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1.
Leukemia ; 18(7): 1252-7, 2004 Jul.
Article in English | MEDLINE | ID: mdl-15208643

ABSTRACT

Data on breast cancer resistance protein (BCRP, MXR, ABCG2) expression in acute myeloid leukemia (AML) have been inconsistent, possibly due to use of different assays in different studies. BCRP mRNA was studied by the reverse-transcription polymerase chain reaction and BCRP protein expression (BXP-21, BXP-34 or anti-ABCG2 antibody, with anti-CD34 and anti-CD33) and function (fumitremorgin C modulation of mitoxantrone retention) by flow cytometry in eight cell lines and in pretreatment blasts from 31 AML patients. BCRP mRNA levels, antibody staining and function correlated strongly in cell lines (Pearson r values, 0.73-0.97), but not in AML samples. AML sample BCRP mRNA levels were between those in parental 8226 and 35-fold mitoxantrone-resistant 8226/MR20 cells in all but one case, and BCRP mRNA had the wild-type sequence at codon 482 in all. In AML, unlike in cell lines, BCRP protein expression or function, when present, was only detected in small subpopulations. BCRP mRNA and protein expression did not correlate, nor did staining with different BCRP antibodies, and function did not correlate with mRNA nor protein expression. Presence of BCRP only in subpopulations and discordance among BCRP measurements suggest complex biology of BCRP in AML and incomplete modeling by cell lines.


Subject(s)
ATP-Binding Cassette Transporters/analysis , ATP-Binding Cassette Transporters/physiology , Leukemia, Myeloid/pathology , Neoplasm Proteins/analysis , Neoplasm Proteins/physiology , ATP Binding Cassette Transporter, Subfamily G, Member 2 , ATP-Binding Cassette Transporters/genetics , Acute Disease , Bone Marrow/pathology , Cell Line, Tumor , Drug Resistance, Multiple , Female , Flow Cytometry , Humans , Leukemia, Myeloid/metabolism , Mutation , Neoplasm Proteins/genetics , RNA, Messenger/analysis , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction
2.
J Natl Cancer Inst ; 93(16): 1234-45, 2001 Aug 15.
Article in English | MEDLINE | ID: mdl-11504769

ABSTRACT

BACKGROUND: The taxane paclitaxel (Taxol) is often of limited efficacy in chemotherapeutic regimens because some cancer cells express high levels of the efflux pump, P-glycoprotein (Pgp), which removes the drug from the cells. The orally active paclitaxel analog IDN-5109 has been reported to overcome Pgp-mediated drug resistance. We tested whether IDN-5109 acts by modulating Pgp activity. METHODS: Human MDA435/LCC6mdr1 and MDA435/LCC6 breast carcinoma cells, which express and do not express Pgp, respectively, were incubated with [3H]IDN-5109 and paclitaxel to determine intracellular drug accumulation. Flow cytometry was used to analyze intracellular retention of two Pgp substrates, rhodamine 123 (Rh-123) and doxorubicin, in both breast carcinoma cell lines and in human colon carcinoma cells (SW-620, DLD1, and HCT-15, whose Pgp levels vary) treated with different taxanes. The effects of IDN-5109 and paclitaxel on tumor growth in vivo were studied with the use of tumors established through xenografts of Pgp-expressing SW-620 and DLD1 cells in severe combined immunodeficiency mice. All statistical tests were two-sided. RESULTS: Pgp-expressing cells treated with IDN-5109 or with the taxane-based drug resistance reversal agent tRA96023, which blocks Pgp activity, retained 8.1- and 9.4-fold more Rh-123 (P =.0001), respectively, and 1.7- and 1.9-fold more doxorubicin (P =.001), respectively, than cells treated with paclitaxel. Non-Pgp-expressing cells treated similarly demonstrated no increased retention of either substrate. MDA435/LCC6mdr1 cells retained 5.3-fold more [3H]IDN-5109 than [3H]paclitaxel after 2 hours (P =.01). IDN-5109 showed statistically significantly higher tumor growth inhibition than paclitaxel against the SW-620 xenograft (P =.003). CONCLUSIONS: IDN-5109 modulates Pgp activity, resulting in superior tumor growth inhibition against Pgp-expressing tumors as compared with paclitaxel. IDN-5109 may broaden the spectrum of taxane use to include colon tumors.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Antineoplastic Agents, Phytogenic/pharmacology , Breast Neoplasms/drug therapy , Bridged-Ring Compounds/pharmacology , Colonic Neoplasms/drug therapy , Paclitaxel/analogs & derivatives , Paclitaxel/pharmacology , Taxoids , ATP Binding Cassette Transporter, Subfamily B, Member 1/drug effects , Administration, Oral , Animals , Antibiotics, Antineoplastic , Antineoplastic Agents, Phytogenic/pharmacokinetics , Breast Neoplasms/metabolism , Bridged-Ring Compounds/administration & dosage , Colonic Neoplasms/metabolism , Doxorubicin/pharmacokinetics , Drug Screening Assays, Antitumor/methods , Female , Flow Cytometry , Fluorescence , Humans , Male , Paclitaxel/administration & dosage , Paclitaxel/pharmacokinetics , Tissue Distribution , Transplantation, Heterologous , Tumor Cells, Cultured
3.
Acad Radiol ; 8(5): 405-8, 2001 May.
Article in English | MEDLINE | ID: mdl-11345271

