODS: Pleural effusions containing primary malignant mesothelioma cells were received from the diagnostic routine. We characterized and tested the chemosensitivity of 18 malignant samples and four benign samples from 16 different patients with pleural effusions. Cells were seeded in a 384-well plate for a robotized ex vivo testing of drug sensitivity to 32 different drugs. The primary cells were further characterized by immunocytochemistry to evaluate the proportion of malignant cells and to study the RRM1 and ERCC1 reactivity, two proteins associated with drug resistance. RESULTS: We observed great individual variability in the drug sensitivity. Primary cell isolates were affected by between one and ten drugs, and resistant to the remaining tested drugs. Actinomycin D and daunorubicin were the two drugs effective in most cases. Adjusting efficiency of individual drugs for varying proportion of tumor cells and to the average effect on benign cells correlated with effect of pemetrexed, cisplatin and survival time. General drug sensitivity, proportion of malignant cells and reactivity to RRM1 correlated to each other and to survival time of the patients. CONCLUSIONS: The proportion of malignant cells and RRM1 reactivity in the pleural effusions correlate to drug sensitivity and survival time. The variability in response to the commonly used chemotherapies emphasizes the need for tests that indicate best individual choice of cytotoxic drugs. The efficiency of the obtained results should preferably be corrected for admixture of benign cells and effects of given drugs on benign cells.


Asunto(s)
Antineoplásicos/farmacología , Proteínas de Unión al ADN/metabolismo , Endonucleasas/metabolismo , Neoplasias Pulmonares/tratamiento farmacológico , Mesotelioma/tratamiento farmacológico , Derrame Pleural Maligno/tratamiento farmacológico , Proteínas Supresoras de Tumor/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Pulmonares/patología , Mesotelioma/patología , Mesotelioma Maligno , Derrame Pleural Maligno/patología , Ribonucleósido Difosfato Reductasa , Análisis de Supervivencia , Células Tumorales Cultivadas
2.
BMC Cancer ; 11: 441, 2011 Oct 12.
Artículo en Inglés | MEDLINE | ID: mdl-21992895

RESUMEN

BACKGROUND: Primary effusion lymphoma (PEL) is a rare KSHV/HHV8-associated high-grade non-Hodgkin's lymphoma (NHL) of B-cell origin, characterized by serous effusions in body cavities. Most patients are HIV-infected men with severe immunosuppression and other HHV8-associated diseases such as Kaposi's sarcoma (KS). The prognosis for those infected is poor, with a median survival of less than 6 months in most cohorts. Sustained complete remission is rare. High-dose chemotherapy regimens are used to improve remission rate and survival. The aim of the present study was to compare the drug sensitivity pattern of the available primary effusion (body cavity based) lymphoma-derived cell lines in order to find additional, potentially effective drugs that are not included in current chemotherapy treatment protocols. METHODS: We have analyzed 11 cell lines against 27 frequently used cytostatic drugs in short term (3 days) survival assays using automated high throughput confocal microscopy. RESULTS: All cell lines showed a distinct, individual drug sensitivity pattern. Considering the in vitro used and clinically achieved drug concentration, Vinorelbine, Paclitaxel, Epirubicin and Daunorubicin were the most effective drugs. CONCLUSIONS: We suggest that inclusion of the above drugs into PEL chemotherapy protocols may be justified. The heterogeneity in the drug response pattern however indicated that assay-guided individualized therapy might be required to optimize therapeutic response.


Asunto(s)
Antineoplásicos/farmacología , Resistencia a Antineoplásicos , Herpesvirus Humano 8 , Linfoma de Efusión Primaria/tratamiento farmacológico , Linfoma de Efusión Primaria/virología , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Análisis por Conglomerados , Infecciones por VIH/complicaciones , Infecciones por Herpesviridae/complicaciones , Humanos , Masculino
3.
Mol Cancer Ther ; 6(2): 644-54, 2007 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-17308061

RESUMEN

Tumors are considered to be possible targets of immunotherapy using stimulated and expanded autologous or allogeneic natural killer (NK) cells mismatched for MHC class I molecules and inhibitory NK receptors. NK cell-based immunoadjuvant therapies are carried out in combination with standard chemotherapeutic protocols. In the presented study, we characterized the effect of 28 frequently used chemotherapeutic agents on the capacity of NK cells to kill target cells. We found that treatment of NK cells with the drugs vinblastine, paclitaxel, docetaxel, cladribine, chlorambucil, bortezomib, and MG-132 effectively inhibited NK cell-mediated killing without affecting the viability of NK cells. On the other hand, the following drugs permitted efficient NK cell-mediated killing even at concentrations comparable with or higher than the maximally achieved therapeutic concentration in vivo in humans: asparaginase, bevacizumab, bleomycin, doxorubicin, epirubicin, etoposide, 5-fluorouracil, hydroxyurea, streptozocin, and 6-mercaptopurine.


