Medicinal Chemistry - Volume 1, Issue 6, 2005

NDP kinase catalyzes the last step in the phosphorylation of nucleotides. It is also involved in the activation by cellular kinases of nucleoside analogs used in antiviral therapies. Adenosine phosphonoacetic acid, a close analog of ADP already proposed as an inhibitor of ribonucleotide reductase, was found to be a poor substrate for human NDP kinase, as well as a weak inhibitor with an equilibrium dissociation constant of 0.6 mM to be compared to 0.025 mM for ADP. The X-ray structure of a complex of adenosine phosphonoacetic acid and the NDP kinase from Dictyostelium was determined to 2.0 Å resolution showing that the analog adopts a binding mode similar to ADP, but that no magnesium ion is present at the active site. As ACP may also interfere with other cellular kinases, its potential as a drug targeting NDP kinase or ribonucleotide reductase is likely to be limited due to strong side effects. The design of new molecules with a narrower specificity and a stronger affinity will benefit from the detailed knowledge of the complex ACP-NDP kinase.

Outer membrane pore proteins such as phosphoporin (PhoE) are important constituents of Gram-negative bacteria such as Escherichia coli. We have studied the interaction of PhoE with the membrane-forming lipids phosphatidylethanolamine (PE) and phosphatidylglycerol (PG) from the inner and lipopolysaccharide (LPS) from the outer leaflet of the outer membrane. These investigations comprise functional aspects of the protein:lipid interaction corresponding to the outer membrane system as well as the activity of LPS:PhoE complexes in the infected host after release from the bacterial surface. The interaction of the lipids PE, PG, and LPS with PhoE was investigated by analysing molecular groups in the lipids originating from the apolar region (methylene groups), the interface groups (ester), and polar groups (phosphates) applying Fourier-transform infrared spectroscopy (FTIR), and by analysing the phase transition behaviour of the lipids using FTIR and differential scanning calorimetry (DSC). The activity of PhoE and LPS:PhoE complexes was investigated in biological test systems (human mononuclear cells and Limulus amebocyte lysate assay) and with phospholipid model membranes using fluorescence resonance energy transfer spectroscopy (FRET). The results show a strong influence of PhoE on the mobility of the lipids leading to a considerable fluidization of the acyl chains of LPS, but much less to those from phospholipids: PhoE released from the outer membrane still contains slight contaminations of LPS, but its strong cytokine-inducing ability in mononuclear cells, which is not found in the LPSspecific Limulus amebocyte lysate test, indicates an LPS-independent mechanism of cell activation.

The complex-formation reactions of the platinum(II) complexes, [Pt(dien)H2O]2+, [PtCl(dien)]+ and [PtBr(dien)]+ (dien is diethylenetriamine) with some biologically relevant ligands such as inosine (INO), inosine-5'- monophosphate (5'-IMP), guanosine-5'-monophosphate (5'-GMP), glutathione (GSH) and L-methionine (S-meth), were studied by UV-Vis (UV-Visible) spectrophotometry and 1H NMR spectroscopy. Reactions of the [PtCl(dien)]+ with Lmethionine were studied in the presence and absence of 5'-GMP. The rate constants clearly showed a kinetic preference toward L-methionine. However, competitive reactions of [PtCl(dien)]+ with L-methionine and 5'-GMP demonstrated initially rapid formation of [Pt(dien)(S-meth)]2+ followed by displacement of L-methionine by 5'-GMP. In the later stages the concentration of [Pt(dien)(N7-GMP )]2+ is predominant. The results are analyzed in reference to the anti-tumour activity of Pt(II) complexes.

Multidrug resistance (MDR) is a major obstacle to successful application of cancer chemotherapy and also a basic problem in cancer biology. Studies on the molecular basis of MDR have revealed that a number of proteins over express in multidrug resistant cells viz., multidrug resistant MDR1 gene product P-glycoprotein, the multidrug resistanceassociated protein (MRP) and enzymes associated with the glutathione (GSH) metabolism. Decreased expression or altered activity of topoisomerase II has also been implicated in MDR. In the present investigation a number of changes in phase II detoxification parameters have been noticed in drug resistant cells but the novel aspect of the present report is the observation that the metal copper is involved in drug resistance. Although copper plays important roles in many human and other biological systems and even in the treatment of cancer but the relation of Cu and drug resistance has not so far been studied in detailed. The present report describes the novel findings that the level of copper increases with the development of drug resistance in Ehrlich ascites carcinoma and in Lewis lung carcinoma cells and also in serum of mice bearing drug resistant cancer cells compared to mice bearing drug sensitive cells; the work indicates the important aspect of treating drug resistant cancer patients by lowering Cu level in the cancerous cells and serum prior to treatment.

