ABSTRACT
BACKGROUND: Isoprostanes are prostaglandin (PG)-like compounds synthesized by oxidative stress, not by cyclooxygenase, and increase in bronchoalveolar lavage fluid of patients with asthma. The airway inflammation implicated in this disease may be amplified by oxidants. Although isoprostanes are useful biomarkers for oxidative stress, the action of these agents on airways has not been fully elucidated. OBJECTIVE: This study was designed to determine the intracellular mechanisms underlying the effects of oxidative stress on airway smooth muscle, focused on Ca(2+) signalling pathways involved in the effect of 8-iso-PGF(2 alpha). METHODS: Using simultaneous recording of isometric tension and F(340)/F(380) (an indicator of intracellular concentrations of Ca(2+), [Ca(2+)]i, we examined the correlation between tension and [Ca(2+)]i in response to 8-iso-PGF(2 alpha) in the fura-2 loaded tracheal smooth muscle. RESULTS: Augmented tension and F(340)/F(380) by 8-iso-PGF(2 alpha) were attenuated by ICI-192605, an antagonist of thromboxane A(2) receptors (TP receptors). Moreover, D609, an antagonist of phosphatidylcholine-specific phospholipase C, markedly reduced both the tension and F(340)/F(380) induced by 8-iso-PGF(2 alpha), whereas U73122, an antagonist of phosphatidylinositol-specific phospholipase C, modestly inhibited them by 8-iso-PGF(2 alpha). SKF96365, a non-selective antagonist of Ca(2+) channels, markedly reduced both tension and F(340)/F(380) by 8-iso-PGF(2 alpha). However, diltiazem and verapamil, voltage-dependent Ca(2+) channel inhibitors, modestly attenuated tension although their reduction of F(340)/F(380) was not different from that by SKF96365. Y-27632, an inhibitor of Rho-kinase, significantly attenuated contraction induced by 8-iso-PGF(2 alpha) without reducing F(340)/F(380), whereas GF109203X and Go6983, protein kinase C inhibitors, did not markedly antagonize them although reducing F(340)/F(380) with a potency similar to Y-27632. CONCLUSION: 8-iso-PGF(2 alpha) causes airway smooth muscle contraction via activation of TP receptors. Ca(2+) mobilization by SKF96365- and D609-sensitive Ca(2+) influx and Ca(2+) sensitization by Rho-kinase contribute to the intracellular mechanisms underlying the action of 8-iso-PGF(2 alpha). Rho-kinase may be a therapeutic target for the physiologic abnormalities induced by oxidative stress in airways.
Subject(s)
Calcium Signaling/physiology , Dinoprost/analogs & derivatives , Muscle, Smooth/drug effects , Trachea/drug effects , Trachea/physiology , 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid/pharmacology , Amides/pharmacology , Animals , Bridged-Ring Compounds/pharmacology , Calcium Channel Blockers/pharmacology , Calcium Signaling/drug effects , Diltiazem/pharmacology , Dinoprost/pharmacology , Dioxanes/pharmacology , Enzyme Inhibitors/pharmacology , Estrenes/pharmacology , Guinea Pigs , Imidazoles/pharmacology , In Vitro Techniques , Indoles/pharmacology , Male , Maleimides/pharmacology , Muscle Contraction/drug effects , Muscle Contraction/physiology , Muscle, Smooth/physiology , Norbornanes , Pyridines/pharmacology , Pyrrolidinones/pharmacology , Receptors, Thromboxane A2, Prostaglandin H2/antagonists & inhibitors , Signal Transduction/drug effects , Signal Transduction/physiology , Thiocarbamates , Thiones/pharmacology , Type C Phospholipases/antagonists & inhibitors , Vasoconstrictor Agents/pharmacology , Verapamil/pharmacology , rho-Associated Kinases/antagonists & inhibitorsABSTRACT
BACKGROUND: Recent studies have revealed that in asthma, mast cells infiltrate to the smooth muscle layer and release tryptase, an enzymatic activator of protease-activated receptor 2 (PAR2). This phenomenon, mast cell myositis, is proposed as a new feature of asthma. However, little is known about the involvement of mast cell myositis in the pathophysiology of asthma. OBJECTIVE: This study was designed to determine whether mast cell degranulation has any functional impact on beta-adrenoceptors via PAR2 in airway smooth muscle. Moreover, we focused on Ca(2+) signalling as a mechanism underlying alteration of smooth muscle tone and responsiveness. METHODS: Isometric tension and F(340)/F(380), an indicator of the concentration of intracellular Ca(2+) ([Ca(2+)](i)), were simultaneously measured using fura-2-loaded tissues isolated from guinea-pig tracheal smooth muscle. RESULTS: Tryptase (1-100 nm) caused tension with elevated F(340)/F(380), and after exposure to tryptase for 15 min the inhibitory effect of isoprenaline (ISO) against methacholine was attenuated without elevating F(340)/F(380) in a concentration-dependent manner. Tryptase (<1 nm) had a modest effect on tension, but prolonged treatment (=120 min) with 0.1 nm tryptase also reduced the effects of ISO in a time-dependent manner. When tissues were incubated with tryptase in the presence of Y-27632, a Rho-kinase inhibitor, reduced responsiveness to ISO by tryptase was reversed without affecting F(340)/F(380). In contrast, pre-treatment with SKF96365, a non-selective inhibitor of Ca(2+) channels, did not antagonize the effect of tryptase. Moreover, pre-treatment with SLIGKV-NH(2), a non-enzymatic activator of PAR2, resulted in a loss of beta-adrenergic efficacy, similar to tryptase. The effect of cAMP-related agents bypassing beta-adrenoceptors was not attenuated after exposure to tryptase. CONCLUSION: In mast cell myositis, tryptase released from mast cells acts on airway smooth muscle, leading to homologous beta-adrenergic desensitization mediated by [Ca(2+)](i)-independent mechanisms via PAR2 activation.
