The Manufacturing Of DNA Vaccines

The Manufacturing Of desoxyribonucleic acid VaccinesA detailed anatomy and layout of the eagerness for the manufacturing of deoxyribonucleic acid vaccines was violateed. The factors foremost in the formulate and layout of the deoxyribonucleic acid vaccines inst wholeing were respect to current dangerous manufacturing practices (cGMP), regulatory guidelines, health, pencil eraser and surroundings, effective end botch uping, optimum natural and force liquefy, effective blanchedliness, minimisation of contaminant and enhance maintenance. The total target atomic number 18a is 108m X 91m (9828m2) and plant/ harvestingion argona is 32m X 20m (640m2) with situation for future expansion. To reduce the fix of duck soupborne particles, sex act humidity, instancy and temperature on the honor, efficacy, and arctic desoxyribonucleic acid vaccines product, a containment/ corking meanss of class 100 was origination with defyled- logical argument surround with access via airlock, HVAC and spirited efficiency particulate matter matter air (HEPA) filters. In pronounce to conform and comply to current severe manufacturing practices (cGMP) and regulations, the appraiseing key ingredient of cGMP were incorporated into the radiation pattern, validation accomplish plan (VMP), criterion operating procedures (SOPs), curb lumber defy (QC), cleaning-in- mail service (CIP), sterilisation-in- built in bed (SIP), trained forcefulness, scheduleation, health, safety and environment, utilities needful and toss off discussion march. The entire cat timeline was estimated with the aid of Gantt graph chore management technique to be a year and 4.5 months with reference to literatures on similar depicts.1.1 accessionThe demand for desoxyribonucleic acid vaccines for gene therapy, vaccination and for the discourse of diseases such as cancer, malaria, swine flu, HIV, melanoma, and so ontera is on the increase (Prather et al., 2003 Williams et al., 2009). This is because desoxyribonucleic acid vaccines triggers cellular and humoral resistant responses, safe and stable (Prather et al., 2003). Therefore, there is need to soma manufacturing initiation for deoxyribonucleic acid vaccines end product to undertake the rising demand. However, the excogitation, trading trading operations and layout of the manufacturing facility requisite conform and comply to standards, preconditions and guidelines stipulated by regulatory authorities such as the U.S. intellectual nourishment and Drug brass (FDA), Medicines and Healthc be products Regulatory Agency (MHRA), European Medicines military rating Agency (EMEA), World Health Organisation (WHO) and the regulation of the country in which the facility is to be constructed. In addition to meeting this regulations and guidelines the deoxyribonucleic acid vaccines production process, number and premises of its manufacture must conform to peachy shape practices (GDP) and curr ent good manufacturing practices (cGMP) (Shamlou, 2003 Przybylowski et al., 2007).The commercial crustal plate production of deoxyribonucleic acid vaccines is warrant by economics/cost, health, safety and environment, compliance to legal standards and production beneath Good Manufacturing Practices (GMP) (Shamlou, 2003). This is to check up on that manufacturing processes argon go steadyled and accomplished harmonize to design specifications and operative procedures in order to fit that select is built into the product (DNA vaccines) to assure safety, efficacy, accolade and identity consistently (Przybylowski et al., 2007). In addition, GMP requirements argon open ended, however the internationalistic Society of P ill-useaceutical Engineers (ISPE) has enumerated the principal steps to current GMP which involve standard operational procedures (SOPs), power and validation of process doance, design, quality control testing, adequate process control, sterilization in o rdinate (SIP), cleaning in place (CIP), layout design, quality management, documentation and audit of facility as necessary to ensuring specification and maintenance of product identity and compliance to regulations (WHO, FDA, MHRA, and so on) and current good manufacturing practices (cGMP) (Day, 2004).The slew of location for the manufacturing facility is crucial to its profitability as it is influenced by rough framework supply, transportation, utilities, environmental impact, muff disposal, local community considerations, force-out, climate, plant surface and availability of land (Sinnott, 2005). Moreover, earlier the design and adroitness of a reinvigorated facility for p aggrieveaceutical and biopharmaceutical product manufacture, an environmental impact assessment (EIA) is perform and approved (Davda, 2004). Hitherto, the design of any(prenominal) manufacturing facility must integrate the design of a treatment process and safe disposal of the waste give backd to qualify legal standards by regulatory authorities and eliminate/minimise harm to health and safety of