Fulvio Gualtieri

The functions and structure of ABC transporters: implications for the design of new inhibitors of Pgp and MRP1 to control multidrug resistance (MDR).

Multidrug resistance (MDR) is a kind of acquired resistance of microorganisms and cancer cells to chemotherapic drugs that are characterized by different chemical structure and different mechanism of action. Classic MDR is the consequence of the over-expression of a variety of proteins that extrude the chemotherapic from the cell, lowering its concentration below the effective one. The ABC (ATP Binding Cassette) is a ubiquitous and important family of such transporter proteins. Members of this super family are present in mammals as well as in prokaryotic organisms and use ATP as the energy source to activate the extrusion process. P-glycoprotein (Pgp) and Multidrug Resistance Proteins (MRP1 and sister proteins) are the most important and widely studied members of ABC super family. Our knowledge about the structures and functions of transporter proteins has definitely improved in recent years, following the resolution of the structure of bacterial pumps which opened the way to the building of homology models for the more complex Pgp and MRP. It can be anticipated that these results will have a strong impact on the design of more potent and safer MDR reverters. A huge number of small molecules, many of natural origin, are able to reverse multidrug resistance by inhibiting the functions of Pgp, MRP1 and sister proteins and their action has been considered a possible way to reverse MDR. However, while a few compounds have reached clinical trials, none of them has, so far, been cleared for therapeutic use. Two main reasons are at the base of this difficulty: i) MDR is a complex phenomenon that may arise from several different biochemical mechanisms, with the consequence that inhibition of transporter proteins may be insufficient to reverse it; ii) the physiological role of Pgp and sister proteins requires more potent modulators with proper selectivity and pharmacokinetic in order to avoid unwanted side effects. This paper first reviews the most recent discoveries on the structures and functions of the ABC super family, in particular Pgp and MRP. Then, the medicinal chemistry of MDR reverters, in light of these findings, is discussed and the molecules that are presently in development are reviewed.

The medicinal chemistry of multidrug resistance (MDR) reversing drugs

Multidrug resistance (MDR) is a kind of resistance of cancer cells to multiple classes of chemotherapic drugs that can be structurally and mechanistically unrelated. Classical MDR regards altered membrane transport that results in lower cell concentrations of cytotoxic drug and is related to the over expression of a variety of proteins that act as ATP-dependent extrusion pumps. P-glycoprotein (Pgp) and multidrug resistance protein (MRP1) are the most important and widely studied members of the family that belongs to the ABC superfamily of transporters. It is apparent that, besides their role in cancer cell resistance, these proteins have multiple physiological functions as well, since they are expressed also in many important non-tumoural tissues and are largely present in prokaryotic organisms. A number of drugs have been identified which are able to reverse the effects of Pgp, MRPI and sister proteins, on multidrug resistance. The first MDR modulators discovered and studied in clinical trials were endowed with definite pharmacological actions so that the doses required to overcome MDR were associated with unacceptably high side effects. As a consequence, much attention has been focused on developing more potent and selective modulators with proper potency, selectivity and pharmacokinetics that can be used at lower doses. Several novel MDR reversing agents (also known as chemosensitisers) are currently undergoing clinical evaluation for the treatment of resistant tumours. This review is concerned with the medicinal chemistry of MDR reversers, with particular attention to the drugs that are presently in development.

Central nicotinic receptors: structure, function, ligands, and therapeutic potential.

The growing interest in nicotinic receptors, because of their wide expression in neuronal and non‐neuronal tissues and their involvement in several important CNS pathologies, has stimulated the synthesis of a high number of ligands able to modulate their function. These membrane proteins appear to be highly heterogeneous, and still only incomplete information is available on their structure, subunit composition, and stoichiometry. This is due to the lack of selective ligands to study the role of nAChR under physiological or pathological conditions; so far, only compounds showing selectivity between α4β2 and α7 receptors have been obtained. The nicotinic receptor ligands have been designed starting from lead compounds from natural sources such as nicotine, cytisine, or epibatidine, and, more recently, through the high‐throughput screening of chemical libraries. This review focuses on the structure of the new agonists, antagonists, and allosteric ligands of nicotinic receptors, it highlights the current knowledge on the binding site models as a molecular modeling approach to design new compounds, and it discusses the nAChR modulators which have entered clinical trials.