The Myxomycetes (true slime molds) are an unusual group of primitive organisms that may be assigned to one of the lowest classes of eukaryotes. As their fruit bodies are very small and it is very difficult to collect much quantity of slime molds, few studies have been made on the chemistry of myxomycetes. Cultivation of the plasmodium of myxomycetes in a practical scale for natural products chemistry studies is known only for very limited species. Here is described a review on the recent results on isolation of bioactive natural products from myxomycetes obtained in these two years in the laboratories. Spore germination experiments were studied of hundreds of field-collected myxomycetes collected in Japan and succeeded in laboratory culture of plasmodia of several myxomycetes in a practical scale for natural products chemistry studies. As a result, pyrroloiminoquinones, polyene yellow pigments, and a peptide lactone from cultured plasmodia of Didymium iridis, Physarum rigidum and P. melleum, respectively were isolated. New naphthoquinone pigments, cycloanthranilylprolines, tyrosine-kinase inhibitory bisindoles, and a cytotoxic triterpenoid aldehyde lactone were also isolated from field-collected fruit bodies of Cribraria purpurea, Fuligo candida, Tubifera casparyi, and Tubifera dimorphotheca, respectively.

The series of quinaldine derivatives were prepared, some of them by means of novel synthetic methods. The synthetic approach, analytical and spectroscopic data of all newly synthesized compounds are presented. The prepared compounds were tested for their in vitro antifungal activity as well as for their photosynthesis-inhibiting activity (the inhibition of photosynthetic electron transport in spinach chloroplasts (Spinacia oleracea L.) and the reduction of chlorophyll content in Chlorella vulgaris Beij.). Structure-activity relationships among the chemical structure, the physical properties and the biological activities of the evaluated compounds are discussed in the article.

A series of N-biarylalkyl-1-phenyl-1,3,8-triazaspiro[4.5]decan-4-ones were prepared and evaluated for biological activity at opioid (μ, δ, κ) and opioid receptor like-1 (ORL-1) G-protein coupled receptors. Substitution on the biaryl moiety produced enhanced affinity for the μ-opioid receptor.

Targeted radiotherapy using radiolabelled meta-iodobenzylguanidine (MIBG) is a promising treatment option for bladder cancer, restricting the effects of radiotherapy to malignant cells thereby increasing efficacy and decreasing morbidity of radiotherapy. We investigated the efficacy of a combined gene therapy and targeted radiotherapy approach for bladder cancer using radiolabelled MIBG. The effectiveness of alternative radiohalogens and alternative preparations of radiolabelled MIBG for this therapeutic strategy were compared. Bladder cancer cells, EJ138, were transfected with a gene encoding the noradrenaline transporter (NAT) under the control of a tumour specific telomerase promoter, enabling them to actively take up radiolabelled MIBG. This resulted in tumourspecific cell kill. Uptake and retention of radioactivity in cells transfected with the NAT gene were compared with that obtained in cells transfected with the sodium iodide symporter (NIS) gene. Substantially greater uptake and longer retention of radioactivity in NAT-transfected cells was observed. Carrier-added (c.a.) [131I]MIBG, no-carrier added (n.c.a.) [131I]MIBG, and [211At]-labelled benzylguanidine (i.e. [211At] meta-astatobenzylguanidine (MABG)) were compared with respect to efficiency of induction of cell kill. N.c.a[131I]MIBG was more cytotoxic than c.a.[131I]MIBG. However, the α- emitter [211At]MABG was, by three orders of magnitude, more effective in causing tumour cell kill than the β-emitter [131I]MIBG. We conclude that NAT gene transfer combined with the administration of n.c.a.[131I]MIBG or [211At]MABG, is a promising novel treatment approach for bladder cancer therapy.

As hospital reports of strains of resistant bacteria are continuing to increase, a new approach is required for the identification of small molecules with antibacterial activity. Natural products that bind covalently to their biological target have been largely unexplored, although in the field of cancer chemotherapy, such molecules have been shown to counter resistance developed through efflux mechanisms. The azinomycins are potent antitumour agents that alkylate DNA and one of the natural products, compound 1, is a mono-alkylator that has been reported to retain potent antitumour activity. All four diastereomers of 1 were synthesized via a route involving late stage introduction of the epoxide stereocentre and separation of the resulting compounds. A non-alkylating analogue and a potential alkylator that cannot intercalate were also made. All four diastereomers are potent antibacterial agents in cell lines containing efflux-based resistance mechanisms. MIC values in the range of 0.25-1.0 μg/ml were observed. Comparison with the antitumour activity of the compounds suggests that the antibacterial activity stems from a similar mechanism of action involving DNA alkylation. As the ultimate molecular target of the azinomycins is unknown, bacterial strains may represent an interesting route for the discovery of the downstream mechanisms affected by DNA alkylation.