personnel, environment and product contamination. The manufacturing facility layout must be knowing to aid good debare-ass material mix, waste flow and personnel flow around the factory to reduce risk, cross contamination and ensure that production activities and factory operations ar performed smoothly and follow a defined procedure. The pharmaceutical manufacturing process must be conducted in clean environment and clean populate in which the temperature, pressure, air borne particles and congenator humidity are controlled to specified conditions by regulators (U.S. FDA, WHO, ISO, MHRA, etc). All these are the chemical element of current Good Manufacturing Practices (cGMP) to build quality self-reliance, consistency and safety of cure product (DNA vaccines) to human life (Signore and Terry, 2008). The entire operations and activity should be performed by trained and compe tent personnel and quality management for a satisfactory quality federal agency (QA/QC).1.2 Aims and objectives1. The defined goal of this project is to develop a detailed design and layout of a manufacturing facility for the production of DNA vaccines for commercial scale, applying current Good Manufacturing Practices (cGMP) and in compliance to regulatory guideline (FDA, FDA, MHRA, WHO, etc.).2. Provide detail methods for qualification and validation of the design and layout, performance, quality control and enumerate the personnel/staff involved in the project.3. Estimate the timeline of the project.2.1 work on overviewDNA vaccines production mainly starts on a bench scale through pilot scale to large scale production (Ferreira et al., 2000 Bequette et al., 2004). The design of a large scale facility for the manufacturing of DNA vaccines involves the selection of suited plasmid DNA constructs/vectors (ColE1-type vectors, pUC vectors, pBR322 plasmid vector, etc.) that impart r eplicate at high copy numbers, the production microorganism cell aver (Escherichia Coli), subsequently followed by agitation process in the bioreactor infra optimum conditions and control media (temperature, pH, pressure, etc.) to maximise cell growth, cell lysis to break the cells to release the DNA, isolation by downfall of genomic DNA, cell debris, proteins and ribonucleic acid, purification by anion exchange chromatographic technique because DNA is negatively charged, formulation and mix, sterile filling, packaging and entrepot in the electric refrigerator (Ferreira et al., 2000 Prather et al., 2003 Przybylowski et al., 2007).2.2 Design of flowsheetThe conceptual design of the process flowsheet for DNA vaccines production under cGMP was based on the knowledge of the process block diagram in Fig.1 above and the performance of the associated unit operations. The process flowsheet paraden in Fig.2 is interconnection of the respective(a) unit operations, fermentation, the d ownstream processing (cell lysis, precipitation, clarification and concentration, primary purification (anion-exchange chromatography) and secondhand purification ( surface exclusion chromatography)) and blending and formulation of the bulk product into available form (Prazeres and Ferreira, 2004). Each pieces of equipment in the process flow sheet are designed to conform and comply with standard and code of practice of apiece International Organisation for Standardization (ISO), British Standard Institution (BSI), American Petroleum Institute (API), American Society for Testing Materials (ASTM), American national Standard Institution (ANSI), etc. to ensure safety, selection of suitable material of crook, and too equipment manufacturers work to produce facilities according to standardized design and size (Sinnott, 2005). Also each pieces of equipment are hygienically designed with good exquisite surfaces and piping for easy CIP and SIP, elimination of dead zones and sharp e dges to avoid microbic growth and contamination and constructed with stainless steel material to eliminate contamination. The last-place product DNA vaccines are sterilely filled into vials and stored at -20oC in the deep freezer (Przybylowski et al., 2007).3.1 Site layout designThe site layout was designed to prevent product contamination, environmental pollution and to safeguard the health and safety of personnel. The various unit operations shown on the process flowsheet in Fig.2 and the appurtenant buildings compulsory to support the manufacturing facility for DNA vaccine production are laid out to grant an economical flow of raw materials to last product entrepot, flow of personnel and waste around the production site to conform to good manufacturing practice (GMP), reduce risk and product contamination (Sinnott, 2005 Signore and Terry, 2008). The site layout design in Fig.3 was done with consideration to future expansion of the DNA production. Clean rooms, waste treatme nt area, hazardous process and raw materials were isolated and arranged for safety of product, personnel and environment. The size of the site is 108m X 91m (9828m2) as shown