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PubMed2005

The practice of Korean medicine: an overview of clinical trials in acupuncture.

Acupuncture, one of the Oriental medical therapeutic techniques that can be traced back at least 2500 years, is growing in popularity all over the world. Korea has continued to develop its own unique tradition of medicine throughout its long history, and has formed different types of acupuncture methods. The purpose of this review is to summarize clinical case studies in acupuncture and related therapies, such as acupressure, electric acupuncture, auricular acupuncture and moxibustion in Korea. A survey of Korean journals revealed that a total of 124 studies were published from 1983 to 2001. Results obtained from the survey showed that most clinical studies using acupuncture, electric acupuncture, moxibustion and other traditional therapies could alleviate a relatively broad range of medical problems. However, it should be emphasized that almost all clinical case studies published in various local journals did not follow the 'good clinical practice' with respect to regulatory aspects. Since they were not conducted using the randomized double-blinded controls with a large sample size, all the results should be considered as therapeutic indications. This review is an attempt to show the scope of acupuncture in our country and the kind of diseases, after many years of clinical experience, that were deemed valid targets for clinical trials.

Journal Article↗

PubMed1997

Cross-protection by anti-core glycolipid antibodies: evidence from animal experiments.

The ability of antibodies against the core glycolipid (CGL) of endotoxin to protect experimentally infected animals against death from Gram-negative sepsis is reviewed. The limitations and confounding factors inherent to animal models of sepsis are also briefly discussed. This review considers 30 studies in mice and 12 in other animal species that investigated protection against heterologous challenge by passive immunization with anti-CGL antibodies. In 28 (67%) of the reviewed studies antibodies were found to be protective, either prophylactically (n = 17) or therapeutically (n = 11). With the possible exception of the type of antibody preparation that was used (monoclonal versus polyclonal antibodies), none of the many differences in the experimental protocols were clearly correlated with success. Convincing proof is still lacking for any of the hypothetical mechanisms of protection by anti-CGL antibodies. Moreover, the evidence that protection by these antibodies is attributable to their anti-CGL specificity is poor. The available data raise serious questions about the validity of the concept underlying the search for broadly cross-protective antibodies raised against the core region of endotoxin. However, continuing research suggests that endotoxin still is a valid target in devising new adjunctive treatment strategies to improve the outcome of serious Gram-negative infections.

Animals↗

PubMed1998

Cationic porphyrins: novel delivery vehicles for antisense oligodeoxynucleotides.

Cationic porphyrins form stable complexes with oligodeoxynucleotides. To evaluate delivery, we used a 20mer phosphorothioate oligomer (Isis 3521) targeted to the 3'-untranslated region of the PKC-alpha mRNA, and complexed it with porphyrin. The expression of PKC-alpha protein and mRNA in T24 bladder carcinoma cells was reduced by approximately 80 +/- 10% at a concentration of oligomer of 3 microM, and 9 microM porphyrin. The expression of PKC-beta1, -delta and -straightepsilon isoforms was unaffected by this treatment, but elimination of PKC-zeta protein and mRNA were observed. However, treatment with the porphyrin complex of Isis 3522, an oligomer which is directed at the 5' coding region of the PKC-alpha mRNA, was equally effective as Isis 3521 with respect to PKC-alpha, but did not affect PKC-zeta protein or mRNA levels. Since Isis 3521 has an 11-base region of complementarity with the PKC-zeta mRNA, wheras Isis 3522 has only a 4-base region, the effect of Isis 3521 on PKC-zeta protein and mRNA expression may be due to irrelevant cleavage. Depending upon the desired application, this new strategy may offer several advantages over other methods of antisense oligodeoxynucleotide delivery including efficiency, stability, solubility, relatively low toxicity and serum compatibility. Porphyrins may thus be a potentially useful delivery vehicle for antisense therapeutics and/or target validation.

Energy Transfer↗

PubMed2001

Statistical prediction of single-stranded regions in RNA secondary structure and application to predicting effective antisense target sites and beyond.