ABSTRACT

RATIONALE AND OBJECTIVES: The authors investigated the use of magnetic resonance (MR) imaging of the brain in adult patients with a primary complaint of chronic headache and no other neurologic symptoms or findings and determined the yield and MR predictors of major abnormalities in these patients. MATERIALS AND METHODS: The medical records and MR images of 402 adult patients with chronic headache were retrospectively reviewed. All patients had been evaluated and referred by the neurology service. The findings were categorized as either negative or positive for major abnormality. Multivariate analysis with a linear logistic regression technique was performed on the clinical data, which included patient age, patient sex, and headache type. RESULTS: Major abnormalities were found in 15 patients (3.7%), consisting of seven women (2.4%) and eight men (6.9%). Major abnormalities were found in 0.6% of those with migraine headaches, 1.4% with tension headaches, none with mixed migraine and tension headaches, 14.1% with atypical headaches, and 3.8% with other types of headaches. Multivariate analysis showed that the atypical headache type was the most significant predictor of major abnormality. CONCLUSION: The yield of major abnormalities found with brain MR imaging in patients with isolated chronic headache is low. However, those patients with atypical headaches have a higher yield of major abnormalities and may benefit from imaging.


Subject(s)
Brain/pathology , Headache Disorders/diagnosis , Magnetic Resonance Imaging , Adult , Aged , Aged, 80 and over , Brain Neoplasms/diagnosis , Female , Humans , Logistic Models , Male , Middle Aged , Retrospective Studies
4.
Cytometry ; 41(4): 252-60, 2000 Dec 01.
Article in English | MEDLINE | ID: mdl-11084610

ABSTRACT

BACKGROUND: Cellular heterogeneity in drug response has important clinical implications, and is believed to develop over many generations during clonal evolution in human tumors. The purpose of this study was to determine the level of heterogeneity exhibited by sister cells soon after their birth. METHODS: Human ileocecal carcinoma cells (HCT-8) were followed up to 11 days in vitro after a 2-h exposure to 1 microM raltitrexed (IC(95)) in a time-lapse video system. RESULTS: Over five experiments, 414 cells were followed after exposure to raltitrexed. Immediate sterility occurred in 74% of treated cells. Only 6% of cells could produce more than two generations of offspring, and heterogeneity in drug response was seen. Comparing sister cells < 24 h old, the more proliferative sibling produced up to 73 times more offspring, with a median ratio of 9.0 (control median = 1.19). Offspring of prolific drug-treated cells had a decreased probability of division (68% compared with 92%) and an increased average interdivision time (19.0 h compared with 15.1 h). CONCLUSIONS: Short-term exposure to raltitrexed resulted in increased interdivision times and production of sterile offspring extending seven generations. Cellular heterogeneity (difference in proliferation potential comparing drug-treated sister cells) was evident without a period of clonal evolution.


Subject(s)
Antimetabolites, Antineoplastic/toxicity , Ileal Neoplasms/drug therapy , Ileal Neoplasms/pathology , Microscopy, Video/methods , Quinazolines/toxicity , Thiophenes/toxicity , Aged , Cell Division/drug effects , Cell Lineage/drug effects , Clone Cells/cytology , Clone Cells/drug effects , Humans , Male , Probability , Tumor Cells, Cultured
5.
Cancer Res ; 60(3): 525-9, 2000 Feb 01.
Article in English | MEDLINE | ID: mdl-10676629

ABSTRACT

At high fluence rates in animal models, photodynamic therapy (PDT) can photochemically deplete ambient tumor oxygen through the generation of singlet oxygen, causing acute hypoxia and limiting treatment effectiveness. We report that standard clinical treatment conditions (1 mg/kg Photofrin, light at 630 nm and 150 mW/cm2), which are highly effective for treating human basal cell carcinomas, significantly diminished tumor oxygen levels during initial light delivery in a majority of carcinomas. Oxygen depletion could be found during at least 40% of the total light dose, but tumors appeared well oxygenated toward the end of treatment. In contrast, initial light delivery at a lower fluence rate of 30 mW/cm2 increased tumor oxygenation in a majority of carcinomas. Laser treatment caused an intensity- and treatment time-dependent increase in tumor temperature. The data suggest that high fluence rate treatment, although effective, may be inefficient.


