Breakthrough Cancer Pain Clinical Features and Differential Opioids Response: A Machine Learning Approach in Patients With Cancer From the IOPS-MS Study.

PURPOSE: A large proportion of patients with cancer suffer from breakthrough cancer pain (BTcP). Several unmet clinical needs concerning BTcP treatment, such as optimal opioid dosages, are being investigated. In this analysis the hypothesis, we explore with an unsupervised learning algorithm whether distinct subtypes of BTcP exist and whether they can provide new insights into clinical practice. METHODS: Partitioning around a k-medoids algorithm on a large data set of patients with BTcP, previously collected by the Italian Oncologic Pain Survey group, was used to identify possible subgroups of BTcP. Resulting clusters were analyzed in terms of BTcP therapy satisfaction, clinical features, and use of basal pain and rapid-onset opioids. Opioid dosages were converted to a unique scale and the BTcP opioids-to-basal pain opioids ratio was calculated for each patient. We used polynomial logistic regression to catch nonlinear relationships between therapy satisfaction and opioid use. RESULTS: Our algorithm identified 12 distinct BTcP clusters. Optimal BTcP opioids-to-basal pain opioids ratios differed across the clusters, ranging from 15% to 50%. The majority of clusters were linked to a peculiar association of certain drugs with therapy satisfaction or dissatisfaction. A free online tool was created for new patients' cluster computation to validate these clusters in future studies and provide handy indications for personalized BTcP therapy. CONCLUSION: This work proposes a classification for BTcP and identifies subgroups of patients with unique efficacy of different pain medications. This work supports the theory that the optimal dose of BTcP opioids depends on the dose of basal opioids and identifies novel values that are possibly useful for future trials. These results will allow us to target BTcP therapy on the basis of patient characteristics and to define a precision medicine strategy also for supportive care.

RNA sequencing reveals PNN and KCNQ1OT1 as predictive biomarkers of clinical outcome in stage III colorectal cancer patients treated with adjuvant chemotherapy.

Five-year overall survival of stage III colorectal cancer (CRC) patients treated with standard adjuvant chemotherapy (ACHT) is highly variable. Genomic biomarkers and/or transcriptomic profiles identified lack of adequate validation. Aim of our study was to identify and validate molecular biomarkers predictive of ACHT response in stage III CRC patients by a transcriptomic approach. From a series of CRC patients who received ACHT, two stage III extreme cohorts (unfavorable vs. favorable prognosis) were selected. RNA-sequencing was performed from fresh frozen explants. Tumors were characterized for somatic mutations. Validation was performed in stage III CRC patients extracted from two GEO datasets. According to disease-free survival (DFS), 108 differentially expressed genes (104/4 up/downregulated in the unfavorable prognosis group) were identified. Among 104 upregulated genes, 42 belonged to olfactory signaling pathways, 62 were classified as pseudogenes (n = 17), uncharacterized noncoding RNA (n = 10), immune response genes (n = 4), microRNA (n = 1), cancer-related genes (n = 14) and cancer-unrelated genes (n = 16). Three out of four down-regulated genes were cancer-related. Mutational status (i.e., RAS, BRAF, PIK3CA) did not differ among the cohorts. In the validation cohort, multivariate analysis showed high PNN and KCNQ1OT1 expression predictive of shorter DFS in ACHT treated patients (p = 0.018 and p = 0.014, respectively); no difference was observed in untreated patients. This is the first study that identifies by a transcriptomic approach and validates PNN and KCNQ1OT1 as molecular biomarkers predictive of chemotherapy response in stage III CRC patients. After a further validation in an independent cohort, PNN and KCNQ1OT1 evaluation could be proposed to prospectively identify stage III CRC patients benefiting from ACHT.

Trabectedin in combination with pegylated liposomal doxorubicin in patients with ovarian tumors.

