REPORTT h e N a t i o n a l I n s t i t u t e f o r B i o t e c h n o l o g y i n t h e N e g e v M a y 2 0 0 74

The NIBN: Bringing the applied potential of basic research to life This NIBN report comes after several NIBN activities and achievements that all of us can and should be proud of. These include the intensive preparations for the recent visit of the International Advisory Committee (IAC). On Dec 10-12, the IAC spent three intensive days at the NIBN hearing a detailed report on NIBN achievements and future plans, holding discussions with Israeli industrialists, and learning of the ongoing research at the NIBN through lectures by several Institute scientists or during visits to some of our labs. Unfortunately, Prof. Phil Needleman could not attend this visit. Prof. Sir Aaron Klug OM FRS and Prof. Raymond Dwek FRS, however, both conveyed their excitement at what is happening at the NIBN. We are on the right pathway, but must continue working towards the NIBN goals of conducting multi-disciplinary, convergent research with clear biotech--nology potential.The NIBN is designed to enable new advances in biotechnology and to serve as the glue that links academia and industry. As such, I believe much effort remains to be invested in developing this aspect of our mandate. As a discovery-driven biotechnology institute, the NIBN must seek out the other side of the coin, namely the realm of product-driven biotech--nology represented by industry. Thus, the last months have witnessed extensive efforts in initiating meetings with repre--sentatives from the industrial sector. While in the beginning, small companies were the target, in the last month, meetings with representatives of major concerns, including Dr. Aharon Schwartz, Vice President, Strategic Business Planning and New Ventures, Teva Pharmaceutical Industries Ltd. and Ruben Krupik Chief Executive Officer of Clal Biotechnology Industries (CBI) and Ofer Goldberg, Business Development Officer, CBI, were carried out, as well as with Dr. David Rosen from Pfizer. At these meetings, the structure and mission of the NIBN, as well as its specific and unique research goals were presented by selected scientists to demonstrate the scientific and technological potential of the NIBN. We hope this will be a start for the establishment of fruitful collaboration.At the international level, the connection of NIBN with Rosetta has been announced in a press release (presented on pages 6-7) and new connections has been established with scientists in many countries.The NIBN was also introduced by the production of the "NIBN: Bringing the applied potential of basic research to life" and "Research at the NIBN" booklets. Finally, the NIBN has also conducted several workshops and conferences, as summarized in this newsletter. I wish all NIBN members continued success. Prof. Varda Shoshan-Barmatz Director

The study of genetic diseases: An opportunity for the identification of novel and unpredictable gene functionsand recognition of novel targets for therapyProf. Ruth Parvari

The interest of our group lies in the identification and characterization of genes causing human diseases. The immediate benefits of such studies include an enabling of the molecular diagnosis of hereditary human diseases, thus permitting carrier detection and prenatal diagnosis. From a broader perspective, knowledge of the affected genes and their molecular cascades would be expected to contribute to better understanding of the pathogggenesis of the related diseases, ultiggmately leading to improved treatment, prevention and opportunities for the identification of novel drug targets.

The high birthrate of the highly consanguineous population of Negev Bedouins treated at the Soroka Univerggsity Medical Center, affiliated with BengGurion University, enables us to search for the genes causing genetic diseases. We are presently studying genes involved in several common diseases, such as autoimmune thyroid diseases, dilated cardiomyopathy and Steroid Sensitive Nephrotic Syndrome, the most common childhood kidney disease. In addition, our research focuses on other diseases with potential for the identification of genes involved in important pathways, such as congenital insensitivity to pain, hypoggphosphatemic rickets and the thoraggcoabdominal syndrome that consists of diaphragmatic hernia. The study of these diseases is expected to provide new strategies for the treatment of pain, osteoporosis and diaphragmatic hernia, respectively.

An example of the identification of an unpredictable gene function

comes from our earlier search for the gene mutated in a severe syndrome reported almost exclusively in Arabs named HRD (OMIM 241410) for congenital hypoparathyroidism, severe growth and mental retardation and facial dysmorphism. In this syndrome,

where the majority of affected indiggviduals die in their teens, hypoparaggthyroidism is caused by the absence of the parathyroid glands that in turn results in the deficiency of parathyroid hormone, causing a derangement of

calcium homeostasis in the blood. The mutated gene unexpectedly turned out to encode TBCE, a protein that acts to fold (chaperone) tubulin. A different mutation found in the same gene in mice leads to progressive motor degeneration and early death, and as

such, may serve as a model for human spinal muscular atrophy. Tubulin is the building block of microtubules, the major structure of the cellular cytoskeleton of all eukaryotic cells and is needed for many essential life

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Figure legend: The cytoskeleton of a patient’s cells (right side) in com--parison to control cells (left side), as revealed by tubulin staining. Upper portion: lymphoblastoid cells. Note the misorganized cytoskeleton. Lower portion: fibroblasts. Note the reduc--tion in density and arrangement of the cytoskeleton in the patient’s cells.

