Are pharmaceuticals with evolutionary conserved molecular drug targets more potent to cause toxic effects in non-target organisms?

The ubiquitous use of pharmaceuticals has resulted in a continuous discharge into wastewater and pharmaceuticals and their metabolites are found in the environment. Due to their design towards specific drug targets, pharmaceuticals may be therapeutically active already at low environmental concentrations. Several human drug targets are evolutionary conserved in aquatic organisms, raising concerns about effects of these pharmaceuticals in non-target organisms. In this study, we hypothesized that the toxicity of a pharmaceutical towards a non-target invertebrate depends on the presence of the human drug target orthologs in this species. This was tested by assessing toxicity of pharmaceuticals with (miconazole and promethazine) and without (levonorgestrel) identified drug target orthologs in the cladoceran Daphnia magna. The toxicity was evaluated using general toxicity endpoints at individual (immobility, reproduction and development), biochemical (RNA and DNA content) and molecular (gene expression) levels. The results provide evidence for higher toxicity of miconazole and promethazine, i.e. the drugs with identified drug target orthologs. At the individual level, miconazole had the lowest effect concentrations for immobility and reproduction (0.3 and 0.022 mg L-1, respectively) followed by promethazine (1.6 and 0.18 mg L-1, respectively). At the biochemical level, individual RNA content was affected by miconazole and promethazine already at 0.0023 and 0.059 mg L-1, respectively. At the molecular level, gene expression for cuticle protein was significantly suppressed by exposure to both miconazole and promethazine; moreover, daphnids exposed to miconazole had significantly lower vitellogenin expression. Levonorgestrel did not have any effects on any endpoints in the concentrations tested. These results highlight the importance of considering drug target conservation in environmental risk assessments of pharmaceuticals.

An NMR investigation of the structure, function and role of the hERG channel selectivity filter in the long QT syndrome.

The human ether-a-go-go-related gene (hERG) voltage-gated K(+) channels are located in heart cell membranes and hold a unique selectivity filter (SF) amino acid sequence (SVGFG) as compared to other K(+) channels (TVGYG). The hERG provokes the acquired long QT syndrome (ALQTS) when blocked, as a side effect of drugs, leading to arrhythmia or heart failure. Its pore domain - including the SF - is believed to be a cardiotoxic drug target. In this study combining solution and solid-state NMR experiments we examine the structure and function of hERG's L(622)-K(638) segment which comprises the SF, as well as its role in the ALQTS using reported active drugs. We first show that the SF segment is unstructured in solution with and without K(+) ions in its surroundings, consistent with the expected flexibility required for the change between the different channel conductive states predicted by computational studies. We also show that the SF segment has the potential to perturb the membrane, but that the presence of K(+) ions cancels this interaction. The SF moiety appears to be a possible target for promethazine in the ALQTS mechanism, but not as much for bepridil, cetirizine, diphenhydramine and fluvoxamine. The membrane affinity of the SF is also affected by the presence of drugs which also perturb model DMPC-based membranes. These results thus suggest that the membrane could play a role in the ALQTS by promoting the access to transmembrane or intracellular targets on the hERG channel, or perturbing the lipid-protein synergy.

Necrosis caused by intra-arterial injection of promethazine: case report.

Promethazine injections have led to necrosis and gangrene of the distal upper extremity when inadvertently injected into an artery. There have been few case reports of this alarming complication in the literature. We report on 2 cases of intra-arterial promethazine injection that led to amputation.

Microcapsule-gel formulation of promethazine HCl for controlled nasal delivery: a motion sickness medication.