Single-stranded regions in RNA secondary structure are important for RNA-RNA and RNA-protein interactions. We present a probability profile approach for the prediction of these regions based on a statistical algorithm for sampling RNA secondary structures. For the prediction of phylogenetically-determined single-stranded regions in secondary structures of representative RNA sequences, the probability profile offers substantial improvement over the minimum free energy structure. In designing antisense oligonucleotides, a practical problem is how to select a secondary structure for the target mRNA from the optimal structure(s) and many suboptimal structures with similar free energies. By summarizing the information from a statistical sample of probable secondary structures in a single plot, the probability profile not only presents a solution to this dilemma, but also reveals 'well-determined' single-stranded regions through the assignment of probabilities as measures of confidence in predictions. In antisense application to the rabbit beta-globin mRNA, a significant correlation between hybridization potential predicted by the probability profile and the degree of inhibition of in vitro translation suggests that the probability profile approach is valuable for the identification of effective antisense target sites. Coupling computational design with DNA-RNA array technique provides a rational, efficient framework for antisense oligonucleotide screening. This framework has the potential for high-throughput applications to functional genomics and drug target validation.

Algorithms↗

PubMed2006

Zinc Finger Tools: custom DNA-binding domains for transcription factors and nucleases.

Individual zinc finger (ZF) domains that recognize DNA triplets with high specificity and affinity can be used to create designer transcription factors and nucleases that are specific for nearly any site in the genome. These domains can be treated as modular units and assembled to create a polydactyl protein that recognizes extended DNA sequences. Deter-mination of valid target sites and the subsequent design of ZF proteins (ZFPs) is error-prone and not trivial, however. As a result, the use of ZFPs have been restricted primarily to those labs with the appropriate expertise. To address these limitations, we have created a user-friendly utility called Zinc Finger Tools (ZF Tools) that can be accessed at the URL http://www.zincfingertools.org. User-supplied DNA sequences can be searched for target sites appropriate for either gene regulation or nuclease targeting. Using a database of experimentally characterized zinc finger domains, the amino acid sequence for a ZFP expected to bind to any chosen target site can be generated. A reverse engineering utility is provided to predict the binding site for a ZFP of known sequence.

Binding Sites↗

PubMed2004

siRNA relieves chronic neuropathic pain.

Double stranded, short interfering RNAs (siRNA) of 21-22 nt length initiate a sequence-specific, post-trancriptional gene silencing in animals and plants known as RNA interference (RNAi). Here we show that RNAi can block a pathophysiological pain response and provide relief from neuropathic pain in a rat disease model by down regulating an endogenous, neuronally expressed gene. Rats, intrathecally infused with a 21 nt siRNA perfectly complementary to the pain-related cation-channel P2X3, showed diminished pain responses compared to missense (MS) siRNA-treated and untreated controls in models of both agonist-evoked pain and chronic neuropathic pain. This form of delivery caused no adverse effects in any of the animals receiving P2X3 siRNA, MS siRNA or vehicle. Molecular analysis of tissues revealed that P2X3 mRNA expressed in dorsal root ganglia, and P2X3 protein translocated into the dorsal horn of the spinal cord, were significantly diminished. These observations open a path toward use of siRNA as a genetic tool for drug target validation in the mammalian central nervous system, as well as for proof of concept studies and as therapeutic agents in man.

Animals↗

PubMed2006

Model for the complex between the insulin-like growth factor I and its receptor: towards designing antagonists for the IGF-1 receptor.

The type-1 insulin-like growth factor receptor (IGF-1R) is the cognate tyrosine kinase receptor for the insulin-like growth factor IGF-I and is expressed widely in many foetal and postnatal tissue cells. IGF-1R is overexpressed in a number of human tumour types and is a valid target for anti-cancer therapeutic efforts. Designing antagonists for IGF-1R would be greatly facilitated by the availability of structural information on the complex between IGF-I and IGF-1R. In the present work we model the three-dimensional structure of the complex between IGF-I and the first three domains of IGF-1R using a macromolecular docking method guided by selected experimental data. Interface metrics indicative of the binding affinity and reliability of the model are computed and compared with other biomolecular complexes. This model is consistent with experimental chimerical and mutagenesis data, provides a structural basis for understanding the primary interaction of IGF-I with its receptor and facilitates design of antagonist ligands.

Algorithms↗

PubMed2005

Developing drugs for cognitive impairment in schizophrenia.

There are strong data suggesting that improvement in the cognitive impairment associated with schizophrenia will contribute to enhanced functional outcomes for patients with this illness. Measurement and Treatment Research to Improve Cognition in Schizophrenia was established to provide a pathway for developing and registering potential cognitive-enhancing agents for this condition by addressing issues related to the content of the cognitive assessment battery and the clinical design features to be used in registration studies. This article examines key challenges related to the actual clinical development of cognitive-enhancing agents. These challenges include improving the probability of technical success and attrition rates of candidate molecules, establishing better animal models of human cognition, and developing biomarkers to decrease development costs and increase the speed of the clinical discovery process. Biomarkers are important for molecular target validation, dose selection, surrogate end points, and population segmentation. Examples of approaches for the development of agents for cognitive impairment associated with schizophrenia are discussed. It is concluded that close collaboration among academia, the National Institutes of Health, regulatory bodies, and industry will be important to advance the goal of developing drugs for this important condition.

