| 1. |
- Lindhagen, Elin, et al.
(författare)
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Significant cytotoxic activity in vitro of the EGFR tyrosine kinase inhibitor gefitinib in acute myeloblastic leukaemia.
- 2008
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Ingår i: Eur J Haematol. - : Wiley. - 1600-0609 .- 0902-4441. ; 81:5, s. 344-353
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVES:Gefitinib inhibits epidermal growth factor receptor (EGFR) signalling, but may also act by non-EGFR dependent mechanisms. We have investigated the activity of gefitinib in haematological tumour cells, in particular acute myeloblastic leukaemia (AML).METHODS:Cytotoxic activity of gefitinib, alone or in combination with standard anti-leukaemic drugs, was assessed by the short-term fluorometric microculture cytotoxicity assay in tumour cells from 117 patients representing five haematological and five non-haematological malignancies. In AML, the EGFR status was analysed by immunochemistry. Gefitinib-induced apoptosis was investigated in a subset of AML samples, as well as in the leukaemia cell line MV-4-11, using a multiparametric high content screening assay. To confirm activation of caspase-3 in cells treated with gefitinib, a blocking test was carried out in which MV4-11 cells were pretreated with the specific caspase inhibitor DEVD-FMK.RESULTS:Gefitinib showed highest cytotoxic activity in AML (n = 19) with many samples being sensitive at concentrations achievable in clinical practice (<10 microM), and no difference between previously untreated and relapsed patients. No correlation between the activity of gefitinib and standard antileukaemic drugs (cytarabine, doxorubicin, etoposide) was observed. Combining gefitinib with these drugs resulted in mainly additive or synergistic (etoposide) effects, with no evidence of sequence dependency. The AML cells did not express the EGFR. Gefitinib induced apoptosis, which was at least partly mediated by activation of the caspase-3 pathway.CONCLUSION:In vitro, gefitinib has significant cytotoxic activity in AML by inducing apoptosis through non-EGFR dependent pathways.
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| 2. |
- Lindhagen, Elin, et al.
(författare)
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The fluorometric microculture cytotoxicity assay
- 2008
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Ingår i: Nature Protocols. - : Springer Science and Business Media LLC. - 1754-2189 .- 1750-2799. ; 3:8, s. 1364-1369
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Tidskriftsartikel (refereegranskat)abstract
- The fluorometric microculture cytotoxicity assay (FMCA) is a nonclonogenic microplate-based cell viability assay used for measurement of the cytotoxic and/or cytostatic effect of different compounds in vitro. The assay is based on hydrolysis of the probe, fluorescein diacetate (FDA) by esterases in cells with intact plasma membranes. The assay is available as both a semiautomated 96-well plate setup and a 384-well plate version fully adaptable to robotics. Experimental plates are prepared with a small amount of drug solution and can be stored frozen. Cells are seeded on the plates and cell viability is evaluated after 72 h. The protocol described here is applicable both for cell lines and freshly prepared tumor cells from patients and is suitable both for screening in drug development and as a basis for a predictive test for individualization of anticancer drug therapy.
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| 3. |
- Mahteme, Haile, et al.
(författare)
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Heterogeneous activity of cytotoxic drugs in patient samples of peritoneal carcinomatosis
- 2008
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Ingår i: European Journal of Surgical Oncology. - : Elsevier BV. - 0748-7983 .- 1532-2157. ; 34:5, s. 547-552
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Tidskriftsartikel (refereegranskat)abstract
- AIMS: To investigate if the pattern of cytotoxic drug sensitivity in vitro in patient samples of peritoneal carcinomatosis (PC) is supportive to the current standardized approach for drug selection for perioperative intraperitoneal chemotherapy (IPC). METHODS: The cytotoxic effect of cisplatin, oxaliplatin, irinotecan, 5-fluorouracil, mitomycin-C, doxorubicin and melphalan was investigated in vitro on tumour cells from 223 patient tumour samples of different PC origins. RESULTS: Considerable differences in cytotoxic drug sensitivity between tumour types of the PC entity and within each tumour type were observed. Cisplatin showed high cross-resistance with oxaliplatin but low cross-resistance with doxorubicin and irinotecan. No cross-resistance was found between irinotecan and doxorubicin. The dose-response relationships for melphalan and irinotecan in individual samples showed great variability. CONCLUSIONS: The activity in vitro of cytotoxic drugs commonly used in IPC for PC is very heterogeneous. Efforts for individualizing drug selection for PC patients undergoing IPC seem justified.
