CANDIDA ALBICANS A M EDICAL D ICTIONARY , B IBLIOGRAPHY , AND A NNOTATED R ESEARCH G UIDE TO I NTERNET R EFERENCES
J AMES N. P ARKER , M.D. AND P HILIP M. P ARKER , P H .D., E DITORS
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ICON Health Publications ICON Group International, Inc. 4370 La Jolla Village Drive, 4th Floor San Diego, CA 92122 USA Copyright ©2004 by ICON Group International, Inc. Copyright ©2004 by ICON Group International, Inc. All rights reserved. This book is protected by copyright. No part of it may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, recording, or otherwise, without written permission from the publisher. Printed in the United States of America. Last digit indicates print number: 10 9 8 7 6 4 5 3 2 1 Publisher, Health Care: Philip Parker, Ph.D. Editor(s): James Parker, M.D., Philip Parker, Ph.D. Publisher's note: The ideas, procedures, and suggestions contained in this book are not intended for the diagnosis or treatment of a health problem. As new medical or scientific information becomes available from academic and clinical research, recommended treatments and drug therapies may undergo changes. The authors, editors, and publisher have attempted to make the information in this book up to date and accurate in accord with accepted standards at the time of publication. The authors, editors, and publisher are not responsible for errors or omissions or for consequences from application of the book, and make no warranty, expressed or implied, in regard to the contents of this book. Any practice described in this book should be applied by the reader in accordance with professional standards of care used in regard to the unique circumstances that may apply in each situation. The reader is advised to always check product information (package inserts) for changes and new information regarding dosage and contraindications before prescribing any drug or pharmacological product. Caution is especially urged when using new or infrequently ordered drugs, herbal remedies, vitamins and supplements, alternative therapies, complementary therapies and medicines, and integrative medical treatments. Cataloging-in-Publication Data Parker, James N., 1961Parker, Philip M., 1960Candida Albicans: A Medical Dictionary, Bibliography, and Annotated Research Guide to Internet References / James N. Parker and Philip M. Parker, editors p. cm. Includes bibliographical references, glossary, and index. ISBN: 0-597-84364-3 1. Candida Albicans-Popular works. I. Title.
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Disclaimer This publication is not intended to be used for the diagnosis or treatment of a health problem. It is sold with the understanding that the publisher, editors, and authors are not engaging in the rendering of medical, psychological, financial, legal, or other professional services. References to any entity, product, service, or source of information that may be contained in this publication should not be considered an endorsement, either direct or implied, by the publisher, editors, or authors. ICON Group International, Inc., the editors, and the authors are not responsible for the content of any Web pages or publications referenced in this publication.
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Acknowledgements The collective knowledge generated from academic and applied research summarized in various references has been critical in the creation of this book which is best viewed as a comprehensive compilation and collection of information prepared by various official agencies which produce publications on Candida albicans. Books in this series draw from various agencies and institutions associated with the United States Department of Health and Human Services, and in particular, the Office of the Secretary of Health and Human Services (OS), the Administration for Children and Families (ACF), the Administration on Aging (AOA), the Agency for Healthcare Research and Quality (AHRQ), the Agency for Toxic Substances and Disease Registry (ATSDR), the Centers for Disease Control and Prevention (CDC), the Food and Drug Administration (FDA), the Healthcare Financing Administration (HCFA), the Health Resources and Services Administration (HRSA), the Indian Health Service (IHS), the institutions of the National Institutes of Health (NIH), the Program Support Center (PSC), and the Substance Abuse and Mental Health Services Administration (SAMHSA). In addition to these sources, information gathered from the National Library of Medicine, the United States Patent Office, the European Union, and their related organizations has been invaluable in the creation of this book. Some of the work represented was financially supported by the Research and Development Committee at INSEAD. This support is gratefully acknowledged. Finally, special thanks are owed to Tiffany Freeman for her excellent editorial support.
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About the Editors James N. Parker, M.D. Dr. James N. Parker received his Bachelor of Science degree in Psychobiology from the University of California, Riverside and his M.D. from the University of California, San Diego. In addition to authoring numerous research publications, he has lectured at various academic institutions. Dr. Parker is the medical editor for health books by ICON Health Publications. Philip M. Parker, Ph.D. Philip M. Parker is the Eli Lilly Chair Professor of Innovation, Business and Society at INSEAD (Fontainebleau, France and Singapore). Dr. Parker has also been Professor at the University of California, San Diego and has taught courses at Harvard University, the Hong Kong University of Science and Technology, the Massachusetts Institute of Technology, Stanford University, and UCLA. Dr. Parker is the associate editor for ICON Health Publications.
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About ICON Health Publications To discover more about ICON Health Publications, simply check with your preferred online booksellers, including Barnes&Noble.com and Amazon.com which currently carry all of our titles. Or, feel free to contact us directly for bulk purchases or institutional discounts: ICON Group International, Inc. 4370 La Jolla Village Drive, Fourth Floor San Diego, CA 92122 USA Fax: 858-546-4341 Web site: www.icongrouponline.com/health
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Table of Contents FORWARD .......................................................................................................................................... 1 CHAPTER 1. STUDIES ON CANDIDA ALBICANS ................................................................................ 3 Overview........................................................................................................................................ 3 The Combined Health Information Database................................................................................. 3 Federally Funded Research on Candida Albicans .......................................................................... 4 E-Journals: PubMed Central ....................................................................................................... 58 The National Library of Medicine: PubMed ................................................................................ 96 CHAPTER 2. NUTRITION AND CANDIDA ALBICANS .................................................................... 143 Overview.................................................................................................................................... 143 Finding Nutrition Studies on Candida Albicans....................................................................... 143 Federal Resources on Nutrition ................................................................................................. 146 Additional Web Resources ......................................................................................................... 146 CHAPTER 3. ALTERNATIVE MEDICINE AND CANDIDA ALBICANS .............................................. 149 Overview.................................................................................................................................... 149 National Center for Complementary and Alternative Medicine................................................ 149 Additional Web Resources ......................................................................................................... 163 General References ..................................................................................................................... 167 CHAPTER 4. DISSERTATIONS ON CANDIDA ALBICANS ................................................................ 169 Overview.................................................................................................................................... 169 Dissertations on Candida Albicans............................................................................................ 169 Keeping Current ........................................................................................................................ 169 CHAPTER 5. PATENTS ON CANDIDA ALBICANS ........................................................................... 171 Overview.................................................................................................................................... 171 Patents on Candida Albicans ..................................................................................................... 171 Patent Applications on Candida Albicans ................................................................................. 196 Keeping Current ........................................................................................................................ 211 CHAPTER 6. BOOKS ON CANDIDA ALBICANS............................................................................... 213 Overview.................................................................................................................................... 213 Book Summaries: Federal Agencies............................................................................................ 213 Book Summaries: Online Booksellers......................................................................................... 214 Chapters on Candida Albicans................................................................................................... 215 CHAPTER 7. MULTIMEDIA ON CANDIDA ALBICANS .................................................................... 219 Overview.................................................................................................................................... 219 Audio Recordings....................................................................................................................... 219 CHAPTER 8. PERIODICALS AND NEWS ON CANDIDA ALBICANS ................................................. 221 Overview.................................................................................................................................... 221 News Services and Press Releases.............................................................................................. 