Subject(s)
Antineoplastic Agents/therapeutic use , Carcinoma, Basal Cell/drug therapy , Dihematoporphyrin Ether/therapeutic use , Hematoporphyrin Photoradiation , Oxygen/metabolism , Carcinoma, Basal Cell/metabolism , Humans
6.
Ann N Y Acad Sci ; 922: 293-7, 2000.
Article in English | MEDLINE | ID: mdl-11193904

ABSTRACT

Preclinical studies at Roswell Park Cancer Institute by Minderman, Cao, and Rustum (unpublished results) showed that a combination of SN-38 and 5-FU against HCT-8 human colon carcinoma cells in vitro was synergistic, with the best interaction occurring when the drugs were added sequentially, SN-38 first. Their in vivo studies using HCT-8 tumor xenografts implanted s.c. in nude athymic mice demonstrated superior efficacy for a sequential i.v. administration of CPT-11, 24 hr before 5-FU. On the basis of these studies, our group has begun to evaluate effects of RFS2000 (9-nitro-20(S)-camptothecin) (9-NC) in combination with a series of other antitumor agents. Using a panel of human tumor cell lines including A121 ovarian cancer, HCT-8 colon cancer, H-460 NSCLC, HT-1080 fibrosarcoma, and MCF7 mammary cancer, we found that a 2-hr exposure to 9-NC resulted in ID50 values of < 1.0 microM, whereas continuous exposure to drug resulted in ID50 values of < 1.0 nM. Tumor growth inhibitory activities of 5-FU, gemcitabine, and paclitaxel were determined for comparison. Combinations of these agents were evaluated with 9-NC using the human HCT-8 colon tumor cell line. Concurrent and sequential combinations of 9-NC with 5-FU had some regions of the concentration-effect surface with local synergy and some with local antagonism. However, sequential combination of 9NC or SN-38 followed by 5-FU, 24 hr later appeared to be highly synergistic at high dose-effect levels (i.e., ID90), suggesting that sequential drug administration may be more efficacious at high effect level and that the order of drug addition is very important. Overall, our results were similar to that found earlier by Rustum's group with CPT11 (or SN-38) and 5-FU, suggesting that sequential combination of 9-NC (or other camptothecin analogues) followed by 5-FU has potential for the treatment of cancer in man.


Subject(s)
Antineoplastic Agents/pharmacology , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Camptothecin/pharmacology , Camptothecin/administration & dosage , Camptothecin/analogs & derivatives , Cell Division/drug effects , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Fluorouracil/administration & dosage , Humans , Tumor Cells, Cultured/drug effects
7.
In Vitro Cell Dev Biol Anim ; 35(5): 270-8, 1999 May.
Article in English | MEDLINE | ID: mdl-10475273

ABSTRACT

In preparing for the routine use of the ubiquitous in vitro cell growth inhibition assay for the study of anticancer agents, we characterized the statistical properties of the assay and found some surprising results. Parabolic well-to-well cell growth patterns were discovered, which could profoundly affect the results of routine growth inhibition studies of anticancer and other agents. Four human ovarian cell lines, A2780/WT, A2780/DX5, A2780/DX5B, and A121, and one human ileocecal adenocarcinoma cell line, HCT-8, were seeded into plastic 96-well plates with a 12-channel pipette, without drugs, and grown from 1-5 d. The wells were washed with a plate washer, cells stained with sulforhodamine B (SRB), and dye absorbance measured with a plate reader. Variance models were fit to the data from replicates to determine the nature of the heteroscedastic error structure. Exponential growth models were fit to data to estimate doubling times for each cell line. Polynomial models were fit to data from 10-plate stacks of 96-well plates to explore nonuniformity of cell growth in wells in different regions of the stacks. Each separate step in the assay was examined for precision, patterns, and underlying causes of variation. Differential evaporation of water from wells is likely a major, but not exclusive, contributor to the systematic well-to-well cell growth patterns. Because the fundamental underlying causes of the parabolic growth patterns were not conclusively found, a randomization step for the growth assay was developed.