The majority of patients with ovarian cancer will experience relapse and thus require second-line therapy. While platinum-based therapies are the primary treatments for refractory disease other options are required, particularly for those with partially platinum-sensitive disease as their response rates are lower. Agents that can resensitize relapsed ovarian cancers to platinum, including trabectedin, are therefore of increasing interest. Trabectedin is a multitarget agent that has a complex, novel mechanism of action and has exhibited promising results in platinum-sensitive ovarian cancer when in combination with pegylated liposomal doxorubicin (PLD). The present study conducted retrospective analysis involving 11 cases (median age 60 years; range 45-75 years) of recurrent ovarian tumors and partial platinum sensitivity undergoing treatment with trabectedin + PLD. The cohort consisted of 7 serous carcinomas, 1 endometrial carcinoma, 2 undifferentiated carcinomas, and 1 mucinous carcinoma. Of the 11 patients, 4 exhibited a complete response, 3 achieved stable disease, and 4 had progression of disease. Mean overall survival was 32.42 months and median progression-free survival was 5.9 months. Trabectedin in combination with PLD was well tolerated in terms of gastrointestinal and hematological toxicity; Grade 3 cutaneous toxicity and grade 3 neutropenia were each observed in 18.2% of patients. There were no grade 4 events. Thus, the present study supports the use of trabectedin + PLD in patients with relapsed ovarian cancer and partial platinum sensitivity, with predictable and manageable toxicity.

Factors Influencing the Clinical Presentation of Breakthrough Pain in Cancer Patients.

Background: The aim of this study was to identify potential variables influencing the clinical presentation of breakthrough cancer pain (BTP). Methods: Cancer patients with a diagnosis of BTP were enrolled. Demographic and clinical characteristics, as well as background pain and BTP characteristics were collected. Multivariate analyses were conducted to assess the correlation between BTP characteristics and the variables examined. Results: Data of 4016 patients were analysed. Average daily number of BTP episodes was 2.4, mean intensity was 7.5, and a mean duration was 43.3 min. A short onset BTP was observed in 68.9% of patients. In 30.5% of patients BTP was predictable. There were 86.0% of participants who reported a marked interference of BTP with their daily activities. Furthermore, 86.8% of patients were receiving opioids for the management of BTP. The average time to meaningful pain relief was 16.5 min and 70.9% of patients were satisfied with their BTP medications. Age, head and neck cancer, Karnofsky, background pain intensity, predictable and fast onset BTP were independently associated with the number of BTP episodes. BTP pain intensity was independently associated with background pain intensity, fast onset BTP, and Karnofsky. Neuropathic pain mechanism was independently associated with unpredictable BTP. Variables independently associated with a longer duration of BTP were age, place of visit, cancer diagnosis, disease-oriented therapy, background pain intensity and mechanism, and unpredictable BTP. Age, Karnofsky, background pain intensity, fast onset, and long duration of BTP were independently associated with interference with daily activity. Conclusions: BTP has a variable presentation depending on interdependent relationships among its different characteristics.

Stable disease in a patient with metastatic leiomyosarcoma treated with trabectedin.

Leiomyosarcomas represent the most common variant of uterine sarcomas, and are also considered to be the least chemosensitive. To date, adriamycin and ifosfamide are believed to be the most effective drugs for its treatment, in addition to docetaxel and gemcitabine. Recently, the introduction of trabectedin has provided clinicians with another treatment option, and the drug may have some benefits for patients as it may allow for long-term treatment. We present the case of a patient who previously failed multiple cycles of chemotherapy and who was subsequently treated with 30 cycles of trabectedin as third-line therapy for multiple metastases of uterine leiomyosarcoma. During the treatment period, the dosage and dose interval of trabectedin were optimized because of the appearance of grade 4 hematological and gastrointestinal toxicity. Dose adjustments led to acceptable tolerability. Trabectedin was associated with a very good partial response, especially at the pulmonary and pancreatic levels, and stable disease was achieved at all metastatic sites. The patient is currently continuing treatment with trabectedin and has clinically stable disease after 2 years of therapy. This case report provides further evidence that trabectedin is a valid and well-tolerated therapeutic option that can be used in the long term in uterine leiomyosarcoma.

