This study examined masitinib employing in vitro and in vivo models of human pancreatic cancer, both as an individual agent and in combination with gemcitabine, with the aim of establishing proof of principle. Molecular systems were CDK inhibition investigated via gene expression profiling. Masitinib was prepared from powder as a 10 or 20 mM stock solution in dimethyl sulfoxide and located at 280uC. Gemcitabine was obtained as a powder and dissolved in sterile 0. 9% NaCl solution and stored as aliquots at 280uC. New dilutions were prepared for every test. Pancreatic cancer cell lines were received from Dr. Juan Iovanna. Cells were managed in RPMI or DMEM medium containing Glutamax 1, supplemented with 100 U/ml penicillin, 100 mg/ml streptomycin, and 10% foetal calf serum. Expression of tyrosine kinases was dependant on RT PCR applying Hot Star Taq in a Thermal CDK4 inhibitor Cycler. All RT PCR primer sequences found in this study are listed in the Information. Mia Paca 2 cells were treated for 6 hours with increasing levels of masitinib in DMEM medium with 0. 5% serum. Cells were washed in PBS, then added to ice, and lysed in 200 ml of ice cold HNTG barrier in the presence of 100 mM Na3VO4 and protease inhibitors. Proteins were resolved by SDS PAGE 10%, adopted by western blotting and immunostaining. The next principal antibodies were used: rabbit anti phospho GRB2 antibody, and anti phosphotyrosine antibody. Primary antibodies were detected with 1:10,000 horseradish peroxidase conjugated anti rabbit antibody or 1:20,000 horseradish peroxidase conjugated anti mouse antibody. Immunoreactive bands were detected using superior chemiluminescent Infectious causes of cancer reagents. Cytotoxicity of masitinib and gemcitabine was evaluated utilizing a WST 1 proliferation/survival analysis in growth medium containing 1% FCS. Treatment was started with the addition of the appropriate drug. For combination treatment, cells were first resuspended in medium containing 0, 5 or 10 mM masitinib and incubated over night before gemcitabine improvement. After 72 hours, WST 1 reagent was added and incubated with the cells for 4 hours before absorbance measurement at 450 nm within an EL800 Universal Microplate Reader. Press alone was used as an empty and expansion in the absence of drug as a positive control served. Results are representative of three to four studies. The masitinib sensitisation index is the relation of the IC50 of gemcitabine against the IC50 of the drug combination. Male Nog SCID mice were obtained from an interior breeding program and were stored at the pet care product SCEA of the Celecoxib molecular weight Centre de Recherche en Cance?rologie de Marseille U891 under specific pathogen free circumstances at 2061uC in a 12 hour light/12 hour dark cycle and ad libitum access to food and filtered water. This study was accepted by the ethical review board at the Centre de Recherche en Cancerolgie de Marseille and carried out in compliance with the INSERM ethical recommendations of animal experimentation.