221 Newsletter Articles .................................................................................................................... 223 Academic Periodicals covering Candida Albicans ..................................................................... 223 APPENDIX A. PHYSICIAN RESOURCES .......................................................................................... 227 Overview.................................................................................................................................... 227 NIH Guidelines.......................................................................................................................... 227 NIH Databases........................................................................................................................... 229 Other Commercial Databases..................................................................................................... 231 APPENDIX B. PATIENT RESOURCES ............................................................................................... 233 Overview.................................................................................................................................... 233 Patient Guideline Sources.......................................................................................................... 233 Finding Associations.................................................................................................................. 235 APPENDIX C. FINDING MEDICAL LIBRARIES ................................................................................ 237 Overview.................................................................................................................................... 237 Preparation................................................................................................................................. 237
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Finding a Local Medical Library................................................................................................ 237 Medical Libraries in the U.S. and Canada ................................................................................. 237 ONLINE GLOSSARIES................................................................................................................ 243 Online Dictionary Directories ................................................................................................... 243 CANDIDA ALBICANS DICTIONARY .................................................................................... 245 INDEX .............................................................................................................................................. 315
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FORWARD In March 2001, the National Institutes of Health issued the following warning: "The number of Web sites offering health-related resources grows every day. Many sites provide valuable information, while others may have information that is unreliable or misleading."1 Furthermore, because of the rapid increase in Internet-based information, many hours can be wasted searching, selecting, and printing. Since only the smallest fraction of information dealing with Candida albicans is indexed in search engines, such as www.google.com or others, a non-systematic approach to Internet research can be not only time consuming, but also incomplete. This book was created for medical professionals, students, and members of the general public who want to know as much as possible about Candida albicans, using the most advanced research tools available and spending the least amount of time doing so. In addition to offering a structured and comprehensive bibliography, the pages that follow will tell you where and how to find reliable information covering virtually all topics related to Candida albicans, from the essentials to the most advanced areas of research. Public, academic, government, and peer-reviewed research studies are emphasized. Various abstracts are reproduced to give you some of the latest official information available to date on Candida albicans. Abundant guidance is given on how to obtain free-of-charge primary research results via the Internet. While this book focuses on the field of medicine, when some sources provide access to non-medical information relating to Candida albicans, these are noted in the text. E-book and electronic versions of this book are fully interactive with each of the Internet sites mentioned (clicking on a hyperlink automatically opens your browser to the site indicated). If you are using the hard copy version of this book, you can access a cited Web site by typing the provided Web address directly into your Internet browser. You may find it useful to refer to synonyms or related terms when accessing these Internet databases. NOTE: At the time of publication, the Web addresses were functional. However, some links may fail due to URL address changes, which is a common occurrence on the Internet. For readers unfamiliar with the Internet, detailed instructions are offered on how to access electronic resources. For readers unfamiliar with medical terminology, a comprehensive glossary is provided. For readers without access to Internet resources, a directory of medical libraries, that have or can locate references cited here, is given. We hope these resources will prove useful to the widest possible audience seeking information on Candida albicans. The Editors
1
From the NIH, National Cancer Institute (NCI): http://www.cancer.gov/cancerinfo/ten-things-to-know.
3
CHAPTER 1. STUDIES ON CANDIDA ALBICANS Overview In this chapter, we will show you how to locate peer-reviewed references and studies on Candida albicans.
The Combined Health Information Database The Combined Health Information Database summarizes studies across numerous federal agencies. To limit your investigation to research studies and Candida albicans, you will need to use the advanced search options. First, go to http://chid.nih.gov/index.html. From there, select the “Detailed Search” option (or go directly to that page with the following hyperlink: http://chid.nih.gov/detail/detail.html). The trick in extracting studies is found in the drop boxes at the bottom of the search page where “You may refine your search by.” Select the dates and language you prefer, and the format option “Journal Article.” At the top of the search form, select the number of records you would like to see (we recommend 100) and check the box to display “whole records.” We recommend that you type “Candida albicans” (or synonyms) into the “For these words:” box. Consider using the option “anywhere in record” to make your search as broad as possible. If you want to limit the search to only a particular field, such as the title of the journal, then select this option in the “Search in these fields” drop box. The following is what you can expect from this type of search: •
Effect of Antimicrobial Mouthrinses on the In Vitro Adhesion of Candida Albicans to Human Buccal Epithelial Cells Source: Clinical Oral Investigations. 5(3): 172-176. September 2001. Contact: Available from Springer-Verlag, New York Inc. Journal Fulfillment Services Department, P.O. Box 2485, Secaucus, NJ 07096-2485. Fax (202) 348-4505. Summary: Oral candidosis (thrush, a fungal infection) is the most frequent opportunistic infection in immunocompromised patients, and Candida albicans represents its primary causative agent. Adhesion to epithelial cells is a critical step in successful oral colonization and infection by Candida albicans. This article reports on a study undertaken to compare three mouthrinse products, containing chlorhexidine 0.2 percent (CHX), cetylpyridinium chloride 0.05 percent (CPC), or triclosan 0.045 percent (TRN) for
4
Candida Albicans
their effects on the in vitro adhesion of C. albicans to human buccal epithelial cells (BEC, the cells of the cheek lining). Candidal adhesion appeared to be significantly reduced by oral rinsing with the CHX containing mouthrinse. In vivo (in the laboratory) exposure of BEC to the CPC mouthrinse also inhibited adhesion of C. albicans. Both CHX and CPC products suppressed adhesion to the same extent. On the other hand, the TRN mouthrinse did not significantly affect epithelial adhesion of C. albicans. These findings suggest that mouthrinses containing CHX or CPC could be of value in the control of candidal colonization and infection. The authors call for clinical trials on the effectiveness of these products in reducing oral Candida infections. 3 tables. 27 references.
Federally Funded Research on Candida Albicans The U.S. Government supports a variety of research studies relating to Candida albicans. These studies are tracked by the Office of Extramural Research at the National Institutes of Health.2 CRISP (Computerized Retrieval of Information on Scientific Projects) is a searchable database of federally funded biomedical research projects conducted at universities, hospitals, and other institutions. Search the CRISP Web site at http://crisp.cit.nih.gov/crisp/crisp_query.generate_screen. You will have the option to perform targeted searches by various criteria, including geography, date, and topics related to Candida albicans. For most of the studies, the agencies reporting into CRISP provide summaries or abstracts. As opposed to clinical trial research using patients, many federally funded studies use animals or simulated models to explore Candida albicans. The following is typical of the type of information found when searching the CRISP database for Candida albicans: •
Project Title: A MODEL OF MORPHOLOGIC SWITCHING IN CANDIDA ALBICANS Principal Investigator & Institution: Gale, Cheryl A.; Pediatrics; University of Minnesota Twin Cities 200 Oak Street Se Minneapolis, Mn 554552070 Timing: Fiscal Year 2002; Project Start 01-JUL-2000; Project End 30-JUN-2003 Summary: Dr. Cheryl A. Gale is a neonatologist at the University of Minnesota. The candidate's long-term goal is to pursue an academic career devoted primarily to basic research. During Dr. Gale's pediatric residency and neonatal fellowship training, she was exposed to the high morbidity and mortality associated with systemic candidiasis in her premature patients on the neonatal intensive care unit. This experience solidified Dr. Gale's interest in understanding fungal pathogenesis as a means toward developing more effective therapies. The proposed application will provide the candidate the additional training necessary to be competitive for independent funding. The rich academic and research environment at the University of Minnesota is extremely well suited for the candidate's career development. Under the direct mentorship of Dr. Judith Berman, Dr. Gale will study the molecular mechanism of morphologic switching in C. albicans. She will focus on characterizing in vivo the localization of cytoskeletal-
2 Healthcare projects are funded by the National Institutes of Health (NIH), Substance Abuse and Mental Health Services (SAMHSA), Health Resources and Services Administration (HRSA), Food and Drug Administration (FDA), Centers for Disease Control and Prevention (CDCP), Agency for Healthcare Research and Quality (AHRQ), and Office of Assistant Secretary of Health (OASH).