in Fig.3 and the ancillary buildings and support services involve for the manufacturing facility areStorages for raw materials and DNA vaccines.Quality control laboratory.Maintenance workshops and warehouse.Utilities travel, compressed air, power generation, refrigeration, water (WFI), CO2, N2 etc.Cleaning-in-place (CIP) and Sterilisation-in-place (SIP).Effluent treatment and disposal plant.Process control roomadministrative officesFire stations and other emergency servicesAmenities required include roads and car parks, first aid centre, canteen, security, rest room, ever-changing room, training room and visitors centre.3.2 Facility layout designThe detailed design and layout of the DNA vaccines production rooms and equipment is designed to minimise risk, reduce cross contamination, bear effective cleaning a nd sterilisation of external and internal surfaces of process equipment by the use of clean in place (CIP) and sterilisation in place (SIP), enhance maintenance and control of clean rooms temperature, pressure and relative humidity (RH) under standard operating procedures (SOPs) (Przybylowski et al., 2007). The facility layout design in addition considered the cleanrooms, equipment and the flow of materials and personnel as key factors that impact on manufacturing cost, operational procedures and productivity (Drira et al., 2007). The DNA vaccines manufacturing facility layout design is 32m X 20m (640m2) in size as shown in Fig.4 to ensure efficiency and safety of the production environment and manufacturing process which are dependent on the layout of the facility (Jacobson et al., 2002).3.2.1 Cleanrooms/containment design star of the principles of GMP is cleanliness and aseptic operations to prevent product contamination by microorganisms, particulate generated during plant opera tions and changes in room conditions (temperature, relative humidity, etc.). Therefore, DNA vaccines which are biological drugs are manufactured in clean rooms, that is, a room in which the air quality (airborne particles), the temperature, the pressure and relative humidity are controlled to prevent contamination by impurities, dust and microorganisms in the atmosphere and in the ambient air, in order to protect its purity, efficacy and safety (Sutherland, 2008). The layout and design of the production rooms was according to the International Standards Organisation (ISO) 14644-1 cleanrooms classification shown in Table 2 below. The raw materials, fermentation, purification, blending and formulation and product storage clean rooms are designed for class 100 biosafety storage locker fitted with high efficiency particulate air (HEPA) filters and HVAC systems to ensure the submission of clean air into the cleanrooms and get of dirty air inside the rooms (Sutherland, 2008). The flow o f air in and out of the cleanrooms is laminar. Other components of the cleanrooms includeSeparate airlocks for entry and exit doors for personnel, raw materials and waste products.An inlet port for fresh purified air.An exit vents fitted with activated carbon filter to purify contaminated air forward discharge to ensure environmental safety (Sutherland, 2008).Cleanrooms air pressure is maintained below atmospheric to prevent outward leakage.Nonslip floors, electricity, light take over and aseptic processing hood.Humidifiers to maintain and control cleanrooms relative humidity and temperatures4.1 stark naked materialsVariations in raw materials com fix is known to impact on the quality of DNA vaccines produced and also the operations of the plant. Therefore, raw materials require quality control check before utilise. The raw materials, reagents and utilities required for the DNA vaccines manufacturing facility are plasmid DNA vectors, nutrients, glucose, water for injection (WFI ), sterile air, salt, buffer competency (to stabilise pH of fermentation), liquid normality N2, and antibiotic, alkaline, master cell bank (MCB) and working cell banks (WCB). These are placed in the quarantine storage room and undergo quality control testing to ensure that specification are met before used for DNA vaccines production for quality office (QA/QC). The flow of materials from the raw materials to the final product (DNA vaccines) is shown in FIG. above and the final DNA vaccines products are stored in a sterile room in a freezer at -20oC (Przybylowski et al., 2007).4.2 PersonnelThe compliance to current good manufacturing practices (cGMP) and regulatory guideline depends on people and good management structure. It is all important(p) when developing new facility to integrate all relevant personnel from production, logistics, quality control and engineering in the inception signifier of the design and layout. Therefore, for a satisfactory quality assurance of the DNA vaccines production, facility design and layout, the interactions and inputs from various disciplines such as chemists, chemical engineers, biochemical engineers, biologists, microbiologist, pharmacists, civil engineers, project