Subject(s)
Cell Division , Drug Screening Assays, Antitumor , Adenocarcinoma/pathology , Cecal Neoplasms/pathology , Colonic Neoplasms/pathology , Female , Humans , Ileal Neoplasms/pathology , Kinetics , Ovarian Neoplasms/pathology , Regression Analysis , Rhodamines , Sensitivity and Specificity , Staining and Labeling , Tumor Cells, Cultured
8.
Biochem Pharmacol ; 57(5): 567-77, 1999 Mar 01.
Article in English | MEDLINE | ID: mdl-9952321

ABSTRACT

Folic acid (PteGlu)-enhanced intense synergy has been observed between nonpolyglutamylatable dihydrofolate reductase (DHFR) inhibitors and polyglutamylatable inhibitors of other folate-requiring enzymes, such as glycinamide ribonucleotide formyltransferase (GARFT) and thymidylate synthase. Since this phenomenon is potentially therapeutically useful, we explored its universality by examining the combined action of a DHFR inhibitor, trimetrexate (TMQ), with a GARFT inhibitor, 4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimidino[5,4,6][1,4]++ +thiazin-6-yl)-(S)-ethyl]-2,5-thienoylamino-L-glutamic acid (AG2034), in eight human cultured cell lines. Using a 96-well plate cell growth inhibition assay, four ileocecal adenocarcinoma cell lines [HCT-8, HCT-8/DW2 (Tomudex-resistant), HCT-8/DF2 (Tomudex-/FdUrd-resistant), and HCT-8/50 (adapted to 50 nM PteGlu)], three head and neck carcinoma cell lines [A253, FaDu, and Hep-2/500 (FdUrd-resistant)], and a non-small cell lung carcinoma cell line [H460] were treated for 96 hr with TMQ + AG2034 in the presence of 23 or 40 microM PteGlu. Cell growth was measured with the sulforhodamine B assay at the end of this period. Drug interactions were assessed by fitting a 7-parameter model including a synergism parameter, alpha, to data with weighted nonlinear regression. Isobologram analysis was also applied. At 23 microM PteGlu, cells exhibited similar intensities of Loewe synergy for the combination of TMQ + AG2034. Loewe synergy was abolished in HCT-8/50 cells cultured and studied in 50 nM PteGlu. At 40 microM PteGlu, the intensity of the combined action in all cell lines was increased However, the most intense Loewe synergy was seen with HCT-8, HCT-8/DF2, H460, FaDu, A253, and Hep-2/500 cells, whereas the HCT-8/50 subculture showed less of the phenomenon, and PteGlu enhancement was the least with HCT-8/DW2, a subline deficient in folylpolyglutamate synthetase (FPGS). The universality of the PteGlu-enhanced intense synergy phenomenon is suggested. Impaired FPGS activity and low-folate adaptation prior to treatment significantly lessen the degree of PteGlu enhancement.


Subject(s)
Antineoplastic Agents/toxicity , Folic Acid Antagonists/toxicity , Folic Acid/pharmacology , Glutamates/toxicity , Hydroxymethyl and Formyl Transferases/antagonists & inhibitors , Pyrimidines/toxicity , Trimetrexate/toxicity , Adenocarcinoma , Carcinoma, Squamous Cell , Cell Division/drug effects , Colonic Neoplasms , Drug Resistance, Multiple , Drug Synergism , Head and Neck Neoplasms , Humans , Kinetics , Lung Neoplasms , Phosphoribosylglycinamide Formyltransferase , Tumor Cells, Cultured
9.
Cancer Res ; 58(24): 5749-61, 1998 Dec 15.
Article in English | MEDLINE | ID: mdl-9865733

ABSTRACT

For potential clinical extrapolation of in vitro findings, it is of interest to relate the measured effect of an anticancer agent to concentration and exposure time. The Hill model (A. V. Hill, J. Physiol., 40: iv-vii, 1910) is commonly used to describe pharmacodynamic (PD) effects, including drug-induced growth inhibition of cancer cells in vitro. The IC(X)n x T = k relationship, in which IC(X) is the concentration of agent required to reduce cell growth by X%, T is the exposure time, and n and k are estimable parameters, was first applied to bacterial disinfectant action and then was successfully used to model anticancer drug potency as a function of exposure time (D. J. Adams, Cancer Res., 49: 6615-6620, 1989). Our goal was to create a new global PD modeling paradigm to facilitate the quantitative assessment of the growth-inhibitory effect of anticancer agents as a function of concentration and exposure time. Wild-type human ovarian A2780 and ileocecal HCT-8 carcinoma cells and sublines that were resistant to cisplatin (A2780/CP3), doxorubicin (A2780/DX5B), and raltitrexed (RTX) (HCT-8/DW2) were exposed to various anticancer agents, cisplatin, doxorubicin, paclitaxel, trimetrexate, RTX, methotrexate, and AG2034, for periods ranging from 1 to 96 h. Cell growth inhibition was measured with the sulforhodamine B protein dye assay. Patterns of time-dependency of drug potency, slope of the concentration-effect curves, and relative degree of resistance were characterized. Empirical mathematical expressions were built into a global concentration-time-effect model. The global PD model was then fit to the concentration-time-effect data with iteratively reweighted nonlinear regression. Under specific treatment conditions, the examination of the slope and the shape of the concentration-effect curves revealed a large heterogeneity in drug response, e.g., shallow concentration-effect curve or double or triple Hill "roller coaster" concentration-effect curve. These patterns, which were observed at intermediate exposure times in parental and resistant cells for paclitaxel and trimetrexate or only in resistant HCT-8/DW2 cells for RTX, methotrexate, and AG2034, revealed mechanistic insights for the former cases but possible methodological artifacts for the latter cases. The comprehensive PD modeling of the cytotoxic effect of anticancer agents showed that it was possible to modulate drug effect, response heterogeneity, and drug resistance by altering the time of exposure to the agents. This approach will be useful for: (a) describing complex concentration-time-effect surfaces; (b) refining biological interpretations of data; (c) providing insights on mechanisms of drug action and resistance; and (d) generating leads for clinical use of anticancer drugs.