CongratulationsThe NIBN congratulates the following members of its International Advisory Committee for the world recognition of their research achievements:

Prof. Ada YonathRecipient of the Wolf Prize, 2007.

Nobel laureate Prof. Aaron Ciechanover,on his election to the National Academy of Sciences of the USA. In addition he has been elected to the Pontificial Academy of Sciences, The Vatican.

processes, including cell division and proper organelle positioning within the cytosol. Thus, tubulin production is a tightly regulated process involving many folding and chaperone proteins.

Most mutations in the genes encoding these proteins are probably lethal at a very early stage of embryogggenesis. However, the genetic lesions in TBCE which cause HRD syndrome in humans and progressive motor neuropggathy in mice could not fully abrogate the function of TBCE. This observaggtion, made in our laboratory in colabggoration with Prof. George Diaz from Mt. Sinai Hospital N.Y., investigating the same mutation in patients from Saudi Arabia, was the first demongg

stration of a disease caused by a defect in the tubulin folding and assembly pathway. The pleiotropic disease maniggfestations of HRD affects tissues with abundant microtubules, such as brain and testis. The specific absence of paragg

thyroid glands, with an accompanying normal development of other organs originating from the same embryggonic derivatives, and the occurrence of special bone thickening (osteoscleggrosis) and recurrent bacterial infecggtions in a subset of affected individuals are unexpected aspects of the derangeggment in tubulin physiology. Thus, we investigated the possibility that TBCE plays additional roles and indeed found that the TBCE protein interacts with proteins involved in other cellular pathways, thus implicating TBCE in a broad spectrum of functions. These findings will lead to a deeper underggstanding of regulatory processes that may be important in a wide range of pathologies, including cancer.

The study of another severe syndrome resulting in high childhood mortality and characterized by cystingguria, neonatal seizures, hypotonia, severe somatic and developmental delay, facial dysmorphism and reduced activity of all mitochondriallygencoded respiratory chain enzymatic complexes revealed that the basis of the disease is a homozygous deletion of 179,311 nucleotide base pairs on chromosome 2, covering a region which includes two known genes and two novel uncharggacterized genes. Ongoing efforts are concentrating on one of the genes in the deletion that turned out to serve an important regulatory activity impliggcated in many cellular pathways. A mouse model is being created in which this gene is missing, so as to verify its specific activity. Such an approach will enable elucidation of various developggmental processes.

The identification of the mutated genes causing the genetic diseases of the Negev population thus not only

enables this population prenatal molecular diagnosis but also holds promise for all mankind for better understanding of the pathogenesis of common diseases, leading to their treatment.

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Combating diseases such as cancer is a major goal for the new millennium. The paradox of so many antigcancer drugs is that their efficiency as killers of cancer cells makes them toxic to normal tissues as well. As such, extenggsive study is being directed at cancergtargeted therapy. In recent years, Prof. Angel Porgador has been explorgging a novel approach to cancergtargggeted therapy and imaging based on the recognition of modified carbohyggdrates expressed in a broad spectrum of cancers. These modified carbohyggdrates are recognized by natural cytoggtoxicity receptors expressed by certain cells of the immune system known as natural killer cells. Prof. Porgador has explored the recognition of human cancers (cell lines and primary tissues) by such receptors to produce recombiggnant receptors fused to a human antiggbody backbone for use in both therapy and imaging of human cancer cell lines growing in mice. In the future, he hopes to translate these studies on cancer therapy and imaging into human trials.

While stategofgthegart cancer theraggpies may hold promise, it is clear that early detection of cancer represents a major strategy for attaining significant reduction in cancergrelated mortalggity. In recent years, Prof. Porgador has become interested in developing techggnologies for the diagnosis of cancerous states. Cancer markers are molecules

New Weapons in the FightAgainst Cancer

found in blood or tissue that are assoggciated with cancer and whose identiggfication or measurement is useful in patient diagnosis or clinical manageggment of the disease. The ideal marker would be detected by a ‘blood test’ for cancer in which a positive result would only be obtained from patients with malignancies, in which the result would correlate with the disease stage or response to treatment and which could be easily and reproducibly meaggsured. At present, no cancer marker meets these criteria. Accordingly, Prof. Porgador serves as the coordinaggtor of a European Community grant intended to develop novel diagnosggtic tools for early diagnosis and monggitoring of cancerous states based on glycome and glycopeptidome analyggsis of blood samples from healthy and cancergbearing patients. For his part, Prof. Porgador is analyzing the pepggtidome with the aim of identifying

Prof. Angel Porgador

Mass spectrum of the blood peptidome from a cancer-bearing patient.