Studies
5
associated proteins during morphologic switching and determining the role of cell cycle checkpoints during morphogenesis. Morphologic switching is postulated to contribute to the success of C. albicans as a pathogen because it gives the fungus versatility in the face of changing environments. Dr. Gale previously found that the C. albicans gene INT1 encodes a protein that is important for the morphologic switch from yeast-form to filamentous-form. Expression of INT1 in the usually non- pathogepic yeast, S. cerevisiae, results in the formation of extremely elongated buds that appear similar to C. albicans germ tubes. Her more recent characterization of these elongated buds in S. cerevisiae has shown that Int1p expression causes a novel localization of cytoskeletal-associated proteins (septins) that mimics that of Int1p itself. She also found that INT1 expression results in a delay in the nuclear cell cycle partially due to the activation of the morphogenesis checkpoint protein, Swe1p. The Specific Aims of this proposal will build on these preliminary results to directly determine, in C. albicans, the role of Int1p, septins, and cell cycle checkpoints in morphologic switching. Dr. Gale will develop a technique of genomic epitope tagging in C. albicans that will allow in vivo localization of Int1p and septins in C. albicans during the cell cycle and during morphologic switching in both wild-type and mutant C. albicans morphologic strains (Specific Aims 1 and 2). The candidate will disrupt the C. albicans SWE1 homolog and study the effect of such a mutation on the viability and ability of the strain to undergo morphologic switching (Specific Aim 3). These studies will allow Dr. Gale to determine the how Int1p, septins and cell cycle control proteins are related in the process of morphologic switching and may identify new therapeutic targets for anti- fungal agents. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: ABC TRANSPORTERS AND PATHOGENECITY OF CANDIDA ALBICANS Principal Investigator & Institution: Kohler, Gerwald A.; Stomatology; University of California San Francisco 500 Parnassus Ave San Francisco, Ca 941222747 Timing: Fiscal Year 2003; Project Start 01-AUG-2003; Project End 31-MAY-2005 Summary: (provided by applicant): ABC Transporters and Pathogenicity of Candida albicans. ABC transporters form a superfamily of ubiquitous membrane proteins involved in the energy-dependent translocation of a wide variety of substrates across cellular membranes. Multiple drug resistance and many human hereditary diseases have been related to ABC transporter function, however, the physiological roles of only a few of these transporters have been elucidated. One of the largest subfamilies of ABC proteins is the MRP/CFTR family (subfamily C); homologues of this group of ABC transporters are also found in the opportunistic fungal pathogen Candida albicans. In preliminary studies we have already shown that the MRP-related ABC transporter gene MLT1 of C. albicans is crucial for its invasion of parenchymal organs in a peritonitis model. This study focuses on a detailed characterization of the involvement of MRPrelated ABC transporters in fungal cell homeostasis networks during in vitro growth and in vivo interaction with the host in commensalism and opportunistic disease. Since the genome sequence of C. albicans is available, genomic expression profiling using DNA microarrays will be used to infer the functional relationships of MLT1 and two of the most closely-related transporters within the MRP/CFTR subfamily. In vitro expression studies in cell culture models will be extended to experimental infection models using innovative methods like in vivo expression technology (IVET) and realtime PCR. Specific ABC transporter gene inactivation mutants derived from wild-type C. albicans strains will be generated by gene disruption or RNA interference and phenotypically characterized. Furthermore, the functional characterization of these
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Candida Albicans
transporters will comprise identification of endogenous and exogenous substrates as well as determination of the subcellular localization. Our findings are likely to contribute to new approaches for treatment of fungal infections with specific compounds inhibiting ABC transporters. Since many human ABC transporters of the MRP/CFTR subfamily are involved in drug resistance or inherited disorders, our findings might help to infer their functional properties from those of their fungal homologues. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: ALBICANS
AIDS:
COMBATTING
DRUG
RESISTANCE
OF
CANDIDA
Principal Investigator & Institution: Cannon, Richard D.; University of Otago Leith St Dunedin, Timing: Fiscal Year 2003; Project Start 01-DEC-2002; Project End 30-NOV-2004 Summary: (provided by applicant): Candida albicans is a pathogenic yeast that causes serious fungal infections in the immunocompromised and opponunistic Candida infections can be the first indication of immunosuppression in HIV+ individuals. AIDS patients frequently suffer from oropharyngeal candidiasis (OPC) and require antifungal therapy. In the 1990s there was a dramatic increase in the failure of fluconazole therapy for AIDS patients with OPC due to C. albicans strains developing fluconazole resistance. The most common mechanism responsible for high level fluconazole resistance in these yeast was over-expression of drug efflux pump Cdrl p. Globally, OFC remains a major opponunistic infection in HIVIAIDS, and the widespread use of fluconazole in the third world is likely to maintain pressure on C. albicans to develop resistance. The overall objective of this research is to use a novel strategy to improve the treatment of AIDS patients with oral candidiasis by combating azoleresistance in C. albicans. Specific objectives are to: 1. Employ a novel heterologous functional hyperexpression system to determine the mechanism of pumping by Cdrl p, using both in vitro mutagenized Cdrl p and Cdrl proteins from clinical C. albicans isolates (obtained from AIDS patients) that demonstrate high and low pump activities. 2. Use the heterologous functional hyper-expression of Cdrlp to screen a unique combinatorial Doctapeptide library for peptides that inhibit the pump. This work will validate a novel approach to combating azole-resistance in C. albicana An understanding of drug pumping mechanisms may indicate new ways to circumvent efflux-mediated resistance. This project is expected to identify a lead compound with the potential to sensitize resistant strains to azole antifungals. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: ANALYSIS OF THE CANDIDA ALBICANS PROTEOME Principal Investigator & Institution: Lopez-Ribot, Jose L.; Assistant Professor; Medicine; University of Texas Hlth Sci Ctr San Ant 7703 Floyd Curl Dr San Antonio, Tx 78229 Timing: Fiscal Year 2003; Project Start 01-AUG-2003; Project End 31-MAY-2005 Summary: (provided by applicant): Candida albicans is by far the most frequently isolated human mycotic agent. In the oral cavity, oropharyngeal candidiasis (OPC) is a significant cause of morbidity in patients with HIV or AIDS. Other forms of mucosal candidiasis are also frequent in different patient populations such as infants, denture wearers, the elderly, and following antibiotic therapy. Azole derivatives, in particular fluconazole, are generally effective in the treatment of mucosal candidiasis. However, resistance has emerged as an important clinical problem. Large-scale DNA sequencing
Studies
7
has provided an important sequence infrastructure for protein analysis. The term "Proteomics" refers to large-scale characterization of the proteins present in a cell, tissue or organism (the proteome) and involves the combined application of techniques to resolve, identify, quantitate and characterize proteins, as well as bioinformatics tools to store, communicate and interlink the resulting information. The experimental design of this proposal takes advantage of the recently completed NIDCR-funded Candida albicans genome sequencing project. The post-genomic era offers unprecedented opportunities to study host-fungal interactions. The specific aims of this proposal include: i) a pilot feasibility study of the analysis of the C. albicans proteome under a wide variety of conditions and development of a searchable proteomic map and database as a resource for the fungal community, ii) analysis of C. albicans azole resistance by proteomics and identification of proteins implicated in the regulatory networks of multidrug resistance. We will expect that these projects will establish the foundations for creating a fundamental tool for the C. albicans research community and for providing a detailed large-scale study of a biological phenomenon (drug resistance) with important clinical repercussions. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: ADJUVANTS
ANTIFUNGALS
FROM
MARINE
INVERTEBRATES--AIDS