Design and Study of Piracetam-like Nootropics, Controversial Members of the Problematic Class of Cognition-Enhancing Drugs

Cognition enhancers are drugs able to facilitate attentional abilities and acquisition, storage and retrieval of information, and to attenuate the impairment of cognitive functions associated with head traumas, stroke, age and age-related pathologies. Development of cognition enhancers is still a difficult task because of complexity of the brain functions, poor predictivity of animal tests and lengthy and expensive clinical trials. After the early serendipitous discovery of first generation cognition enhancers, current research is based on a variety of working hypotheses, derived from the progress of knowledge in the neurobiopathology of cognitive processes. Among other classes of drugs, piracetam-like cognition enhancers (nootropics) have never reached general acceptance, in spite of their excellent tolerability and safety. In the present review, after a general discussion of the problems connected with the design and development of cognition enhancers, the class is examined in more detail. Reasons for the problems encountered by nootropics, compounds therapeutically available and those in development, their structure activity relationships and mechanisms of action are discussed. Recent developments which hopefully will lead to a revival of the class are reviewed.

Central cholinergic antinociception induced by 5HT4 agonists: BIMU 1 and BIMU 8

The antinociceptive effect of two 5-HT4 agonists, BIMU 1 and BIMU 8, were examined in mice and rats by using the hot-plate, abdominal constriction and paw-pressure tests. In both species, BIMU 1 (10-20 mg kg-1 s.c. and 40-60 mg kg-1 p.o. in mice; 20 mg kg-1 i.p. in rats) and BIMU 8 (20-30 mg kg-1 s.c. and 60 mg kg-1 p.o. in mice; 20 mg kg-1 i.p. in rats), produced significant antinociception which was prevented by atropine (5 mg kg-1 i.p.), hemicholinium-3 (1 microgram per mouse i.c.v.), SDZ 205-557 (10 mg kg-1 i.p.), GR 125487 (20 mg kg-1 i.p.) but not by naloxone (1 mg kg-1 i.p.), CGP 35348 (100 mg kg-1 i.p.) and reserpine (2 mg kg-1 i.p.). Moreover, BIMU 1 and BIMU 8 increase of pain threshold, is abolished by nucleus basalis magnocellularis (NBM) lesions in rats. SDZ 205-557 and GR 125487 which totally antagonized BIMU 1 and BIMU 8 antinociception did not modify morphine (7 mg kg-1 s.c.) or baclofen (4 mg kg-1 s.c.) antinociception. Intracerebroventricular injection in mice of BIMU 1 (3 micrograms per mouse) and BIMU 8 (10 micrograms per mouse), doses which were largely ineffective by parenteral routes, induces an antinociception whose intensity equaled that obtainable s.c., i.p. or p.o. In the antinociceptive dose-range, neither 5HT4 agonist impaired mice motor coordination evaluated by rota-rod test. On the basis of the above data, it can be postulated that BIMU 1 and BIMU 8 exerted an antinociceptive effect mediated by a central amplification of cholinergic transmission.

