2017 MiniBioman Drug Product Manufacturing Day 2 Aseptic

2017 Mini-Bioman Drug Product Manufacturing Day 2. Aseptic Vial Filling and Sealing Ivy Tech Community College Bloomington Sengyong Lee, Ph. D Professor and Chair of Biotechnology

Drug Filling Operation : https: //www. youtube. com/watch? v=20 Hbxky. W_p. M

1. Sterile Product Filling Operation • Most of the compounded drug products are in the forms of a liquid, suspension, emulsion and as powders. A liquid is more readily subdivided uniformly and introduced into a container having a narrow mouth than is a solid. Mobile liquids are considerably easier to transfer and fill than viscous, sticky liquids, which require heavy-duty machinery for rapid production filling. • Although many devices are available for filling containers with liquids, certain characteristics are fundamental to them all. All provide for repetitively forcing a measured volume of the liquid through the orifice of a delivery tube that is introduced into the container. The size of the delivery tube will vary from that of about a 20 -gauge hypodermic needle to a tube 1/2 inches or more in diameter. The size required is determined by the physical characteristics of the liquid, the desired delivery speed, and the inside diameter of the neck of the container. The tube must enter the neck and deliver the liquid well into the neck to eliminate spillage, allowing sufficient clearance to permit air to leave the container as the liquid enters.

• The delivery tube should be as large in diameter as possible to reduce the resistance and decrease the velocity of flow of the liquid. Product surface tension, viscosity, and temperature dictate the potential of product dripping or the formation of “threads” of product on the sealing surface of the vial or syringe wall. To reduce the possibility of the product splashing out of the container, most automated filling systems fill “bottom up” with the filling tube inserted to its greatest depth at the start of the filling cycle and withdrawing the tube as the product is dosed into the container. • For smaller volumes of mobile liquids, the delivery usually is obtained from the stroke of the plunger of a syringe or rotation of a peristaltic pump, forcing the liquid through a delivery tube/needle combination and into the final container/closure system. • For heavy, viscous liquids, a sliding piston, the turn of an auger in the neck of a funnel, or the oscillation of a rubber diaphragm may be used. For large volumes, the quantity delivered usually is measured in the container by the level of fill in the container, the force required to transfer the liquid being provided by gravity, a pressure pump, or a vacuum pump.

Ampoule filling: https: //www. youtube. com/watch? v=A 5 qoa. RLGrtg

• The narrow neck of an ampoule limits the clearance possible between the delivery tube and the inside of the neck. Since a drop of liquid normally hangs at the tip of the delivery tube after a delivery, the neck of an ampoule will be wet as the delivery tube is withdrawn, unless the drop is retracted. Therefore, filling machines should have a mechanism by which this drop can be drawn back into the lumen of the tube, called a “suck-back feature”. • Since the liquid will be in intimate contact with the parts of the machine through which it flows, these must be constructed of non-reactive materials such as borosilicate glass or stainless steel. Modern coatings such as Advanced Materials Components Express AMCX 2286 are used to coat stainless steel needles for products that are affected by contact with metals, e. g. formulations containing chelating agents or having very acidic or alkaline p. H values. Coated Disposable Filling Needles

2. Filling Mechanisms • Filling machines are classified by the type of driving device or filling mechanism used to deliver the drug-containing formulation into the primary package. There at least four driving devices and four filling mechanisms: Driving Device Filling Mechanism(s) Gravity (solids and liquids) Gravimetric, time-pressure, fill by weight Piston (liquids and gases) Rotary piston, Rolling diaphragm pump Rotary Pump (liquids and gases) Rotary peristaltic Auger screw or vibrator (solids) Vibratory/Mechanical force

A. Gravity/Time Pressure Filling – it is the oldest and the most economical filling machine. The filling principle is simple; the amount of product flowing through the filling nozzle is driven by gravity and will always be the same for a fixed amount of time. The finished bulk solution is pumped into a holding tank above a set of pneumatically operated valves. Each valve is independently timed by a master computer for the filling machine so that precise amounts of liquid will flow by gravity into the container. The amount of product dispensed is controlled by adjusting the time for closing the valve. • These systems can range bulk bag systems working with 1000 of kilograms to more precise systems where weight feedback is used to control the volume of dispensed product. Independent timing of each filling valve/nozzle corrects for minor variations in flow rates so that each container is filled accurately and uniformly.