Subject(s)
Calcium/metabolism , Mast Cells/enzymology , Muscle, Smooth/metabolism , Receptors, Adrenergic, beta/metabolism , Trachea/metabolism , Tryptases/metabolism , Adrenergic beta-Agonists/pharmacology , Animals , Calcium Signaling/drug effects , Guinea Pigs , Humans , Male , Mast Cells/drug effects , Muscle, Smooth/drug effects , Receptor, PAR-2/metabolism , Trachea/drug effects , rho-Associated Kinases/metabolismABSTRACT
The L-myc gene was first isolated from a human small-cell lung cancer (SCLC) cell line on the basis of its amplification and sequence similarity to c-myc and N-myc. A new mechanism of L-myc activation which results from the production of rlf-L-myc fusion protein was recently reported. On the basis of our earlier observation of a rearrangement involving amplified L-myc in an SCLC cell line, ACC-LC-49, we decided to investigate this rearrangement in detail along with the structure of L-myc amplification units in five additional SCLC cell lines. We report here the identification of a novel genomic region, termed jal, which is distinct from rlf and is juxtaposed to and amplified with L-myc during the process of DNA amplification of the region encompassing L-myc. Long-range analysis using pulsed-field gel electrophoresis revealed that the amplified L-myc locus is involved in highly complex intrachromosomal rearrangements with jal and/or rlf. Our results also suggest that the simultaneous presence of rearrangements both in rlf intron 1 and in regions immediately upstream of L-myc may be necessary for the expression of rlf-L-myc chimeric transcripts.
Subject(s)
Carcinoma, Small Cell/genetics , Chromosomes, Human, Pair 1 , Gene Rearrangement , Genes, myc/genetics , Lung Neoplasms/genetics , Chromosome Mapping , Electrophoresis, Gel, Pulsed-Field , Gene Amplification , Gene Expression , Humans , RNA, Messenger/metabolism , Transcription, GeneticABSTRACT
Accumulating evidence suggests that altered DNA methylation may play a role in the oncogenesis of human neoplasms, including lung cancer. The presence of aberrant hypermthylations at 3p, 9p, 11p, ad 17p, which are known to be hot spots for allele loss in lung cancers, is suggested to be a reflection of the existence of tumor suppressor genes in these chromosomal regions. In the present study, we investigated the methylation status of the Rb locus at 13q14 as well as that of the bcl-2 locus at 18q21 in 134 lung cancer specimens, representing all major histological subtypes. As a result, 18q21 was identified to be the fifth chromosomal region affected by frequent tumor-specific aberrant hypermethylation in lung cancers. The occurrence of aberrant hypermethylation at the bcl-2 locus at 18q21 was restricted to non-small cell lung cancers, and among non-small cell lung cancers, such epigenetic aberrations were observed most frequently in adenocarcinomas without any association with bcl-2 expression. Interestingly, allelic loss at the bcl-2 locus was also seen in 40% (7 of 17 informative cases) of adenocarcinomas; this frequency was also the highest among values for the various histological subtypes of lung cancers. These results suggest that aberrant hypermethylation at the bcl-2 locus may be a reflection of a putative tumor suppressor gene residing at 18q21, and aberrant hypermethylation might play a role in its inactivation. In contrast, altered methylation status of the Rb locus appears to be quite rare in lung cancers, if present at all.