Resveratrol, a polyphenolic phytoalexin, is a very effective antioxidant that also exhibits strong antiproliferative and anti-inflammatory properties. Recent studies have provided support for the use of resveratrol in human cancer chemoprevention, in combination with either chemotherapeutic drugs or cytotoxic factors for a most efficient treatment of drug refractory tumor cells. Resveratrol is also widely used in topical preparations, as a chemoprotective compound against development of several cutaneous disorders, including skin cancer. Nevertheless, the combined effect of resveratrol and UVA irradiation on cellular toxicity and DNA damage has never been assessed. The aim of this work was to investigate the effect of resveratrol on cell fate in immortalized human keratinocytes HaCaT cells. The results indicated that resveratrol potentiates the production of significant amounts of 8-oxo-7,8-dihydro-2'- deoxyguanosine in UVA-irradiated genomic DNA. Moreover, the combination of resveratrol with UVA significantly enhances the induction of DNA strand breaks and cell death in HaCaT keratinocytes. The conclusion is a potential hazardous effect of topical application of resveratrol, particularly on regions exposed to sunlight.

Fewer than one million HIV infected individuals are currently receiving antiretroviral therapy. Present antiretroviral therapy costs between $10,000 and $20,000 per year, which provides excellent value for money in developed countries with a cost of about $10,000 per life year saved; this compares very favourably with other therapies in chronic use. Recent studies have demonstrated a dramatic decline in HIV and AIDS related morbidity and mortality across developed countries and these reductions have been sustained since the introduction of highly active antiretroviral therapy (HAART) since 1996. The use of HAART has been associated with specific toxicities related to the drug class, problems with adherence with the subsequent emergence of viral isolates and resistance associated mutations. The replacement of older therapies with newer drugs that avoid cross resistance even within the same class of antiretroviral, represents a new hope in retroviral targeting.

The intestine is one of the major organs that are involved in sepsis. The inducible isoform of nitric oxide synthase (iNOS) is known to play a critical role in the pathogenesis of septic tissue injury by generating excess amount of nitric oxide (NO) in response to cytokines and endotoxin. In this study, we examined changes in gene expression of iNOS in various regions of the intestine as well as the distribution of iNOS protein in the intestinal cells in a rat model of endotoxemia produced by intraperitoneal injection of lipopolysaccharide (LPS; 10 mg/kg). While iNOS mRNA was undetectable in the intestine of untreated control animals, it underwent marked induction following LPS treatment. Induction of iNOS mRNA in the ileum was marked and biphasic, while it was also marked but monophasic in the jejunum. The induction of iNOS mRNA was maximal in the ileum. The administration of interleukin-6 (IL-6) upregulated intestinal iNOS gene expression specifically in the ileum. Consistent with enhanced iNOS gene expression, iNOS protein was markedly expressed in the ileum after LPS treatment, exclusively in the mucosal epithelium both at crypt and villus cells, although more prominently in the former. These findings suggested that intestinal iNOS expression was upregulated both at transcriptional and protein levels not only in a site-specific, but also in a cell typespecific manner in a rat model of endotoxemia, possibly through increasing serum IL-6 levels. Differential regulation of iNOS expression along the longitudinal and crypt-villus axes of the gut might be a determinant of the pattern of sepsisinduced intestinal damage.

The economic case for fundamental changes that are required to ensure long term viability of the pharmaceutical industry demands a close look at which compounds are advanced into clinical development. This perspective will cover recent efforts that have had the greatest influence on defining the optimal range of physical properties of compounds that are intended to act as human therapeutic agents. Our focus will be on models and properties that are most amenable to change via synthetic design, are potentially fixable in the lead optimization process, and have the greatest impact on overall attrition in clinical development. In particular, we will examine the optimal physicochemical properties for oral absorption based on solubility, permeability, and a few easily computed parameters. Additionally, the fate of compounds that have entered clinical trials provides a compelling case for adhering to the defined properties ranges. Finally, emerging data suggests that there has been a shift in the leading causes of compound attrition, and attention should now be focused on building toxicological models to guide drug discovery efforts.