The current method of choice for astronauts to treat space motion sickness is an intra-muscular injection of promethazine hydrochloride (PMZ HCl) which is invasive and causes considerable local irritation and discomfort at the site of injection. Intra-nasal delivery is considered a feasible alternative route for administration of medications to treat space motion sickness. The purpose of this research is to develop a PMZ HCl formulation that can be administered intra-nasally without irritation (i.e. leukocyte infiltration) in the nasal epithelium when dosed at PMZ HCl concentrations greater than the cytotoxic limit. The biocompatibility of PMZ HCl was tested in vitro and was shown to be cytotoxic at concentrations greater than 10(-5) molar regardless of pH. A controlled-release microencapsulated dosage formulation was developed using spinning disk atomization and release rates for the PMZ HCl microcapsules were determined in phosphate buffered saline. An animal study was conducted to determine the irritation response of rat nasal mucosa when dosed with encapsulated and non-encapsulated PMZ HCl.

Development of a highly sensitive in vitro phototoxicity assay using the SkinEthic reconstructed human epidermis.

The reconstituted human epidermis model SkinEthic was used to evaluate the phototoxicity of topically applied chemicals. For comparison with published data, we first tested a library of 13 nonphototoxic (NPT) and phototoxic (PT) compounds, applied onto SkinEthic reconstituted human epidermal tissues, in a protocol as close as possible to the one described by Liebsch using another skin tissue model. The results showed that, under these nonoptimized conditions, the SkinEthic model was already able to fully discriminate between known NPT and PT compounds. Furthermore, these epidermal tissues being highly resistant to UVA irradiation, it was possible to increase irradiation by (at least) 3-fold without decrease in tissue viability. In such conditions, the phototoxicity assay is much more sensitive, so that the model is expected to be of great interest for the detection not only of strong but also of weak phototoxic compounds.

Peripheral nerve injection injury with antiemetic agents.

Antiemetics are widely used drugs, frequently administered to alleviate postoperative and postchemotherapeutic nausea and vomiting. While antiemetics do not induce peripheral neurotoxicity when administered systemically, it is not known whether peripheral nerve injury can occur as a result of inadvertent intraneural injection during intramuscular administration. The purpose of this study was to characterize the neurotoxic effect of three commonly used antiemetic agents (promethazine, dimenhydrinate, and prochlorperazine) as compared to saline in the rat sciatic nerve model. Intrafascicular and extrafascicular injection as well as direct application of the antiemetic drugs were performed. Nerves were harvested at 2 weeks postoperatively for histology and morphometry, with an additional sacrifice point at 8 weeks for the intrafascicular injection group. Injection injuries caused by antiemetic drugs differed depending on the agent injected and the location of injection. Extrafascicular injection and direct application caused no damage. Intrafascicular injection caused diffuse axonal injury in the promethazine and dimenhydrinate groups, while prochlorperazine caused only focal injury. Regeneration was prominent at 8 weeks in all intrafascicular injection groups in this rat model. Prochlorperazine thus appears to be less neurotoxic when injected intraneurally and should preferentially be used for intramuscular injections.

Photo hen's egg test: a model for phototoxicity.

The aim of this investigation was to establish a new model for phototoxicity which is more advanced than the widely used cultures of yeasts, bacteria or cells of various origin, and at the same time to avoid animal testing. We studied the extraembryonal vasculature of the incubated hen's egg. This model was originally introduced by toxicologists as an alternative to the rabbit's eye irritation test (Draize test). In the photo hen's egg test, substances are applied to the embryo's yolk-sac blood vessel system at a non-toxic concentration and are irradiated with 5 J/cm2 ultraviolet A (UVA) (320-400 nm). Promethazine, haematoporphyrin, ciprofloxacin and 8-methoxypsoralen were tested in this system. Death of the embryo, membrane discoloration and haemorrhage are parameters for phototoxic damage, which were recorded during an observation period of 24 h. These well-known phototoxic substances induced pronounced damage of the yolk-sac membrane and blood vessels which was not found in the controls (test substance alone, UVA alone or untreated) using a 2 x 2 factorial test design. The photo hen's egg test serves as a valid screening model for substances supposed to be photosensitizers owing to a phototoxic mechanism.

