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Technology Available

Potent vaccine candidate against T.B

Summary/Background/Description:
The BCG vaccine provides immunity to TB only during childhood, this protection lapses as time goes by and the vaccinated children reach adulthood. Furthermore BCG vaccine as per clinical trials does not confer any protection in the TB endemic areas. The present stratagem seems to overcome this problem by providing long term immunity in population and being a lipid –peptide conjugate requiring minimal antigen processing, also bypasses immune-mechanical hurdles to give immunity even in areas where there is a high occurrence of TB, mycobacterial loads.

Advantages/Value Proposition: Synthetic lipid-peptide conjugate as a potent vaccine candidate for developing long lasting immunity against especially TB pathogen and potentially against other wide range of intracellular parasites.

  • Technology can be adapted to create protective immunity against a wide range of intra-cellular pathogens
  • Peptide based vaccines work independent of pathogen load, hence are functional even in endemic areas of diseases
  • No autoimmune disease side effect due to absence of autoreactive parts
  • Self-adjuvating as it is potent enough to elicit a immune response by itself
  • Completely synthetic construct
  • Constructs can be used as prophylactics as well for population not previously vaccinated

Applications:
Its utility lies in development of a vaccine against the intracellular pathogens, which are causative agents of tuberculosis, brucellosis, leishmaniasis, listeriosis, leprosy, malaria, typhoid, trypanosomiasis, streptococcus, HIV, cancer, allergy, autoimmune diseases, etc.

Plasma Gelsolin Diagnostic Kit for Prematurity and Sepsis

CSIR-IMTECH has developed two mass-production ready plate based kits to estimate plasma gelsolin levels in humans. Plasma gelsolin is fast emerging as a health condition biomarker and its repletion in compromised cases is under Phase 1 and 2 trials. While, most other labs including commercial set-ups are focusing on improving bulk production of this protein, IMTECH has developed bonsai versions of this protein by structure-based insights. IMTECH successfully demonstrated the anti-sepsis properties of miniaturized versions in LPS-induced sepsis model of mice (Peddada et al., 2013). This project is part of the objective proposed in 12 FYP BioDiscovery where IMTECH proposed to design mini-proteins or peptides capable of rescuing sepsis condition. The kits which have been developed in IMTECH will help in determining a quantitative measure of the sepsis. Alongside, we are testing the kits viability in predicting cases of preterm birth. Both kits come with a cell phone based application for measuring the gelsolin value and providing a prognosis on the patient status.

Three patents have been filed in this regard and negotiations are underway with commercial partners for mass production and marketing of our kits.

Process for the production of peglated staphylokinase (SAK)

Summary/Background/Description:
New derivatives of Staphylokinase, having increased stability and extended plasma half-life have been developed. The gene encoding SAK has been modified for the PEG attachment and protocols for the purification and PEGylation, have been optimized. PEG-conjugated staphylokinase may prove better for the treatment of cardiovascular complications.

Advantages/Value Proposition:
As a clot bluster drug, staphylokinase is considerably more fibrin-specific than streptokinase. PEGylated staphylokinase derivatives have better potential for the efficient and cost-effective treatment of cardio-vascular diseases

Applications: To be used as thrombolytic drug for clot-dissolution.

Promising Novel Therapeutics for Strokes and Cardiac Maladies

Summary/Background/Description:
A lab-scale process for the preparation of PEGylated-streptokinase has been standardized. The process entails the culturing of E. coli carrying appropriate plasmid DNAs encoding for mutant streptokinase, followed by cell-lysis, refolding to their biologically active states, PEGylation reaction and their isolation by chromatographic means. The PEGylated clot-specific protein display plasminogen activation property as well as few additional superior properties listed below:

  • Prevention of excessive plasmin generation.
  • Less fibrinogen depletion as compare to streptokinase in clot dissolution experiments.
  • Up to five-fold increased half-life in test animals.
  • Upwards of 50% reduction in immune reaction.
  • Clot specific activation mechanism.

Advantages/Value Proposition:
This is a cutting-edge technology that has a truly international potential since it can compete with the most advanced clot-buster drug currently available (TPA, tissue plasminogen activator). The technology will be sufficiently scaled-up in-house so that after transfer to a commercial partner it has a minimum take-off period.

Applications:
A blood clot dissolving life-saver drug which is used for the treatment of myocardial infarction, stroke and other circulatory disorders. These clot-specific variants of streptokinase have been developed by protein engineering with increased half-life, decreased immune reactivity, less systemic plasmin generation and less fibrinogen depletion, and therefore highly promising therapeutic lead molecules with global reach.