Asunto(s)
Antineoplásicos/farmacología , Supervivencia Celular/efectos de los fármacos , Citotoxicidad Inmunológica , Células Asesinas Naturales/inmunología , Cromo/metabolismo , Humanos , Células Tumorales Cultivadas/efectos de los fármacos
4.
Leuk Lymphoma ; 48(9): 1835-45, 2007 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-17786721

RESUMEN

Depending on stage and risk factors, up to 30% of patients with advanced Hodgkin lymphoma (HL) progress or relapse. Patients with pleural effusions have a particularly poor prognosis and this stage of HL is regularly resistant to chemotherapy. All currently available HL cell lines are derived from late stage HL patients. In the present study we measured the sensitivity of these HL lines against the 26 most frequently used cytostatic drugs. We used a novel fluorescent short-term survival assay where the cell was incubated with the drugs for 4 days. The precise number of differentially stained live and dead cells was determined using a custom-built automated laser confocal fluorescent microscope. We found that HL cells, independently of their origin, showed very similar sensitivity patterns for several of the drugs. All HL cell lines were highly sensitive to dactinomycin, paclitaxel and etoposide. Our data suggest that the inclusion of dactinomycin and paclitaxel into chemotherapy protocols against late stage Hodgkin lymphoma with pleural effusion may be justified.


Asunto(s)
Antineoplásicos/farmacología , Dactinomicina/farmacología , Enfermedad de Hodgkin/tratamiento farmacológico , Paclitaxel/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Enfermedad de Hodgkin/patología , Humanos
5.
BMC Cancer ; 6: 265, 2006 Nov 13.
Artículo en Inglés | MEDLINE | ID: mdl-17101045

RESUMEN

BACKGROUND: Epstein-Barr virus (EBV) is the causative agent of immunosuppression associated lymphoproliferations such as post-transplant lymphoproliferative disorder (PTLD), AIDS related immunoblastic lymphomas (ARL) and immunoblastic lymphomas in X-linked lymphoproliferative syndrome (XLP). The reported overall mortality for PTLD often exceeds 50%. Reducing the immunosuppression in recipients of solid organ transplants (SOT) or using highly active antiretroviral therapy in AIDS patients leads to complete remission in 23-50% of the PTLD/ARL cases but will not suffice for recipients of bone marrow grafts. An additional therapeutic alternative is the treatment with anti-CD20 antibodies (Rituximab) or EBV-specific cytotoxic T-cells. Chemotherapy is used for the non-responding cases only as the second or third line of treatment. The most frequently used chemotherapy regimens originate from the non-Hodgkin lymphoma protocols and there are no cytotoxic drugs that have been specifically selected against EBV induced lymphoproliferative disorders. METHODS: As lymphoblastoid cell lines (LCLs) are well established in vitro models for PTLD, we have assessed 17 LCLs for cytotoxic drug sensitivity. After three days of incubation, live and dead cells were differentially stained using fluorescent dyes. The precise numbers of live and dead cells were determined using a custom designed automated laser confocal fluorescent microscope. RESULTS: Independently of their origin, LCLs showed very similar drug sensitivity patterns against 29 frequently used cytostatic drugs. LCLs were highly sensitive for vincristine, methotrexate, epirubicin and paclitaxel. CONCLUSION: Our data shows that the inclusion of epirubicin and paclitaxel into chemotherapy protocols against PTLD may be justified.