Principal Investigator & Institution: Molinski, Tadeusz F.; Professor; Chemistry; University of California Davis Sponsored Programs, 118 Everson Hall Davis, Ca 95616 Timing: Fiscal Year 2002; Project Start 01-MAR-1997; Project End 31-MAY-2005 Summary: (provided by applicant): AIDS-related fungal infections are important targets for reduction of mortality and improvement in the quality of life for people living with AIDS. Each new generation of azole drugs, such as fluconazole, have succumbed to recurrent cross resistance. Natural products are represented among clinically useful antifungal agents. Marine invertebrates, particularly Porifera (sponges) that produce chemically diverse libraries of natural products, some of which show antifungal activity. The general goal of this competitive renewal is to find and identify small molecules from marine organisms that are active against fluconazole-resistant strains of Candida albicans and inherently fluconazole-resistant non-albicans species, including Candida glabrata and Candida krusei and use these as prototypes leads for antifungal drugs. We plan to prepare and screen extracts for antifungal agents using mechanism-selective approach that may be useful identifying new leads for antifungal therapy. This program embodies a rational search for compounds with unique mechanisms of action, including inhibition of fungal sphingolipid biosynthesis, that complement current therapies and intervene at strategic points in fungal cell metabolism or life cycle. Active components will be isolated by a combination of solvent-partitioning, chromatography, liquid-liquid centrifugal counter current chromatography and other techniques. The in vitro antibiotic susceptibilities of pathogenic fungi will be evaluated in a panel of fluconazole-resistant fungi. Selected leads will be advanced to in vivo evaluation in murine models of C. albicans, Cryptococcus neoformans and C. glabrata. The structures of novel compounds will be determined by a combination of spectroscopic techniques including mass spectrometry, nuclear magnetic spectroscopy, circular dichroism and X-ray crystallography. Absolute stereochemistry of chiral molecules will be determined using a combination of chiroptical techniques and chemical degradation. Derivatives of existing leads, including the C. glabrata-specific dimeric sphingolipid, oceanapiside, will be synthesized de novo or by semi-synthetic modification to prepare limited libraries of analogs for structure-activity studies. Optimized leads identified from those libraries
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Candida Albicans
will be advanced to in vitro and in vivo evaluation. The strengths of this program include a successful track record in targeting emergent pathogenic fungi, including fIuconazole-resistant Candida species that are of importance in human health, and maximization of chemical diversity to enhance the chances of discovery of natural product antifungal agents. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: ANTIMICROBIAL COATING FOR BIOFILM INHIBITION Principal Investigator & Institution: Rawls, H Ralph.; Professor; Biomedical Development Corporation 500 Sandau, Ste 200 San Antonio, Tx 78216 Timing: Fiscal Year 2002; Project Start 01-SEP-2002; Project End 31-AUG-2004 Summary: Description(provided by applicant): Candida is the third leading cause of catheter-related nosocomial bloodstream infections. Indwelling devices have been shown to support colonization and biofilm formulation by Candida. Intravenous lines are the most frequent indwelling device and are the single most common cause of candidemia. Once a Candida biofilm forms in vivo, removal of the substrate that is supporting the biofilm growth is almost always required to eliminate the infection. Unfortunately, in many instances removal is impossible due to deteriorated patient condition, anatomical location, or underlying disease. The goal of this project is to incorporate an antifungal agent into unique, patented formulations to create a coating for medical devices that is resistant to C. albicans. A coating and antifungal delivery system that could extend the time and/or increase the rate of delivery and achieve a higher concentration of antifungal agents to a C. albicans-susceptible site would have enormous therapeutic advantages over systemic and inefficacious topical routes of delivery. To demonstrate the feasibility of this approach, the specific aims are to optimize the coating formulation for use as short-term antifungal coatings for medical devices, evaluate the coatings for antifungal activity in vitro, and to perform an intramuscular implantation study in rabbits. PROPOSED COMMERCIAL APPLICATION: Candida albicans is by far the most frequently isolated human mycotic agent. The large number of immunosuppressed patients with indwelling devices combined with the extensive social and economic costs associated with treating C. albicans-related infections serve as justification for the pursuit of alternative therapies. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
•
Project Title: BIOFILMS
ANTIMICROBIAL
RESISTANCE
IN
CANDIDA
ALBICANS
Principal Investigator & Institution: Tyler, Bonnie J.; Associate Chemical/Fuels Engineering; University of Utah Salt Lake City, Ut 84102
Professor;
Timing: Fiscal Year 2001; Project Start 15-AUG-1999; Project End 30-JUN-2004 Summary: Oral candidias is a biofilm based fungal infection that can have severe implications particularly in the growing population of immunocompromised patients. Despite their clincal importance, very little research has been done on fungal biofillms including candida. Few if any studies involving in situ analysis of C. albincans biofilms have been published. One ex situ study has shown that Candida biofilms, like those of bacteria, are resistant to antimicrobial agents. Although a variety of mechanisms have been proposed to explain the recalcitrance of bacterial biofilms to antibiotics, the mechanisms by which Candida biofilms resist the action of antifungal agents are unknown. The goal of this proposed research is to delineate the relationship between the structure of C. albicans biofilms and their resistance to antimicrobial agents. In this
Studies
9
study a combination of in situ methods will be used to quantitatively investigate the structure of C. albicans biofilms. These methods will include Attenuated Total Reflectance Infrared Spectroscopy (ATR/FTIR), Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS), electrochemical microsensors, fluorescent microscopy and scanning laser confocal microscopy. The influence of a variety of environmental factors including fluid shear, growth media, serum and saliva proteins and substratum chemistry on the biofilm structure will be investigated. These factors will then be used to manipulate the biofilm structure in order to determine the influence of structural features on the antifungal resistance of the biofilms. The efficacy of three antimicrobial agents, hydrogen peroxide, chlorhexidine gluconate, and fluconazole, against both biofilms and planktonic C. albicans will be measured. In addition, penetration of these agents into the biofilm will be measured with a combination of ATR/FTIR, ToF-SIMS imaging of cryosections, and microelectrodes. Multivariate statistical modeling will be used to identify relationships between the biofilm structure and antimicrobial resistance. Mechanistic differential equation based models will be used to identify possible causal relationships between biofilm structure and antifungal efficacy. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: AZOLE-RESISTANT CANDIDA IN MARROW TRANSPLANT PATIENTS Principal Investigator & Institution: Marr, Kieren A.; Fred Hutchinson Cancer Research Center Box 19024, 1100 Fairview Ave N Seattle, Wa 98109 Timing: Fiscal Year 2002; Project Start 01-APR-1999; Project End 31-MAR-2004 Summary: This abstract is not available. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
•
Project Title: BIOLOGY AND DRUG RESISTANCE OF CANDIDA BIOFILMS Principal Investigator & Institution: Ghannoum, Mahmoud A.; Professor and Center Director; Dermatology; Case Western Reserve University 10900 Euclid Ave Cleveland, Oh 44106 Timing: Fiscal Year 2002; Project Start 01-MAR-2002; Project End 31-DEC-2005 Summary: (provided by the applicant): Candida-associated denture stomatitis (chronic atrophic candidiasis) is the most prevalent superficial oral infection and the most common form of Candida-associated disease. The etiology of denture stomatitis involves dental plaque. Dental plaque consists of a complex biofilm of bacteria and yeasts, predominantly Candida albicans. C. albicans biofilms have received much less attention than bacterial biofilms, and our present knowledge of their biology and drug resistance is at a rudimentary stage. Frequent denture stomatitis treatment failures combined with a steadily increasing population of elderly people, many of whom will be endentulous, make this area of study particularly important. The long-range goal of our work is to understand the biology and drug resistance of C. albicans biofilms. Our preliminary work in this new area resulted in the successful development of a reproducible model of C. albicans-associated biofilms (Publication #1). Since the last submission, we used this model to: 1) define the three stages of C. albicans associated biofilm development, 2) demonstrate that C. albicans biofilm is a highly heterogeneous structure, 3) show that the antifungal resistance of C. albicans biofilm increases in conjunction with biofilm development, 4) show that C. albicans has greater ability than the less pathogenic C. parapsilosis and Saccharomyces cerevisiae to form denture biofilm, and 5) show that C. albicans genes are differentially expressed under biofilm