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| 4. |
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| 5. |
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| 6. |
- Nygren, Yvonne, 1965-, et al.
(författare)
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Hydrophilic interaction liquid chromatography (HILIC) coupled to inductively coupled plasma mass spectrometry (ICPMS) utilizing a mobile phase with a low-volatile organic modifier for the determination of cisplatin, and its monohydrolyzed metabolite
- 2008
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Ingår i: Journal of Analytical Atomic Spectrometry. - : Royal Society of Chemistry. - 0267-9477 .- 1364-5544. ; 23:7, s. 948-954
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Tidskriftsartikel (refereegranskat)abstract
- This study demonstrates the on-line coupling of hydrophilic interaction liquid chromatography (HILIC) to inductively coupled plasma mass spectrometry (ICPMS). A low volatile organic solvent, dimethylformamide (DMF), was used as organic modifier of the mobile phase to minimize the solvent loading of the ICP and thereby improve analyte sensitivity and method robustness. The concept was illustrated by the selective determination of cisplatin (cis-DDP), and its mono-hydrolyzed metabolite (MH-DDP). Species were separated on a 2.1 × 150 mm ZIC-HILIC column and detected on-line by selective platinum (m/z 194 and 195) monitoring, giving a detection limit of 2 pg Pt (0.2 ng ml-1). Compared to the use of the conventional solvent acetonitrile (AcN), with DMF, cis-DDP sensitivity was enhanced as much as 36 times and no addition of oxygen to the plasma was needed to avoid carbon depositions on instrumental parts. Furthermore, several non-identified platinum containing compounds were observed when using AcN as a result of unwanted reactions between this solvent and the analytes. No such species were observed when DMF was used. The molecular structures of eluting compounds were verified by electrospray ionization mass spectrometry. Combined with a simple sample treatment protocol, the HILIC-ICPMS system allowed determination of free intracellular cis-DDP in in-vitro grown T289 human malignant melanoma cells up to 60 min after exposure to 50 g ml-1 cis-DDP for 1 h. This work shows that HILIC-ICPMS is a potent hyphenated technique for the analysis of hydrophilic metal compounds and that the use of chromatographic mobile phases with low-volatile organic solvents may be a generic approach to improve analyte sensitivity and system robustness of HPLC-ICPMS when mobile phases with high amount of organic solvent are used.
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| 7. |
- Rickardson, Linda, 1980-
(författare)
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New Methods to Screen for Cancer Drugs and to Evaluate their Mechanism of Action
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cancer is a common disease and due to problems with resistance against cancer drugs and the limited benefit from chemotherapy in many diagnoses, there is a need to develop new cancer drugs. In this thesis new methods to screen for cancer drugs and to evaluate their mechanism of action are discussed. In Paper I, it was found that by studying the gene expression of a cell line panel and combining the data with sensitivity data of a number of cytotoxic drugs, it was possible to cluster compounds according to mechanism of action as well as identifying genes associated with chemosensitivity. In Paper II, studies of compounds with selective activity in drug-resistant cell lines revealed the glucocorticoids as a group of interesting compounds. The glucocorticoid receptor was overexpressed in 8226/Dox40 and the difference in sensitivity was abolished when the cells were treated with a glucocorticoid receptor antagonist. In Paper III, an image-based screening method for new proteasome inhibitors was successfully developed and the compounds disulfiram, PDTC and NSC 95397 were identified as inhibitors of the proteasome. In Paper IV, disulfiram and PDTC were shown to induce cytotoxic activity, to inhibit the activation of the transcription factor NFkappaB and to inhibit the degradation of proteins normally degraded by the proteasome. In Paper V, NSC 95397 was shown to be cytotoxic to all cells in the resistance-based cell line panel as well as to patient samples from a variety of cancer diagnoses. Connectivity Map was successfully used as a tool to propose a new mechanism of action of NSC 95397. The gene expression induced by NSC 95397-treatment was similar to that induced by several proteasome inhibitors not present in the Connectivity Map.