Sublethal effects of hexavalent chromium on the body growth rate and liver function enzymes of phenobarbitone-pretreated and promethazine-pretreated rabbits.

Hepatotoxic effects of chromium have been studied on the liver function enzymes of male New Zealand white rabbits, Oryctolagus cuniculus, with and without pretreatment with phenobarbitone (PB) and promethazine (PM). The total body weight was decreased under all experimental conditions. After PB administration (5 mg/kg body wt/day for 5 days), the serum glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), lactate dehydrogenase (LDH), and isocitrate dehydrogenase (ICDH) activities decreased 21%, 65%, 25%, and 37%, respectively, whereas the alkaline phosphatase (AP) activity increased 70%. After PM treatment (5 mg/kg body wt/day for 5 days) the serum GPT was inhibited 73%, whereas LDH activity was increased 37%. The hepatic GPT and AP activities decreased after PB (52% and 31%, respectively), and PM (48% and 44%, respectively) treatments, whereas the activities of LDH and ICDH increased (after PB: 817% and 109%, respectively, and after PM: 136% and 44%, respectively). Potassium dichromate, administered at a dose of 8 mg/kg body wt/day for 5 days, decreased serum GOT (44%), GPT (61%), LDH (63%), and AP (44%) activities. The hepatic GOT, GPT and AP activities were likewise decreased (86%, 51%, and 46%, respectively), whereas hepatic LDH and ICDH activities increased 667% and 193%, respectively. When administered to PB-pretreated animals, the serum GOT and AP activities were decreased (50% and 68%), whereas ICDH was increased (29%). The hepatic GOT, LDH, and ICDH activities increased 79%, 221%, and 130%, respectively. In the PM-pretreated animals, the chromium treatment inhibited the activities of serum GOT (48%), GPT (44%), and LDH (43%). The hepatic GPT, LDH, and ICDH activities increased 90%, 133%, and 52%, respectively.

Preclinical skin sensitization testing of antihistamines: guinea pig and local lymph node assay responses.

Preclinical test methods for allergic contact sensitivity have been widely used for sensitization hazard identification and, with consideration of human exposure conditions, have also been valuable tools for sensitization risk assessment. For many years, the guinea pig has been the test species of choice with a variety of test methods developed to assess the sensitization response. More recently the local lymph node assay (LLNA) in mice has been developed to provide a more objective index of sensitization potential. The standardized methods have proven to be very well suited to most situations in which potential skin sensitization of a chemical needs to be assessed before human exposure. A potential difficulty with all these relatively limited exposure preclinical test methods, however, is in the ability to detect weak contact allergens that prove to be significant clinical allergens due to chronic topical exposure, exposure to compromised skin, and/or highly exaggerated exposure through transdermal delivery. This has been shown with the transdermal drug clonidine and might also be the case for topical antihistamines. The latter are considered significant clinical contact allergens, although predictive preclinical test data are minimal or lacking. A series of guinea pig (modified Buehler) tests with two common antihistamine compounds (triprolidine and diphenhydramine) and LLNA on these and two other compounds (chlorpheniramine and promethazine) was conducted. Positive Buehler test results required use of penetrating vehicle systems and a modified nine-induction patch regimen. Positive LLNA responses were obtained with all four materials (to varying degrees) only if the application site was pre-abraded or a penetrating vehicle (dimethylformamide) was used. These data support the notion that preclinical sensitization test methods can be modified to increase sensitivity. This may be critical for preclinical assessment of topical/transdermal drugs or other materials with chronic or high-concentration exposures in man.

Irreversible binding with biological macromolecules and effects in bacterial mutagenicity tests of the radical cation of promethazine and photoactivated promethazine. Comparison with chlorpromazine.

