基礎講座 電気泳動(Electrophoresis)

F. Vinyl Sulfones and Other Michael Acceptors 4683 G. Azodicarboxamides 4695 IV. Acylating Agents 4695 A. Aza-peptides 4695 B. Carbamates 4699 C. Peptidyl Acyl Hydroxamates 4700 D. â-Lactams and Related Inhibitors 4704 E. Heterocyclic Inhibitors 4714 1. Isocoumarins 4715 2. Benzoxazinones 4722 3. Saccharins 4725 4. Miscellaneous Heterocyclic Inhibitors 4728 V. Phosphonylation Agents 4728 A. Peptide Phosphonates 4728 B. Phosphonyl Fluorides 4734 VI. Sulfonylating Agents 4735 A. Sulfonyl Fluorides 4735 VII. Miscellaneous Inhibitors 4736 VIII. Summary and Perspectives 4737 IX. Acknowledgments 4740 X. Note Added in Proof 4740 XI. References 4740

Irreversible inhibitors of serine, cysteine, and threonine proteases.

Increased efforts in antimalarial drug discovery are urgently needed. The goal must be to develop safe and affordable new drugs to counter the spread of malaria parasites that are resistant to existing agents. Drug efficacy, pharmacology and toxicity are important parameters in the selection of compounds for development, yet little attempt has been made to review and standardize antimalarial drug-efficacy screens. Here, we suggest different in vitro and in vivo screens for antimalarial drug discovery and recommend a streamlined process for evaluating new compounds on the path from drug discovery to development.

https://www.mmv.org/sites/default/files/uploads/docs/publications/7_-_ScreeningArticlePDF_7.pdf

Antimalarial drug discovery: efficacy models for compound screening

Since the first reports of chloroquine-resistant falciparum malaria in southeast Asia and South America almost half a century ago, drug-resistant malaria has posed a major problem in malaria control. By the late 1980s, resistance to sulfadoxine-pyrimethamine and to mefloquine was also prevalent on the Thai-Cambodian and Thai-Myanmar (Thai-Burmese) borders, rendering them established multidrug-resistant (MDR) areas. Chloroquine resistance spread across Africa during the 1980s, and severe resistance is especially found in east Africa. As a result, more than ten African countries have switched their first-line drug to sulfadoxine-pyrimethamine. Of great concern is the fact that the efficacy of this drug in Africa is progressively deteriorating, especially in foci in east Africa, which are classified as emerging MDR areas. Urgent efforts are needed to lengthen the lifespan of sulfadoxine-pyrimethamine and to identify effective, affordable, alternative antimalarial regimens. Molecular markers for antimalarial resistance have been identified, including pfcrt polymorphisms associated with chloroquine resistance and dhfr and dhps polymorphisms associated with sulfadoxine-pyrimethamine resistance. Polymorphisms in pfmdr1 may also be associated with resistance to chloroquine, mefloquine, quinine, and artemisinin. Use of such genetic information for the early detection of resistance foci and future monitoring of drug-resistant malaria is a potentially useful epidemiological tool, in conjunction with the conventional in-vivo and in-vitro drug-sensitivity assessments. This review describes the various features of drug resistance in Plasmodium falciparum, including its determinants, current status in diverse geographical areas, molecular markers, and their implications.

https://www.thelancet.com/pdfs/journals/laninf/PIIS1473-3099(02)00239-6.pdf

Epidemiology of drug-resistant malaria.

Continued and sustainable improvements in antimalarial medicines through focused research and development are essential for the world's future ability to treat and control malaria. Unfortunately, malaria is a disease of poverty, and despite a wealth of scientific knowledge there is insufficient market incentive to generate the competitive, business-driven industrial antimalarial drug research and development that is normally needed to deliver new products. Mechanisms of partnering with industry have been established to overcome this obstacle and to open up and build on scientific opportunities for improved chemotherapy in the future.

Medical need, scientific opportunity and the drive for antimalarial drugs

http://www.jbc.org/content/275/37/29000.full.pdf

Characterization of native and recombinant falcipain-2, a principal trophozoite cysteine protease and essential hemoglobinase of Plasmodium falciparum.

In the malaria parasite Plasmodium falciparum, erythrocytic trophozoites hydrolyse haemoglobin to provide amino acids for parasite protein synthesis. Cysteine protease inhibitors block parasite haemoglobin hydrolysis and development, indicating that cysteine proteases are required for these processes. Three papain-family cysteine protease sequences have been identified in the P. falciparum genome, but the specific roles of their gene products and other plasmodial proteases in haemoglobin hydrolysis are uncertain. Falcipain-2 was recently identified as a principal trophozoite cysteine protease and potential drug target. The present study characterizes the related P. falciparum cysteine protease falcipain-3. As is the case with falcipain-2, falcipain-3 is expressed by trophozoites and appears to be located within the food vacuole, the site of haemoglobin hydrolysis. Both proteases require a reducing environment and acidic pH for optimal activity, and both prefer peptide substrates with leucine at the P(2) position. The proteases differ, however, in that falcipain-3 undergoes efficient processing to an active form only at acidic pH, is more active and stable at acidic pH, and has much lower specific activity against typical papain-family peptide substrates, but has greater activity against native haemoglobin. Thus falcipain-3 is a second P. falciparum haemoglobinase that is particularly suited for the hydrolysis of native haemoglobin in the acidic food vacuole. The redundancy of cysteine proteases may offer optimized hydrolysis of both native haemoglobin and globin peptides. Consideration of both proteases will be necessary to evaluate cysteine protease inhibitors as antimalarial drugs.