Immunohistochemical staining of ligands to NCR1 on primary human melanoma (upper panel) and dysplastic nevus (lower panel), employing NCR1-Ig as the detecting reagent.

blood peptides (specific peptides or peptide patterns) that are associated with the specific cases of human prosggtate and pancreatic cancers.

Prof. Porgador was awarded an M.Sc. with distinction and a Ph.D. from the Weizmann Institute of Science. He subsequently trained as a postdoctoral fellow at Duke University Medical Center and at the National Institute of Allergy and Infectious Disease, National Institutes of Health, USA. Upon arriving at BengGurion Univerggsity in 1998, Prof. Porgador established a laboratory for the study of tumor immunology in the Department of Microbiology and Immunology in the Faculty of Health Sciences. Today, the Porgador laboratory works towards (i) elucidating the mechanism of pattern recognition by natural killer cells and, in particular, the tumoral and viral ligands recognized by natural cytoggtoxicity receptors, (ii) early detection

and monitoring of cancerous states by analyzing the blood peptidome and glycopeptidome, and (iii) developing subunit vaccines against viral infecggtions based on virusgspecific epitopes and carrier peptides from heat shock proteins. In these studies, Prof. Porgaggdor has established longgstanding colgglaborations, in particular with Prof. Ofer Mandelboim (natural killers) from the Hebrew University, and with Dr. Rachel G. Lichtenstein (glyggcopeptidome) and Prof. Bracha Rager (subunit vaccines) from BengGurion University.

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postgdoctoral training was performed at the Weizmann Institute of Science in the laboratory of Prof. Dan Tawfik. During this period, I focused on the directed evolution of a particularly interesting enzyme family called serum paraoxonases (PON). These mammalian enzymes protect against organophoaphate toxicity and atheroggsclerosis, thus serving as protectors of human arteries. We evolved PONs for efficient expression in E. coli and later, for increased specific catalytic activities towards various substrates, including organophosphates. My second postgdoctoral training was performed in the laboratory of Prof. Steve Withers in Vancouver, Canada. During this period, I focused on the directed evolution of glycosyltransferggases and developed novel technologies to allow for high throughput screening of extremely large mutant libraries for enzymatic activities.

At the beginning of December 2006, I joined the faculty of Ben Gurion University and the NIBN and movedggwith my family to the Negev. The NIBN recently acquired a fully autogg

The engineering of proteins to include new functions represented a major challenge for biochemists over decades. In the past few years, however, a breakthrough in the field was achieved with the development of a novel approach for protein engiggneering termed directed protein evoluggtion. Directed evolution methodologg

Engineering Proteins withNew Functions

gies are based on natural Darwinian evolution in which the fittest variggants are selected from a population containing wide genetic diversity. To create such genetic diversity under laboratory conditions, we diversify the target gene and create DNA libraries. These libraries are then subjected to a powerful selection or screening assay in order to identify and isolate variggants with improved specific funcggtions. The selected genes then undergo iterative diversification and selection steps so as to identify and isolate variggants with optimized new functions. Recently, this approach has proven to be highly successful, yielding proteins showing increased stability under extreme conditions, increased soluggbility following expression in heterggologous systems, and enzymes with novel reaction and substrate speciggficities. This approach is particularly advantageous in those cases where no prior knowledge of a protein’s mechaggnism and structure is available.

During my postgdoctoral training, I focused on the directed evolution of enzymes with new properties. My first

Dr. Amir Aharoni

Novel methodology that allows the high throughput screening of large mutant libraries of glycosyltransferases. This methodology is based on selective entrapment of the fluorescently labeled transfer product in the cell and will be used for the directed evolution of cytosolic sulfotransferases with novel properties (adopted from Nature Methods, News Views, Aug 2006), highlightingDr. Aharoni’s paper.