Breakthrough Cancer Pain: Preliminary Data of The Italian Oncologic Pain Multisetting Multicentric Survey (IOPS-MS).

INTRODUCTION: An ongoing national multicenter survey [Italian Oncologic Pain multiSetting Multicentric Survey (IOPS-MS)] is evaluating the characteristics of breakthrough cancer pain (BTP) in different clinical settings. Preliminary data from the first 1500 cancer patients with BTP enrolled in this study are presented here. METHODS: Thirty-two clinical centers are involved in the survey. A diagnosis of BTP was performed by a standard algorithm. Epidemiological data, Karnofsky index, stage of disease, presence and sites of metastases, ongoing oncologic treatment, and characteristics of background pain and BTP and their treatments were recorded. Background pain and BTP intensity were measured. Patients were also questioned about BTP predictability, BTP onset (≤10 or >10 min), BTP duration, background and BTP medications and their doses, time to meaningful pain relief after BTP medication, and satisfaction with BTP medication. The occurrence of adverse reactions was also assessed, as well as mucosal toxicity. RESULTS: Background pain was well controlled with opioid treatment (numerical rating scale 3.0 ± 1.1). Patients reported 2.5 ± 1.6 BTP episodes/day with a mean intensity of 7.5 ± 1.4 and duration of 43 ± 40 min; 977 patients (65.1%) reported non-predictable BTP, and 1076 patients (71.7%) reported a rapid onset of BTP (≤10 min). Higher patient satisfaction was reported by patients treated with fast onset opioids. CONCLUSIONS: These preliminary data underline that the standard algorithm used is a valid tool for a proper diagnosis of BTP in cancer patients. Moreover, rapid relief of pain is crucial for patients' satisfaction. The final IOPS-MS data are necessary to understand relationships between BTP characteristics and other clinical variables in oncologic patients. FUNDING: Molteni Farmaceutici, Italy.

Pharmacological aspects of the antibiotics used for urological diagnostic procedures.

Surgical antimicrobial prophylaxis is the use of an antibiotic before, during, or shortly after a urological procedure to prevent postoperative infections such as urinary tract or wound infection. The optimal antimicrobial drug must be microbiologically active against the most frequent potential pathogens and have good pharmacological properties. Correct timing of antimicrobial prophylaxis is the first critical issue in determining treatment efficacy. The antibiotic must be administered before the start of the surgical procedure in order to ensure a high tissue level at the time of microbial contamination. If using an oral antibiotic, this must be administered 1-3 hours before the operation and a parenteral antibiotic should be administered at the induction of anaesthesia. The antibiotics potentially useful for antimicrobial prophylaxis are the beta-lactams, cotrimoxazole, fluoroquinolones, and fosfomycin trometamol. The criteria for choosing the optimal antibiotic include an appropriate antimicrobial spectrum, favourable pharmacokinetic parameters (especially good tissue penetration), and elevated safety or tolerability. The use of cotrimoxazole must be restricted due to increasing chemoresistance. Unfortunately fluoroquinolone-based regimens, once the mainstay of prophylaxis guidelines, are increasingly ineffective due to a constant increase in multidrug-resistant (MDR) Gram-negative bacteria. The same concerns apply with regard to the second and third generation cephalosporins that have problems of resistance and, if administered orally, do not sufficiently penetrate prostatic tissue. An appropriate beta-lactam could be an aminopenicillin combined with a beta-lactamase inhibitor. Fosfomycin trometamol can also be a good potential choice due to its elevated activity against MDR Gram-negative bacteria and its favourable pharmacokinetic parameters, including an elevated penetration into prostatic tissue.

Overcoming tumor multidrug resistance using drugs able to evade P-glycoprotein or to exploit its expression.

