General discussion I

Kwan I want to describe some of the studies we have done in Glasgow. The hypothesis is that refractory epilepsy is associated with a localized overexpression of P glycoprotein (Pgp), which restricts the access of drugs to their intended site of action. To test this hypothesis we carried out two parallel sets of experiments. The first set involved gene expression studies. We measured the expression of Mdrla and Mdrlb by quantitative RT-PCR, using an internal standard to allow absolute...

Contribution to drug resistance by MRP5

MPR5 is ubiquitously expressed in almost all tissues with relatively high levels of gene transcripts in skeletal muscle and various segments of the brain (Kool et al 1997, McAleer et al 1999). MRP5 is an organic anion transporter of molecules such as glutathione conjugates (e.g. dinitrophenyl glutathione) and fluorochromes, and can be inhibited by typical inhibitors of organic anion transport (e.g. sulfinpyrazone, benzbromarone and probenecid) (Jedlitschky et al 2000, McAleer et al 1999,...

Reversal of multidrug resistance lessons from clinical oncology

Bates, Clara Chen, Robert Robey, Min Kang, William D. Figg and Tito Fojo Molecular Therapeutics Section, Medicine Branch, National Cancer Institute, Bethesda, MD 20892, USA Abstract. Modulation of P glycoprotein (Pgp) in clinical oncology has had limited success. Contributing factors have included the limitation in our understanding of the tumours in which Pgp overexpression is mechanistically important in clinical drug resistance the failure to prove that concentrations of modulators...

Drug resistance caused by multidrug resistanceassociated proteins

Department of Ophthalmogenetics, The Netherlands Ophthalmic Research Institute, Meibergdreef47, 1105 BA, Amsterdam and Divisioon oof Molecular Biology, The Netherlands Cancer lnstitute,Plesmanlaan 121, 1066 CX, Amsterdam, The Netherlands Abstract. Three types of drug efflux pumps, the multidrug resistance gene 1 (MDR1 or ABCB1)-encoded P glycoprotein, the multidrug resistance-associated protein (MRP or ABCC1) and breast cancer resistance protein (BCRP or ABCG2) may play an important part in the...

Drug resistance in epilepsy the role of the bloodbrain barrier

Khan, Christopher M. S. Rollinson, Andreas Reichel, Damir Janigro*, Stephen M. Dombrowski*, Michael S. Dobbie and David J. Begley Blood-Brain Barrier Research Group, Centre for Neuroscience Research, King's College London, London SE1 1UL, UK and * Cerebrovascular Research, Cleveland Clinic Foundation, Cleveland, Ohio OH 44195, USA Abstract. The blood brain barrier (BBB) is formed by the endothelial cells lining the brain microvessels. Complex tight junctions linking...

Expression and activity in vivo

Resistance Epileptic

Epidauros Biotechnology, Pharmacogenetics Laboratory, Am Neuland 1, 82347 Bernried, Germany A bstract. The human MDR1 gene encodes an integral membrane protein, P glycoprotein Pgp , whose function is the energy dependent export of substances from the inside of cells, and from membranes, to the outside. Its physiological role is the protection of cells from toxic substances or metabolites. Many drugs that have been developed for the treatment of human diseases are substrates of Pgp. Because of...

Positron emission tomography

Positron Emission Tomography Particles

To study the Pgp function in vivo, several invasive techniques, like Northern blotting and PCR for detection of mRNA , and immunohistochemistry and Western blotting for detection of the gene product are now available. However, due to ethical limitations, tumour biopsies are often hard to obtain. Furthermore, these methods give no insight in the dynamic function of Pgp. PET offers the possibility of probing the physiological function of Pgp in humans in vivo. PET can provide information on drug...

Protection of brain and testis tissues by MRP1

The contribution of the multidrug resistance gene 1 MDRI -encoded P glycoprotein to the protection of tissues e.g. brain has been clearly demonstrated. Analysis of mice lacking Mdrla established an important role for MDRla in the intestine, where it actively excretes drugs from the bloodstream into the intestinal lumen, and limits the entry of these MDR1 substrates from the intestinal lumen for review see Schinkel 1999 . The presence of MDR1 in the endothelium of the blood brain capillaries...