Subject(s)
Antineoplastic Agents/pharmacology , Drug Resistance, Neoplasm , Models, Biological , Antineoplastic Agents/administration & dosage , Cell Division/drug effects , Dose-Response Relationship, Drug , Statistics as Topic , Time Factors
10.
Pharm Res ; 15(7): 1069-76, 1998 Jul.
Article in English | MEDLINE | ID: mdl-9688062

ABSTRACT

PURPOSE: A new mathematical approach was developed to quantify convulsant interaction between pefloxacin and theophylline in rats. METHODS: Animals received each compound separately or in different combination ratios. Infusion was stopped at the onset of maximal seizures. Cerebrospinal fluid (CSF) and plasma samples were collected for HPLC drug determination. The nature and intensity of the pharmacodynamic (PD) interaction between drugs was assessed with a new modeling approach which includes (a) data transformation to create an essentially error-free X-variable and (b) estimation of an interaction parameter a by fitting a nonlinear hyperbolic model to the combination data with unweighted nonlinear regression. RESULTS: Drug disposition to the biophase was linear within the range of administered doses. The estimates of a suggested a Loewe antagonistic interaction between pefloxacin and theophylline at the induction of maximal seizures in rats. Similar intensity of PD interaction was observed at the dose and biophase level (alpha was -0.415 +/- 0.069 and -0.567 +/- 0.079, respectively). CONCLUSIONS: The suitability of the proposed model was assessed by Monte Carlo simulation. This new mathematical approach enabled the characterization of the Loewe antagonistic nature of the PD (convulsant) interaction between pefloxacin and theophylline, whereas previously used methodologies failed to do so.


Subject(s)
Anti-Infective Agents/pharmacokinetics , Anti-Infective Agents/toxicity , Models, Biological , Pefloxacin/pharmacokinetics , Pefloxacin/toxicity , Seizures/chemically induced , Theophylline/pharmacokinetics , Theophylline/toxicity , Vasodilator Agents/pharmacokinetics , Vasodilator Agents/toxicity , Animals , Drug Interactions , Male , Mathematical Computing , Monte Carlo Method , Rats , Rats, Sprague-Dawley
11.
Cancer Res ; 58(14): 3036-50, 1998 Jul 15.
Article in English | MEDLINE | ID: mdl-9679969

ABSTRACT

The combined action among polyglutamylatable and nonpolyglutamylatable antifolates, directed against dihydrofolate reductase (DHFR), glycinamide ribonucleotide formyltransferase (GARFT), 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase (AICARFT), and thymidylate synthase (TS), in human ileocecal HCT-8 cells was examined in a 96-well plate growth inhibition assay (96-h continuous drug exposure). An interaction parameter, alpha, was estimated for each of 95 experiments by fitting a seven-parameter model to data with weighted nonlinear regression. In a representative experiment, raising the folic acid concentration in the medium dramatically increased the Loewe synergy for the combination of trimetrexate (TMTX) and the GARFT inhibitor AG2034 (from a mean alpha +/- SE of 1.50 +/- 0.25 at 2.3 microM folic acid to 146 +/- 20 at 78 microM folic acid). Enhancements were also found for combinations of TMTX with the GARFT inhibitors AG2032, Lometrexol, and LY309887, the AICARFT inhibitor AG2009, and the TS inhibitors LY231514 and Tomudex but not with the GARFT inhibitor LL95509 or with the TS inhibitors AG337, ZD9331, and BW1843U89. Replacing TMTX with methotrexate in two-drug mixtures decreased the intensity of Loewe synergy. Examination of isobolograms at different effect levels revealed informative reproducible changes in isobol patterns. No two-drug combinations among inhibitors of GARFT, AICARFT, and TS exhibited Loewe synergy at either 2.3 or 78 microM folic acid. Thus, the ideal requirement for the folic acid-enhanced synergy is that a nonpolyglutamylatable DHFR inhibitor be combined with a polyglutamylatable inhibitor of another folate-requiring enzyme. A hypothesis to explain this general phenomenon involves the critical role of folylpoly-gamma-glutamate synthetase and the effect of the DHFR inhibitor in decreasing the protection by folic acid of cells to the other antifolates.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Folic Acid Antagonists/pharmacology , Polyglutamic Acid/metabolism , Tetrahydrofolate Dehydrogenase/drug effects , Antineoplastic Combined Chemotherapy Protocols/metabolism , Drug Synergism , Folic Acid Antagonists/metabolism , Glutamates/metabolism , Glutamates/pharmacology , Humans , Methotrexate/metabolism , Methotrexate/pharmacology , Pyrimidines/metabolism , Pyrimidines/pharmacology , Thymidylate Synthase/antagonists & inhibitors , Trimetrexate/metabolism , Trimetrexate/pharmacology , Tumor Cells, Cultured/drug effects
12.
J Pharmacokinet Biopharm ; 26(6): 717-33, 1998 Dec.
Article in English | MEDLINE | ID: mdl-10485082