Multidrug resistance (MDR) is a major obstacle to the effective treatment of cancer. Cellular overproduction of P-glycoprotein (P-gp), which acts as an efflux pump for various anticancer drugs (e.g. anthracyclines, Vinca alkaloids, taxanes, epipodophyllotoxins, and some of the newer antitumor drugs) is one of the more relevant mechanisms underlying MDR. P-gp belongs to the superfamily of ATP-binding cassette transporters and is encoded by the ABCB1 gene. Its overexpression in cancer cells has become a therapeutic target for circumventing MDR. As an alternative to the classical pharmacological strategy of the coadministration of pump inhibitors and cytotoxic substrates of P-gp and to other approaches applied in experimental tumor models (e.g. P-gp-targeting antibodies, ABCB1 gene silencing strategies, and transcriptional modulators) and in the clinical setting (e.g. incapsulation of P-gp substrate anticancer drugs into liposomes or nanoparticles), a more intriguing strategy for circumventing MDR is represented by the development of new anticancer drugs which are not substrates of P-gp (e.g. epothilones, second- and third-generation taxanes and other microtubule modulators, topoisomerase inhibitors). Some of these drugs have already been tested in clinical trials and, in most of cases, show relevant activity in patients previously treated with anticancer agents which are substrates of P-gp. Of these drugs, ixabepilone, an epothilone, was approved in the United States for the treatment of breast cancer patients pretreated with an anthracycline and a taxane. Another innovative approach is the use of molecules whose activity takes advantage of the overexpression of P-gp. The possibility of overcoming MDR using the latter two approaches is reviewed herein.

Thymidylate synthase expression and genotype have no major impact on the clinical outcome of colorectal cancer patients treated with 5-fluorouracil.

BACKGROUND AND OBJECTIVES: Thymidylate synthase (TS) expression levels appear to be related to response to 5-fluorouracil-(5-FU)-based chemotherapy in colorectal cancer (CRC) patients. Three polymorphisms have been proposed as modulators of TS expression: a tandemly repeated sequence (2R/3R) in the 5' UTR, a SNP (G>C) within the 3R allele and a 6bp deletion in the 3' UTR. To evaluate the influence of TS expression and polymorphisms on clinical outcome of 5-FU-treated patients we performed a comprehensive genetic analysis on 63 CRC patients. METHODS: TS expression levels were analyzed in normal and tumor tissues. TS coding sequence and UTR polymorphisms were investigated on DNA from normal tissue. LOH analysis was performed to determine tumor genotype. RESULTS: A difference in disease-free survival (DFS), although not statistically significant, was observed between high and low mRNA expression levels: patients with low levels showed longer DFS. The 2R2R genotype showed significantly lower expression than the 3R3R and 2R3R genotypes in normal tissue. No other TS polymorphism was associated with mRNA expression or clinical outcome. CONCLUSIONS: The results obtained in this pilot study indicate that the number of 5' UTR repeats is the major genetic determinant of TS expression. The lack of association with other polymorphisms might be partially explained by the existence of linkage disequilibrium in the TS gene. Our data support the growing evidence that TS control may require multiple mechanisms acting in close coordination with one another and suggest that TS genotyping alone in tumor samples is not sufficient to accurately predict response to 5-FU.

Identification of potential pharmacogenomic markers of clinical efficacy of 5-fluorouracil in colorectal cancer.