Technology development for production of pullulan

Summary/Background/Description:

  • In CSIR-IMTECH, we have developed a process for pullulan production using an osmotolerant strain of Aureobasidium pullulans. It was possible to obtain more than 70g/L pullulan after process optimization. The yield and productivity of our process is higher as compared to already published literature. Moreover, the process of CSIR-IMTECH was developed using low cost agri-industrial residues and would result in significant reduction in the raw material cost vis a vis cost of production. Hence, our process is actually about creating wealth from waste.
  • The process has already been optimized in 5L fermenter scale. Initial testing about the scalability of the process up to 1500L fermenter scale has also been done and preliminary data suggested the process can be scaled up to 1500L scale.

Advantages/Value Proposition:
Pullulan is an extremely versatile ingredient with capability of providing a technology platform for product innovation. This has been used as a food ingredient for over 20 years in Japan. It has Generally Regarded As Safe (GRAS) status in the US for a much wider range of applications and thus higher intakes than the current application. It is an excellent film-former, producing a film which is heat sealable with good oxygen barrier properties and which can also be printed. Pullulan can be formed into capsules for use with pharmaceutical and nutraceutical products. Its non-animal origin ensures there are no safety concerns with the consumers and it is suitability for all the consumers groups. Chemically, pullulan is a homopolysaccharide ( neutral glucan) consisting of maltotriose units. In this polymer, three glucose units in maltotriose are connected by an α -1,4 glycosidic bond, whereas consecutive maltotriose units are connected to each other by an α -1,6 glycosidic bond.

Applications:

  • Due to several unique physicochemical properties pullulan has found wide application in food, pharmaceuticals and cosmetic industries. Recent literature suggested its potential towards application in the field of drug and gene delivery . The global market size of pullulan was 10,000 TPA ( $ 250 million) in 2009. It was expected to grow @ 6.75% per annum. More importantly, its major competitive products (gelatin and xanthan gum,) also have cumulative market size of more than $ 2 billion, which can also be tapped to enhance the total market of pullulan. The major manufacturer of this molecule are Hayashibara , NOF Corporation and Yushiseihin Co., Ltd ( all from Japan).
  • As of best of our knowledge, at present, there is no industrial manufacturer of this molecule in India.

Quantification of specific protein in sample for Diabetes diagnosis

Summary/Background/Description:
High blood glucose concentration, cause glucose to bind to Hemoglobin. Unlike fasting plasma glucose, the amount of HbA1c is indicative of average blood glucose levels over the last several months. Thus Hb1Ac level is a reliable indicator for Diabetes. A bifurcated device has been engineered that can segregate the total glycated and nonglycated Hemoglobin. This device can separately compute the relative amounts in percent, of the two types of protein, based on the strength of the protein-antibody binding signal.

Advantages/Value Proposition:
Method & device for quantitative measurement of the amount of Glycated protein (Hemoglobin) in blood serum.

  • Minimized Error: As assay detects both non-glycated and total hemoglobin using single Hb Antibody, thus further decreasing the chances of error.
  • Minimal False Positives: Anti-HbA1c antibody has been generated in chickens showing very less cross reactivity with other mammalian biomolecules.
  • Low cost, simple and field applicable

Applications:
Development of potential new kit for the detection and quantification of HbA1c for Diabetes patients.

Protein production in yeast for therapeutic products or Low cost production of therapeutic Proteins in Yeast

Summary/Background/Description:
Using Recombinant DNA Technology, single celled yeasts have been genetically modified to produce high levels of functional human proteins. Molecular cloning techniques were used to produce a system for the synthesis of the protein using the yeast's molecular machinery. These unicellular gene expression systems can produce the desired protein in its final functional state, as shown with HBS protein that can be used as a vaccine. Furthermore it is projected that they can match, if not exceed the highest protein yield so far generated in yeasts that to in less than half the time, thus reducing the overall cost of protein production in comparison with the existing systems.

Advantages/Value Proposition: Genetically engineered fission yeast for biosynthesis of proteins at commercially viable levels of production.

