Advanced therapies are rapidly becoming a reality, targeting unmet patient needs and treatments for diseases with limited therapeutic options. Some advanced therapies, such as gene therapy and mRNA, require frozen storage due to their fragile nature at ambient temperature, putting unprecedented expectations on the vial-stopper-seal combination to maintain container closure integrity (CCI) during storage at low temperature conditions. The quality, patient safety, and high value of these therapeutics can be jeopardized if the chosen vial-stopper-seal combination doesn’t maintain a sterile barrier from when the drug is packaged through to administration to the patient. During cooling from ambient temperature to temperatures below -80°C, packaging components such as glass vials, elastomeric closures, and aluminum seals exhibit contraction and changes in physico-mechanical properties. These changes in material properties and component dimensions may affect their ability to maintain CCI, resulting in the formation of transient leaks.

The objective of this joint study conducted by West and Corning Incorporated was to determine if the chosen vial-stopper-seal combination will remain integral while frozen at -80°C, under studied conditions, when the appropriate materials, processing, and sealing conditions are applied. The study was conducted in four separate experiments to explore the influences of capping force, cooling rate, dimensional changes, and long-term storage at -80°C on the vial-stopper-seal combination’s ability to maintain closure integrity. Study 1 assessed component compatibility using existing stack-up tolerance analysis and determined the processing and sealing conditions (capping force) required for CCI at -80°C (tested by helium leak rate). Study 2 characterized the impact of cooling rate on ability to maintain CCI after conditioning. Study 3 examined vial-stopper combinations with varying critical dimensions and their effect on CCI. Study 4 assessed the CCI of the vial-stopper combination after three months of extended storage at -80°C.

The vial-stopper-seal combination was comprised of the following components:

• West NovaPure® 4432/50G 13mm and 20mm stoppers
• West Flip-Off® 13mm and 20mm seals with TruEdge® technology
• Corning® Valor® Glass vials 2R and 10R vials (EU blowback)

The following equipment was used during the study:

• Genesis Model RW Rail Capper
• Genesis Model AWG RSF Tester
• Leak Detection Associates/PTI SIMS 1284+ Helium Leak Detector
• Lighthouse Carbon Dioxide Headspace Analyzer

Study 1 – Effect of the Capping Force on CCI

In this first study, component dimensional compatibility was assessed through a stack-up tolerance analysis. Then the chosen vial-stopper-seal combination was capped within a range of forces (13 N – 111 N; 3 lb – 25 lb) to determine the threshold residual seal force (RSF) needed to achieve CCI at -80°C. A total of (n=360) 10R vials was produced using capping forces that represent six (6) different RSF values (n=60 vials per each RSF value) and held at ambient temperature (23°C ± 3°C). At T=4 Hours, T=7 Days, and T=21 Days, the RSF values of capped vials were measured prior to performing the helium leak test (n=20 per RSF per time point). Samples held at ambient temperature were allowed to equilibrate to -80°C for at least 3 hours and then all vials were tested at -80°C by helium leak detection. Leaks were both expected and observed in vial populations with a capping force < 27 N (6 lb). The helium leak rates for all other vials were below the Kirsch limit of 6.0 x 10-6 mbar*L/sec (correlating with a low probability of microbial ingress), which was adjusted to 3.9 x 10-6 mbar*L/sec, the equivalent leak rate at -80°C. CCI was consistently demonstrated at -80°C with the chosen combination of glass vial, rubber stopper, aluminum seal and a capping force ≥ 27 N (6 lb).

Study 2 – Effects of Cooling Process on CCI

In the second study, the goal was to investigate the impact of cooling rate on the ability of the sealing materials to maintain CCI at -80°C (e.g., as seen during shipping on dry ice). Negative controls, positive controls, and 10R vial systems were capped to produce an RSF value of approximately 67 N (15 lbf). Positive controls were created with nominal 2 µm, 5 µm, and 15 µm cold ablation laser-drilled defects on the sidewall of the vials, and gross defects were created by puncturing the stopper of assembled vials with a 23G needle. A Corning CoolCell® container was used for controlled freezing at -1°C/sec to -80°C using dry ice (n=30 plus n=9 positive controls). A cardboard box was used for uncontrolled freezing to -80°C using dry ice (n=30 plus n=9 positive controls). Ten negative controls were stored at ambient temperature and were not exposed to dry ice.

A CO2 headspace analyzer was used to measure the CO2 partial pressure of samples at T=0 Days (prior to freezing), T=7 Days (stored at -80°C and thawed to ambient temperature), and T=10 Days (3 days post-thaw to ambient temperature). Any vial with a CO2 partial pressure > 0.13 kPa (1 Torr) was considered a leaking vial. All test samples and negative controls had CO2 partial pressures < 0.13 kPa and all positive controls had CO2 partial pressures > 0.13 kPa. Regardless of cooling rate, the chosen container closure combinations remained effective at maintaining CCI after 7 days of storage in dry ice at -80°C.