Asunto(s)
Antineoplásicos/toxicidad , Antivirales/farmacología , Linfocitos B/virología , Transformación Celular Viral , Herpesvirus Humano 4/fisiología , Linfoma/tratamiento farmacológico , Trastornos Linfoproliferativos/tratamiento farmacológico , Linfocitos B/efectos de los fármacos , Trasplante de Médula Ósea/efectos adversos , Línea Celular Tumoral , Herpesvirus Humano 4/efectos de los fármacos , Humanos , Trastornos Linfoproliferativos/inmunología , Complicaciones Posoperatorias/inmunología
6.
Exp Hematol ; 44(1): 38-49.e1, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26325331

RESUMEN

New drugs targeting important cellular signaling pathways are currently being developed for chronic lymphocytic leukemia (CLL). It is therefore of interest to analyze their in vitro killing capacity in manufacturer-independent, comparative experiments. We here report on the sensitivity of CLL cells to a panel of emerging targeted therapeutics using high-throughput screening based on an automated fluorescence digital scanning system. Fresh CLL cells from 42 patients with indolent or progressive CLL were cultured for 72 hours on microtiter plates in a unique primary cell culture medium. Antitumor effects of 31 small therapeutic molecules (and, as controls, 29 cytostatic agents) at equimolar concentration were compared in a fluorescence survival assay. In vitro sensitivity to each drug exhibited considerable interpatient variability. The highest mean direct killing was observed for one survivin inhibitor (YM-155), two bcl-2 inhibitors (ABT-199, ABT-737), and one selective CDK inhibitor (dinaciclib). Their killing capacity was, in contrast to most cytostatic agents, similarly high in refractory versus untreated CLL patients and was significantly higher on cells with the 17p deletion/TP53 mutation than on cells with other cytogenetic abnormalities (p = 0.02). Sensitivity of bone marrow and lymph node cells was highly correlated with that of blood cells. Even though direct killing may not be the only therapeutic effector function in vivo, results from this head-to-head comparison may help to identify drugs of particular interest for intensified clinical development.


Asunto(s)
Antineoplásicos/uso terapéutico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Anciano , Anciano de 80 o más Años , Análisis por Conglomerados , Resistencia a Antineoplásicos , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Técnicas In Vitro , Masculino , Persona de Mediana Edad , Células Tumorales Cultivadas
7.
Genes Cancer ; 6(3-4): 119-128, 2015 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-26000095

RESUMEN

We propose to assess the therapeutic value of biomarker-guided individualized chemotherapy in patients with metastasizing lung adenocarcinoma. In this study, we used primary cells from pleural effusions from sixteen patients diagnosed with adenocarcinomas originating in the lung and from four patients with no malignant diagnosis. The ex vivo drug sensitivity of primary cells was assessed for 32 chemotherapeutical drugs. Linear regression analyses were performed to examine possible correlations between the drug sensitivity, overall survival and expression of ERCC1 and RRM1. The ex vivo drug sensitivity profiles of the patients revealed considerable heterogeneity in drug response. Vinblastine, vinorelbine, paclitaxel and actinomycin D showed high efficiency against 50% of the tested primary cells. Significant correlation was detected between the ex vivo sensitivity to platinum based drugs and gemcitabine and the level of ERCC1 and RRM1. No significant correlation was however seen between overall survival and drug sensitivity. The heterogeneity of the drug response suggests that optimal care of the adenocarcinoma patients should include the determination of drug sensitivity of the primary cells and would benefit to use personalized therapy.

8.
Exp Hematol ; 38(12): 1219-30, 2010 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-20837094

RESUMEN

OBJECTIVE: To generate a comprehensive map of the drug sensitivity of chronic lymphoid leukemia cells (CLL) using a newly developed in vitro drug-sensitivity assay based on automated evaluation of cell viability on single-cell level. MATERIALS AND METHODS: Primary CLL cells from 77 patients were tested using automated digital fluorescence microscopy. The effect of 27 frequently used chemotherapeutic agents was measured in short-term fluorescence survival assay. To avoid typical in vitro artifacts such as growth factor depletion and oxidative damage, the cell were cultured in a novel, total human blood lysate-based medium (OmniSanguine) in order to preserve the composition of growth factor flora and redox conditions of the in vivo environment. RESULTS: CLL cells from different patients showed considerable heterogeneity in their drug-sensitivity patterns. This pattern was stable even after in vitro activation of cell proliferation. Half of the samples were sensitive to fludarabine and chlorambucil. Daunorubicin was the most potent drug. It was effective in 75 of 77 cases. In addition, daunorubicin and prednisolone showed a strong synergistic effect. CONCLUSIONS: We suggest that the combination of low-dose daunorubicin and prednisolone might be an additional treatment option for therapy-resistant cases of CLL.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Ciclo Celular/efectos de los fármacos , Análisis por Conglomerados , Daunorrubicina/administración & dosificación , Doxorrubicina/administración & dosificación , Epirrubicina/administración & dosificación , Humanos , Leucemia Linfocítica Crónica de Células B/patología , Prednisolona/administración & dosificación
9.
Biomed Pharmacother ; 63(6): 413-20, 2009 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-18834693