• Improvements in holding tank headspace pressure control and feedback control have made time pressure filling machines more accurate than pump systems for many applications. • The disadvantage of this type of technology is that the dynamics of the fluid path and nozzle actuation characteristics continuously change over time. This requires the operator to make adjustments to the machine's stored parameters more frequently than other filling mechanisms. Automated check-weighing with a consistent feedback to the filler control system reduce the number of operator interventions.

B. Piston filling - piston filling includes pumps with lapped rotary or check valves and pumps that use a rolling diaphragm. Lapped rotary pumps involve a cylinder that is lapped by both the piston and the rotary valve to produce an exceedingly tight fit. Pumps with check valves are not typically used for injectable filling because the valves are difficult to clean and have a tendency to seize.

• Pumps with the rolling diaphragm use a flexible membrane attached to the pump at its outside diameter and to the piston at its inside diameter. A space between the piston and the body internal cylinder allows the diaphragm to be doubled and to roll as the piston moves up and down. Vacuum may be used to maintain the shape of the diaphragm and to pull the piston downward on the refill part of the filling cycle. Piston pumping machines are very popular for filling liquid but are not the best choice as a filling mechanism for shearsensitive liquids (especially, biologics) and suspensions because of the tight clearances between the piston and the cylinder.

C. Peristaltic Filling: https: //www. youtube. com/watch? v=AN_n. Sz. Bap. Gk Peristalsis describes movement of ingested food in the gastrointestinal tract. The same principle is used for filling machines. Peristaltic filling involves positive displacement where the solution contained within a flexible tube that is fitted inside a circular (rotary) or elongated (linear) pump casing. A rotor with a number of "rollers", "shoes" or "wipers" attached to the external circumference compresses the flexible tube. As the rotor turns or moves, the part of tube under compression closes (or "occludes") thus forcing the fluid to be pumped to move through the tube. Additionally, as the tube opens to its natural state after the passing of the cam ("restitution") fluid flow is induced to the pump. Typical tubing systems used for filling machines, regardless of mechanism, are silicone rubber, polyvinyl chloride (fallen out of favor due to potential leachables), and fluoropolymer.

Since there are no moving parts in contact with the fluid, peristaltic pumps are inexpensive to manufacture. Their lack of valves, seals and glands makes them comparatively inexpensive to maintain, and the use of a hose or tube makes for a relatively low-cost maintenance item compared to other pump types. Peristaltic pumps also minimize shear forces experienced by the product solution, good for shear sensitive protein products. However, they are not as good for high viscosity liquids and cannot match rotary piston machines for small volume filling precision.

Principles Generating High Accuracy in Filling • Volumetric filling machines employing pistons or peristaltic pumps are most commonly used and are best suited for small batch filling of 2 m. L vials (13 mm openings) to 100 m. L vials or bottles with 20 mm openings. Filling speeds for 2 m. L to 20 m. L vials are dependent on size of the vial and the vial handling equipment. When the product is sensitive to metals, a peristaltic-pump filler may be used because the product comes in contact only with silicone rubber tubing. • Time-pressure (or time-gravity) filling machines are also commonly found. A product tank is connected to the filling system with a pressure sensor which continuously measures pressure and transmits values to the PLC system which controls the flow of product from tank to filling manifold. Product flow occurs when tubing is mechanically un-pinched and stops when tubing is mechanically pinched. The main advantage of time/pressure filling operations is that they do not contain mechanical moving parts in the product stream. The product is driven by pressure (usually nitrogen) with no pumping mechanism involved. Thus, especially for proteins that are quite sensitive to shear forces, time/pressure filling is preferable.

3. Filling Materials • Pre-filled Syringe Processing and Filling: https: //www. youtube. com/watch? v=t-5 I 95 RTKKM, Syringes are cleaned, sterilized (by ethylene oxide or radiation) and sealed with a “puncture proof” lid by the syringe manufacturer before delivering to the finished product manufacturer. Syringes are contained in a plastic tub system double wrapped that maintains sterility of the syringes. The transfer of these tubs containing sterile syringes from a receiving area into the aseptic filling area presents a challenge with respect to maintaining sterility. Typically, the outer bag wrap is removed within a Grade C/ISO 8 area and the inner bag wrap is sanitized (alcohol or hydrogen peroxide vapor) before moving into the aseptic area. Low energy e-beam radiation is becoming a new alternative as a surface decontamination process that increases the level of sterility assurance in the transfer of pre-sterilized syringe tubs into the aseptic area. In the Grade A area, an operator removes the lid of the tub and the tub is placed on the filling line. Syringes are filled row by row with precise filling volumes (can be accurate within 0. 01 m. L) and then the rubber plunger is accurately inserted at the predetermined location within the syringe barrel to assure accurate delivery volume.