  • Cost Advantage: Many fold decrease in cost of production as compared to other yeast sp.
  • Proteins so produced are readily functional, needing no further chemical conditioning to work
  • Potential for up to 12-fold increase in productivity towards HBS protein biosynthesis
  • Inexpensive media and better solubility of the protein and ease of purification

Applications:

  • Biopharmaceutical production of therapeutic proteins, enzymes and vaccines
  • Adaptable for metabolic engineering application for synthesizing high value biochemicals

Designer cells for biocatalysis

Summary/Background/Description:
A strategy has been devised to design microbial whole cell biocatalysts for the structural isofrom selective preparation of a wide variety of complex alcohols. This technology combines the best features of pure enzyme usage along with whole cell genetically engineered biocatalysts. Metabolic engineering approaches were employed to drive the reaction towards the production of a product with a specific structural isoform and optical activity. The genetically modified biocatalysts attached to outer membranes showed exponential increase in activity, high turnover and accumulation of concentrated product was thus achieved with no membrane permeability issues for substrate or product.

Advantages/Value Proposition: Highly efficient cells designed for the biocatalytic synthesis of structurally complex alcohols and other class of organic molecules

  • Can be used with a wide variety of chemically related substrates that to at high concentrations
  • Automatic regeneration of cofactors built in, enhanced conversation rate
  • No osmotic, active transport limitation of substrate or product as enzyme on outer membrane
  • High quality product yield in a short processing time period and small space
  • Structurally pure, optically active product

Applications: Preparation of a large variety of important pharmaceutical intermediates with specific chirality, the technology platform can be modified for the production of other class of organic compounds used in pharmaceutical industry.

Novel Uricases, their mutants and their applications thereof

Summary/Background/Description:
Human Uricase gene is non-functional and thus the body becomes diseased at high serum uric acid levels caused by metabolic disturbances, which deposited as crystals in joints, kidneys and soft tissues. Using Recombinant DNA Technology, Uric Acid oxidizing enzymes (Uricases) with a novel sequences have been engineered for their use as Serum Uric Acid (SUA) lowering agents. Their protein sequence has been changed such that they are quite stable at biological pH of humans and thus have maximum activity in breaking down uric acid at physiological pH.

Advantages/Value Proposition: 
Genetically modified Uricase enzymes which are optimized for maximum biocatalytic activity at human physiological pH for treatment of Hyperuricemia and Hyperuricosuria. These recombinant enzymes show high stability at biological pH itself These novel enzymes have many fold higher biocatalytic efficiency at physiological pH than their pre-existing counter parts.

Applications:

  • The uricase mutants can be used in clinical diagnosis kits to measure the SUA levels
  • It theoretically has a great potential to be used as a fast acting therapeutic agent in the future, retaining nearly complete activity at physiological pH to significantly reduce SUA levels. They could be employed for the treatment of chronic gout and tumor lysis syndrome.

Biomolecule drug against T.B. pathogen

Summary/Background/Description:
A small genetic molecule has been constructed that can specifically bind to the gene responsible for the expression of a critically important protein based biocatalyst (enzyme) –Peptide deformylase in Mycobacterium tuberculosis. By blocking the particular part of the genetic machinery responsible for the stability and proper functioning of the pathogen enzyme, the bacteria growth is inhibited. Thus this novel molecule validates peptide deformylase as a potential drug target for new mycobacterial antibiotics.

Advantages/Value Proposition: Synthesis and targeting of a novel nucleotide biomolecule based gene silencing drug against T.B. microbial pathogen –Mycobacterium spp.

  • Specifically targeted to pathogen genetic material
  • Establishes the genetically suppressed peptide deformylase enzyme as a drug target in this microbe

Applications:
A novel biopharmaceutical drug to combat T.B.

Production of novel high redox potential laccase enzyme

Summary/Background/Description:
In CSIR-IMTECH, we have developed a process for production of a novel high redox potential laccase using a fungal strain isolated in CSIR-IMTECH. Laccase fermentation is carried out in a special surface culture tray reactor. The laccase produced is extracellular and easy to purify from the broth. Laccase fermentations generally have extended duration making it expensive to run. The CSIR-IMTECH laccase fermentation process is low energy intensive and based on moderate cost not complex medium. The process has been demonstrated on 10 L scale and has yield of 15,000 IU per liter.(1 unit of enzyme activity is the enzyme quantity that produce 1 μM of ABTS cation per min in 50 mM Citrate phosphate buffer, pH 4.0). The process can be easily scaled up to semi pilot or pilot scale.