The irreversible binding of the radical cation of promethazine (PMZ+.) to DNA and protein in vitro and bacterial macromolecules in situ has been studied. Binding experiments were performed with synthesized [35S] promethazine. The results are compared to those with the chlorpromazine radical cation (CPZ+.). Secondary reaction products which result from fission of the alkylamino side chain are involved in the macromolecular binding of PMZ+. Compared to CPZ+. the covalent DNA binding of PMZ+. is significantly less. A larger amount of PMZ+. binds to single-stranded DNA than to double-stranded DNA. The extent of binding to proteins and RNA is of the same order as that of CPZ+. Bacterial mutagenicity tests show that the low genotoxicity of PMZ+. is related to the low DNA binding. The bacterial cytotoxicity is possibly related to the covalent protein binding. Similar results have been obtained with photoactivated promethazine (PMZ) and chlorpromazine (CPZ). The role of radical cations in the photosensitization and metabolic activation of phenothiazine drugs is discussed.

[Evaluation of the effect of the antihistaminic preparations dimebon and diprazin on embryogenesis in rats]. / Otsenka vliianiia antigistaminnykh preparatov dimebona i diprazina na émbriogenez krys.

A single administration of dimebon (800 mg/kg) to rats per os (2/3 of the LD50 isoeffective for females) raises the preimplantation death, whereas pipolphen increases the intrauterine lethality and inhibits the development of fetuses. Pipolphen in a dose of 175 mg/kg and dimebon in doses of 300-150 mg/kg (exceeding 25- and 300-150-fold, respectively, the therapeutic dose for man) do not exert any specific embryotropic action.

Critical biochemical functions of isolated hepatocytes as sensitive indicators of chemical toxicity.

Isolated hepatocytes from adult male Wistar rats are a suitable experimental model to study the cytotoxicity of chemicals. Indeed, the isolated cells incubated in suspension in a Waymouth medium supplemented with 10% newborn calf serum maintain critical biochemical functions such as cytochrome P-450-dependent monooxygenase activity, glycogen, and protein synthesis capacities. This cellular model is used to detect the early biochemical effects of various xenobiotics, i.e., chlorpromazine, promethazine, bromobenzene, paracetamol, and isoniazid. Both cellular lysis (measured by the LDH leakage) and metabolic competence of the hepatocytes (glycogen deposits and protein synthesis) are modified as a function of both the duration of exposure to, and the concentration of, the chemicals. These results point out that the evaluation of metabolic functions of isolated cells surviving in suspension might be a sensitive test to predict early cell injury. Indeed, changes in the cellular behavior may occur before or without cell death. Furthermore, since both the cytochrome P-450 content and its dependent monooxygenase activity together with critical biochemical functions of the isolated cells remain stable, this model is of significant interest in ascertaining the mechanisms of toxicity.

The Scandinavian multicenter photopatch study. Preliminary results.

The Scandinavian photopatch test procedure has been applied to 745 patients with suspected photodermatoses during the years 1980-1981. Our experience has been encouraging with the recording of several relevant reactions. A total of 132 positive photocontact reactions and 120 ordinary contact reactions were seen. Photocontact reactions to musk ambrette (19 cases) and PABA (19 cases) were surprisingly frequent. The next most common photocontact reactions were to promethazine (24), chlorpromazine (22 cases) and fentichlor (12). Ordinary contact reactions were observed to balsam of Peru (30), PABA (23), lichen mix (21), wood mix (14) and to perfume mix (10).

The effects of cimetidine and ranitidine on psychomotor function in healthy volunteers.

Single oral doses of cimetidine (400 mg), ranitidine (150 mg), promethazine (25 mg) or placebo were administered to 8 healthy volunteers in a double-blind study. Cimetidine and ranitidine did not cause any significant change in critical flicker frequency (c.f.f.), reaction time, pursuit rotor of the visual analogue scale scores for sedation. Promethazine significantly lowered c.f.f., prolonged reaction time and increased sedation when compared with placebo. It is concluded that in this study cimetidine and ranitidine had little, if any, effect on psychomotor function.