RESUMEN

Rituximab is a humanized chimeric monoclonal antibody, targeted against the pan B cell marker CD20. It is frequently used to treat a variety of B cell lymphomas and immunosuppression associated lymphoproliferations such as posttransplant lymphoproliferative disorder (PTLD). The response rate of rituximab treatment is 65%, but the exact in vivo mechanism of action is not yet fully understood, although antibody-dependent cell-mediated cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), and direct induction of apoptosis have been suggested as effector mechanism. Rituximab may affect different types of lymphomas through different mechanisms. As lymphoblastoid cell lines (LCLs) are well-established in vitro models of PTLD, we investigated the effect of rituximab on these cells using a custom built automated laser confocal fluorescent microscope. We found that rituximab alone was not effective at inducing cell death of EBV-transformed B cells. The antibody was effective in the complement-mediated CDC. Rituximab could induce NK cell-mediated ADCC but it was more effective in the presence of untreated fresh human plasma compared to heat-inactivated human plasma. Our data suggest that complement-enhanced NK-mediated ADCC is required for effective rituximab mediated killing of EBV-transformed B cells. Determining and monitoring of serum complement levels and in vitro killing efficacy of NK cells of PTLD patients might help to predict resistant cases to rituximab therapy. On the other hand our results suggest a possibility that rituximab should be combined only with cytotoxic drugs that spare NK function when treating PTLD patients.


Asunto(s)
Anticuerpos Monoclonales/farmacología , Antineoplásicos/farmacología , Linfocitos B/efectos de los fármacos , Células Asesinas Naturales/metabolismo , Anticuerpos Monoclonales de Origen Murino , Linfocitos B/inmunología , Linfocitos B/virología , Línea Celular Transformada , Línea Celular Tumoral , Transformación Celular Viral , Infecciones por Virus de Epstein-Barr/inmunología , Calor , Humanos , Trastornos Linfoproliferativos/tratamiento farmacológico , Microscopía Confocal/métodos , Plasma/metabolismo , Rituximab
10.
J Immunother ; 31(3): 283-93, 2008 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-18317359

RESUMEN

Tumors are considered to be possible targets of immunotherapy using stimulated and expanded cytotoxic T-lymphocytes (CTL). It is important to consider the drug-induced effects when chemotherapeutic regimens and CTL-mediated immunotherapy is planned to be used in parallel. In this study, we characterized the effect of 29 frequently used chemotherapeutic agents on the cytotoxic activity of autologous and allogeneic CTLs. We found that treatment of CTLs with the following drugs: docetaxel, vincristine, chlorambucil, mitomycin C, oxaliplatin, doxorubicin, and bleomycin effectively inhibited CTL-mediated killing, without affecting their viability. On the other hand, the following drugs enhanced or permitted efficient CTL-mediated killing in vitro at concentrations comparable with the maximally achieved therapeutic concentration in vivo in humans: daunorubicin, prednisolone, vinorelbine, cisplatin, methotrexate, hydroxyurea, cytarabine, cyclophosphamide, topotecan, epirubicin, fluorouracil, carboplatin, asparaginase, 6-mercaptopurine, and bortezomib. Our results could potentially be used in the future to design new CTL-based adjuvant immunotherapy protocols.


Asunto(s)
Antineoplásicos/farmacología , Citotoxicidad Inmunológica/efectos de los fármacos , Linfocitos T Citotóxicos/inmunología , Antineoplásicos/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica , Línea Celular Transformada , Supervivencia Celular/efectos de los fármacos , Pruebas Inmunológicas de Citotoxicidad , Humanos , Inmunoterapia Adoptiva , Microscopía Confocal , Neoplasias/tratamiento farmacológico , Neoplasias/inmunología , Linfocitos T Citotóxicos/efectos de los fármacos
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