Syringe fillers are designed to first fill the product into sterile syringes, then the sterile stopper is inserted. If the stopper insertion rods or tubes are not properly aligned then the product could potentially contact the rods and tubes and glass will break. Syringe fillers typically can fill 0. 5 to 20 m. L syringe at rates between 60 to 600 syringes per minute. Inova Syringe Filling Machine Needle and Plunger Assemblies Image Courtesy of OPTIMA Corporation

Cartridge Filling - Bausch + Stroebel cartridge filling machines may fills up to 3 m. L cartridges at rates of 300 per minute. With cartridges, the rubber plunger is first inserted to a predetermined place within the barrel of the siliconized cartridge. The product is filled, typically with a two, even three-shot fill so that there is no significant head space, and then the cartridge is sealed with a sterile, rubber septum within an aluminum cap. Excessive air space in a cartridge will affect dose accuracy when the contents of the cartridges are ejected through a pen delivery system. Cartridges contains a specific amount of medication. Cartridge must be placed in or attached to a special holder which serves as a plunger. Most have additional space to allow compatible drugs to be added.

Suspensions and Other Dispersed System Filling. The main issues or potential problems that may occur in the filling of dispersed systems include: Maintaining dose homogeneity container-to-container Validation of dose homogeneity especially with higher product viscosities Clogging of filling needles/nozzles Batch size Aseptic formulation Particle size reduction under aseptic conditions. Maintaining dose homogeneity during filling operations is a huge challenge. Dose homogeneity is a function of the ability of recirculation system supporting the filling system to prevent suspension particle settling or emulsion globule interaction and growth. The primary way is filling of the recirculated suspension. Some form of in-process check is performed to assure suspension homogeneity during the filling process.

Check Weighing - All filling operations must be checked for accurate dose filling, both prior to the start of the filling operation to make proper initial adjustments and during filling by checking fill volumes periodically to ensure that pre-determined volumes or weights are within specifications. There a number of check weighing methods (focus on vials): Manual check weighing Robotic check weighing of a single container Robotic check weighing of a full container set 100% non-contact check weighing Each filling operation has a target fill volume or weight with upper and lower acceptance limits. Typical fill requirements are 0. 5% of the target fill volume for each and every filling nozzle, however the smaller the fill volume the more difficult it is to maintain those tight tolerances. For example, a target fill weight might be 5. 0 grams with the upper limit being 5. 1 grams and the lower limit being 4. 9 grams. A fill volume of 0. 2 grams would require a fill tolerance of +/- 0. 001 grams.

4. Stoppering The stoppering operations must occur under Grade A/B (ISO 5) clean room conditions. Ampoules do not require rubber closures and are sealed with a flame. Vials are closed with rubber stoppers (or, for vials containing solution to be freeze-dried, the stopper is partially inserted into the vial opening). Syringes and cartridges are closed with rubber plungers at the distal end (with rubber septa sealing the proximal end except for staked-needle syringes). Rubber stoppers and plungers need to be lubricated either with applied silicone oil or emulsion or with special coatings (fluropolymer coating place on the closure by the manufacturer) that permit and facilitate rubber units to move easily from the hopper along stainless steel tracks or rails to the openings of the primary containers. For syringes and cartridges, the placement of the rubber plunger is dictated by the desired position of the plunger within the barrel of the syringe or cartridge to deliver the claimed volume of product.

For vial openings, the closure must fit snugly, not “pop out”. Often, filling efficiencies are dependent more on the stoppering process than on the actual filling process as there are tendencies for rubber closures to slip or pop off the openings of vials. The closing of primary containers will affect the final integrity of the container/closure interface. For syringes and cartridges, no further sealing is done although units are either placed in secondary packaging for unit dosing or part of a tray system. For vials and bottles, aluminum seals are crimped around the rubber closure and top of the container. Seal force integrity is measured by a torque-testing device. Problems encountered during stoppering include: Too little or too much silicone on stoppers Misaligned or bent syringe stopper insertion rods or tubes Stoppers become jammed on the track Improper head space (syringes) Stoppers are not completely seated

5. Sealing Ampoule-filled containers should be sealed as soon as possible to prevent the contents from being contaminated by the environment. Ampoules are sealed by melting a portion of the glass neck. Tip-seals are made by melting enough glass at the tip of the neck of an ampoule to form a bead and close the opening. These can be made rapidly in a high-temperature gas-oxygen flame. To produce a uniform bead, the ampoule neck must be heated evenly on all sides, such as by burners on opposite sides of stationary ampoules or by rotating the ampoule in a single flame. Care must be taken to adjust the flame temperature and the interval of heating properly to completely close the opening with a bead of glass. Excessive heating will result in the expansion of the gases within the ampoule against the soft bead seal and cause a bubble to form. If it bursts, the ampoule is no longer sealed; if it does not, the wall of the bubble will be thin and fragile.