ABSTRACT

Modeling of nonlinear pharmacodynamic (PD) relationships necessitates the utilization of a weighting function in order to compensate for the heteroscedasticity. The structure of the variance was studied for concentration-effect data generated in an in vitro 96-well plate cell growth inhibition assay, where data are numerous (480 data points per experiment) and replication is easy. From the five candidate models that were considered, the power function S2Y = phi 2Y phi 3, where Y is the sample mean and S2Y is the sample variance, was shown to be the most appropriate to describe the nonuniformity of the variance along the range of measured effect for 253 sets of (Y; S2Y) data. The Hill model was fit to the concentration-effect data with weighted nonlinear regression, where the weights were equal to the reciprocal of the predicted variance. The examination of the distribution of the 253 sets of parameters of the PD model showed that IC50 was lognormally distributed whereas the distribution of gamma was normal. The characterization of the appropriate variance function and concentration-effect function in a simple in vitro experimental setting with a large number of experiments, with each experiment including a large number of data points, will be useful for guiding similar in vitro concentration-effect studies where data are plentiful and for guiding PD modeling in complex clinical settings in which extensive data for model characterization is impossible to obtain.


Subject(s)
Antineoplastic Agents/pharmacology , Cell Division/drug effects , Humans , Models, Biological , Tumor Cells, Cultured
13.
Cancer Res ; 57(18): 4000-7, 1997 Sep 15.
Article in English | MEDLINE | ID: mdl-9307285

ABSTRACT

An in vivo quantitative structure-activity relationship (QSAR) study was carried out on a congeneric series of pyropheophorbide photosensitizers to identify structural features critical for their antitumor activity in photodynamic therapy (PDT). The structural elements evaluated in this study include the length and shape (alkyl, alkenyl, cyclic, and secondary analogs) of the ether side chain. C3H mice, harboring the radiation-induced fibrosarcoma tumor model, were used to study three biological response endpoints: tumor growth delay, tumor cell lethality, and vascular perfusion. All three endpoints revealed highly similar QSAR patterns that constituted a function of the alkyl ether chain length and drug lipophilicity, which is defined as the log of the octanol:water partition coefficient (log P). When the illumination of tumor, tumor cells, or cutaneous vasculature occurred 24 h after sensitizer administration, activities were minimal with analogs of log P < or = 5, increased dramatically between log P of 5-6, and peaked between log P of 5.6-6.6. Activities declined gradually with higher log P. The lack of activity of the least-lipophilic analogs was explained in large part by their poor biodistribution characteristics, which yielded negligible tumor and plasma drug levels at the time of treatment with light. The progressively lower potencies of the most lipophilic analogs cannot be explained through the overall tumor and plasma pharmacokinetics of photosensitizer because tumor and plasma concentrations progressively increased with lipophilicity. When compensated for differences in tumor photosensitizer concentration, the 1-hexyl derivative (optimal lipophilicity) was 5-fold more potent than the 1-dodecyl derivative (more lipophilic) and 3-fold more potent than the 1-pentyl analog (less lipophilic), indicating that, in addition to the overall tumor pharmacokinetics, pharmacodynamic factors may influence PDT activity. Drug lipophilicity was highly predictive for photodynamic activity. QSAR modeling revealed that direct antitumor effects and vascular PDT effects may be governed by common mechanisms, and that the mere association of high levels of photosensitizer in the tumor tissue is not sufficient for optimal PDT efficiency.


