In Silico Approaches In Carcinogenicity Hazard Assessment: Current Status and Future Needs.

Historically, identifying carcinogens has relied primarily on tumor studies in rodents, which require enormous resources in both money and time. In silico models have been developed for predicting rodent carcinogens but have not yet found general regulatory acceptance, in part due to the lack of a generally accepted protocol for performing such an assessment as well as limitations in predictive performance and scope. There remains a need for additional, improved in silico carcinogenicity models, especially ones that are more human-relevant, for use in research and regulatory decision-making. As part of an international effort to develop in silico toxicological protocols, a consortium of toxicologists, computational scientists, and regulatory scientists across several industries and governmental agencies evaluated the extent to which in silico models exist for each of the recently defined 10 key characteristics (KCs) of carcinogens. This position paper summarizes the current status of in silico tools for the assessment of each KC and identifies the data gaps that need to be addressed before a comprehensive in silico carcinogenicity protocol can be developed for regulatory use.

Feasibility of 5-fluorouracil pharmacokinetic monitoring using the My-5FU PCM™ system in a quaternary oncology centre.

PURPOSE: 5-Fluorouracil (5FU) drug exposure correlates with treatment response and toxicity in cancer patients. Dosing is based upon body surface area which does not correlate with 5FU pharmacokinetics (PK)/pharmacodynamics. Therapeutic drug monitoring has enabled real-time 5FU dose adjustments: reducing toxicity with increased efficacy. The aim of this study was to assess feasibility of a 5FU monitoring service utilising a commercial kit in a quaternary cancer centre and to compare PK parameters to previously published studies. METHODS: Cancer patients receiving continuous infusional (CI) 5FU with ECOG PS 0-2, and adequate organ function, were eligible. Patients had blood samples taken at t = 0, mid infusion (if feasible) then 2 h pre infusion end. 5FU levels were measured using a commercial kit (My-5FU PCM™). A feasibility questionnaire was completed by trial nurses and toxicity data were recorded at baseline and at the commencement of the next cycle. 5FU pharmacokinetic exposure parameters were calculated. RESULTS: Twenty patients (12 male; 8 female), median age 62, (range 37-71) had samples taken. Twenty (100%) feasibility forms were available for assessment. Blood samples were taken at 51/69 (74%) specified time points. Ease of sample processing was recorded as easy in all 20 patients. Patient compliance with scheduled visits was 18/20 (90%). One form noted other difficulties with predicting end of infusion time. 19/20 patients had blood samples analysed. Mean measured 5FU AUC (0-Tlast) for 5FU 1 g/m2 with platinum: 35.8 h mg/L (range 28.56-44.26), mean Css 372.2 µg/L (range 297.5-461.0); 5FU 600 mg/m2 with platinum: 12.42 h mg/L (range 6.91-18.29), mean Css 111.0 µg/L (72.0-190.5) and 5FU 2400 mg/m2 as part of FOLFOX ± bevacizumab: 14.75 h mg/L (range 6.74-22.93), mean Css 320.70 µg/L (range 146.5-498.5). One patient had grade 4 neutropenia and one patient without PK parameters experienced febrile neutropenia (grade 4 neutropenia). Mucositis was observed in two patients: [5FU/platinum (1), grade 1, FOXFOX ± bevacizumab (1) grade 1]. Diarrhoea was reported in three patients [5FU/platinum (2) grade 1-2, FOXFOX ± bevacizumab (1) grade 1]. CONCLUSION: Therapeutic 5FU drug monitoring was feasible using commercial kits and analysers and hence warrants development as a routine standard of care in cancer patients. The variability in the 5FU exposure parameters is consistent with other studies using the My 5FU PCM kit.

Combination anti-CD137 and anti-CD40 antibody therapy in murine myc-driven hematological cancers.

In order to stimulate antigen presentation and T cell activity against cancer, we treated three different tumor models in mice with the monoclonal antibodies anti-CD40 plus anti-CD137 (BiMab). In a subcutaneous transplantable MC38 colon cancer model, there was significant enhancement in the survival of mice following BiMab treatment. Anti-CD40 has shown considerable success against lymphoma in previous studies by other investigators, and we also showed in this study that, in a model of Eµ-Myc lymphoma, there was a statistically significant enhancement of survival of mice following BiMab treatment. Following the success of the BiMab treatment in the previous two models, we wished to determine if it would be successful in a mouse model of multiple myeloma. Firstly, we tested a transplantable model of disease in which multiple myeloma cells derived from Vk*MYC mice were injected intravenously. A minor proportion of anti-CD137 and BiMab treated mice experienced prolongation of life beyond 250 days. Then we tested the therapy in a spontaneously occurring multiple myeloma model, in Vk*MYC transgenic mice. The majority of mice treated survived longer than control mice, although statistical significance was not demonstrated.