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Candida Albicans
and planktonic conditions. Additionally, we initiated efforts to construct a C. albicans DNA array, and developed a bioprosthetic associated candidal biofilm model. Importantly, our studies showed that biofilm grown in our in vitro model has similar morphology as that growing in vivo on a catheter obtained from a patient with catheterassociated infection. Specific aims of the current proposal are: Specific Aim 1: Use our established biofilm model to determine the antifungal susceptibility profiles of C. albicans isolates obtained from denture stomatitis patients, and to study the effect of antifungal agents on the growth kinetics of C. albicans bioflims. Specific Aim 2: Investigate the mechanism(s) responsible for increased antifungal resistance of biofilmassociated C. albicans. Specific Aim 3: Identify genes that are involved in the formation and contribute to the development of antifungal resistance of C. albicans biofilms. By studying biofilm model systems and applying this knowledge to the patient population, we will gain a wealth of data about the biology and drug resistance of C. albicans in biofilms. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: C. ALBICANS FROM HIV+ INDIVIDUALS&ITS ROLE IN DRUG RESIS Principal Investigator & Institution: Vargas, Kaaren G.; Dows Inst for Dental Research; University of Iowa Iowa City, Ia 52242 Timing: Fiscal Year 2002; Project Start 01-JUL-2001; Project End 31-MAY-2005 Summary: (provided by applicant) As a recent graduate from a PhD program and newly appointed assistant professor at The University of Iowa, I feel that I would benefit significantly from the Faculty Transition portion of the K22. In conjunction with the completion of the research proposed, the PI will receive training in the ethics of biomedical research. The research project itself will receive guidance from a number of very experienced researchers. Drs. Michael Pfaller, Christopher Squier, Georgia Johnson, Philip Wertz and David Soll. The research experience that is outlined in this proposal will enable me to develop into an independent investigator and contributor to the advancement of science. I will receive extensive support from my advisors and the College of Dentistry. Research facilities are available for my use and courses are offered that will enable me to conduct research in a responsible manner. The. experimental portion sets out to answer the following questions: 1. What is the antifungal susceptibility of C. albicans switch phenotypes isolated from HIV-positive individuals? 2. Are there differences in uptake of antifungals among different switch phenotypes or differences in ergosterol content in the presence of antifungals and are there differences in expression of known multi-drug resistance genes (MDR1, CDR1 and ERG11) among the different switch phenotypes? and 3. Are different switch phenotypes better able to survive under in vivo conditions of antifungal drug therapy for candidiasis? For this, a collection of samples from HIV+ and HIV- individuals previously collected by the PI and Dr. Michael Pfaller will be used. From these studies we hope to increase our understanding of the role that phenotypic switching plays in antifungal drug resistance, which is an increasing problem in severely immunosuppressed individuals. This increased knowledge could lead to improvements in treatment of oral candidiasis in all immunocompromised individuals. In the long-term, I would like to make significant contributions to science in the area of antifungal drug resistance. It is an exciting area of research and I feel strongly that I have the motivation to accomplish this goal. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
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Project Title: EPITHELIA
C.ALBICANS
REGULATION
BETA-DEFENSINS
IN
11
ORAL
Principal Investigator & Institution: Weinberg, Aaron; Associate Professor; Periodontics; Case Western Reserve University 10900 Euclid Ave Cleveland, Oh 44106 Timing: Fiscal Year 2002; Project Start 28-SEP-2000; Project End 31-JUL-2004 Summary: (adapted from the applicant's abstract) Oropharyngeal candidiasis (OPC) is an emerging disorder owing to the prevalence of AIDS, misuse of antibiotics, and host immunosuppression in general. Candida albicans is the most common fungal species isolated from OPC lesions. Recent findings show that mucosal epithelial cells synthesize and secrete antibacterial and antifungal agents, belonging to a family of small, cationic peptides. These molecules, human beta-defensins 1 and 2 (hBD-1, hBD-2) are predicted to function as a first line of host defense against microbial pathogenesis. The PI has discovered that these peptides are expressed in normal human gingival epithelial cells and associated with differentiated epithelium of oral tissues. Moreover, they found that the non oral, yet disseminating isolate C. albicans strain SC5314 stimulates betadefensin expression in oral epithelial cells, but a clinical OPC isolate does not. This proposal intends to test hypotheses relevant to oropharyngeal candidiasis emanating from the postulate that oral epithelial cells can be stimulated to produce beta-defensins that protect the host from candidal challenges at the oral mucosal barrier. The objectives of this proposal are (1) to determine beta-defensin expression in oral epithelial cells in response to challenge with OPC derived C. albicans isolates, (2) to characterize key virulence factors of C. albicans SC5314 and OPC isolates that lead to beta-defensin response, (3) to examine beta-defensin protection against C. albicans, and (4) to identify genes in oral epithelial cells associated with C. albicans modulation of beta-defensin expression, using microarray technology. The PI hypothesizes that peptide-based antimicrobial defense may be a way in which the gingival epithelium resists invasion of potential pathogens. In light of the frequent adjunctive use of antibiotics and antimycotics in treating oral diseases, with the threat of microbial resistance, investigations into novel eukaryotic peptides, such as beta-defensins, are highly significant and offer the potential for future clinical promise. The PI states that this research direction may be significant in leading to future studies with potential application to oral disorders, therapeutic use, and technology development. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: CANDIDA ALBICANS ORAL BIOFILM Principal Investigator & Institution: Chaffin, Welda L.; Professor; Microbiology and Immunology; Texas Tech University Health Scis Center Health Sciences Center Lubbock, Tx 79430 Timing: Fiscal Year 2002; Project Start 01-SEP-2001; Project End 31-AUG-2005 Summary: (provided by applicant): Candida albicans is a commensal that colonizes skin and mucosal surfaces including the oral cavity. The organism is also an agent of opportunistic disease of these surfaces as well as internal disseminated disease. Oral candidiasis is associated with derangements of the oral flora related with the acquisition of microbes by neonates and anti-bacterial therapy, oral prostheses, and host factors such as diabetes mellitus and HIV infection. Oral manifestations include pseudomembraneous candidiasis (thrush) and denture stomatitis. Oropharyngeal infection is virtually an inescapable consequence of AIDS (96 percent patients) and frequently reoccurs. Denture stomatitis may affect 50 percent of complete denture wearers. The organism forms biofilms on mucosa, teeth and oral devices such as
12
Candida Albicans
dentures, generally in association with oral bacteria. Compared to planktonic cells, organisms in biofilms have characteristics such as reduced susceptibility to antifungal drugs and the presence of an extracellular matrix. This study will test the hypothesis that unique characteristics associated with C. albicans biofilms are the result of altered gene expression in general cellular metabolism as well as bioflim specific gene expression. A model of saliva-coated denture acrylic established in this laboratory will be used. About 230 alterations in general cellular metabolism have been identified in biofilm compared to planktonic cells by exploiting the high homology between Saccharomyces cerevisiae and C. albicans and the commercial availability of gene arrays for S. cerevisiae. In Aim 1 this approach will be applied to examine expression temporally during biofilm formation and to other conditions of biofilm development using C. albicans DNA chips. In Aim 2, expression in in vitro biodiverse models will also be examined to identify genes inherently associated with biofilms as differentiated from those influenced by the biofilm environment. Expression of selected genes from the inherent biofilm expression class will be determined in vivo in organisms recovered from human saliva. Aim 3 will examine the role of biofilm-regulated genes such as TUP1and EFG1 using genetically modified strains. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: CANDIDA INVASION OF ENDOTHELIUM AND VIRULENCE Principal Investigator & Institution: Filler, Scott G.; Professor; Harbor-Ucla Research & Educ Inst 1124 W Carson St Torrance, Ca 90502 Timing: Fiscal Year 2004; Project Start 20-JAN-2004; Project End 31-DEC-2008 Summary: (provided by applicant): The incidence of bloodstream infections caused by Candida species has increased dramatically; these organisms now account for 10% of all bloodstream isolates. The mortality associated with candidemia approaches 40%, even with therapy. Because of this unacceptably high mortality, new strategies to treat and prevent this infection are necessary. Our goal is to identify new Candida albicans virulence genes and to determine the mechanisms by which they contribute to pathogenicity. This information holds promise to identify new targets for antifungal strategies. Our hypothesis is that the ability of C. albicans to invade and damage host cells is critical for the organism to establish and maintain a deep-seated infection. Support for this hypothesis comes from our finding that many C. albicans mutants with decreased virulence in the mouse model of hematogenously disseminated infection also have reduced ability to invade and damage endothelial cells in vitro. Furthermore, we have screened a collection of random C. albicans homozygous insertion mutants to identify strains with impaired capacity to damage endothelial cells in vitro. We discovered that zed1/zed1 and cka2/cka2 insertion mutants caused much less endothelial cell damage than did the isogenic control strain. Also, the zed1/zed1 insertion mutant had significantly attenuated virulence in mice (the virulence of the cka2/cka2 strain has not yet been tested). These results strongly suggest that C. albicans genes required for in vitro endothelial cell damage are also required for virulence. Our objective is to use in vitro studies of the interactions between C. albicans and endothelial cells to define mechanisms of host-pathogen interaction. In this project, we will 1) identify the endothelial cell receptors that C. albicans uses to invade this host cell; 2) determine the mechanisms by which C. albicans Zed1p, Cka2p, and other newly identified gene products contribute to virulence; 3) use defined C. albicans mutants to elucidate functional relationships among C. albicans virulence regulators, endothelial cell receptors, and endothelial cell damage; and 4) use a random insertional mutagenesis
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approach to identify new C. albicans genes that promote endothelial cell damage in vitro and virulence. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: CENTROMERE STRUCTURE AND FUNCTION Principal Investigator & Institution: Carbon, John A.; Professor of Biochemistry; Molecular, Cellular & Dev Biol; University of California Santa Barbara 3227 Cheadle Hall Santa Barbara, Ca 93106 Timing: Fiscal Year 2003; Project Start 01-JAN-1977; Project End 31-DEC-2007 Summary: (provided by applicant): The long-range objective of this research program is to understand in molecular terms how the centromere-kinetochore functions in eukaryotic cell division. Our previous research in this area emphasized centromere structure/function studies in the budding yeast Saccharomyces cerevisiae and fission yeast Schizo-saccharomyces pombe. Building on the valuable information gained in these groundwork studies, we will now extend the work to investigate centromerekinetochores in the two most common and clinically significant pathogenic budding yeasts, Candida albicans and Candida glabrata. All budding yeast species examined to date contain relatively small point centromeres (CEN less than 400 bp in length), whereas other organisms contain large regional centromeres characterized by the presence of long stretches of heterochromatic repeated DNA sequences. It is postulated that this fundamental difference in centromere structure could be exploited eventually to develop agents selectively toxic to the budding yeasts. Specific aims are: (1) combined molecular and genetic strategies will be used to isolate and characterize CEN DNAs of C. albicans; (2) stable CEN-based plasmid and artificial chromosome vector systems will be constructed to facilitate molecular genetic research in C. albicans; and (3) inner kinetochore-associated proteins from both C. albicans and C. glabrata will be identified and characterized as potential targets for specific antifungal drug therapy. We have identified, epitope-tagged, and expressed an evolutionarily conserved, centromerespecific histone H3 protein (CaCse4p) in C. albicans. Chromatin immunoprecipitation (CHIP) with antibodies directed against this protein will be used to isolate and clone CEN DNAs. Putative CEN DNAs will be identified and characterized in terms of their nucleotide sequence and ability to mitotically stabilize plasmid and artificial chromosome vectors in actively dividing cells. Inner kinetochore proteins of Candida species will be isolated by CEN DNA affinity chromatography and the corresponding genes will be cloned and characterized. One-hybrid and two-hybrid screens currently under development for use with C. glabrata will be applied to a search for additional genes specifying CEN-associated proteins. Kinetochore association of candidate proteins will be confirmed by ChIP analysis and by studying the effects of gene deletion and/or mutagenesis on cell division and chromosome segregation. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
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Project Title: CHARACTERIZATION OF THE S. POMBE CAMP PATHWAY Principal Investigator & Institution: Hoffman, Charles S.; Professor; Biology; Boston College 140 Commonwealth Ave Newton, Ma 02467 Timing: Fiscal Year 2002; Project Start 01-JUL-1991; Project End 31-MAR-2005 Summary: (Adapted from the Investigator's abstract): One way cells respond to their environment is by creating internal signals that regulate gene expression. Human and yeast cells employ homologous signaling pathways to control cell growth, stress response, metabolic pathways and differentiation. Therefore, the relatively simple and
14
Candida Albicans
genetically pliable budding and fission yeasts are valuable model organisms providing important insights to mechanisms of signal transduction in mammalian cells. My laboratory studies the glucose/cAMP signal pathway that is central to the transcriptional regulation of the fission yeast fbpl gene. Environmental glucose triggers the activation of adenylate cyclase, and the resulting cAMP signal activates protein kinase A to repress fbpI transcription. Many, but not all, of the genes we have identified in this pathway in fission yeast encode proteins whose human homologues carry out similar functions in cAMP signaling. Therefore this model system has the potential both to advance our structural understanding of conserved signaling mechanisms and to lead to novel discoveries. In addition, the catalytic domain of fission yeast adenylate cyclase enzyme closely resembles those of several pathogenic organisms including the human pathogens Trypanosoma brucei and Candida albicans. While the cAMP pathway appears to be important to growth and differentiation of these organisms, nothing is known about the regulation of their adenylate cyclase enzymes. Therefore, our studies may suggest potential targets for inhibiting the growth or invasiveness of these pathogens. We propose to continue our work on adenylate cyclase activation by conducting genetic, molecular and biochemical studies of the fission yeast glucose/cAMP pathway. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: CONTROL OF BIOFILMS BY NATURAL PRODUCTS Principal Investigator & Institution: Costerton, John William.; Professor & Director; Sequoia Sciences 11199 Sorrento Valley Rd, Ste H San Diego, Ca 92121 Timing: Fiscal Year 2003; Project Start 01-APR-2001; Project End 31-JAN-2005 Summary: (provided by applicant): Chronic bacterial infections are serious medical problems in the United States. In chronic bacterial infections, biofilms protect bacteria from antibiotics and immune response mechanisms, thus increasing the rates of reoccurring symptoms and resistance to antibiotics. We discovered four novel compounds in Phase I under this STTR project that prevent the formation and disrupt biofilms, and we expect to identify additional novel compounds in Phase II. We propose to use the strategies developed in Phase I to prioritize the other active samples that have been identified. We will elucidate the structures of the active compounds and characterize their biological activity as biofilm inhibitors or antibacterials. We will also continue the discovery process for additional active samples. This work will enable us to commercialize these compounds that regulate biofilms and to further optimize or methods and strategies by which to discover more novel compounds that regulate formation of biofilms that are needed for a wide range of applications. In the United States, the market for microbial biofilm inhibitors is contained within the $8.5 billion market for antibiotics. Biofilms are involved in 65% of human bacterial infections; accordingly, biofilm inhibitors could capture a $4 to $6-billion segment of the antibiotic market. Biofilm inhibitors will have the greatest medical impact by treating many chronic infections, reducing catheter- and medical device-related infections, and in treating cystic fibrosis patients. Research has clearly established that biofilms play a significant role in these areas, representing a large market whose needs are unmet. The potential market penetration for potent biofilm inhibitors is exemplified by the sheer number of cases in which biofilms cause medical problems. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