Subject(s)
Antineoplastic Agents/chemistry , Chlorophyll/analogs & derivatives , Photochemotherapy/methods , Photosensitizing Agents/chemistry , Animals , Chlorophyll/chemistry , Chlorophyll/pharmacology , Female , Lipids/chemistry , Mice , Mice, Inbred C3H , Regional Blood Flow/drug effects , Solubility , Structure-Activity Relationship
14.
Photochem Photobiol ; 66(2): 237-44, 1997 Aug.
Article in English | MEDLINE | ID: mdl-9277143

ABSTRACT

A method for determining the concentration of Photofrin in tissues and biological fluids was developed. The procedure is based on the dissolution of biological material with Solvable, a commercially available tissue solubilizer, followed by porphyrin-specific fluorescence detection and measurement. It was found necessary to use a quadratic standard curve for the estimation of unknown Photofrin concentrations. While this method is limited to compounds that are stable in strong base, it has the advantages of being sensitive, rapid and low cost.


Subject(s)
Antineoplastic Agents/analysis , Dihematoporphyrin Ether/analysis , Spectrometry, Fluorescence/methods , Animals , Calibration , Mice , Reproducibility of Results , Solubility
15.
Cancer Chemother Pharmacol ; 40(6): 495-505, 1997.
Article in English | MEDLINE | ID: mdl-9332464

ABSTRACT

PURPOSE: The combination of paclitaxel (PTX) and cisplatin (DDP) shows good clinical efficacy against ovarian cancer. In order to examine the potential cellular basis for this, and provide leads as to how to optimize the combination, we examined the role of sequence of exposure to PTX and DDP on cell growth in vitro. METHODS: Four human ovarian carcinoma cell lines, A121, A2780/WT, A2780/DX5B and A2780/CP3, two human head and neck carcinoma cell lines, A253 and FaDu, and the human ileocecal carcinoma cell line, HCT-8, were treated with PTX + DDP with seven schedules: (A) 96 h exposure to PTX + DDP; (B) 24 h PTX alone, then 72 h PTX + DDP; (C) 4 h DDP alone, then 92 h PTX + DDP; (D) 24 h PTX alone, 4 h DDP alone, then 68 h drug-free; (E) 4 h DDP alone, 24 h PTX alone, then 68 h drug-free; (F) 3 h PTX alone, 1 h DDP alone, then 92 h drug-free; and (G) 1 h DDP alone, 3 h PTX alone, then 92 h drug-free. Each of 66 two-drug experiments included five plates (440 randomly treated wells per experiment). Cell growth was measured by the sulforhodamine B assay. The nature and the intensity of the drug interactions were assessed by fitting a seven-parameter model to data with weighted nonlinear regression, enabling the estimation of an interaction parameter, alpha, with its standard error. RESULTS: Overall there was very little departure from Loewe additivity: 43 experiments showed Loewe additivity, 10 showed Loewe antagonism, and 13 showed slight Loewe synergy. In vitro Loewe synergy was rare, was small when present, and reproducible only for the A121 and HCT-8 cells exposed to schedule D (24 h PTX prior to 4 h DDP). Isobolographic analysis showed complex combined-action surfaces with regions of local Loewe synergy and antagonism. CONCLUSION: It appears unlikely that the good clinical efficacy of the combination is primarily caused by a synergistic interaction at the cellular level.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Ovarian Neoplasms/pathology , Cell Division/drug effects , Cisplatin/administration & dosage , Colonic Neoplasms/pathology , Drug Resistance, Neoplasm , Female , Head and Neck Neoplasms/pathology , Humans , Paclitaxel/administration & dosage , Tumor Cells, Cultured
17.
Photochem Photobiol ; 62(5): 896-905, 1995 Nov.
Article in English | MEDLINE | ID: mdl-8570729

ABSTRACT

The therapeutic effect of photodynamic therapy (PDT: photodynamic sensitizer + light) is partly due to vascular damage. This report describes a new vascular photodamage assay for PDT agents and a validation of the assay. The method described here quantitates changes in tissue blood perfusion based on the relative amount of injected fluorescein dye in treated and untreated tissues. A specially designed fluorometer uses chopped monochromatic light from an argon laser as a source for exciting fluorescein fluorescence. The fluorescent light emitted from the tissue is collected by a six element fiberoptic array, filtered and delivered to a photodiode detector coupled to a phase-locked amplifier for conversion to a voltage signal for recording. This arrangement permits a rather simple, inexpensive construction and allows for the simultaneous use of the argon laser by other investigators. The routine assay for characterizing a specific photosensitizer at a standard dose consists of the sequential allocation of eight mice to a set of different light doses designed to span the dose-response range of fluorescein fluorescence exclusion (measured 8-10 min after fluorescein injection). The assay validation experiment used an anionic photosensitizer, 2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide-a at a dose of 0.4 mumol/kg. The parameter estimates (n = 34 mice) from fitting the standard Hill dose-response model to the data were: median fluorescence exclusion light dose FE50 = 275 +/- 8.3 J/cm2 and Hill sigmoidicity parameter m = -3.66 +/- 0.28. Subsets of the full data set randomly selected to simulate a standard eight mice experiment yielded similar parameter estimates. The new assay provides reliable estimates of PDT vascular damage with a frugal sequential experimental design.