Subject(s)
Chromosomes, Human, Pair 18 , Genes, Retinoblastoma , Lung Neoplasms/genetics , Proto-Oncogene Proteins/genetics , Proto-Oncogenes , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Blotting, Southern , Carcinoma, Large Cell/genetics , Carcinoma, Large Cell/pathology , Carcinoma, Small Cell/genetics , Carcinoma, Small Cell/pathology , Carcinoma, Squamous Cell/genetics , Carcinoma, Squamous Cell/pathology , Cell Line , Chromosome Mapping , Chromosomes, Human, Pair 13 , DNA, Neoplasm/chemistry , DNA, Neoplasm/isolation & purification , Exons , Humans , Lung Neoplasms/pathology , Methylation , Proto-Oncogene Proteins c-bcl-2 , Restriction MappingABSTRACT
As an initial step to understand rapid growth of small cell lung cancer (SCLC), a complementary DNA library prepared from a SCLC cell line was screened with viral oncogene probes encoding protein-tyrosine kinases, which are known to play an important role in regulation of cell growth. Fifteen clones hybridizing with v-fms probe were isolated, and, by partial sequence analysis, four of them were identified to be c-kit protooncogenes. Northern blot study demonstrated that most of the SCLC tumors and cell lines expressed c-kit transcripts, while non-SCLC tumors and cell lines did not. Neither amplification nor rearrangement of the c-kit gene was demonstrated in SCLC cell lines by Southern blot analysis, however. Our results suggested that c-kit expression in SCLC reflects the unique biological nature of the tumor cells different from non-SCLC and further suggested that the c-kit product may participate in autocrine or paracrine stimulation of SCLC growth.
Subject(s)
Carcinoma, Small Cell/genetics , Gene Expression Regulation, Neoplastic/genetics , Lung Neoplasms/genetics , Protein-Tyrosine Kinases/genetics , Proto-Oncogene Proteins/genetics , Receptor Protein-Tyrosine Kinases , Transcription, Genetic , Base Sequence , Carcinoma, Non-Small-Cell Lung/genetics , Gene Library , Humans , Molecular Sequence Data , Oncogene Protein gp140(v-fms)/genetics , Oncogene Proteins, Viral/genetics , Proto-Oncogene Proteins c-kit , Tumor Cells, CulturedABSTRACT
We evaluated the prognostic significance of p53 mutations and an allelic loss of chromosome 3p in 71 patients with non-small cell lung cancer who underwent potentially curative resection. p53 mutations were detected in 35 cases (49%), while 3p deletions were observed in 34 of 70 informative cases (49%). The presence of the p53 mutation was associated with a shortened survival in all patients (P = 0.014 by log rank test), including those in early stages of the disease (stage I or II, n = 48) (P = 0.016 by log rank test). Multivariate analysis by the Cox proportional hazards model also revealed that p53 mutation was an independent yet unfavorable prognostic factor (P = 0.013). Patients with 3p deletion tended to have a poorer prognosis, but not to a statistically significant extent.
Subject(s)
Carcinoma, Non-Small-Cell Lung/genetics , Chromosomes, Human, Pair 3/physiology , Gene Deletion , Genes, p53/genetics , Lung Neoplasms/genetics , Aged , Alleles , Base Sequence , Carcinoma, Non-Small-Cell Lung/surgery , Female , Humans , Lung Neoplasms/surgery , Male , Middle Aged , Molecular Sequence Data , Mutation , PrognosisABSTRACT
Irinotecan unexpectedly causes severe toxicity of leukopenia or diarrhea. Irinotecan is metabolized to form active SN-38, which is further conjugated and detoxified by UDP-glucuronosyltransferase (UGT) 1A1 enzyme. Genetic polymorphisms of the UGT1A1 would affect an interindividual variation of the toxicity by irinotecan via the alternation of bioavailability of SN-38. In this case-control study, retrospective review of clinical records and determination of UGT1A1 polymorphisms were performed to investigate whether a patient with the variant UGT1A1 genotypes would be at higher risk for severe toxicity by irinotecan. All patients previously received irinotecan against cancer in university hospitals, cancer centers, or large urban hospitals in Japan. We identified 26 patients who experienced severe toxicity and 92 patients who did not. The relationship was studied between the multiple variant genotypes (UGT1A1*28 in the promoter and UGT1A1*6, UGT1A1*27, UGT1A1*29, and UGT1A1*7 in the coding region) and the severe toxicity of grade 4 leukopenia (< or =0.9 x 10(9)/liter) and/or grade 3 (watery for 5 days or more) or grade 4 (hemorrhagic or dehydration) diarrhea. Of the 26 patients with the severe toxicity, the genotypes of UGT1A1*28 were homozygous in 4 (15%) and heterozygous in 8 (31%), whereas 3 (3%) homozygous and 10 (11%) heterozygous were found among the 92 patients without the severe toxicity. Multivariate analysis suggested that the genotype either heterozygous or homozygous for UGT1A1*28 would be a significant risk factor for severe toxicity by irinotecan (P < 0.001; odds ratio, 7.23; 95% confidence interval, 2.52-22.3). All 3 patients heterozygous for UGT1A1*27 encountered severe toxicity. No statistical association of UGT1A1*6 with the occurrence of severe toxicity was observed. None had UGT1A1*29 or UGT1A1*7. We suggest that determination of the UGT1A1 genotypes might be clinically useful for predicting severe toxicity by irinotecan in cancer patients. This research warrants a prospective trial to corroborate the usefulness of gene diagnosis of UGT1A1 polymorphisms prior tb irinotecan chemotherapy.
Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Antineoplastic Agents, Phytogenic/toxicity , Camptothecin/analogs & derivatives , Camptothecin/pharmacology , Camptothecin/toxicity , Glucuronosyltransferase/genetics , Polymorphism, Genetic , Adult , Aged , Case-Control Studies , Female , Genotype , Heterozygote , Homozygote , Humans , Irinotecan , Leukopenia/chemically induced , Male , Middle Aged , Multivariate Analysis , Promoter Regions, Genetic , Retrospective Studies , Risk FactorsABSTRACT
The chromosome region 18q21 has been shown to be frequently deleted in lung cancers. Recent identification at this locus of DPC4, a gene whose inactivation has been suggested to play a role in pancreatic carcinogenesis, prompted as to examine whether it might also be altered in lung cancers. Two missense and 2-bp frameshift somatic mutations in DPC4 were detected among 42 lung cancer specimens taken directly from patients. DPC4 mutations, however, were not present in all lung cancers carrying l8q21 deletions. These findings suggest that DPC4 may play a role in a limited fraction of lung cancers and that another tumor suppressor gene may also exist in this chromosome region.
Subject(s)
Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Small Cell/genetics , Chromosomes, Human, Pair 18/genetics , DNA-Binding Proteins , Gene Deletion , Genes, Tumor Suppressor/genetics , Lung Neoplasms/genetics , Trans-Activators , Base Sequence , Humans , Molecular Sequence Data , Polymerase Chain Reaction , Proteins/genetics , Sequence Analysis, DNA , Smad4 ProteinABSTRACT
Accumulating evidence suggests that c-kit and its ligand, stem cell factor (SCF), play an important role in the regulation of at least three lineages of stem cell growth and possibly in leukemogenesis, while only limited data are available that suggest possible involvement of c-kit/SCF in the development of human solid tumors such as lung cancer. We have recently reported that c-kit is aberrantly expressed almost exclusively in small-cell lung cancer (SCLC) among various types of solid tumors. The present study revealed that c-kit protein ectopically expressed in SCLC is indistinguishable from that in leukemia cell lines with megakaryocytic characteristics with respect to amount, molecular size, and autophosphorylation status in response to recombinant human SCF. Furthermore, significant chemotactic response as well as moderate in vitro cell growth was induced in SCLC cell lines by the addition of recombinant human SCF, suggesting that c-kit/SCF may play an important biological role in the development of SCLC. Our extensive search for activating mutations naturally occurring in the c-kit gene revealed an amino acid substitution in the transmembrane domain of an SCLC cell line, although the functional consequences of this variant allele are yet to be determined.
Subject(s)
Carcinoma, Small Cell/genetics , Chemotaxis/drug effects , Hematopoietic Cell Growth Factors/pharmacology , Lung Neoplasms/genetics , Proto-Oncogene Proteins/analysis , Carcinoma, Small Cell/chemistry , Carcinoma, Small Cell/pathology , Cell Division/drug effects , Humans , Leukemia/genetics , Lung Neoplasms/chemistry , Lung Neoplasms/pathology , Phosphorylation/drug effects , Proto-Oncogene Proteins/metabolism , Proto-Oncogene Proteins c-kit , Recombinant Proteins/pharmacology , Sequence Analysis , Stem Cell Factor , Tumor Cells, CulturedABSTRACT
Germline mutations of p53 have been implicated as a cause of cancer susceptibility in the Li-Fraumeni syndrome. Since inactivation of p53 has been suggested to play an important causative role in lung cancer, the present study of the prevalence of germline mutations in 148 patients with this neoplasm was performed. None of 138 randomly chosen patients were found to carry such mutations, while a single patient had a nonsense mutation at codon 213 among 10 patients selected for early onset and/or occurrence of multiple primary cancers. In contrast to the previous report of biallelic expression of p53 in a case with a germline missense mutation, preferential expression of the wild-type allele was observed in the heterozygous state in both normal lung and peripheral blood lymphocytes of our case, whereas expression of mutant mRNA was readily detectable in her lung cancer in the absence of the remaining wild-type allele. Interestingly, the family history of the proband showed a mild aggregation of adulthood cancers and a high prevalence of stomach cancer, a rare component in American families affected by the syndrome. These observations suggest the presence of heterogeneity with regard to molecular and clinical features of germline p53 mutations.