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Project Title: DENDRITIC CELL DEFENSIN AND TYPE I IFN RESPONSES IN HIV Principal Investigator & Institution: Pope, Melissa J.; Scientist; Population Council 1 Dag Hammarskjold Plaza New York, Ny 10017 Timing: Fiscal Year 2003; Project Start 01-SEP-2003; Project End 30-JUN-2007 Summary: (provided by applicant): The dendritic cell (DC) system, comprising the myeloid (MDC) and plasmacytoid (PDC) subsets, orchestrates innate and adaptive immunity to pathogens. DCs are located within the epithelial tissues (Langerharis cells, LCs) and in the underlying lymphoid follicles (PDCs and MDCs) that line the oral cavity. Therefore, DCs are in prime positions to encounter oral pathogens and may play an important role in sustaining a healthy oral mucosa. Increasing evidence highlights how DCs both produce and respond to innate factors such as type I IFNs and defensins that possibly provide important barriers against HIV infection as well as in controlling commensal organisms in the oral cavity. We hypothesize that unlike HIV, organisms like herpes simplex virus (HSV) or Candida albicans will trigger strong innate type I IFN and defensin responses in DCs that contribute to the resistance of the oral mucosa to HIV infection as well as controlling HSV and Candida infections in healthy people. However, prior exposure to HIV will impede these innate DC responses rendering individuals more susceptible to HSV infection and reactivation as well as candidiasis. Three major questions will be addressed to investigate this. 1. Are defensin responses in LCs induced by HIV and organisms present in the oral cavity? 2. What are the innate type I IFN and defensin responses of PDCs and MDCs to HIV and oral pathogens? 3. Do epithelial cells and keratinocytes influence the innate responses of DCs to HIV and oral pathogens? By investigating these issues we will reveal the innate responses of distinct DC subsets that are found in the tissues of the oral cavity to HIV and related copathogens, how these responses are influenced by cell-cell (DCs, epithelial cells, and keratinocytes) contact, and whether prior exposure to one organism alters a DCs' response to another. The involvement of specific receptors (toll like receptors, TLRs and C-type lectin receptors, CLRs) will be examined to elucidate their role in capture of a pathogen vs signaling of cellular responses. These extensive in vitro studies will uncover pertinent information about the innate responses of DCs to various infections and how these contribute to a healthy oral environment that is perturbed in HIV infected individuals. This will afford critical insight as to how such responses could be boosted to prevent opportunistic infections and also identify potential targets for the development of strategies to prevent HIV transmission across the oral and other mucosal surfaces. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
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Project Title: DETERMINANTS IN PLATELET MICROBICIDAL PROTEINS Principal Investigator & Institution: Yeaman, Michael R.; Professor; Harbor-Ucla Research & Educ Inst 1124 W Carson St Torrance, Ca 90502 Timing: Fiscal Year 2002; Project Start 01-JUL-2000; Project End 30-JUN-2005 Summary: (Adapted from the Applicant's Abstract): Human and rabbit platelets contain platelet microbicidal proteins (PMPs). The investigators' data show that PMPs play key roles in platelet antimicrobial functions. PMPs exert potent microbicidal actions against antibiotic-resistant bloodstream pathogens, including Staphylococcus aureus and Candida albicans. Specific PMPs are released from platelets exposed to pathogens or agonists at sites of endovascular infection, and intensify locally at these sites. PMPsusceptible pathogens are less virulent in animal models than their isogenic PMPresistant counterparts. These compelling facts support their hypothesis that PMPs
16
Candida Albicans
significantly contribute to antimicrobial host defense. PMPs are minimally toxic to human vascular endothelial cells or erythrocytes, and differ markedly in structure and mechanism from antimicrobial peptides that are not released into the bloodstream. These facts suggest PMPs have key functional determinants that optimize microbicidal activity without concomitant host cytotoxicity. Beyond its microbicidal effects, they have discovered that PMP-2 also potentiates neutrophil chemotaxis, phagocytosis and intracellular killing of S. aureus. PMP-2 has a cystine-X-cystine (CXC) motif distinctive of alpha-chemokines such as human platelet factor-4 (hPF-4) that amplify antimicrobial mechanisms of neutrophils. These facts indicate that PMP-2 is a unique molecule that exerts both direct microbicidal and neutrophil-potentiating effects. The investigators' central hypothesis contends that PMP-2 has specific determinants responsible for these distinct host defense functions. They further hypothesize these determinants can be defined, modeled, and used to establish key structure-activity relationships (SARs) governing specific functions. The proposed studies are designed to explore these hypotheses. Defining SARs in PMP-2 functional determinants is crucial to their eventual goal of designing anti-infective agents with potent and/or selective activity against antibiotic-resistant pathogens. Therefore, their Specific Aims are: i) to define the structural determinants responsible for direct microbicidal functions of PMP-2; ii) define the PMP-2 structural determinants that potentiate the antimicrobial functions of neutrophils; and iii) establish the key SARs in antimicrobial determinants of PMP-2. Comparison of PMP-2 and hPF-4 determinants responsible for their potent and/or discriminative antimicrobial functions will enable future studies of human PMPs in the rabbit model that cannot be conducted in humans. Moreover, SAR themes discovered in PMP-2 will accelerate discovery of novel anti-infective strategies against pathogens resistant to conventional agents. Thus, these studies will significantly advance our understanding of antimicrobial host defense, and may yield new modes for its amplification. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: DEVELOPMENT OF ANTIFUNGALS OF CLINICAL IMPORTANCE Principal Investigator & Institution: Judd, Amrit K.; President; Synvax 969-C Industrial Rd San Carlos, Ca 94070 Timing: Fiscal Year 2004; Project Start 15-FEB-2004; Project End 31-JAN-2005 Summary: (provided by applicant): Fungal infections have increased dramatically in recent years to become important causes of mortality in hospitalized patients. The increase in life-threatening fungal infections has brought about an increased use of antifungal drugs and a pressing need for new, broad-spectrum, fungicidal agents that can be used empirically in immunocompromised patients e.g., AIDS and organ transplant patients. There is still a treatment failure of more than 50% among patients with acute invasive aspergillosis, and a 20 to 30% failures with candidemia. Current available therapies for treating fungal infections often suffer from drug-related toxicity, hazardous drug-drug interactions, non-optimal pharmacokinetics, and development of drug resistance. Preliminary studies have shown that several peptides from our chemical library have antifungal activity against Rhodotorula pilimanae, a nonpathogenic fungus. Three of the compounds tested so far showed activity against Candida albicans. These peptides are found to be nontoxic in several human and monkey cell lines and therefore show promise for further pursuing. It is proposed to conduct antifungal activity studies on all the peptides against Candida, Aspergillus, and Cryptococcus, the fungi of clinical importance. The specific aims of Phase I studies are (a) resynthesize all the compounds, (b) evaluate compounds for antifungal activity
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17