Animals↗

PubMed2000

Telomerase inhibitors: targeting the vulnerable end of cancer?

In the past decade, a great deal has been learnt about the maintenance of telomeres in mammalian cells by the specialized reverse transcriptase, telomerase, and its associated proteins. The catalytic component of telomerase, hTERT, appears to be selectively activated in the vast majority of tumors relative to most somatic cells suggesting that its inhibition may result in antitumor effects. Although beset with some unusual issues as a drug target, recent 'target validation' studies using hTERT dominant-negative and antisense approaches strongly support the view that potent and selective telomerase inhibitors will induce inhibitory effects on tumors, especially in those possessing relatively short telomeres. Inhibitory strategies have focused on three main areas: antisense molecules (oligonucleotides, RNA molecules, ribozymes and peptide nucleic acids) directed against the hTR RNA component of telomerase, small molecule reverse transcriptase inhibitors (e.g. azidothymidine), and, probably most advanced, small molecules capable of interacting with and stabilizing four-stranded (G-quadruplex) structures formed by telomeres. G-quadruplex interactive agents that inhibit telomerase at sub-micromolar concentrations in cell-free assays have been described. Lead optimization and preclinical whole-cell and animal antitumor and pharmacology studies are now progressing which should result in the first generation of telomerase inhibitors being evaluated in the clinic within the next few years.

Animals↗

PubMed2004

Protein biomarkers and drug design for cancer treatments.

The development of new cancer treatments is quickly evolving away from traditional practices of the last 25 years. This change is occurring not only at the technical level, but also conceptually as the human genome is unravelled and decades of research contribute to our understanding of the molecular complexity of this disease. It is anticipated that disease initiation and progression is dictated by an understandable set of acquired capabilities. Knowledge of the molecular events associated with these acquired capabilities will allow the development of targeted agents coupled with new biomarkers for the prevention of cancer progression. This will have a profound influence on how drugs are developed, approved, and used by the medical community. The Food and Drug Administration (FDA) has over 400 Investigational New Drug (IND) applications for cancer in its portfolio, which increasingly involve molecular targets and genomic applications. However, only one-fifth of IND agents succeed in New Drug Application (NDA) and there is more expense and uncertainty around successful drug development than ever before. Biomarkers should help the success rate of INDs by enhancing the link between target and disease as well as in improving patient selection and monitoring response. In this review, we discuss how biomarkers can be used for target validation and pharmacodynamic modeling in preclinical drug discovery. We then explore the use of biomarkers in clinical development from proof of mechanism to proof of concept studies, as well as their use in the prevention setting.

Animals↗

PubMed2004

Identifying the combination of the transcriptional regulatory sequences on prostate specific antigen and human glandular kallikrein genes.

PURPOSE: The combination of prostate specific antigen (PSA) and human glandular kallikrein (KLK2) promoters and/or enhancers was used to establish a new model to determine the feasibility of tissue specific expression for prostate cancer. MATERIALS AND METHODS: In vitro studies used the construction of PSA and KLK2 promoters/enhancers vectors to elucidate the link between the promoter/enhancer of PSA and KLK2. Reverse transcriptase-polymerase chain reaction assays were used to determine cell specific expression. Therefore, an attractive tissue specific expression vector for PSA and KLK2 gene was identified. RESULTS: The reporter vectors driven by KLK2 promoter had much lower luciferase activities than those of the reporter vectors driven by PSA promoter in LNCaP cells. Furthermore, the most efficient and cell specific reporter activity after 5alpha-androstan-17beta-ol-3-one treatment among the reporter vectors constructed in this study was that of pKLK2EPSABHE, which was driven by KLK2 enhancer and PSA promoter/enhancer. The pKLK2EPSABHE reporter vector could induce 800-fold higher than the KLK2 basic promoter and its reporter activity was 16 times that of the enhancer/promoter element of KLK2 following induction by androgen. CONCLUSIONS: The results verify that the PSA promoter/enhancer must be combined with KLK2 to ensure the full activity and cell specificity of the gene. These expressions coupled with mechanic target validation yield valuable clues regarding the model of action of complex mixtures. This model is a potentially useful tool in gene therapy for metastatic prostate cancer.