Subject(s)
Genes, p53 , Germ-Line Mutation , Lung Neoplasms/genetics , Adult , Alleles , Female , Humans , Immunohistochemistry , Japan , Pedigree , Polymerase Chain ReactionABSTRACT
The beta-catenin gene (CTNNB1) has been shown to be genetically mutated in various human malignancies. To determine whether the beta-catenin gene is responsible for oncogenesis in thoracic malignancies, we searched for the mutation in 166 lung cancers (90 primary tumors and 76 cell lines), one blastoma and 10 malignant mesotheliomas (two primary tumors and eight cell lines). Among the lung cancers, including 43 small cell lung cancers (SCLCs) and 123 non-small cell lung cancers (NSCLCs), we identified four alterations in exon 3, which is the target region of mutation for stabilizing beta-catenin. One primary adenocarcinoma had a somatic mutation from C to G, leading to an amino acid substitution from Ser to Cys at codon 37. Among the cell lines, SCLC NCI-H1092 had a mutation from A to G, leading to an Asp to Gly substitution at codon 6, NSCLC HCC15 had a mutation from C to T, leading to a Ser to Phe substitution at codon 45, and NSCLC NCI-H358 had a mutation from A to G, leading to a Thr to Ala substitution at codon 75. One blastoma also had a somatic mutation from C to G, leading to a Ser to Cys substitution at codon 37. Among the 10 malignant mesotheliomas, we identified a homozygous deletion in the NCI-H28 cell line. Cloning of the rearranged fragment from NCI-H28 indicated that all the exons except exon 1 of the beta-catenin gene are deleted and that the deletion junction is 13 kb downstream from exon 1. Furthermore, Northern blot analysis of 26 lung cancer and eight mesothelioma cell line RNAs detected ubiquitous expression of the beta-catenin messages except NCI-H28, although Western blot analysis showed that relatively less amounts of protein products were expressed in some of lung cancer cell lines. Our findings suggest that the beta-catenin gene is infrequently mutated in lung cancer and that the NCI-H28 homozygous deletion of the beta-catenin gene might indicate the possibility of a new tumor suppressor gene residing in this region at 3p21.3, where various types of human cancers show frequent allelic loss.
Subject(s)
Chromosome Deletion , Chromosomes, Human, Pair 3 , Cytoskeletal Proteins/genetics , Homozygote , Lung Neoplasms/genetics , Mesothelioma/genetics , Mutation , Trans-Activators , Base Sequence , DNA, Neoplasm , Exons , Gene Rearrangement , Humans , Molecular Sequence Data , beta CateninABSTRACT
PURPOSE: The antitumor effect of etoposide is increased by maintaining a low blood level, whereas high peak levels may cause myelotoxicity. We investigated whether a constant low blood level could be obtained by the administration of oral etoposide three times daily. PATIENTS AND METHODS: Nineteen patients with non-small-cell lung cancer were treated with oral etoposide (25 mg three times daily for 21 days) as monotherapy or in combination with cisplatin 80 mg/m2. A pharmacokinetic model that predicted the mean blood concentration (Cmean) was developed in the 10 patients on etoposide monotherapy and validated in the nine patients on combination chemotherapy. Pharmacodynamic relationships were evaluated in each group. RESULTS: Etoposide dose per body-surface area ranged from 45 to 63 mg/m2/d (median, 53), but did not correlate with plasma level. Cmean was 1.1 +/- 0.3 micrograms/mL. Peak concentrations ranged from 0.6 to 2.5 micrograms/mL. The intrapatient coefficient of variation for plasma etoposide concentrations was 22% +/- 10%. Cmean was accurately estimated as follows: Cmean = 0.098 + 0.413 x C0 + 0.458 x C2 (r = .97, P = .0001), where C0 and C2 represent concentrations before and 2 hours after administration. This model was unbiased (mean predictive error [MPE], 0.0 microgram/mL) and precise (root mean square error [RMSE], 0.1 microgram/mL). Leukopenia was the major toxicity. The surviving fraction of leukocytes (SF; nadir count/pretreatment count) was correlated to Cmean as follows: SF = 0.87 - 0.34 x Cmean (r = .67, P = .03) in the monotherapy group and SF = 0.64 - 0.33 x Cmean (R = .77, P = .03) in the combination chemotherapy group. Two and four patients treated with monotherapy and combination chemotherapy showed responses, respectively. All responders had a Cmean > or = 1.0 microgram/mL. CONCLUSION: Hyperfractionated oral etoposide achieveda stable plasma level that could be predicted by measurement at only two times.