against Candida, Aspergillus, and Cryptococcus, (c) determine minimum inhibitory concentrations and minimum fungicidal concentrations, and (d) conduct in vivo experiment on the most promising compound against Candida, Aspergillus, and Cryptococcus. These experiments will include inhibition of colony forming incidences and survival of mice. Based on the data obtained in Phase I studies, Phase II studies will focus on designing metabolically stable analogs for oral activity using computer-based molecular modeling, developing pharmacologically-based and physiologic-based appropriate administration strategies, conduct extensive in vivo studies, determine the mechanism of action; conduct toxicology and pharmacokinetic studies, and file IND. The data generated by these studies will provide important information to raise this technology to a level of maturity where it can compete successfully for commercial funding to bring a new class of antifungal drugs to clinical use. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: DRUGS AND DELIVERY SYSTEMS FOR OPPORTUNISTIC INFECTIONS Principal Investigator & Institution: Miller, Marvin J.; George & Winifred Clark Chair Professor; Chemistry and Biochemistry; University of Notre Dame 511 Main Bldg Notre Dame, in 46556 Timing: Fiscal Year 2002; Project Start 01-FEB-1991; Project End 31-JAN-2004 Summary: (Adapted from Applicant's Abstract) The general goal of this proposed research is to develop new methods and agents for the treatment of opportunistic infections associated with AIDS and other diseases in which the immune system is compromised. Emphasis will be on the design and synthesis of new antifungal drugs and the development of new microbe selective antifungal delivery agents based on active iron transport processes that are necessary for assimilation of physiologically essential iron by pathogens. The general hypotheses to be tested are that the recently developed synthetic methods of the applicant will lead to the design of new antifungal agents (and perhaps other therapeutic agents), and that conjugates of antifungal agents with species selective microbial iron transport agents (siderophores) can be used to actively transport antifungal agents into cells of pathogenic organisms or that selective blockage of fungal iron trasport will inhibit fungal growth. Specific individual aims include: 1) Utilize methods developed by the applicant to prepare a library of siderophores and components to determine which can be recognized and used by opportunistic pathogens such as Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus (though many others will be included in broad screening). 2) Determine if modified forms of siderophores can block iron assimilation by select pathogens and lead to the development of new antimicrobial agents with a novel mode of action by inducing iron starvation. 3) Synthesize and study siderophore- antifungal agent conjugates, including "multiwarhead" conjugates, to determine if they can actively transport antifungal agents (drugs) into the cell or anchor the siderophore-drug conjugate in the cell membrane and, in either case, exert a lethal effect. Known and novel antifungal agents with various modes of action will be conjugated to the siderophores to determine a) optimal microbial selectivity and b) if mammalian toxicity of some drugs can be reduced by siderophore-mediated targeting of the drug to fungi. 4) Study the influence and importance of linkers [covalent (including amide, ester, hydrazone, oxime), ionic and novel "microbe triggered" release processes] between the siderophores and antifungal agents and determine if drug release is necessary. 5) Develop efficient syntheses of novel antifungal agents (and conjugates) including a) forms of potent antifungal neoenactins, b) carbocyclic nucleosides, c) peptidyl
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Candida Albicans
nucleosides and carbocyclic analogs, and d) novel "self delivering" antisense oligonucleotides. Detailed broad screen biological evaluation of all the antifungal agents and conjugates will help define important structure-activity relationships to demonstrate the therapeutic potential of microbially- targeted iron transport-mediated design of antifungal agents while enhancing the understanding of the essential role of iron assimilation and metabolism for virulence of opportunistic pathogens. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: ENDODONTIC INFECTIONS IN TYPE 1 DIABETIC HOSTS Principal Investigator & Institution: Fouad, Ashraf F.; Associate Professor; Restorative Dentistry; University of Connecticut Sch of Med/Dnt Bb20, Mc 2806 Farmington, Ct 060302806 Timing: Fiscal Year 2002; Project Start 30-SEP-2001; Project End 31-AUG-2004 Summary: (provided by applicant): The causative microbial pathogens and the fundamental host responses in teeth with pulp necrosis and periapical (PA) lesions have not been adequately characterized. Much less is known about these host/pathogen interactions in patients with type 1 diabetes mellitus (DM). We hypothesize that patients with type 1 DM have more symptomatic and/or therapy-resistant PA lesions, which may or may not be related to the degree of their glycemic control. We also hypothesize that in root canals with necrotic pulp of these patients, more virulent and more numerous species of endodontopathic microorganisms are present. The aims of this research project are: (1) Determine the effects of type 1 DM on the development of symptoms in patients with pulp necrosis and apical periodontitis, and on the resolution of the periapical lesion after one year, and (2) Determine the effects of type 1 DM on the prevalence of pathogenic bacteria and Candida albicans before and after root canal preparation in these teeth, using sensitive molecular techniques. Endodontic patients who have type 1 Dm or are non-diabetic, and who have at least one tooth with pulp necrosis and a periapical lesion, will be recruited for this study. Documentation of perioperative symptoms will be done using visual analog scale measure s for pain and swelling. Endodontic treatment will be completed in a standardized manner. Microbial samples form root canals of the teeth treated will be subjected to PCR amplification of the 16SrRNA gene of selected pathogenic bacteria or 18SrRna gene Candida. Analysis will include universal eubacterial identification, followed by species-level identification of the selected pathogenic organisms using specific oligonucleotide PCR primers. Molecular sequencing will be performed on the PCR product generated with universal bacterial primers, in order to identify other root canal bacterial species present. Sampling and molecular identification will be repeated just before oburation of the root canals. Patients will have follow-up examinations one year post-operatively, with standardized periapical radiographs. Type 1 diabetic and non-diabetic patients will be compared as to peri-operative symptoms, treatment outcome, number of microbial species in the root canals preoperatively and following canal instrumentation, and the association of microbial species and their quantitative measures with symptoms and treatment outcome. Microbial and clinical factors will also be related to the degree of glycemic control of the diabetic patients. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen
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Project Title: ALBICANS
ESSENTIAL
PROTEIN
SECRETION
GENES
OF
CANDIDA
Principal Investigator & Institution: Wong, Brian; Associate Professor; Internal Medicine; Yale University 47 College Street, Suite 203 New Haven, Ct 065208047
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Timing: Fiscal Year 2002; Project Start 01-APR-2000; Project End 31-MAR-2004 Summary: Description (Adapted from abstract): Candida albicans causes more serious infections in humans that any other fungus. The C. albicans genome program is making steady progress and it is expected that > 6000 genes will be sequenced within the next year. The function of many of these genes can be studied by gene disruption and phenotypic analysis, but essential genes cannot be disrupted without loss of viability. The overall goals of this project are to: 1) develop new approaches for studying essential C. albicans genes using two secretion pathway genes as models and 2) use these approaches to study intracellular transport and secretion of two virulence associated C. albicans proteins. In Saccharomyces cerevisiae, SEC4 and YPT1 encode small ras-like GTPases that are required, respectively for fusion of post-Golgi secretory vesicles to the plasma membrane and for ER-to-Golgi protein transport. The SEC4 and YPT1 genes of C. albicans have been cloned and sequenced. When gene disruption experiment suggested that C. albicans SEC4 was essential, it was found that over expressing a mutant sec4 allele similar to those encoding dominant inhibitors of other ras-like GTPases inhibited growth, protein secretion and fusion of secretory vesicles to the plasma membrane in C. albicans. These results demonstrated the feasibility of using molecular approaches to study essential C. albicans genes. Aim 1 is to i) generate C. albicans strains with temperature sensitive and/or inducible dominant-negative sec4 mutations and ii) determine if double-stranded RNAs can block expression of SEC4 and other C. albicans genes. Aim 2 will define the functions of C. albicans SEC4. The C. albicans strains from Aim 1 will be tested for growth an survival, morphology and germ tube formation, ultrastructure, and the ability to transport and secrete aspartyl protease and phospholipase B. Aim 3 will generate C. albicans strains with loss-of-function ypt1 mutations and to use these mutants to define the functions of C. albicans YPT1. Website: http://crisp.cit.nih.gov/crisp/Crisp_Query.Generate_Screen •
Project Title: FOCUSED PARALLEL SYNTHESIS OF DICATION ANTIFUNGAL AGENTS Principal Investigator & Institution: Tidwell, Richard R.; Professor of Pathology and Laboratory Me; Pathology and Lab Medicine; University of North Carolina Chapel Hill Office of Sponsored Research Chapel Hill, Nc 27599 Timing: Fiscal Year 2002; Project Start 01-FEB-2001; Project End 31-JAN-2004 Summary: The proposed studies stem from our previous research on the antifungal activity of dicationic molecules. These initial in vitro studies on over 300 dication molecules showed that leading compounds were both inhibitory and fungicidal against Candida albicans and Cryptococcus neoformans with MIC80S of