Although adjuvant chemotherapy has significantly increased overall survival in resected Stage III colorectal cancer, disease recurrence is still high (30-40%). 20-25% of Stage II patients also develop recurrent disease. Thus, high-risk patients may benefit from chemotherapy. As patient response to standard chemotherapy varies, the study of molecular differences in the expression of pharmacologically relevant genes may help clinicians to understand variability and tailor therapy. The expression of 5-fluorouracil (5-FU) pathway genes in tumors from 53 Stages II-III colorectal cancer patients who underwent 5-FU adjuvant chemotherapy was investigated by reverse transcription quantitative real-time polymerase chain reaction. Patients were dichotomized into high- and low-mRNA expression level groups using median values of gene mRNA levels. Then, a threshold analysis to identify a cut-off distinguishing recurrent- or nonrecurrent-disease was used. A high degree of interpatient variation in relative tumor expression of study genes was observed. Multiple gene correlations were found, which suggest possible coregulation mechanisms. No statistically significant relationship between experimental data and baseline clinical/pathological characteristics or clinical outcome was observed using gene expression median values. Threshold analysis indicated significant inverse relationships between deoxyuridine triphosphatase (DUT), ferrodoxin reductase (FDXR) or tumor protein p53 (TP53) and disease-free survival (DFS) in the entire case series and between DUT or NM23-H1 and DFS in Stage III patients: higher gene expression was associated with shorter DFS. This study provides data on relationships between expression of 5-FU pathway genes and clinical outcome of colorectal cancer patients undergoing 5-FU adjuvant chemotherapy and underscores the predictive role of specific genes. Validation in an independent case series is warranted.

The ARESC study: an international survey on the antimicrobial resistance of pathogens involved in uncomplicated urinary tract infections.

The ARESC (Antimicrobial Resistance Epidemiological Survey on Cystitis) study is an international survey to investigate the prevalence and susceptibility of pathogens causing cystitis. Female patients (n=4264) aged 18-65 years with symptoms of uncomplicated cystitis were consecutively enrolled in nine European countries as well as Brazil during 2003-2006. Pathogens were identified and their susceptibility to nine antimicrobials was determined. Escherichia coli accounted for 76.7% of isolates. Among E. coli, 10.3% of the isolates were resistant to at last three different classes of antimicrobial agents. Resistance was most common to ampicillin (48.3%), trimethoprim/sulfamethoxazole (29.4%) and nalidixic acid (18.6%). Fosfomycin, mecillinam and nitrofurantoin were the most active drugs (98.1%, 95.8% and 95.2% susceptible strains, respectively) followed by ciprofloxacin, amoxicillin/clavulanic acid and cefuroxime (91.7%, 82.5% and 82.4%, respectively). Resistance to ciprofloxacin was >10% in Brazil, Spain, Italy and Russia. Overall, Proteus mirabilis were more susceptible to beta-lactams and less susceptible to non-beta-lactams than E. coli, whereas Klebsiella pneumoniae strains, which are intrinsically resistant to ampicillin, were less susceptible to mecillinam (88.8%), fosfomycin (87.9%), cefuroxime (78.6%) and nitrofurantoin (17.7%). Resistance was rare in Staphylococcus saprophyticus, with the exception of ampicillin (36.4%) and trimethoprim/sulfamethoxazole (10.2%). In Italy, Spain, Brazil and Russia, the countries most affected by antimicrobial resistance, extended-spectrum beta-lactamase (ESBL) enzymes (mainly CTX-M type) were detected in 48 strains (39 E. coli, 6 K. pneumoniae and 3 P. mirabilis). Despite wide intercountry variability in bacterial susceptibility rates to the other antimicrobials tested, fosfomycin and mecillinam have preserved their in vitro activity in all countries investigated against the most common uropathogens.

Ertapenem peritoneal fluid concentrations in adult surgical patients.

Ertapenem, a novel carbapenem, is approved for the treatment of mild to severe intra-abdominal infections (IAIs), although its in vivo concentrations in peritoneal fluid are unknown. The purpose of this study was to determine the peritoneal fluid concentration of ertapenem after a single 1 g intravenous dose. After informed consent, 21 patients (9 females and 12 males; mean+/-standard deviation (S.D.) age 50.2+/-17.7 years) requiring intra-abdominal surgery were enrolled. Plasma and peritoneal fluid samples were taken at fixed times during surgery. Drug concentrations were determined by high-performance liquid chromatography (HPLC) with ultraviolet detection. Mean+/-S.D. ertapenem peritoneal fluid concentrations were 64.3+/-23.4 mg/L at 1h and 31.3+/-26.5 mg/L at 3 h after administration. The mean tissue/plasma ratio ranged from 46.7% to 83.1%. The mean peritoneal fluid concentrations were well above the MIC(90) (minimum inhibitory concentration for 90% of the organisms) for susceptible bacteria found in IAIs, especially Escherichia coli, viridans streptococci, Enterobacteriaceae, Klebsiella spp. and Bacteroides fragilis, during the entire sampling time. These pharmacokinetic results support the assumption that ertapenem might be suitable for the treatment of IAIs.