/pdf/expression-and-characterization-of-the-plasmodium-falciparum-1z3tf6s48o.pdf

Expression and characterization of the Plasmodium falciparum haemoglobinase falcipain-3.

The molecular mechanism of chloroquine resistance in Plasmodium falciparum remains uncertain. Polymorphisms in the pfcrt and pfmdr-1 genes have been associated with chloroquine resistance in vitro, although field studies are limited. In evaluations of known polymorphisms in parasites from patients with uncomplicated malaria in Kampala, Uganda, the presence of 8 pfcrt mutations and 2 pfmdr-1 mutations did not correlate with clinical response to therapy with chloroquine. Most notably, the pfcrt lysinerthreonine mutation at position 76, which recently correlated fully with chloroquine resistance in vitro, was present in 100% of 114 isolates, of which about half were from patients who recovered clinically after chloroquine therapy. These results suggest that, although key pfcrt polymorphisms may be necessary for the elaboration of resistance to chloroquine in areas with high levels of chloroquine resistance, other factors, such as host immunity, may contribute to clinical outcomes. Resistance of the malaria parasite Plasmodium falciparum to commonly used antimalarial agents is a large and growing problem. In particular, resistance to chloroquine, which remains the standard therapy for malaria in most of Africa, is an urgent concern [1]. The molecular basis of chloroquine resistance remains uncertain. Chloroquine resistance has been correlated with mutations in a number of P. falciparum genes, although some results have been inconsistent [2]. In initial studies of the P. falciparum multidrug resistance gene (pfmdr-1), an AsnrTyr mutation at amino acid 86 (N86Y) and other mutations in this gene correlated with chloroquine resistance [3]. However, in several field studies, associations between pfmdr-1 point mutations and in vivo or in vitro chloroquine resistance were not consistent [2]. More recently, transfection studies showed that the replacement of mutant pfmdr-1 with the wild-type sequence in resistant parasites decreased chloroquine resistance from high to moderate levels [4]. Thus, it appears that, although mutations

Polymorphisms in the Plasmodium falciparum pfcrt and pfmdr-1 Genes and Clinical Response to Chloroquine in Kampala, Uganda

Visual impairment associated with diabetes mellitus (DM) and diabetic eye disease may result from DME (DME), macular ischemia, vitreous hemorrhage, or diabetic tractional retinal detachment. Of all these causes, DME is the most common cause of vision loss in this patient population. Data from the Wisconsin Epidemiologic Study of Diabetic Retinopathy suggest that the 14-year incidence of DME is 26%. DME is clinically seen as thickening of the retina. The Early Treatment Diabetic Retinopathy Study has further classified it to assist physicians in decision-making regarding its treatment. On angiography, it is seen as leakage in the macula with dye pooling in areas of longstanding macular edema. The reasons for visual loss in DME are multifactorial and include light scattering, impaired cell-to-cell interaction and disturbances of normal ionic balances in the retina. The study of pathophysiology looks at the detailed malfunctioning that arises from or-alternately-causes disease. This article will attempt to explain the still elusive pathophysiology of DME.

Pathophysiology of diabetic macular edema.

Falcipain-2, a cysteine protease and essential hemoglobinase of Plasmodium falciparum, is a potential antimalarial drug target. We compared the falcipain-2 sequences and sensitivities to cysteine protease inhibitors of five parasite strains that differ markedly in sensitivity to established antimalarial drugs. The sequence of falcipain-2 was highly conserved, and the sensitivities of all of the strains to falcipain-2 inhibitors were very similar. Thus, cross-resistance between cysteine protease inhibitors and other antimalarial agents is not expected in parasites that are now circulating and falcipain-2 remains a promising chemotherapeutic target.

Ajay Singh

Papers

Characterization of native and recombinant falcipain-2, a principal trophozoite cysteine protease and essential hemoglobinase of Plasmodium falciparum.

Expression and characterization of the Plasmodium falciparum haemoglobinase falcipain-3.

Polymorphisms in the Plasmodium falciparum pfcrt and pfmdr-1 Genes and Clinical Response to Chloroquine in Kampala, Uganda

Pathophysiology of diabetic macular edema.

Comparison of Efficacies of Cysteine Protease Inhibitors against Five Strains of Plasmodium falciparum