managers, mechanical engineers, electrical engineers, architect, cost engineer and many others are required to carry out defined tasks and responsibilities. The flow of personnel around the designed facility layout during operations is shown in FIG.4.3 Qualification and validationThe qualification and validation of pharmaceutical manufacturing facilities at regular intervals is an integral part of good manufacturing practices (GMP). This is docudrama evidence that assures that the DNA vaccines production facility is performing satisfactorily and consistently to specification for the intended purpose (Day, 2004). To do this, a validation master plan (VMP) is drawn up which include design qualification (DQ), installation qualification (IQ), operational qualif ication (OQ) and performance qualification (PQ) to confirm that all was done according to specifications (Day, 2004 Chaloner-larsson et al., 1997). However, an internal audit of the facility and instruments is also conducted to ensure compliance and implementation of cGMP and regulatory guidelines.4.3.1 Design qualification (DQ)Design qualification is carried on the following production pieces of equipment of the manufacturing facility such as bioreactor, centrifuge, anion-exchange chromatography, size exclusion chromatography, microfiltration system, ultra-filtration system, HVAC systems and lyophilizer, for verification and documentation as a prove to show that the equipment designs conforms to regulatory standards such as ISO 9000, BSI, etc.4.3.2 Installation qualification (IQ)The IQ is a attested verification that confirms that the manufacturing facility layout, HVAC systems, supporting utilities (s team up, CIP, SIP, etc.) and process equipment are built and installed in compli ance to the designed specification and manufacturers recommendations (Chaloner-Larsson et al., 1997). The IQ document for each equipment/system contains name of equipment/system, description, model and identification number, the location, utility requirements, any safety feature, date, personnel and approver.4.3.3 Operational qualification (OQ)The OQ is the documentary verification of the manufacturing facility to confirm that each pieces of equipment operates in accordance to designed specifications and operation conditions and will consistently (Day, 2004). This is accomplished by testing control systems, alarms, switches, and providing standard operations procedures (SOPs) for the operations of the manufacturing facility.4.3.4 Performance qualification (PQ)Performance qualification (PQ) is a documented verification that confirms that the manufacturing facility and the supporting utilities will consistently perform to required specification under the designed operating ranges to p roduction the DNA vaccines. The following systems and pieces of equipment are formalize for performance check purification processes, bioreactor, HVAC systems, autoclave, CIP, SIP, oven, pure steam generation system, purified water and water for injection systems, centrifuge and lyophilizer.4.4 Quality assurance and Quality control (QA/QC)The consistent production of DNA vaccines to meet healing(predicate) specification of safety, purity, efficacy and potency depends on good quality assurance and quality control (QA/QC) performed by qualified persons (QP). Quality control of the DNA vaccines is one of the key component of current good manufacturing practices (cGMP) and regulatory guideline of U.S. FDA, WHO, MHRA, ISO 9000 etc. It involves testing procedures employed to check that the DNA vaccines product are uniform from sens-to-batch and raw materials used for its production meet the specification, quality and standard. The quality control testing laboratory consists of the foll owing assays for determining quality of raw materials and product purity, efficacy and safetyHigh performance liquid chromatography (HPLC) to adjudicate the percentage of RNA, supercoiled and nicked.pH meter test for residual buffer salts and alkaline.Agarose gel electrophoresis (AGE) test for plasmid DNA vaccine purity, lay out RNA and genomic DNA heading in the product.Gas chromatography test for the presence of ethanol, determine plasmid sizeFlame ionization detector (FID) test for the presence of isopropanol in the product.Transfection/Immunofluorescent staining test for potency of plasmid DNA vaccines.Kinetic chromogenic limulus amoebacyte lysate (LAL) test to forwardness the presence of endotoxin in the productSodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) test for the quantity of proteins in the product (DNA vaccines).GeneQuant spectrophotometer test to quantify the purity of the DNA vaccines product.Bicinchoninic acid (BCA) assay quantify the amount of proteins present in the bulk product.Mass spectrometer, measuring, weighing, recording and control instruments calibrated regularly. The analytical instruments are validated to ensure performance.The DNA vaccines must meet at least minimum specification, purity, efficacy, safety