Surveillance study in Europe and Brazil on clinical aspects and Antimicrobial Resistance Epidemiology in Females with Cystitis (ARESC): implications for empiric therapy.

BACKGROUND: Uncomplicated cystitis in females is among the most frequent infections in community. OBJECTIVE: To determine clinical aspects, epidemiology, and antimicrobial susceptibility of uropathogens. INTERVENTION: Patients were investigated clinically and with urinalysis and urine culture. MEASUREMENTS: This survey started in 2003 and ended in 2006 including 68 centres in nine European countries and in Brazil. Female patients between 18 and 65 yr with symptoms of uncomplicated cystitis were consecutively enrolled and clinically evaluated. Uropathogens were identified and their susceptibility tested for nine antimicrobials. RESULTS AND LIMITATIONS: Clinical data of 4264 eligible patients were analysed. A positive urine culture was found in 74.6%. Within the 3018 pathogens, Escherichia coli (E. coli) was most frequent (76.7%), followed by Enterococcus faecalis (4.0%), Staphylococcus saprophyticus (3.6%), Klebsiella pneumoniae (3.5%), and Proteus mirabilis (3.5%). E. coli showed the highest rate of susceptibility to fosfomycin (98.1%) followed by mecillinam (95.8%), nitrofurantoin (95.2%), and ciprofloxacin (91.8%). The lowest rate was found for ampicillin (45.1%). For the total spectrum the order was fosfomycin (96.4%), mecillinam (95.9%), ciprofloxacin (90.3%), and nitrofurantoin (87.0%). In all countries a susceptibility rate to E. coli above 90% was found only for fosfomycin, mecillinam, and nitrofurantoin. The susceptibility rates varied significantly from country to country (p<0.0001), except for fosfomycin, mecillinam, and nitrofurantoin. CONCLUSIONS: Despite wide cross-country variability of bacterial susceptibility/resistance rates to the other antimicrobials tested, fosfomycin, mecillinam, and nitrofurantoin have preserved their in vitro activity in all countries investigated. They may represent good options for the empiric therapy of female patients with uncomplicated cystitis.

Relationships between promoter polymorphisms in the thymidylate synthase gene and mRNA levels in colorectal cancers.

Thymidylate synthase (TS) intratumoural expression may be a prognostic marker and predict outcome of 5-fluorouracil (5-FU)-based chemotherapy in colorectal cancer patients. The TS gene promoter enhancer region contains two different polymorphisms which can influence TS mRNA transcriptional and translational efficiency: a polymorphic tandem repeat sequence (2 or 3 repeats; 2R and 3R) and a single nucleotide polymorphism (SNP), G > C, within the second repeat of the 3R alleles. We studied the relationship between tumoural TS mRNA expression levels and TS gene polymorphisms in the colonic mucosa of 48 colorectal cancer patients. The 3R/3R genotype was characterised by higher TS mRNA levels in the tumour than the 2R/2R-2R/3R genotypes (P = 0.071). Regarding the relationship with the SNP polymorphism, a statistically significant difference in TS gene expression between the 3RG/3RG genotype and 2R/2R-2R/3RC-2R/3RG genotype subset was observed (P = 0.017). No statistically significant correlation was observed between experimental data and baseline clinical-pathological characteristics as well as clinical outcome in the relatively small patient series investigated. This is the first study reporting an association between the TS intra-repeat SNP and gene expression levels in colorectal cancer patients. These results suggest that in 3R/3R patients, the G > C polymorphism may be an important factor in determining TS mRNA expression levels, and warrant further investigation of the role of TS promoter polymorphisms as predictors of sensitivity to 5-FU-based chemotherapy in larger case series.