Insufficient heating will leave an open capillary through the center of the bead. An incompletely sealed ampoule is called a leaker. Pull-seals are made by heating the neck of the ampoule below the tip, leaving enough of the tip for grasping with forceps or other mechanical devices. The ampoule is rotated in the flame from a single burner. When the glass has softened, the tip is grasped firmly and pulled quickly away from the body of the ampoule, which continues to rotate. The small capillary tube thus formed is twisted closed. Pull-sealing is slower, but the seals are surer than tip-sealing. Powder ampoules or other types having a wide opening must be sealed by pull-sealing. Fracture of the neck of ampoules during sealing may occur if wetting of the necks occurred at the time of filling. Also, wet necks increase the frequency of bubble formation and unsightly carbon deposits if the product is organic. To prevent decomposition of a product, it is sometimes necessary to displace the air in the space above the product in the ampoule with an inert gas. This is done by introducing a stream of the gas, such as nitrogen or carbon dioxide, during or after filling with the product. Immediately thereafter the ampoule is sealed before the gas can diffuse to the outside. This process should be validated to ensure adequate displacement of air by the inert gas in each container.

Vials and bottles are sealed by a rubber closure (stopper). This must be done as rapidly as possible after filling to prevent contamination of the contents. The large opening makes the introduction of contamination much easier than with ampoules. Therefore, during the critical exposure time the open containers should be protected with a blanket of HEPAfiltered laminar airflow. The closure must fit the mouth of the container snugly enough so that its elasticity will seal rigid to slight irregularities in the lip and neck of the container. Closures preferably are inserted mechanically using an automated process, especially with high-speed processing. To reduce friction so that the closure may slide more easily through a chute and into the container opening, the closure surfaces are halogenated or treated with silicone. When the closure is positioned at the insertion site, it is pushed mechanically into the container opening. For lyophilized drugs, the filled and partially stoppered vials would be transferred to a sterile lyophilizer for the completion of the lyophilization cycle. It is normal for the stoppers to be seated (pressed in) in the vials inside the sterile drier at the end of the lyophilization cycle prior to opening the door. The stoppered vials are then removed from the sterile drier and immediately capped. The delay in sealing the container, immediately after the filling process, allows the drug to be exposed to the environment is an additional risk that occurs with sterile lyophilization.

When small lots are encountered, manual stoppering with forceps may be used, but such a process poses greater risk of introducing contamination than automated processes. This is a good test for evaluation aseptic operator aseptic techniques, but not recommended for any product filling and stoppering. Container-closure integrity testing has become a major focus for the industry because of emphasis by regulatory agencies. Container-closure integrity measures the ability of the seal between the glass or plastic container opening and the rubber closure to remain tight and fit and to resist any ingress of microbial contamination during product shelf life. A relatively recent trend, although now standard practice, is the requirement that sealing of vials and other containers be accomplished under Class 100/Grade A/ISO 5 clean room air (the air meets Grade A requirements only under static conditions). Formerly such sealing occurred in unclassified environments.

Rubber closures are held in place by means of aluminum caps. The caps cover the closure and are crimped under the lip of the vial or bottle to hold them in place. The closure cannot be removed without destroying the aluminum cap; it is tamperproof. Therefore, an intact aluminum cap is proof that the closure has not been removed intentionally or unintentionally. Such confirmation is necessary to ensure the integrity of the contents as to sterility and other aspects of quality. The aluminum caps are so designed that the outer layer of double-layered caps, or the center of single-layered caps, can be removed to expose the center of the rubber closure without disturbing the band that holds the closure in the container. Rubber closures for use with intravenous administration sets often have a permanent hole through the closure. In such cases, a thin rubber disk overlaid with a solid aluminum disk is placed between an inner and outer aluminum cap, thereby providing a seal of the hole through the closure.