Subject(s)
Carcinoma, Non-Small-Cell Lung/blood , Etoposide/administration & dosage , Etoposide/blood , Lung Neoplasms/blood , Administration, Oral , Adult , Aged , Aged, 80 and over , Carcinoma, Non-Small-Cell Lung/drug therapy , Cisplatin/administration & dosage , Drug Administration Schedule , Etoposide/adverse effects , Female , Humans , Leukopenia/chemically induced , Lung Neoplasms/drug therapy , Male , Middle Aged , Regression AnalysisABSTRACT
PURPOSE: A phase I study was conducted to determine the maximum-tolerated dose (MTD) of a 14-day continuous infusion of etoposide, and to evaluate the pharmacokinetics in patients with lung cancer. PATIENTS AND METHODS: Etoposide was administered continuously through a central venous catheter using a pump. The starting dose level was 300 mg/m2 over 14 days, with dose escalations of 100 mg/m2 over 14 days until unacceptable toxicities occurred. Pharmacokinetic studies were performed in all patients. RESULTS: Twenty-one patients, 20 with non-small-cell lung cancer and one with refractory small-cell lung cancer, received 37 courses. No World Health Organization (WHO) grade III or greater toxicity occurred at doses up to 400 mg/m2 over 14 days. At 700 mg/m2 over 14 days, all four patients experienced grade III or IV leukocytopenia, and two developed grade III stomatitis. No cumulative toxicity was observed. A steady concentration of etoposide was achieved 24 hours after the start of chemotherapy, and it was significantly correlated with surviving fractions of leukocytes (r = -.64, P = .001) and platelets (r = -.68, P < .001). The leukocyte count at the termination of chemotherapy predicted the nadir count (r = .93, P < .001). CONCLUSION: Steady blood levels of etoposide were maintained for prolonged periods, during 14-day continuous infusions. Leukocytopenia and stomatitis were dose-limiting. Nadir counts and surviving fractions of leukocytes were predicted by the leukocyte count at the end of chemotherapy and the concentration of etoposide, respectively. The recommended dose for phase II trials is 600 mg/m2 over 14 days.
Subject(s)
Etoposide/administration & dosage , Etoposide/pharmacokinetics , Lung Neoplasms/drug therapy , Adult , Aged , Catheterization, Central Venous , Drug Administration Schedule , Etoposide/adverse effects , Etoposide/therapeutic use , Female , Hematologic Diseases/chemically induced , Humans , Infusion Pumps , Infusions, Intravenous , Lung Neoplasms/metabolism , Male , Middle AgedABSTRACT
This study was performed to determine whether pleural effusion in patients with advanced non-small cell lung cancer (NSCLC) has a negative impact on survival. We evaluated 12 prognostic factors in 197 patients with stage IIIB or IV NSCLC. Each factor was dichotomized, and survival curves calculated by the Kaplan-Meier technique were compared using the log-rank test. The Cox proportional hazards regression model was used to confirm the significance of each prognostic factor selected by univariate analysis. We compared the survival times for stage IIIB with pleural effusion with those of stage IIIB without effusion and stage IV. To determine the impact of the cytological results of the effusion on survival, we compared the survival times for cytologically positive and negative effusions. Univariate analysis identified eight significant prognostic factors: pleural effusion, node status, stage, performance status, weight loss, hemoglobin, albumin, and lactate dehydrogenase. Pleural effusion was selected as a prognostic factor in the multivariate analysis, together with stage, performance status, albumin, and node status. Median survival times for stage IIIB without effusion, stage IIIB with effusion, and stage IV were 15.3, 7.5, and 5.5 months, respectively (P < 0.0001). Survival time for stage IIIB with effusion was significantly different from that of stage IIIB without effusion (P = 0.0129) but not from that of stage IV (P = 0.0797). Among patients with effusion, no significant difference in survival time was observed between cytologically positive and negative effusions. We conclude that pleural effusion in advanced NSCLC is a prognostic factor. Survival time for stage IIIB with pleural effusion is more similar to that of stage IV rather than that of stage IIIB without effusion.
Subject(s)
Carcinoma, Non-Small-Cell Lung/diagnosis , Lung Neoplasms/diagnosis , Pleural Effusion , Adult , Aged , Aged, 80 and over , Carcinoma, Non-Small-Cell Lung/physiopathology , Female , Humans , Lung Neoplasms/physiopathology , Male , Middle Aged , Multivariate Analysis , Neoplasm Staging , Pleural Effusion/pathology , Prognosis , Survival AnalysisABSTRACT
Creatinine clearance (Ccr) is widely used as a practical substitute for glomerular filtration rate (GFR) in the Calvert formula: carboplatin dose (mg) = target area under the concentration versus time curve (AUC, mg ml(-1) min) x [GFR (ml min(-1)) + 25]. However, it causes systematic overdosing when the creatinine levels are measured by an enzymatic peroxidase-antiperoxidase method (PAP-Cr). We previously suggested an amended dosing formula to adjust this overdosing: carboplatin dose (mg) = AUC (mg ml(-1) min) x [adjusted Ccr (ml min(-1)) + 25], where the Ccr was adjusted by adding 0.2 (mg dl(-1)) to serum PAP-Cr. In this study, we prospectively validated this formula in 55 patients from six institutions. Target AUC ranged from 3 to 7 mg ml(-1) min, and Ccr was measured by 24-h urine collection. Estimation of carboplatin clearance with the amended formula was unbiased [mean prediction error (MPE) +/- SE = 2.9 +/- 3.4%] and acceptably precise [root mean squared error, (RMSE) = 24.7%], whereas the Calvert formula using non-adjusted Ccr overpredicted carboplatin clearance systematically (MPE +/- SE = 24.9 +/- 4.9% and RMSE = 36.1%). The improvement in the bias and precision of the estimation was seen in all of the participating institutions as shown by decrease in the absolute value of MPE and RMSE for each institution. The Chatelut formula also highly overestimated carboplatin clearance when PAP-Cr was used, but the adjustment of PAP-Cr yielded a decrease in MPE by 30.4% and in RMSE by 21.3%. These results confirmed the necessity of adjusting the serum PAP-Cr in carboplatin dosing formulas.