Limitations to use of Pgp modulation in oncology

There are several cautionary points that have to kept in mind when considering other potential roles for Pgp modulation, summarized below The therapeutic window may be narrow for Pgp modulation. The agents may change pharmacology of other drugs. The modulators may affect drug metabolism, particularly through cytochrome P450 effects. Multiple resistance mechanisms that are now recognized in cancer cells will limit efficacy of even the most potent Pgp modulator. Resistance to cytotoxic agents may...

Discussion

Ling When you get these tissue slices and put them on a plate, how long is this piece of tissue viable for Deisz The slices are stored in 100 ml beakers with artificial CSF which is equilibrated with carbogen at room temperature. Judging from the electrophysiological properties of the neurons, the slices are viable up to 24 h. Ling So 24 h later the tissue is giving the same pattern. Deisz-' Yes, the tissue seems stable for about 24 h. We ascertained the stability of the preparation in the...

References

Ambudkar SV, Lelong IH, Zhang J, Cardarelli CO, Gottesman MM, Pastan I 1992 Partial purification and reconstitution of the human multidrug-resistance pump characterization of the drug-stimulatable ATP hydrolysis. Proc Natl Acad Sci USA 89 8472 8476 Armstrong S, Tabemero L, Zhang H, Hermodsen M, Stauffacher C 1998 Powering the ABC transporters the 2.5A of the ABC domain of RbsA. Pediatr Pulmonol 17 91-93 Azzaria M, Schurr E, Gros P 1989 Discrete mutations introduced in the predicted...

The problem of the drugresistant epilepsies

Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK Abstract. For the majority of patients with epilepsy, the prognosis for seizure control is very good however, refractory epilepsy develops in about 20 30 of patients and represents a significant challenge for both clinical management and for research. Physicians treating epilepsy are often asked to predict prognosis and make decisions about commencing and withdrawing treatment. Practice can be guided by...

Normal tissue expression of

High levels ofPgp expression are seen in the adrenal cortex, kidney, liver, placenta, colon, small bowel, brain blood brain barrier , testes, pancreas and haemopoietic precursor cells Fojo et al 1987, Goldstein et al 1989, Ro et al 1990 . Adrenal medulla, trachea, lung and prostate show moderate expression skin, muscle, heart, spleen, oesophagus, stomach, ovary, spinal cord and bone marrow have low levels of Pgp. Diverse functions have been ascribed to Pgp, including steroid transport in the...

The bloodbrain barrier

Bbb And Log

The blood brain barrier BBB consists of endothelial cells lining the capillaries in the brain. It is considered as an important barrier to drug transport into the brain. In contrast to the endothelium in other tissues, the endothelial cells in the brain are closely interconnected by tight junctions. Furthermore, the BBB contains very few fenestrations and pinocytosis is virtually absent. Consequently, only very small hydrophilic molecules can enter the brain by diffusion via tight junctions...

Results

Generation of epothilone A-resistant cells MDA 435 breast adenocarcinoma cells were incubated in the presence of 10 nM of epothilone A for 5 weeks. Although majority of the cells died, several clones survived the drug selection. Each clone was further expanded in media containing 10 nM epothilone A but only one clone expanded well resulting in a resistant cell line named EA10. EA10 cells served as founder cells for the selection of cells with higher epothilone A resistance that were maintained...

Use of Pgp modulators in oncology

Before considering specific Pgp modulators and clinical trials in oncology, it would be useful to review the general principles that governed early studies. Some of the earliest modulators first generation were agents that were in clinical use and were found to have a general mild Pgp modulating ability. Examples are verapamil, cyclosporin and tamoxifen. The advantages with these agents were the general familiarity with longer-term use and knowledge of chronic toxicity. Limitations were the...

Drug resistance in epilepsy human epilepsy

Harding , M. V. Squier and M. Thom Epilepsy Research Group, University Department of Clinical Neurology, Department of Neuropathology, Institute of Neurology, and Department of Neuropathology, Institute of Child Health, University College London, Queen Square, London, and Department of Neuropathology, Radcliffe Infirmary, Oxford, UK Abstract. The basis of drug resistance in human epilepsy is not understood. Parallels with resistance in cancer suggest that drug...

