| In classical phase-I trials with novel (investigational) anticancer drugs the agents are administered to patients starting with a very low dose (based on mouse toxicology data). Next, dose escalation is performed to find the highest dose that a patient can reasonably tolerate. Clinical toxicity is thus the pharmacodynamic study end-point. In oncology the belief persists that one can expect the highest antitumor activity of a drug when administered at the highest dose that a patient can tolerate. For alkylating drugs this approach makes sense. However, it can be seriously questioned whether this also holds for the many new anticancer drugs currently in the pipeline and which can be classified as 'biochemical modulators' e.g. the farnesyltransferase inhibitors (FTIs). Administration of a higher dose than necessary to block the involved biochemical pathway may not lead to a better antitumor activity, but exposes patients unnecessarily to unwanted side-effects. We will test the hypothesis that in dose-finding studies of biochemical modulators the drug target, or its biochemical effect is a better pharmacodynamic end-point than toxicity. FTIs, currently in clinical development, have been selected as model drugs. The pharmacodynamic, biochemical end-points to be tested in this project are the post-translational prenylation states of Rho and (mutant) Ras proteins as well as selected biomarker proteins during treatment with FTIs, including downstream MAPK and PI3-Akt pathways involved in apoptosis and cell survival. In our approach target (activated) proteins will be measured quantitatively by a novel advanced mass spectrometry method in vitro, in in vivo model systems and in patient tumor and surrogate tissues. PLAN OF INVESTIGATION: In the first, preclinical part of the project we will further optimize our analytical methodologies to measure the target proteins quantitatively in biological matrices. Advanced new mass spectrometric technologies e.g (micro/nano)-electrospray ionization (ESI) mass spectrometry, combined with selective sample pretreatment procedures will be used to analyze the prenylation status of the target proteins. In vitro models using cell lines with normal RhoB and normal and mutated K- or H-ras and in vivo models including K-ras and Alb-H-ras transgenic tumor bearing mice and K-ras containing Calu1 xenografts in nude mice will be used to quantitate in detail biochemical modulation of the Rho/Ras signalling proteins with FTIs. These investigations aim to identify dose/concentration effect (biochemically, antitumor activity, toxicity) relationships (pharmacokinetics-pharmacodynamics, PK-PD) and scheduling effects. Protein analyses will be performed both in tumor and normal tissues. Possibly, the determination of farnesylated proteins other than Rho and Ras can serve as a reliable substitute to estimate prenylation states, and this possibility will be investigated. Candidates include downstream elements Akt, ERK1/2, and chaperone protein HDJ-2, peroxisomal protein PxF, RhoE and intranuclear filament protein lamin A. In the second, clinical part of the project and based on the preclinical results we will test FTIs (e.g R115777, SCH66336 and AZD3409) in dose-escalating and activity trials in patients. Biopsies of accessible tumors and normal tissues (buccal smears and blood leukocytes) will be collected to identify PK-PD relationships. The outcome of this clinical investigation may serve as the basis for a randomized study comparing FTIs at biochemically optimal dosages and at maximum tolerated dosages. Pharmacokinetic drug monitoring will be essential in both the preclinical and clinical parts of the project. POSSIBLE RESULTS: This project, in which a quantitative method is developed for protein analysis, should result in the identification of new pharmacodynamic end-points to be used in the preclinical and clinical development of FTIs. This approach may also be of major importance for the clinical development of other agents that modulate signal transduction pathways. Finally, this non-precedent, innovative approach in anticancer drug treatment may optimize and rationalize clinical development of FTIs given as single agent or in combination and prevent patients from exposure to unnecessary high, toxic dosages. |