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SEATTLE, Washington, U.S.A. and BEERgSHEVA, Israel — Rosetta Biosoftware announced today that the National Institute for Biotechnology in the Negev (NIBN) at BengGurion University of the Negev is deploying two unique, computer systems for the study of Bioinformatics. The two systems – the Rosetta Resolver® system for gene expression analysis and the Rosetta Elucidator® system for differential protein expression analysis – will be used to conduct a study on plant response to stress. Both systems will be used in conjunction to integrate transcriptomic and metabggolomic data for systems biology projggects carried out at the NIBN and the University’s Department of Microbiggology and Immunology, in collaboggration with the Weizmann Institute of Science and the Volcani Center for Agriculture Research. In addition, both systems are being considered for a national testgbed program, in which computational biotechnology tools will be made available to academic and industrial researchers for prolonged evaluation periods. NIBN plans to establish a testgbed program to assist researchers in identifying and prioritizing softggware and databases for biotechnology research. “We are very excited about the opporggtunities that the Rosetta Resolver and Rosetta Elucidator systems open to us,” said Eitan Rubin, Ph.D., the research scientist at the NIBN and Department of Microbiology and Immunology who is leading the pilot project. “The combination of industrial strength and easegofguse that both systems offer allows us to create a resource that will not only help us in our own research, but one we hope could be used by the entire research community in Israel.”“We are pleased to provide the Resolver and Elucidator systems for a

Ben-Gurion University of the Negev Deploys Unique BioSoftware System for a Pilot Study in Bioinformatics

pilot study to BengGurion University, and to have our product be considggered for a national testgbed program,” said Yelena Shevelenko, vice presiggdent and general manager of Rosetta Biosoftware. “The integration capaggbilities built into our products will be valuable for enabling crossgplatform research that is being done at BengGurion University. We are honored to support their systems biology research and provide the university with the robust data management and analysis capabilities for gene expression and comparative proteomics research.”

About the RosettaResolver SystemThe Rosetta Resolver system is a highgcapacity data storage, retrieval, and analysis solution for gene expresggsion data. The system is ideal for life science research organizations that need to assess compound specificity or toxicity, identify new genes or theraggpeutic targets or compare and analyze large databases of expression profiles. The Rosetta Resolver system combines flexibility and ease of use with highgperformance algorithms to produce a comprehensive solution for rapid analysis of gene expression data. The Rosetta Resolver system can accept and analyze data from a wide variety of expression profiling formats and applies Rosetta Biosoftware’s propriggetary error models to yield quality statistics for every gene expression measurement within the system. These statistics are automatically leveraged by Rosetta Resolver system analysis tools for optimal results. Life science research organizations that have licensed the Rosetta Resolver system include many of the top pharggmaceutical companies in the world, such as Aventis Pharmaceuticals, BristolgMyers Squibb, GlaxoSmithgg

The Tecan Evo 150 fully automated liquid handling and robotic system that was recently purchased by the NIBN. This system will facilitate the high throughput research that will be performed in Dr. Aharoni’s laboratory.

mated liquid handling robotics system to facilitate the high throughput research that will be performed in my laboratory. The robot will signifiggcantly speed up the screening process for proteins with novel functions and will open the possibility of managing projects which are extremely labor intensive, if performed manually. Our group will focus on engineering enzymes with broad specificity, such as cytosolic sulfotransferases (SULTs), in order to understand the molecular basis for their ‘promiscuity’. These enzymes play important roles in a variety of biological functions, such as modulating hormone and neurotransggmitter levels. They are also of signifggicant biotechnological importance, as they could provide a means for the detoxification of a wide variety of xenobiotics that possess steroidglike activity and that are increasingly being detected in drinking water. Using directed evolution, we aim to improve the detoxification properties of SULTs by implementing a new high throughput screening methodology that will allow for the screening of millions of mutant enzymes in parallel for increases in catalytic efficiency. Improved mutants enzymes may find ex vivo biotechnological applications, such as in bioremediation.

From Press Releases - February 13, 2007

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Kline and Merck & Co., Inc., as well as premier academic institutions such as the Harvard University Center for Genomics Research and the California Institute of Technology.

About the RosettaElucidator SystemThe Elucidator system’s advanced data management capabilities allow users to effectively organize, search for, and retrieve raw and processed protein expression data. The application serves as a core integrator for proteomics research by working directly with select instrument APIs, sample annoggtation systems, and common protein ID search engines. In addition, it provides a suite of advanced statistical analysis and visualization tools for data mining and discovery of potenggtial protein biomarkers and proteins that are differentially expressed

among different phenotypes or drug treatments. The Elucidator system combines these data management and analysis capabilities with the flexibility of a highly customizable environment: users can incorporate organizationgspecific workflows into customized visual scripts that leverage Elucidator system statistical tools and customergproprietary algorithms, as well as those available in the public domain.