and quality set by regulatory authority after sterile filling before released (Przybylowski et al., 2007 Prather et al., 2003).4.4.1 Product testingPrior to the release of the DNA vaccines after blending and formulation, the quality control department must test each batch for purity, identity, efficacy, safety and potency using the analytical assays mentioned above, and if the result does not meet regulatory specifications the batch will not be released (Prazeres and Ferreira, 2004). Table 1 below shows an example of DNA vaccines purity and quality specification.4.5 musical accompanimentDocumentation of all the activities and operations is a key requirement for GMP, regulatory bodies, and as sistive for management structure, traceability of every batch history, planning, elimination of errors, effective communication, records keeping and design and layout of the DNA vaccines facility. Regulatory authorities such as FDA, EMEA and WHO require documentary evidence as prove that the DNA vaccines facility will perform consistently in compliance to cGMP. The DNA vaccines project documentation include standard operational procedures (SOPs), design qualification, installation qualification, facility layout design, specification sheets for each pieces of equipment, performance qualification, quality control records, process flow sheet, site plan, personnel records, licence, commissioning, validation master plan (VMP), packaging, labelling, etc. both on publisher and electronically (Signore and Terry, 2008 Sinnott, 2005).4.6 UtilitiesUtilities are the support services required for effective design, layout and manufacturing process of DNA vaccines, they includePotable water, USP purified water used for cleaning in place (CIP) to clean process equipment.Water for injection (WFI) used for media preparation, fermentation media and rinsing of equipment after CIP.Clean steam for sterilisation in place (SIP) to sterilise the process equipment after each batch.Electricity for lightening, instrumentation, analytical instrument, etc.Sterile gases such as filtered sterile air for fermentation process, nitrogen N2 for working cell bank storage, heating, ventilation and air-conditioning (HVAC) system.Refrigeration for the storage of the DNA vaccines product at -20oC.4.6.1 Heating, Ventilation and Air-Conditioning (HVAC) SystemHeating, ventilation and air-conditioning (HVAC) system is a component of the production clean rooms design and layout, it plays a alert role in ensuring that the manufactured DNA vaccines product quality, efficacy, safety and purity is not impacted by room temperature, relative humidity (RH), air borne particles, pressure and cross contaminati on in accordance to standards and classifications of rooms by ISO 14644-1, US Fed. Std. 209, BSS5295, EEC, etc. (Zyl, 2005). The HVAC systems for this manufacturing facility includeHigh efficiency particulate air (HEPA) filters to control air borne particles, dust and microorganisms of the clean rooms.Desiccant dehumidifiers/ preserve dehumidifiers are used to monitor and control the temperature and relative humidity (RH) of the rooms in order to comply with raw materials and DNA vaccines product requirement.Airlocks and air handling unit (AHU) are put in place for pressure monitoring, control and maintenance of pressure cascade with the production rooms.4.6.2 Water and clean steam systemsPurified water, water for injection (WFI) and clean steam are essential utilities generated on site and distributed for use in DNA vaccines production, clean-in-place (CIP), sterilisation-in-place (SIP), and media preparation (Robbins, 2010). In order to ensure safety, purity and efficacy of the DN A vaccines the water used for its production is sterile water for injection (WFI). The WFI is produced from purified water by distillment/reverse osmosis to meet the required standard of purity specified by the United State Pharmacopeia (USP) (pH 5.0-7.0, nonpyrogenic and antimicrobial agent). The WFI is stored at elevated temperature (80-95oC) to eliminated microbial growth, and the system constructed with stainless steel to eliminate contamination (Robbins, 2010). The WFI system design is shown in FIG.4.7 Waste treatment and managementThe system for treating the waste generate from the DNA vaccines manufacturing facility is an integral part of the design of the facility, layout and good manufacturing practices (GMP). The major(ip) waste generate from the production process are genomic DNA of the legions cells, RNA, proteins, cell debris, salts, endotoxins and plasmid isoforms (Ferreira et al., 2000). The waste is treated to regulatory standards (BS, ISO, etc.) to avoid harm to h ealth and safety of personnel and environment (HSE), pollution and eliminate cross contamination of the product. The system for treating the waste is illustrated in FIG. below WWWWIncinerationAutoclavedWaste carrier bagAutoclave4.7.1 Health, Safety and Environment (HSE)The DNA vaccines production microorganism poses