Advantages/Value Proposition:
Laccase (EC 1.10.3.1) has broad enzyme specificity that creates very high application potential in various fields such as bioremediation, textile, pulp and paper, food and beverage, cosmetics and organic synthesis. Laccase production is difficult as it is produced in the idiophase when cell metabolism is slow. Low intrinsic production and long fermentation cycle increase laccase production cost.

The CSIR-IMTECH process has addressed some issues in the production of laccase and using a novel strain, a reliable process for the production of high redox potential laccase has been developed.

The laccase has been tested in industrial environment and has been found to give good performance in denim bleaching.

Applications:

  • Bioremediation: degradation of textile dyes, removal of endocrine disrupting substances (EDS)such as Bisphenol, polyaromatic hydrocarbons (PAH)
  • Textile industry (Bleaching of denim)
  • Pulp and paper industry ( Kraft pulp brightening)
  • Biofuel (increasing efficiency of cellulolytic enzymes.
  • Cosmetic (deodorants)
  • Medical (in situ release of iodine)
  • Organic synthesis 9Polymerization and synthesis of drug intermediates)

The availability of laccase is an issue. There is no manufacturer of laccase in India. Currently commercial laccase is available from the only firm Amano Enzymes, USA

IP status: Method for obtaining laccase enzyme from Arthrographis sp. PCT Patent Appn,No.WO 2012023021 A1

Awarded in China and Bangladesh.

Know-how for Alkaline Protease

Alkaline proteases have found increasing applications in detergent and leather industry. A strain of Bacillus sphaericus, isolated by IMTECH scientists showed protease activity upto 70,000 AU/litre in 100 L fermenter with a medium containing starch and soyatose. This know-how can be used for the production of a better quality enzyme for use in detergents and in leather industry for bating etc. The process is available for licensing. The advantage of IMTECH process is its thermotolerance at higher temperatures.

Know-how for Alpha Amylase

This enzyme is used in textiles and food industry and in many other applications. IMTECH scientists have cloned and expressed in B. subtilis, alpha-amylase gene from a thermophilic strain. The know-how is available for transfer at 100 L, process scale.

Contact Details:

Director
CSIR-Institute of Microbial Technology
Sector 39-A, Chandigarh- 160 036 (India)
Fax Nos.: 2690632, 2690585; Tel. Nos.: 2690785, 2690684;
Email: director@imtech.res.in
Web Page: http://www.imtech.res.in

Technology for production of novel high redox potential laccase enzyme

Summary/Background/Description:
In CSIR-IMTECH, we have developed a process for production of a novel high redox potential laccase using a fungal strain isolated in CSIR-IMTECH. Laccase fermentation is carried out in a special surface culture tray reactor. The laccase produced is extracellular and easy to purify from the broth. Laccase fermentations generally have extended duration making it expensive to run. The CSIR-IMTECH laccase fermentationprocess is low energy intensive and based on moderate cost not complex medium. The process has been demonstrated on 10 L scale and has yield of 15,000 IU per liter.(1 unit of enzyme activity is the enzyme quantity that produce 1 μM of ABTS cation per min in 50mM Citrate phosphate buffer, pH 4.0). The process can be easily scaled up to semi pilot or pilot scale.

Advantages/Value Proposition:
Laccase (EC 1.10.3.1) has broad enzyme specificity that creates very high application potential in various fields such as bioremediation, textile, pulp and paper, food and beverage,cosmetics and organic synthesis. Laccase production is difficult as it is produced in the idiophase when cell metabolism is slow. Low intrinsic production and long fermentation cycle increase laccase production cost.

The CSIR-IMTECH process has addressed some issues in the production of laccase and using a novel strain, a reliable process for the production of high redox potential laccase has been developed.

The laccase has been tested in industrial environment and has been found to give good performance in denim bleaching.

Applications:

  • Bioremediation: degradation of textile dyes, removal of endocrine disrupting substances (EDS)such as Bisphenol, polyaromatic hydrocarbons (PAH)
  • Textile industry (Bleaching of denim)
  • Pulp and paper industry (Kraft pulp brightening)
  • Biofuel (increasing efficiency of cellulolytic enzymes.
  • Cosmetic (deodorants)
  • Medical (in situ release of iodine)
  • Organic synthesis 9Polymerization and synthesis of drug intermediates)

The availability of laccase is an issue. There is no manufacturer of laccase in India. Currently commercial laccase is available from the only firm Amano Enzymes, USA

IP status:Method for obtaining laccase enzyme from Arthrographis sp. PCT Patent Appn,No.WO 2012023021 A1

Awarded in China and Bangladesh.