S-(-)-ET 126 : A POTENT AND SELECTIVE M1 ANTAGONIST IN VITRO AND IN VIVO

Abstract The pharmacological profile of the competitive muscarinic antagonist S-(−)-α-(hydroxymethyl)benzeneacetic acid 1-methyl-4-piperidinyl ester (S-(−)-ET 126) was evaluated on M 1 (rabbit vas deferens; pA 2 = 8.99), M2 (rat left atrium; pA 2 = 8.21) and M3 (rat ileum; pA 2 = 6.84) muscarinic receptors in comparison with pirenzepine. The drug shows a subtype selectivity ( M 1 M 2 = 8 ; M 1 M 3 = 178 ; M 2 M 3 = 22 ) that proposes it as a useful pharmacological tool for receptor studies. S-(−)-ET 126, like pirenzepine, prevents the antinociception induced by M 1 agonists (McN-A-343 and AF-102B). Unlike pirenzepine and spirotramine, the compound is able to cross the blood brain barrier which makes it useful for in vivo investigations.

Antisense 'knockdowns' of M1 receptors induces transient anterograde amnesia in mice

The effect on memory processes of inactivation of the M1 gene by an antisense oligodeoxyribonucleotide (aODN) was investigated in the mouse passive avoidance test. Mice received a single intracerebroventricular (i.c.v.) injection of M1 aODN (0.3, 1.0 or 2.0 nmol per injection), degenerated ODN (dODN) or vehicle on days 1, 4 and 7. An amnesic effect, comparable to that produced by antimuscarinic drugs, was observed 12, 24, 48 and 72 h after the last i.c.v. aODN injection, whereas dODN and vehicle, used as controls, did not produce any effect. Reduction in the entrance latency to the dark compartment induced by aODN disappeared 7 days after the end of aODN treatment, which indicates the absence of any irreversible damage or toxicity caused by aODN. Quantitative reverse transcription-polymerase chain reaction analysis demonstrated that a decrease in M1 mRNA levels occurred only in the aODN-treated group, being absent in all control groups. Furthermore, a reduction in M1 receptors was observed in the hippocampus of aODN-treated mice. Neither aODN, dODN nor vehicle produced any behavioral impairment of mice. These results indicate that the integrity and functionality of M1 receptors are fundamental in the modulation of memory processes.

Benzisoxazole and benzisothiazole analogs of auxin

Abstract In order to ascertain to what extent benzisoxazole and benzisothiazole can substitute the indole nucleus in auxin, several benzisoxazole and benzisothiazole analogs of indole-3-acetic acid were synthesized and their activity as plant hormones tested. The results of the slit-pea-stem curvature test are reported; among other compounds, 5-chloro-1, 2-benzisothiazole-3-acetic acid was the most active, showing about 330 per cent of the activity of indole-3-acetic acid.

Hybridized and isosteric analogues of N1-acetyl-N4-dimethyl-piperazinium iodide (ADMP) and N1-phenyl-N4-dimethyl-piperazinium iodide (DMPP) with central nicotinic action

A series of piperazine derivatives, obtained by hybridization of N1-acetyl-N4-dimethyl-piperazinium iodide (1, ADMP) and N1-phenyl-N4-dimethyl-piperazinium iodide (3, DMPP) or of the corresponding tertiary bases (2, 4) with arecoline (5) and arecolone (6) or by isosteric substitution of the phenyl ring of DMPP, has been synthesized. Hybridization afforded compounds that, both as tertiary bases and as iodomethylates, have no affinity for the nicotinic receptor. On the contrary, isosteric substitution gave compounds that maintain affinity for the receptor; among them, two tertiary bases (37, 38), show affinity in the nanomolar range for the nicotinic receptor. The pharmacological profile of these isomeric compounds is quite interesting as they present differences in their peripheral and central effects, suggesting that they interact with different subtypes of the nicotinic receptor.

Negative inotropic activity of para-substituted diethyl benzylphosphonates related to fostedil☆

Abstract Several p -substituted diethyl benzylphosphonates related to the calcium antagonist diethyl 4-(2-benzothiazolyl)benzylphosphonate (fostedil) have been studied. They produce a dose-dependent negative inotropic effect on left atrial muscle isolated from guinea pig heart. Some of the compounds are equipotent or slightly more potent than fostedil and diltiazem taken as reference drugs. Structure—activity relationships are discussed.