Genes, Regulator↗

PubMed2002

Impact of insulin-like growth factor receptor-I function on angiogenesis, growth, and metastasis of colon cancer.

Insulin-like growth factors and their principal receptor, IGF-I receptor (IGF-IR), are frequently expressed in human colon cancers and play a role in preventing apoptosis, enhancing cell proliferation, and inducing expression of vascular endothelial growth factor (VEGF). The role of IGF-IR in regulating angiogenesis and metastases of human colon cancer has not been elucidated. To determine the in vitro and in vivo effects of IGF-IR in human colon cancer growth and angiogenesis, human KM12L4 colon cancer cells were transfected with a truncated dominant-negative form of IGF-IR (IGF-IR dom-neg). IGF-IR dom-neg-transfected cells demonstrated markedly decreased constitutive expression of VEGF mRNA and protein. Subcutaneous injections of IGF-IR dom-neg-transfected cells in nude mice led to significantly decreased tumor growth (p < 0.05) that was associated with decreased tumor cell proliferation, VEGF expression, and vessel count and with increased tumor cell apoptosis (p < 0.05 for all parameters compared with controls). In addition, pericyte coverage of endothelial cells was significantly decreased in tumors from IGF-IR dom-neg-transfected cells. Following this observation, we demonstrated in vitro that vascular smooth muscle cells migrated significantly less in conditioned medium derived from IGF-IR dom-neg-transfected cells compared with medium from control cells. After splenic injections, IGF-IR dom-neg transfectants failed to produce liver metastases, in contrast to parental cells and mock transfectants (p < 0.05). In addition, IGF-IR dom-neg-transfected cells failed to form liver tumors after direct injection into the liver. These studies demonstrate that the IGF-IR plays an important role in multiple mechanisms that mediate the growth, angiogenesis, and metastasis of human colon cancer. IGF-IR is a valid target for the therapy of human colon cancer.

Cell Division↗

PubMed2006

Do metalloproteinases destabilize vulnerable atherosclerotic plaques?

PURPOSE OF REVIEW: Atherosclerotic plaque rupture and thrombosis underlie most myocardial infarctions. Matrix metalloproteinases are a family of enzymes that remodel the extracellular matrix. Metalloproteinases could stabilize rupture-prone plaques by promoting smooth muscle cell migration and proliferation. Alternatively, metalloproteinases could destabilize vulnerable plaques by promoting matrix destruction, angiogenesis, leucocyte infiltration, and apoptosis. Evidence is reviewed from genetically modified mice and human biomarker and genetic studies that sheds light on this dual role of metalloproteinases. RECENT FINDINGS: Inhibition of metalloproteinases in mice using tissue inhibitors of metalloproteinases increases plaque stability; however, double knockouts of apolipoprotein E with matrix metalloproteinase 2, 3, 7, 9, 12, and 13 have more or less stable plaques, consistent with harmful or protective effects of individual metalloproteinases. Overexpression studies in mice or rabbits show that high activities of matrix metalloproteinase 9 and 12 decrease stability. Biomarker and human genetic studies demonstrate that increased metalloproteinase activity is associated with vascular repair or myocardial infarction. SUMMARY: Recent studies reinforce evidence for a dual role of matrix metalloproteinases in plaque stabilization and rupture, which probably depends on the stage, site, and severity of disease. Dysregulated metalloproteinase activity in end-stage coronary artery disease appears a valid target for therapy.

Animals↗

PubMed2005

A double mutation of Escherichia coli2C-methyl-D-erythritol-2,4-cyclodiphosphate synthase disrupts six hydrogen bonds with, yet fails to prevent binding of, an isoprenoid diphosphate.