P glycoprotein and the mechanism of multidrug resistance

Andras Varadi , Gergely Szakacsf, Eva Bakos and Balazs Sarkadif lnstitute of Enzymology, Hungarian Academy of Sciences, Karolina ut 29, Budapest H-1113, and National Institute of Hematology and Immunology, Membrane Research Group of SEB-Hungarian Academy of Science, Daroczi ut 24, Budapest H-1113, Hungary Abstract. The human P glycoprotein Pgp MDR1 is an ATP-driven transporter for hydrophobic drugs and causes multidrug resistance in cancer. Our knowledge related to the mechanistic details of...

Imaging Pgp transport in vivo with PET

Radiolabelled MDR-associated cytostatic agents can be used to study drug efflux pumps in vivo. One example is colchicine, a Pgp substrate which is a naturally occurring alkaloid Ford amp Hait 1990 . Mehta et al 1992 studied the biodistribution of 3H colchicine in mice xenografted with Pgp-negative and Pgp-positive human neuroblastoma tumours Mehta et al 1992 . The pharmacokinetics of colchicine are possibly favourable for PET studies due to their limited metabolism Hunter amp Klaassen 1975 ....

OC144093 a novel P glycoprotein inhibitor for the enhancement of antiepileptic therapy

Newman, Ross Dixon and Barry Toyonaga Ontogen Corporation, 6451 El Camino Real, Carlsbad, CA 92009, USA Abstract. Inhibitors of P glycoprotein Pgp may be useful for the enhancement of blood-brain barrier penetration of anti-epileptic drugs AEDs . Due to polypharmacy and the need for chronic treatment, Pgp inhibitors used in epilepsy should be highly specific and non-toxic. In particular, it may be essential to use compounds that produce minimal inhibition of enzymes involved in...

Unverricht Lundeborg

Rasmussen's encephalitis 8 redundancy 96 97 relapse, after anti-epileptic drug withdrawal 10 predictors 9 spontaneous 17 remote symptomatic seizures 10 rhodamine efflux 23, 232 rifampin 184 river blindness 45 RNA isolation 121 rolandic epilepsy 6 schizophrenia, seizures following ECT 162 self-sustained limbic status epilepticus model 197,202 Sestamibi 91,96-97 severe myoclonic epilepsy of childhood tolerance to 164,165 single-gene defects 13,14 So324 76 sodium channel modulation 48, 155-156...

Pgp in cancer

Cancer Multidrug Resistance

Drug resistance, either intrinsic or acquired, is a frequently encountered problem in the failure of antineoplastic agents. Pgp, an efflux pump that extrudes hydrophobic cytotoxic drugs from cancer cells, plays a key role in multidrug resistance MDR and may contribute to clinical drug resistance. Pgp is a 170 kDa cell-surface glycoprotein, encoded for by the MDR1 gene Riordan amp Ling 1985 . The presence of MDR has been correlated with poor outcome in acute myeloid leukaemia, non-Hodgkin's...

Gene expression profiling of epothilone Aresistant cells

Peter Atadja, Yan Yan-Neale, Harry Towbin, Frank Buxton and Dalia Cohen Functional Genomics, Novartis Corporation, Summit, NJ 07901, USA Abstract. In the current study, we isolated sublines of the human breast adenocarcinoma cell line MDA 435 that exhibited increasing resistance to epothilone A, a microtubule-stabilizing cytotoxic agent. The resistant cells did not express P glycoprotein or multidrug resistance-associated protein MRP which are known mediators of multidrug resistance MDR . Two...

Oc144093

Pgp, with an average EC50 of 32 nM Table 1 . Full reversal of MDR is observed at doses between 0.25 and 1.0 mM Newman et al 2000 . OC144-093 is at least as potent as other third generation Pgp inhibitors currently in development, such as LY335979 Dantzig et al 1996, Starling et al 1997 and XR9576 Roe et al 1999, Mistry et al 1999 . OC144-093 is not cytotoxic by itself against 15 normal, non-transformed or tumour cell lines, regardless of Pgp status, with an average cytostatic IC50 of 460 mM,...