The drug vehicle and solvent N-methylpyrrolidone is an immunomodulator and antimyeloma compound.

N-methyl-2-pyrrolidone (NMP) is a common solvent and drug vehicle. We discovered unexpected antineoplastic and immunomodulatory activity of NMP in a cMYC-driven myeloma model. Coincident to this, NMP was identified as an acetyllysine mimetic and candidate bromodomain ligand. Accordingly, NMP-treated cells demonstrated transcriptional overlap with BET-bromodomain inhibition, including downregulation of cMYC and IRF4. NMP's immunomodulatory activity occurred at sub-BET inhibitory concentrations, and, despite phenotypic similarities to lenalidomide, its antimyeloma activity was independent of the IMiD targets cereblon and Ikaros-1/3. Thus, low-affinity yet broad-spectrum bromodomain inhibition by NMP mediates biologically potent, cereblon-independent immunomodulation and at higher doses targets malignant cells directly via BET antagonism. These data reveal that NMP is a functional acetyllysine mimetic with pleotropic antimyeloma and immunomodulatory activities. Our studies highlight the potential therapeutic benefits of NMP, the consequences of current human NMP exposures, and the need for reassessment of scientific literature where NMP was used as an "inert" drug-delivery vehicle.

Carboplatin dosing in ovarian cancer: problems and pitfalls.

OBJECTIVE: Carboplatin is one of the most effective chemotherapeutic drugs for the treatment of ovarian cancer. It has simple pharmacokinetics and a predictable toxicity profile. The dose can be calculated effectively based on a patient's renal function as defined by the glomerular filtration rate (GFR). The measurement of the GFR is best done using radioisotopes, but this is expensive and not widely available, so many centers use equations to estimate GFR based on serum creatinine and other easily measured data. Recent changes in the measurement of serum creatinine, and a move toward isotope dilution mass spectrometry standardized values, have highlighted the difficulty in safely and effectively calculating doses of carboplatin in patients with ovarian cancer. METHODS: We have evaluated the currently available evidence for the most common methods of estimating and measuring GFR. We explored the problems and pitfalls with using each of these methods or equations and examined the effects of small changes in clinical parameters and the effect on carboplatin dose. RESULTS: Previous studies evaluating carboplatin's toxicity and efficacy used various different methods of GFR estimation and older methods of creatinine measurement. These may not translate to use with newer laboratory methods and may result in higher delivered doses than anticipated. CONCLUSIONS: The lack of consistency in carboplatin dosing, and changing creatinine values are a cause for concern if patient toxicity is a possible outcome. The need for new studies using new standard methods that can be widely used are urgently required to provide clarity in this area.

Genotyping African haplotypes in ATM using a co-spotted single-base extension assay.

Human genetic analysis, including population genetic studies, increasingly calls for cost-effective, high-throughput methods for the rapid screening of single nucleotide polymorphisms (SNPs) across many individuals. The modified single-base extension assay described here (arrayed SBE) is a highly accurate and robust method for SNP genotyping that can deliver genotypes at 3.5 cents each, following PCR. Specifically, amino-modified probe/target pairs were prehybridized, then co-spotted in a microarray format prior to enzymatic addition of allele-specific nucleotides. Probe/target identity was determined solely by its physical location on the array rather than by hybridization to a complementary target, resulting in a call rate of 99-100%. These innovations result in an inexpensive, accurate assay with exceptional signal-to-noise ratios, depending on the glass surface employed. Comparison of glass slides from three different manufacturers indicated that aldehyde-based Zyomyx slides provided superior performance for this assay. Arrayed SBE was applied to study the geographic distribution of three African-specific haplotypes in the human ATM gene. Four selectively neutral markers, which define the haplotypes H5, H6, and H7, were screened in a total of 415 individuals. Region-specific haplotype frequencies were consistent with patterns of human migration across and outside of Africa, suggesting a possible haplotype origin in East Africa. Arrayed SBE was a robust tool for this analysis that could be applied to any situation requiring the genotyping of a few SNPs in many individuals.