Study 3 – Effects of Stopper Flange Height and Vial Crown Dimensional Variability on CCI

In this study, the vials and stoppers were intentionally selected or produced to represent a range of dimensional variability. The 10R vial crown and the 20mm stopper flange height were chosen as dimensional variables for the study. The large vial crown samples were selected from Corning Valor® Glass Vials that were within tolerance of standard ISO specification (ISO 8362-1) but measured on the high side of the tolerance. The large stopper flange height samples were custom made by West. Four vial-stopper configurations were capped using the 20mm Flip-Off® seal with TruEdge® technology with a force to produce an RSF value of approximately 67 N (15 lbf):

• Configuration 1:  Nominal vial crown – Nominal stopper flange height
• Configuration 2:  Nominal vial crown – Large stopper flange height
• Configuration 3:  Large vial crown – Nominal stopper flange height
• Configuration 4:  Large vial crown – Large stopper flange height
  
   

Helium leak testing was conducted at -80°C on T=0 Days and T=7 Days. CO2 headspace analysis was conducted on T=0 Days prior to freezing and T=7 Days stored at -80°C in dry ice. Test samples were equilibrated to ambient temperature prior to CO2 headspace analysis. The testing was performed on n=30 vials per configuration per CCI technology.

All samples exhibited helium leak rates below the Kirsch limit, correlating with a low probability of microbial ingress, and no significant CO2 ingress. CCI was maintained with all large vial crown and stopper flange height configurations for 7 days of storage at -80°C for the vial-stopper-seal combination evaluated in this study.

Study 4 – Effects of 3 months storage time under CO2 atmosphere at -80°C on CCI

The final experiment of the study assessed the ability of the sealing materials to maintain CCI after an extended storage of three months at -80°C under a CO2 atmosphere. All 2R vial combinations were capped to produce an RSF value of approximately 44 N (10 lbf) and all 10R vial combinations were capped to produce an RSF value of approximately 67 N (15 lbf). Different capping forces were used to seal the 2R and 10R vials because the seal diameters between the two vial sizes are different. The actual RSF values were measured before the experiment was started. Vial-stopper-seal combinations were stored for 90 days at -80°C under a CO2 atmosphere. CO2 headspace analysis was performed for vial-stopper-seal combinations that were stored for 90 days at -80°C under a CO2 atmosphere as per the following two scenarios:

• Cycled samples (n=30): T=0, 30, 60, and 90 Days: samples were returned to cold storage between time points to simulate freeze-thaw cycles
• Non-cycled samples (n=100): samples were tested at T=0 and 90 Days 

CCI was maintained after three months storage at -80°C under a CO2 atmosphere for both the thermally cycled and non-cycled 2R and 10 R vials.

Conclusions

The Corning® Valor® Vial and NovaPure® 4432/50G stopper with Flip-Off® seal with TruEdge® technology combination is a suitable and reliable container closure combination for frozen drug storage at temperatures as low as -80°C. Reliable maintenance of CCI was achieved with the appropriate capping process, RSF, and good seal quality. The freezing process did not influence the integrity of the container, which maintained CCI after 7 days of storage at -80°C in dry ice conditions. CCI was demonstrated for a range of vial and stopper dimensions and was maintained when the combinations were stored at -80°C for three months under a CO2 atmosphere. The container closure combination used in these studies is successfully being used in the packaging and distribution of marketed frozen drug products.

More information specific to the Valor® Glass vial and NovaPure® 4432/50G stopper with Flip-Off® seal with TruEdge® technology study is detailed in the presentation titled Selection of a Primary Container Closure System Capable of Maintaining Container Closure Integrity at -80°C and in TR 2023/259 titled Container Closure Integrity of Corning® Valor® Glass Vials with NovaPure® Closures and Flip-Off® Seals at -80°C. Learn more on container closure integrity testing or refer to West’s Knowledge Center for a comprehensive library of supporting information regarding West’s products and services. Contact us for more information on partnering with West for your CCI testing needs or if you have questions about our Corning glass offering: West.GlassSystems@westpharma.com

 

NovaPure and TruEdge are registered trademarks of West Pharmaceutical Services, Inc. in the United States and other jurisdictions. Corning® and Valor® Glass are registered trademarks of Corning Incorporated. West Pharmaceutical Services Inc. is the exclusive distributor of Corning® Valor® Glass. All other trademarks are the properties of their respective owners.