Manual Aseptic Drug Filling and Sealing Training • Flexicon PF 6 Filler - The PF 6 is a tabletop peristaltic filling machine. It is able to fill volumes from 0. 2 ml to more than 250 ml, with a filling accuracy of +/- 0. 5%. 27

Group Operation on Aseptic Vial Filling & Sealing • https: //www. youtube. com/watch? v=Zj. WSvg-p. RZI • Each group will have at least three operators responsible for filling, stoppering, and crimping of vials. • Each group will aseptically fill and seal 30 glass vials with each filled with 10 ml of LB solution. • The entire operation should be streamlined and aseptically executed in a biosafety hood.

1. Filling tube set up • The filling tubing lines (3. 2 mm diameter silicone tubing), the vials, the stoppers, the WFI and the LB solution were pre-sterilized for your aseptic operation and each group will have to use new sets for their operation. • http: //www. watson-marlow. com/us-en/range/flexicon/tabletop-filling/pf 6/ • The split intake tube ends should be aseptically placed into the WFI water bottle for washing tubes. The WFI and LB solution bottles should be placed on a test tube lack (the position higher than the pump head level) inside of the biosafety hood to help efficient feeding of the solution into the tubing. • The dispensing tube end should be securely held by a clamp holder attached to a stand inside of the biosafety hood. • The sterile tube should be mounted into the dispenser head roller part of the Flexicon filler. First, the two locking pins should be unlocked (by lifting them up) and the tube bridge should be lifted up. Second, the tubes should be situated in the notches in the tube lock sitting on a dowel pin and the dispenser head. The Y-connector in the tubing should be located to the right of the dispenser head. Third, the tube bridge is placed back and the locking pins should be reengaged.

2. WFI wash and priming of the tube • Once installed, the entire tubing should be washed with WFI water. Place a beaker under the dispensing tube end. Press the “Pump” and the “Go” buttons to start washing the tubes with WFI. Press “Stop” when about 50 ml of WFI was dispensed into the beaker. • Once WFI wash is done, aseptically transfer the feeding tube ends from the WFI bottle to the LB solution bottle. • Before filling vials, the tube should be primed with the LB solution. Press the “Prime” button until the LB solution replaces WFI in the tube. The entire tubing should be primed with the LB solution and no air bubble should be found from the tube. Once the tube is primed with the LB solution, the beaker under the dispensing end of the tube should be replaced with the first vial to be filled.

3. Vial filling operation • The Flexicon filler should be preprogramed to fill 30 vials with 10 ml fill in each. • The filling operator is responsible for switching vials in and out of the filling position. • There is a 3 second delay between fills and a vial switch should be made by the filling operator during the delay. • Once the first vial is placed into the filling position under the dispensing tube end on the stand, press the “Disp” button on the Flexicon filler to have it entered into the filling mode. • Press the “Go” button to initiate the operation for 30 vial filling. • Once the entire operation is completed, the filler will stop automatically. • If the filling should be stopped before the entire program is done, press “Stop”. • The programed filling can be resumed by pressing “Go”. • The filled vial should be passed to the stoppering operator for immediate stoppering. A forceps should be used for placing and pushing a sterile stopper into a vial. • Once closed with a stopper, the vial is passed to the crimping operator who will be placing and crimping aluminum crimp on the vial with a crimper. • This operation should continue until the entire batch of 30 vials are filled and sealed.

4. WFI wash, drain and removal of the tube • Once the filling operation is done, the WFI washing step should be done again to rinse the tubes. The intake ends of the tube should be placed back to the WFI water bottle and a beaker should be placed under the dispensing end of the tube. • Wash the tube with WFI by pressing “Pump” and “Go”. After collecting about 100 ml of WFI in the beaker, stop the pump by pressing “Stop” and remove the intake ends of the tube from the WFI water bottle. • Press “Prime” until all WFI in the tube is drained into the beaker. • Once the tube is empty, remove the tube from the dispenser head after unlocking the pins. • Never leave a filling tube in the dispenser head after a filling operation done. It causes a damage to the tube.

5. Inspection. • The entire batch of filled vials will be inspected for any spillage, splash, over- or underfilling, misplaced or jammed stoppers, under- or over-crimping. Vials with any of such conditions will be considered as “defects”. • The filled vial batches will be incubated at 37 o. C overnight for the aseptic validation. Since the LB solution is a microbial growth medium, any contamination during your operation will result a significant microbial growth in the vials. • Cross inspection between the two operation groups can identify operation errors and defects from the entire filled batch. • The group with less number of operation errors and defects will win the competition.

Inspection Report Form Vials filling and sealing operation performed by: Inspection performed by: Number of Defects Identified The nature of the defects Microbial Contamination Under or over filled vials Spillage during filling Stopper defects Crimp defects for under- or over crimping Other operation errors Total number of defects