Subject(s)
Carboplatin/administration & dosage , Carboplatin/therapeutic use , Creatinine/blood , Creatinine/urine , Neoplasms/blood , Neoplasms/drug therapy , Adult , Aged , Aged, 80 and over , Area Under Curve , Bias , Female , Humans , Male , Middle Aged , Peroxidase/blood , Time FactorsABSTRACT
To analyze the pharmacological characteristics of etoposide in elderly patients, we conducted a Phase I trial of a 14-day administration of oral etoposide on 12 chemotherapy-naive patients, ages 75 years or older, with lung cancer. The pharmacological profiles of etoposide in elderly patients were compared with those of younger patients in our previous studies (H. Minami et al., J. Clin. Oncol., 11: 1602-1608, 1993; H. Minami et al., J. Clin. Oncol., 13: 191-199, 1995; Y. Ando et al., Jpn. J. Cancer Res., 87: 200-205, 1996). The sigmoid Emax model and logistic regression model were used for pharmacodynamic analysis. The maximum tolerated dose for elderly patients was 75 mg/body/day. The apparent oral clearance in elderly patients was 37+/-10 (mean +/- SD) ml/min, which was not different from that in younger patients (44+/-12 ml/min). The area under the concentration-versus-time curve of etoposide over the treatment period (total AUC) that produced a 50% decrease in absolute neutrophil counts was significantly different between elderly and younger patients, 14.3+/-2.5 and 21.6+/-2.7 mg x min/ml, respectively (P = 0.048). The incidence of grade 3 or 4 neutropenia at total AUC of 30 mg x min/ml (corresponding to a plasma concentration of 1.5 microg/ml for 14 days) was 81% in elderly patients but only 48% in younger patients. Although there was no pharmacokinetic difference between elderly and younger patients, equivalent exposure to etoposide resulted in severer myelosuppression in elderly patients. These findings suggest that prolonged etoposide administration with plasma concentration maintained at 1-2 microg/ml may cause severe myelotoxicity in elderly patients.
Subject(s)
Antineoplastic Agents, Phytogenic/therapeutic use , Etoposide/therapeutic use , Lung Neoplasms/drug therapy , Administration, Oral , Aged , Aged, 80 and over , Antineoplastic Agents, Phytogenic/adverse effects , Antineoplastic Agents, Phytogenic/pharmacokinetics , Etoposide/adverse effects , Etoposide/pharmacokinetics , Humans , Lung Neoplasms/metabolism , Treatment OutcomeABSTRACT
We used a recombinant retrovirus as one of the potential vectors for human gene therapy to transfer a drug sensitivity gene into human lung cancer cells. The gene encoding the thymidine kinase (TK) of herpes simplex virus type 1 (HSV1) was used as the drug sensitivity gene. The antiherpes drugs acyclovir (ACV) and ganciclovir (GCV) were chosen to test the HSV1-TK activity transferred into the human lung cancer cell lines. The rationale for this approach was that ACV and GCV are nucleoside analogs specifically converted by HSV1-TK to a toxic form capable of inhibiting DNA synthesis or disrupting cellular DNA replication. The results obtained from our experiments demonstrate that the retroviral vector-mediated HSV1-TK gene transfer leads to ACV- and GCV-dependent cytotoxicity in human lung cancer cell lines, including both small-cell carcinoma and non-small-cell carcinoma. Although the gene transfer of HSV1-TK gene into tumor cells would be one model for gene therapy to control lung cancer, further investigations are necessary for the proper choice of the therapeutic gene and vector targeting such as tumor cell specific delivery of the gene or tumor cell specific expression of the transduced gene.