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| 8. |
- Wickström, Malin, et al.
(författare)
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The novel alkylating prodrug J1: diagnosis directed activity profile ex vivo and combination analyses in vitro
- 2008
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Ingår i: Investigational new drugs. - : Springer Science and Business Media LLC. - 0167-6997 .- 1573-0646. ; 26:3, s. 195-204
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Tidskriftsartikel (refereegranskat)abstract
- Objective: The dipeptide J1 acts as a prodrug of melphalan with a significant increased potency in vitro resulting from activation by cellular aminopeptidases. The current study was performed to evaluate the ex vivo profile of J1 using 176 primary tumor cell cultures from patients. In addition, the activity of J1 in combination with eight standard drugs, representing different mechanistic classes, was studied in nine different human tumor cell lines of different histopathological origin. Methods: Ex vivo evaluation of tumor type selectivity, was performed using the established fluorometric microculture cytotoxicity assay (FMCA). Combinations between J1 and eight standard chemotherapeutic drugs were analyzed using the median-effect method. Results: The prodrug J1 expressed approximately 50- to 100-fold higher potency but similar activity profile as that of its metabolite, melphalan. The difference was greater in some diagnoses (e.g. breast cancer, NHL and AML), and exceptionally high in some breast cancer samples with aggressive phenotypes. Combination analysis of J1 and standard chemotherapeutics yielded several potentially additive and synergistic interactions, most striking for etoposide with significant synergism in all studied cell lines. Conclusions: In conclusion, the ex vivo profile suggests that further evaluation of J1 as the alkylating agent in for example aggressive breast cancer might be of particular interest, preferentially in combination with DNA-topoisomerase II inhibitors like etoposide.
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| 9. |
- Åleskog, Anna, et al.
(författare)
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Rapamycin shows anticancer activity in primary chronic lymphocytic leukemia cells in vitro, as single agent and in drug combination
- 2008
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Ingår i: Leukemia and Lymphoma. - : Informa UK Limited. - 1042-8194 .- 1029-2403. ; 49:12, s. 2333-2343
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Tidskriftsartikel (refereegranskat)abstract
- The mammalian target of rapamycin inhibitor rapamycin and its analogues show promising anticancer activity in various experimental tumor models and are presently evaluated in clinical trials. We, here, evaluated the in vitro activity of rapamycin with regard to tumor-type specificity and possible mechanisms of drug resistance in 97 tumor cell samples from patients and in a resistance-based cell line panel, using the fluorometric microculture cytotoxicity assay. Rapamycin was dose-dependently cytotoxic in patient tumor cells and in cell lines. In primary cells, rapamycin was more active in hematological than in solid tumor samples, with chronic lymphocytic leukemia (CLL) and acute lymphocytic leukemia being the most sensitive tumor types. Considerable inter-individual differences in sensitivity were apparent among CLL samples, but no difference was observed between IGHV mutated and unmutated CLL samples, whereas a tendency to lower rapamycin sensitivity was indicated for samples displaying poor-prognostic genomic markers. Combination experiments in CLL cells indicated that rapamycin acted synergistically with vincristine, cisplatin, chlorambucil and taxotere. These results and the clinically-experienced good tolerance to rapamycin analogues encourage clinical studies of rapamycin in CLL treatment as single agent but also in combination with, e.g., vincristine and chlorambucil.
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