Cytotoxic activity of 3-nitropyrazolo[5,1-c][1,2,4]benzotriazine derivatives: a new series of anti-proliferative agents.

We report the synthesis and biological evaluation of a new series of 3-nitropyrazolo[5,1-c][1,2,4]benzotriazine derivatives (compounds 1-4) bearing appropriate substitutions in positions 7 and/or 8. The objective of this investigation was to study the effects of these substitutions on the cytotoxic activity of four new compounds against established human cancer cell lines (i.e. HT29 and HCT-8, colon carcinoma, MCF7, breast carcinoma, and A549, lung carcinoma cells). The inhibitory effects of compounds 1-4 on cell growth were assessed by the sulforhodamine B assay. Also, the effects of these compounds on cell cycle distribution of human colon carcinoma cells (HCT-8) were analyzed by flow cytometry. 3-Nitropyrazolo[5,1-c][1,2,4]benzotriazine derivatives displayed IC(50) values in the micromolar range on the growth of the four cell lines tested. Cell cycle perturbations induced on HCT-8 cells by study compounds at the IC(50) values consisted prevalently of a slight accumulation of cells in G(0)/G(1) phase and a slight decrease in G(2)/M phase. However, compound 3 induced a marked accumulation of cells into S phase with concomitant decrease in G(0)/G(1) and G(2)/M phases. Cytotoxicity data, compared to those obtained with 3-cyano-8-chloropyrazolo[5,1-c][1,2,4]benzotriazine 5-oxide (compound 5, NSC 683334) and other compounds previously synthesized in our laboratory, demonstrated a similar or even improved cytotoxic potency. Cell cycle perturbations caused by these compounds support the hypothesis that they may act by a direct or an indirect inhibition of DNA synthesis.

Molecular characterization of established human colon carcinoma cell lines (HCT-8) made resistant to 5-fluorouracil by different selection schedules.

To provide some insight into molecular mechanisms of 5 fluorouracil (5-FU) clinical resistance in colorectal cancer, we hypothesized that different in vitro exposure schedules of human colorectal cancer cell lines mimicking clinical infusion or bolus regimens could lead to differential gene expression. Resistant HCT-8 colon cancer cell lines (HCT-8/FUI/15R and HCT-8/FUB/2R) were selected from parental sensitive HCT-8 cells by long-term and short-term exposure schedules, respectively. Expression levels of the 437 genes evaluated by the Atlas Select cDNA Expression Human Tumor Array were not substantially different between HCT-8/FUB/2R and HCT-8 cell lines except for three genes downregulated in the resistant subline. Several genes were differentially expressed in HCT-8/FUI/15R cells compared to the parental cell line: 43 genes, including three chemoresistance-related genes, were upregulated, and three genes were downregulated. HCT-8/FUB/2R cells were substantially more resistant to 5-FU in comparison to HCT-8/FUI/15R cells after both 4- and 72-h exposures. No substantial differences were observed among resistant and parental cells in sensitivity to SN-38, the active metabolite of irinotecan, and oxaliplatin. Analysis of the mRNA levels of thymidylate synthase, thymidine phosphorylase, and bcl-2 genes evaluated by reverse transcription and real time PCR (RT-PCR) assay showed comparable results in resistant sublines and sensitive parental cells, whereas expression of the dihydropyrimidine dehydrogenase gene was markedly increased in both resistant cell lines compared to parental cells.

Clinical pharmacokinetics of depot leuprorelin.