Subject(s)
Gene Transfer Techniques , Genetic Therapy , Genetic Vectors , Retroviridae/genetics , Simplexvirus/genetics , Thymidine Kinase/genetics , Acyclovir/pharmacology , Base Sequence , Cloning, Molecular , DNA, Recombinant/genetics , Ganciclovir/pharmacology , Humans , Lung Neoplasms , Molecular Sequence Data , Retroviridae/chemistry , Simplexvirus/enzymology , Transfection/genetics , Tumor Cells, CulturedABSTRACT
The infection of recombinant adenovirus expressing wild-type p53 (Ad-p53) to lung cancer cells that harbor mutant p53 genes improves their response to cis-diamminedichloroplatinum(II). In this study, we tested whether this improvement in response is also seen in wild-type p53 (wt-p53)-containing cancer cells and whether this phenomenon is universal with other commonly used chemotherapeutic agents, including etoposide, 7-ethyl-10-hydrocycamptothecin, paclitaxel, and docetaxel. Using a panel of 7 non-small cell lung cancer cell lines with wild-type (2) or abnormal (2 null, 3 point-mutated) p53, we examined in vitro cytotoxicity using a tetrazolium-based colorimetric assay (3-(4,5-diethylthiazoyl-2-yl)-2,5-diphenyltetrazolium bromide assay) and analyzed the combined effects of Ad-p53 and chemotherapeutic agents using the isobologram method. Ad-p53 and DNA-damaging agents (cis-diamminedichloroplatinum(II), etoposide, and 7-ethyl-10-hydrocycamptothecin) showed synergistic effects in six of seven cell lines but additive effects against a p53-mutated cell line. In contrast, Ad-p53 showed additive effects with the antitubulin agents (paclitaxel and docetaxel) in all four of the cell lines tested. Furthermore, we examined this synergistic interaction between Ad-p53 and DNA-damaging agents by flow cytometric analysis and DNA fragmentation analysis. Both analyses revealed that a sublethal dose of Ad-p53 augmented the apoptotic response induced by DNA-damaging agents in six of seven cell lines. Our results suggest that Ad-p53 may synergistically enhance the chemosensitivity of the majority of non-small cell lung cancers to DNA-damaging agents due to augmentation of apoptosis.
Subject(s)
Cisplatin/toxicity , Genes, p53 , Tumor Suppressor Protein p53/physiology , Adenoviridae/genetics , Carcinoma, Non-Small-Cell Lung , Cell Survival/drug effects , Humans , Lung Neoplasms , Mutagenesis , Transfection , Tumor Cells, CulturedABSTRACT
Gene therapy using the herpes simplex virus thymidine kinase (HSV-TK) gene combined with an anti-herpes drug, ganciclovir (GCV), has been applied for human diseases, especially for cancer treatment. However, bone marrow toxicity has been the most consistent adverse effect of GCV treatment in clinical settings. We evaluated the cytotoxic activity of a novel guanosine analog, (1'S,2'R)-9[[1',2'-bis(hydroxymethyl)cycloprop-1'-yl]methyl]guanin e (A-5021), against retrovirus-mediated HSV-TK gene-transduced human lung cancer cells. The bone marrow toxicity of A-5021 and GCV was studied by colony formation assay in both rodent and human bone marrow specimens. We demonstrated that A-5021 had potent cytotoxic activity equal to that of GCV against the retroviral vector-mediated HSV-TK gene-transduced lung cancer cell lines. Further, phosphorylated A-5021 could be transferred to neighboring cells, and this analog killed HSV-TK- neighboring cells, as was the case for GCV. In contrast, A-5021 did not exhibit an inhibitory effect on bone marrow progenitor cells and colony formation (the 50% inhibitory concentration of the colony-forming units in culture = >100 microg/mL in human bone marrow specimens and >66 microg/mL in rodent bone marrow specimens). These results indicate that A-5021 has potent cytotoxic activity as a nucleoside analog for gene therapy using HSV-TK gene, and can be used much more safely than GCV.
Subject(s)
Antiviral Agents/toxicity , Bone Marrow Cells/drug effects , Cell Survival/drug effects , Genetic Therapy/methods , Guanine/analogs & derivatives , Herpesvirus 1, Human/genetics , Thymidine Kinase/genetics , Transfection , Adenocarcinoma , Animals , Bone Marrow Cells/cytology , Carcinoma, Small Cell , Cells, Cultured , Dose-Response Relationship, Drug , Ganciclovir/toxicity , Genetic Vectors , Guanine/toxicity , Humans , Lung Neoplasms , Mice , Retroviridae , Thymidine Kinase/metabolism , Tumor Cells, CulturedABSTRACT
CD40-CD40 ligand (CD40L) interaction is an important costimulatory signaling pathway in the crosstalk between T cells and antigen-presenting cells. This receptor-ligand system is known to be essential in eliciting strong cellular immunity. Here we demonstrate that murine lung cancer cells (3LLSA) transduced with the CD40L gene (3LLSA-CD40L) were rejected in syngeneic C57BL/6 mice, but grew in CD40-deficient mice to the same extent as control tumor cells. Immunohistochemical study showed that inflammatory cells, including CD4+, CD8+ T cells and NK cells, infiltrated into the inoculated 3LLSA-CD40L tumor tissue. Inoculation of 3LLSA-CD40L cells into mice resulted in the induction of 3LLSA-specific cytotoxic T-cell immunity, and the growth of parental 3LLSA tumors was inhibited when 3LLSA cells were inoculated into C57BL/6 mice mixed with 3LLSA-CD40L cells or when they were rechallenged 4 weeks after 3LLSA-CD40L cells were rejected. Furthermore, co-inoculation of interferon (IFN)-gamma-transduced cells (3LLSA-IFNgamma) with 3LLSA-CD40L cells enhanced the antitumor immunity efficiently in vivo. These results indicate that the in vivo priming with CD40L- and IFN-gamma gene-transduced lung cancer cells is a promising strategy for inducing antitumor immunity in the treatment of lung cancer.