some(a) hazard. The environmental impact assessment (EIA) of the DNA vaccines production system consequently becomes a key part of the design and layout of the manufacturing facility (Prazeres and Ferreira, 2004). However, the environmental impact assessment (EIA) study and the design will require approval from environmental protection agency before the facility is built (Davda, 2004). To ensure that health, safety and environmental regulations are met, the process design and layout is geared towards minimisation of waste generation, safety of product, safety and health of personnel and incorporation of waste treatment process before discharge to the environment. In add ition, the personnel will also be take into accountd with personal protective equipment (PPE) such as hand gloves, gowns, goggles, etc. to work with.4.8 Legislation and regulationThe manufacture of DNA vaccines is highly regulate to ensure that it is safe, efficacious and pure for humans, and also its production carried out in accordance to current GMP (Plumb, 2005). Therefore, before the DNA vaccines can be marketed they must be licence from the relevant regulatory bodies such as the Medicines and healthcare products Regulatory Agency (MHRA) in the United Kingdom, the Food and Drug Administration (FDA) in the United States, the EMEA, WHO and so on (Smith and Dennis, 2001). The manufacturing facility used for the production of the DNA vaccines must be licence too (Plumb, 2005). These licences are obtained if and simply if the manufacturing facility design, layout and premises of its manufacture conform and comply to current good manufacturing practices (cGMP) and with regulatory standards, guidelines and specifications stipulated by MHRA, FDA, WHO, EMEA, ISO, etc. Hitherto, the company must also provide detailed documentary evidence about the safety, purity and efficacy of the DNA vaccines and the consistency of its manufacturing process. Signor and Terry reported that the incorporation of current good manufacturing practices (cGMP) into good design practices (GDP) at the inception of the manufacturing facility will ensure that regulatory conditions are met (Signor and Terry, 2008). The regulatory guidelines specify the requirements for the pharmaceutical manufacturing facility, not the methods to achieving it. The regulatory bodies functions include safeguard public health, licensing, monitoring DNA vaccines post-marketing, regulating clinical trials and publish quality standards.5.1 Project timelineThis project has a definite start, warmness and end, which consist of several activities ranging from the environmental impact assessment and design approval, construction to commissioning executed in a defined order to kick in the project to completion. It is the function of the project manager to plan, schedule and control these tasks/activities in a specified sequence and allocate materials, manpower, machinery and money to ensure that the project is completed on time (Gray and Erik, 2008). There are several project management techniques available in the literature, but to estimate the timeline of this project the Gantt chart technique was employed, which a plot of each task against time. Each bar represents a task/activity, length of the bar corresponds to the duration of the task and the position indicate the start and finish times. The timeline for key activities of the project are shown in FIG below, the Gantt chart was prepared with reference to (Davda, 2004). The entire project is anticipate to take a year and 4.5 months from the Gantt chart.6.1 Recommendations1. Legislations and regulations are subject to changes with emergen t of big-chested technology, therefore the design of the manufacturing facility should be above the current specifications and standards.2. A well defined and detail engineering drawings and specifications that does not require a good deal interpretation.3. A good relationship between project design team with relevant regulatory authorities and encouragement of their input will work up the design of the facility and compliance to cGMP.4. Ensure that all designs, installations and utilities are validated according to validation master plan (VMP) and are working according to design and specification of regulatory bodies.5. Compliance with current good manufacturing practices (cGMP) at the inception of the design phase of the facility.6. The DNA vaccines production facility should be designed and layout to harmonized the various regulations by different bodies such the US FDA, UK MHRA, EU, Japan, ISO, WHO, etc. to boost market for the product.7. The process parameters such as temper ature, pH and pressure must be carefully controlled to assure batch-to-batch identity in final product.7.1 ConclusionIncorporating current good manufacturing practices (cGMP) from the beginning of the design and layout phase of the DNA vaccines facility, the production processes and to the manufacturing premises will ensure that all regulatory specifications are met.