The National Computational Biology Test BedIn this press release, the acquisition by NIBN of two new top-of-the-line computational platforms is described: Rosetta Resolver for microarray analysis and Rosetta Elucidator for Mass-spectrometry. With the addition of these two platforms to the range of computational tools available at NIBN, we were able to establish the National Bioinformatics Test-Bed. The test-bed is an initiative to facilitate the adaptation of computational biotechnology by providing expert knowledge and access tools to researchers in academia and industry throughout Israel. Rosetta Resolver – an industrial strength platform for microarray data analysis – is immediately available for any researcher in Israel who would benefit from its use. Rosetta Elucidator is currently being added to the test-bed. With these packages, we are able to offer state-of-the-art computational biotechnology to the research community in Israel.The purchase of the two packages was made possible by generous funding from NIBN and the Israeli Ministry of Science, and with a lot of help from members of the computer support group at the Faculty of Health Sciences, Ben-Gurion University.

Dr. Eitan Rubin

The Rosetta Elucidator product line is designed to meet the needs of both large and small organizations.

About RosettaBiosoftware RosettaBiosoftware is a leading provider of informatics solutions for life science research. Its comprehensive softggware solutions, including the Rosetta

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Screenshots from the Elucidator proteomics analysis platform

Resolver and Rosetta Elucidator systems, empower life scientists with advanced, scalable, and easygtoguse analysis platforms that accelerate discovery research. Rosetta Biosoftggware is a business unit of Rosetta Inpharmatics LLC, a wholly owned subsidiary of Merck & Co., Inc. (NYSE MRK).

Special EventsNovember - AprilConferences

ProteOn XPR36 Protein InteractionArray Systemorganized by Dr. Sharon EtzionBGU. Nov. 9, 2006

From Shrimp to Cancer Therapy –Pharmapox annual meetingorganized by Prof. Amir SagiEin Gedi, Israel. Nov. 9-12, 2006

GLYFDIS – The European Commission6th Framework Program meetingorganized by Prof. Angel PorgadorDead Sea, Israel. Dec. 7-9, 2006

NIBN workshop No. 4. Sede BoqerFeb. 19, 2007

The Negev Systems BiologyGet Together. A mini- conferenceorganized by Dr. Eitan RubinBGU. Feb. 20, 2007

Visit of the NIBN InternationalAdvisory Committee (IAC) includingProf. Sir A. Klug and Prof. R. DwekDec. 10-12, 2006One day was devoted to a workshop, divided into two sessions:

A. Establishing collaborations with industryThis session included scientists fromthe NIBN and BGU, as well asrepresentatives of biotechnologicalindustries and focused its discussionson how to bridge the gap betweenacademic research at the NIBN andvarious branches of industry involvedin biotechnology and innovation.

B. Pharma and NIBN collaboration: A new approach for protein crystallizationIn this session, examples of novelinnovations with potential applicabilityto biotechnology involvingbiotechnology companies, NIBNscientists and the use of NIBN corefacilities were presented

A meeting with delegations ofvisiting industrialists:Teva Pharmaceutical IndustriesDr. Aharon Schwartz, Vice President,Strategic Business Planning andNew Ventures

CBI (Clal Biotechnology Industries)Mr. Ruben Krupik, Chief ExecutiveOfficer, Mr. Ofer Goldberg, BusinessDevelopmentB.G. Negev TechnologiesMr. Neta Cohen, CEO, Dr. Ora Horovitz,Business DevelopmentAt these meetings, the NIBN was

presented by Prof. VardaShoshan-Barmatz and NIBNresearchers: Dr. Ohad Medalia,Dr. Amir Aharoni, Dr. Ohad Birk,Prof. Ruth Parvari, Dr. Eitan Rubin,Dr. Ariel Kushmaro, Dr. Vered Caspiand Dr. Micha Volokita, who presentedtheir projects, followed by a discussionon future collaboration betweenCBI and NIBN (March 18, 2007)

Ongoing meetings with businessdevelopment consultants:Dr. A. Eitan, M. Shavit andDr. D. Graeser

Meeting with Dr. James C. Stevens from The Dow Chemical Company, Performance Plastics & Chemicals (December 13, 2006)

A delegation from the Natural Science Foundation of China (NSFC) visited the

NIBN (January 11, 2007)

Industry Contacts and Visits to NIBN

The National Institute forBiotechnology in the NegevBengGurion University of the NegevBeergSheva, 84105, IsraelTel: 972g8g6461912Fax:972g8g6272983Website: nibn.bgu.ac.ilEgmail: nibn@bgu.ac.il