Leuprorelin acetate is a synthetic agonist analogue of gonadotropin-releasing hormone. Continued leuprorelin administration results in suppression of gonadal steroid synthesis, resulting in pharmacological castration. Since leuprorelin is a peptide, it is orally inactive and generally given subcutaneously or intramuscularly. Sustained release parenteral depot formulations, in which the hydrophilic leuprorelin is entrapped in biodegradable highly lipophilic synthetic polymer microspheres, have been developed to avoid daily injections. The peptide drug is released from these depot formulations at a functionally constant daily rate for 1, 3 or 4 months, depending on the polymer type [polylactic/glycolic acid (PLGA) for a 1-month depot and polylactic acid (PLA) for depot of >2 months], with doses ranging between 3.75 and 30mg. Mean peak plasma leuprorelin concentrations (C(max)) of 13.1, 20.8 to 21.8, 47.4, 54.5 and 53 microg/L occur within 1 to 3 hours of depot subcutaneous administration of 3.75, 7.5, 11.25, 15 and 30 mg, respectively, compared with 32 to 35 microg/L at 36 to 60 min after a subcutaneous injection of 1mg of a non-depot formulation. Sustained drug release from the PLGA microspheres maintains plasma concentrations between 0.4 and 1.4 microg/L over 28 days after single 3.75, 7.5 or 15mg depot injections. Mean areas under the concentration-time curve (AUCs) are similar for subcutaneous or intravenous injection of short-acting leuprorelin 1mg; a significant dose-related increase in the AUC from 0 to 35 days is noted after depot injection of leuprorelin 3.75, 7.5 and 15mg. Mean volume of distribution of leuprorelin is 37L after a single subcutaneous injection of 1mg, and 36, 33 and 27L after depot administration of 3.75, 7.5 and 15mg, respectively. Total body clearance is 9.1 L/h and elimination half-life 3.6 hours after a subcutaneous 1mg injection; corresponding values after intravenous injection are 8.3 L/h and 2.9 hours. A 3-month depot PLA formulation of leuprorelin acetate 11.25mg ensures a C(max) of around 20 microg/L at 3 hours after subcutaneous injection, and continuous drug concentrations of 0.43 to 0.19 microg/L from day 7 until before the next injection. Recently, an implant that delivers leuprorelin for 1 year has been evaluated. Serum leuprorelin concentrations remained at a steady mean of 0.93 microg/L until week 52, suggesting zero-order drug release from the implant. In general, regular or depot leuprorelin treatment is well tolerated. Local reactions are more common after application of the 3- or 4-month depot in comparison with the 1-month depot.

Gold(III) complexes with bipyridyl ligands: solution chemistry, cytotoxicity, and DNA binding properties.

Gold(III) compounds generally exhibit significant cytotoxic effects on cancer cell lines and are of potential interest as antitumor drugs. We report here on the solution chemistry, the cytotoxicity, and the DNA binding properties of two new bipyridyl gold(III) compounds: [Au(bipy)(OH)(2)][PF(6)] (1) and the organometallic compound [Au(bipy(c)-H)(OH)][PF(6)] (2) (bipy(c) = 6-(1,1-dimethylbenzyl)-2,2'-bipyridine). Both compounds are sufficiently soluble, and stable for hours, within a physiological buffer at 37 degrees C; [Au(bipy)(OH)(2)][PF(6)], at variance with [Au(bipy(c)-H)(OH)][PF(6)], is quickly and quantitatively reduced by ascorbate. Both compounds showed relevant cytotoxic effects toward the A2780S, A2780R, and SKOV3 tumor cell lines; lower effects were detected on the CCRF-CEM/S and CCRF-CEM/R lines. In most cases the mechanisms of resistance to CDDP are only marginally effective against these gold(III) complexes. The interactions of [Au(bipy)(OH)(2)][PF(6)] and [Au(bipy(c)-H)(OH)][PF(6)] with calf thymus DNA were investigated in vitro by various techniques to establish whether DNA represents a primary target for these compounds. Addition of saturating amounts of DNA did not affect appreciably the visible spectra of these gold(III) complexes. Some slight modifications of the CD spectra of calf thymus DNA and of the DNA melting parameters were observed; in any case, ultrafiltration experiments showed that binding of these gold(III) complexes to DNA is weak and reversible. The mechanistic implications of these findings are discussed.