Subject(s)
Blood Vessels/drug effects , Photochemotherapy , Animals , Blood Vessels/radiation effects , Chlorophyll/analogs & derivatives , Chlorophyll/pharmacology , Fluorescein , Fluoresceins , Mice , Mice, Inbred C3H , Photosensitizing Agents/pharmacology , Reproducibility of Results , Skin/blood supply , Skin/drug effects , Skin/radiation effects , Spectrometry, Fluorescence/methods , Spectrometry, Fluorescence/statistics & numerical data
18.
Cancer Res ; 55(14): 3078-84, 1995 Jul 15.
Article in English | MEDLINE | ID: mdl-7606730

ABSTRACT

Two human pancreatic cell lines, MIA PaCa 2 and Capan 2, were treated by photodynamic therapy in vitro with Photophrin (0.01-25 micrograms/ml; 24 h) and then light (1-50 J/cm2; lambda = 630 nm). The following model was fit to 6 datasets with weighted nonlinear regression: [sequence: see text] The symbols are: E, cell growth; Econ, control growth in the absence of the combination; B, background signal; m, slope parameter; gamma, interaction parameter; D, concentration of Photofrin; L, light dose; F, fraction of Photofrin not photobleached by the light dose; k, k1, k2, bleaching parameters; A, distribution parameter for biexponential bleaching equation. Simple reciprocity of photosensitizer concentration and light dose was not found; compensation for photobleaching was critical. MIA PaCa2 required the monoexponential bleaching factor, whereas Capan 2 required the biexponential bleaching factor. The greater photosensitivity of MIA PaCa2 over Capan 2 can be best explained not by differences in the interaction parameter but rather by differences in the photobleaching pattern and rate. It may be possible to further enhance the selectivity of photodynamic therapy if differences in photobleaching between different cell types can be exploited by adequate dosimetry.


Subject(s)
Adenocarcinoma/drug therapy , Hematoporphyrin Derivative/pharmacology , Pancreatic Neoplasms/drug therapy , Photochemotherapy/methods , Adenocarcinoma/metabolism , Computer Simulation , Dose-Response Relationship, Drug , Hematoporphyrin Derivative/pharmacokinetics , Humans , Models, Biological , Pancreatic Neoplasms/metabolism , Thymidine/metabolism , Tritium , Tumor Cells, Cultured/drug effects
20.
Cancer Res ; 53(12): 2796-802, 1993 Jun 15.
Article in English | MEDLINE | ID: mdl-8504422

ABSTRACT

A liposome-encapsulated form of doxorubicin (TLC D-99), which was shown in preclinical toxicology to be less toxic to the gastrointestinal tract and myocardium than free doxorubicin, was administered by constant infusion (1.00-1.80 h) to 38 patients in single doses of 20, 30, 45, 60, 75, and 90 mg/m2 every 3 weeks and daily for 3 days at doses of 20, 25, and 30 mg/m2/day. The dose-limiting toxicity was leucopenia: the maximum tolerated doses were one at 90 mg/m2 and three at 25 mg/m2/day. Nausea, vomiting, and stomatitis were minimal or absent at each dose; alopecia was minor. Fever and chills were noted at most of the doses, and malaise was seen in some patients, especially at the higher doses. No hepatic, renal, or other organ toxicities were observed. Clinical cardiac toxicity was not observed in any patient; however, the cumulative doxorubicin dose was greater than 400 mg/m2 in only one patient. There was large variation among patients in estimated pharmacokinetic parameters and profiles. Higher plasma levels and dose intensities were achieved with TLC D-99 than were predicted for free doxorubicin. Liposome-encapsulated doxorubicin was well tolerated and produced less nausea, vomiting, and stomatitis than would be expected with free doxorubicin administered at equally myelosuppressive doses.


Subject(s)
Doxorubicin/administration & dosage , Neoplasms/drug therapy , Adult , Aged , Doxorubicin/adverse effects , Doxorubicin/pharmacokinetics , Drug Administration Schedule , Drug Carriers , Female , Humans , Leukopenia/chemically induced , Liposomes , Male , Middle Aged , Nausea/chemically induced , Neoplasms/blood , Neoplasms/metabolism , Thrombocytopenia/chemically induced , Vomiting/chemically induced
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