The essential enzyme 2C-methyl-D-erythritol-2,4-cyclodiphosphate (MECP) synthase, found in most eubacteria and the apicomplexan parasites, participates in isoprenoid-precursor biosynthesis and is a validated target for the development of broad-spectrum antimicrobial drugs. The structure and mechanism of the enzyme have been elucidated and the recent exciting finding that the enzyme actually binds diphosphate-containing isoprenoids at the interface formed by the three subunits that constitute the active protein suggests the possibility of feedback regulation of MECP synthase. To investigate such a possibility, a form of the enzyme was sought that did not bind these ligands but which would retain the quaternary structure necessary to create the active site. Two amino acids, Arg142 and Glu144, in Escherichia coli MECP synthase were identified as contributing to ligand binding. Glu144 interacts directly with Arg142 and positions the basic residue to form two hydrogen bonds with the terminal phosphate group of the isoprenoid diphosphate ligand. This association occurs at the trimer interface and three of these arginines interact with the ligand phosphate group. A dual mutation was designed (Arg142 to methionine and Glu144 to leucine) to disrupt the electrostatic attractions between the enzyme and the phosphate group to investigate whether an enzyme without isoprenoid diphosphate could be obtained. A low-resolution crystal structure of the mutated MECP synthase Met142/Leu144 revealed that geranyl diphosphate was retained despite the removal of six hydrogen bonds normally formed with the enzyme. This indicates that these two hydrophilic residues on the surface of the enzyme are not major determinants of isoprenoid binding at the trimer interface but rather that hydrophobic interactions between the hydrocarbon tail and the core of the enzyme trimer dominate ligand binding.

Anti-Infective Agents↗

PubMed2006

Heparanase expression in nasopharyngeal carcinoma inversely correlates with patient survival.

AIMS: To determine the expression and prognostic significance of heparanase in nasopharyngeal carcinoma (NPC). METHODS: Immunohistochemistry was performed on formalin-fixed paraffin-embedded sections of 46 patients with NPC. Clinical and immunohistochemical data were correlated with gender, age, histological type, Epstein-Barr virus (EBV) status, stage and survival. RESULTS: Heparanase immunoreactivity was found in 35% (16/46) of specimens. The cumulative survival of patients diagnosed as heparanase negative (n = 30) at 10 years was 70%. In contrast, the cumulative survival of patients diagnosed as heparanase positive (n = 16) at 10 years was 25%, differences that are highly statistically significant (P = 0.03). No significant correlations were found between heparanase immunoreactivity and gender, age, EBV status, tumour histology or tumour stage. CONCLUSION: Heparanase expression is inversely correlated with survival of NPC patients, clearly indicating that heparanase is a reliable prognostic factor for this malignancy, and further supports the notion that heparanase is a valid target for the development of anti-cancer drugs.

Adolescent↗

PubMed1995

A brief description of the methods of economic appraisal and the valuation of health states.

The interest of nurses in methods of economic evaluation appears limited to cost-effectiveness analysis, with an apparent unawareness of other methods of economic appraisal and the types of efficiency they consider. The main methods of economic appraisal are discussed, and linked to different kinds of efficiency. Methods for the valuation of health states, an important accompaniment to the methods of economic appraisal, are briefly described along with some of the practical difficulties. If skilled nursing care--alone or with other disciplines--changes health status then the measurement and valuation of such states may be used to inform resource allocation decisions involving nursing. It could be argued that the main impact of nursing is on quality of life, and if so this suggests cost utility analysis, and not cost-effectiveness analysis, as the natural level of appraisal for nursing. The use of these methods in research, and participation in their future development, are both suggested as valid targets for nurses to aim for.

Cost-Benefit Analysis↗

PubMed2006

In vivo pharmacological disease models for psoriasis and atopic dermatitis in drug discovery.

In order to perform relevant in vivo pharmacological investigations in drug discovery within dermatology it is fundamental to master or have access to relevant skin disease models that makes it possible to identify and validate targets and to screen and discover drugs in vivo. There is a strong need for highly predictive in vivo models in order to introduce the right drug candidates into the clinical phases of development. Recent advances in dermatological in vivo pharmacological disease models for psoriasis and atopic dermatitis are reviewed together with a discussion of the rationale for their application in drug discovery.

Animals↗

PubMed2007

Exploring in vitro roles of siRNA in cardiovascular disease.

RNA interference (RNAi) is an adaptive defense mechanism through which double stranded RNAs silence cognate genes in a sequence-specific manner. It has been employed widely as a powerful tool in functional genomics studies, target validation and therapeutic product development. Similarly, the application of small interfering RNA (siRNA) to the silencing of the disease-causing genes involved in cardiovascular diseases has made great progress. In this overview, we attempt to provide a brief outline of the current understanding of the mechanism of RNAi and its potential application to the cardiovascular system, with particular emphasis on its ability to identify the pathophysiological function of genes related to several important cardiovascular disorders. The prospects of RNAi-based therapeutics, as well as the advantages and potential problems, are also discussed.

Animals↗