Abstract 4492: Novel carborane based inhibitors of carbonic anhydrase IX

Carborane-based compounds have emerged as promising lead structures for the development of carbonic anhydrase (CA) inhibitors. The aim of this study is to evaluate the effect of new carboranes with functional sulfonamide residues on CAIX function. CAIX is a transmembrane isoform of carbonic anhydrase with an extracellular-facing catalytic site and therefore is well positioned to act in the control of tumor pH, and is overexpressed in various solid tumors. The function of CAIX can be inhibited by CAIX selective sulphonamides, and the inhibition perturbs the in vitro survival, under hypoxia conditions. To investigate the potency of carboranes to inhibit CAIX, at cellular level, carboranes were chosen based on their enzymatic activity against CAIX and CAII, in vitro from a library of 28 new carboranes with sulphonamide residues (IOCB, ASCR, v.v.i., Czech Republic). 3 carboranes, viz., CB-30, CB-31, and CB-33, showed high binding constant (10, 16, and 64 nM, respectively) and high selectivity for CAIX, and were studied further. Extracellular pH in cell cultures was measured, in parallel with measurements of cellular cytotoxicity, in both 2D and 3D culture systems. At cellular level, the activity of CB-30 and CB-31 was significantly lower than CB-33, due to increased binding of CB-30 (99.7%) and CB-31 (99.3%) to the plasma proteins, compared with 93.3% binding for CB-33. This indicated that the activity at cellular level was preserved by the free fraction of CB-33. Moreover, CB-33 comparatively showed the highest change of extracellular pH under hypoxia conditions, and decreased migration of cells in anti-metastatic assay. Further, we developed a new method of Raman spectroscopy, to locate the distribution of carboranes in cells, and determined the distribution pattern of CB-30 and CB-31 in HT-29 cells, under hypoxia conditions. The highest Raman signal was detected on the cell membrane. Furthermore, we investigated the pharmacodynamics and pharmacokinetics profiles of carboranes. All tested carboranes were distributed using ADME methods, according to their plasma and microsomal stability, plasma protein binding, and presence of passive and active transport in cells. The rapidity of drug metabolism, suitability of carboranes for per-oral administration, and the ability of carboranes to penetrate through the blood brain barrier were then evaluated. Pharmacokinetic analysis is currently ongoing process, based on which anti-tumor studies will be performed, followe