Strategies for Induced Nucleation

A Review and Scale-up Case-Study for a 56 m2

Freeze-Dryer with 195,960 Vials using VERISEQ® Nucleation

April 22nd, 2016

VERISEQ® Nucleation is a registered trade mark of The Linde Group

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Motivation for Controlled Nucleation

Introduction to VERISEQ® Nucleation

VERISEQ® Nucleation on freeze dryers of varying sizes:

demonstrated scalability

15m2 shelf area – Test at IMA Life Tonawanda

39m2 shelf area – 45,540 vials at customer site

56m2 shelf area – 195,960 vials at customer site

Key findings

IMA Life and THE LINDE GROUP – a strong partnership

OUTLINE

NUCLEATION BASICS

Process by which ice crystals (seed crystals) are formed from originating sites (nuclei) under proper temperature conditions (super-cooling)

Naturally, suspended particles or container defects provide nucleation sites

In production settings, nuclei form from water, but only after significant super-cooling

Because nucleation is an exothermic event, it can be detected with temperature measuring devices

Natural (uncontrolled) Nucleation:

Occurs randomly, once nuclei

are formed

Over extended period of time

Within extended temperature range

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Without Nucleation Control:

Larger degree of super-cooling = smaller ice crystals

Smaller ice crystals = higher resistance to vapor flow

High resistance to vapor flow = longer cycles, possible vial breakage

Larger vial to vial variability

With Nucleation Control:

Reduced vial to vial variance – scale-up prediction

Better prediction of product stability

Quite often leads to reduced reconstitution time

Regulatory bodies are putting pressure on manufacturers to better control and understand their processes. Until now, control of the nucleation step has not been

possible.

WHY CONTROLLED NUCLEATION?

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REVIEW

“…determine the influence of ice nucleation temperature on the primary drying rate during lyophilization for samples in vials that were frozen on a lyophilizer…”

James A. Searles, John F. Carpenter, Theodore W. Randolph

Higher nucleation temperature faster drying

Nucleation temperature affects drying rate:

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“[…] effect of the ice nucleation temperature on the primary drying process using an ice fog technique for temperature-

controlled nucleation. In order to facilitate scale up of the freeze-drying process, this research seeks to find a correlation of the

product resistance and the degree of supercooling with the specific surface area

of the product.”

Abstract from “Heat and Mass Transfer Scale-Up Issues during Freeze Drying: II. Control and

Characterization of the degree of Supercooling” AAPS PharmSciTech 2004 (S. Rambhatla, R.

Ramot, C. Bhugra, M. J. Pikal)

Nucleation temperature affects product mass transfer resistance:

REVIEW

Higher nucleation temperature lower product resistance to water vapor

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“…fundamentals of ice nucleation in the field of freeze-drying are presented. Furthermore, the impact of

controlled ice nucleation on product qualities is discussed, and methods to achieve controlled ice

nucleation are presented…”

R. Geidobler, G. Winter (European Journal of Pharmaceutics

and Biopharmaceutics, May 2013)

REVIEW

Importance of controlled nucleation is emphasized by various researchers

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VIAL

FORMULATION

Partially seated stopper

VERISEQ® NUCLEATION REVIEW

1. VERISEQ® Nucleation uses liquid nitrogen and water vapor (steam) to create a cryogenic fog or “ice fog”.

2. This fog permeates the product chamber and enters the vials.

3. Ice crystals contact (seed) the surface of the product, nucleation proceeds rapidly from the top downwards in the vial.

VERISEQ® NUCLEATION REVIEW

*Note: same product, same magnification

Advantages of VERISEQ® nucleation:

Ability to induce nucleation at a fixed (reduced) degree of super-cooling (higher product temperature)

Increased robustness Laboratory (non-aseptic, stand-alone) and production scale (aseptic, fully

integrated) versions No pressure rating requirement Scalability Ability to retrofit practically any freeze dryer

Natural nucleation* VERISEQ® Nucleation*

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SYSTEM TYPES

Lab-scale (non-aseptic) units Designed for use with laboratory

freeze dryers

Stand alone device that operates at or below atmospheric pressure using its own PLC for system control.

The system is designed to work on pressure rated as well as non-pressure rated freeze dryers.

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SYSTEM TYPES

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Production scale (aseptic) units Sterile liquid nitrogen is obtained

from condensing filtered gaseous nitrogen.

The aseptic system uses clean steam normally used to sterilize the freeze dryer.

The VERISEQ® Nucleation system is sterilized prior to use.

This system is fully integrated into the freeze dryer control system.

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SUMMARY OF LAB-SCALE TESTING

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SCALE-UP CHALLENGES

Key challenges

Uniformity of ice-fog distribution.

Ensuring that every vial is seeded.

Duration of ice-fog injection.

Product temperature control. Nucleation is based on two factors: presence of nucleation sites (nuclei), and proper degree of super-cooling. If the product is too warm, nucleation will not occur even if nuclei are present.

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SUBLIMATION

TRANSPORT

Larger heat transfer on top shelves (vials run warmer)

THERMAL GRADIENT

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Optimization Study on 15 m2 Freeze Dryer

IMA Life, Tonawanda

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SETUP: LAYOUT

OUTLET: 2” MAIN DOOR

INLET: 2”

SINGLE INLET – SINGLE OUTLET

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SHELF 4 (S4)

SHELF 8 (S8)

SHELF 1 (S1)

S4, Bl

S4, FL

S4, CENTER

S4, BR S4, FR

S8, FL S8, FR

15 m2 shelf area Cluster of 8-10 vials in each location Each pack mounted with 1 internal thermocouple and 1 external thermocouple

LAYOUT: VIAL FROM MAIN DOOR

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1 vial nucleates during ramp down on shelf 1 (directly facing chamber wall)

VIAL ON SHELF 1 DOES NOT NUCLEATE

WALL TEMPERATURE: ~16 ˚C

POSITION DEPENDENCE: 2˚C SPREAD IN PRODUCT

TEMPERATURE

STANDARD CONFIGURATION

INTERNAL THERMOCOUPLES

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100% nucleation on all shelves Wall temperature can be a crucial factor in success of inducing nucleation

INTERNAL THERMOCOUPLES

WALL TEMPERATURE: ~10 ˚C

WITH CHAMBER WALL COOLING

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~150 vials with 1 ml water fill in 3 ml vials, used in cluster of ~10 vials each,

1 external and 1 internal thermocouple in each cluster

Shelf 1,4,8 used with clusters located in 4 corners and center, Tsh hold time

~90 minutes

Successful run (no nucleation during ramp down) obtained with chamber wall

cooling for reduced radiation (65%)

Optimized process and injection configuration successfully tested with

placebo in dryer

SUMMARY AND LEARNINGS

Thermal gradients in large production freeze dryers should be considered when designing the freezing protocol

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Full Scale Test Without Jacketed Cooling 45,540 vials in a 39m2 machine at

Client Site

Shelf area=39 m2,

Chamber volume=9 m3

14+1 shelves

Machine has no jacket cooling

Number of vials: 45,540 including:

15,840 vials of 20x dose

29,700 vials of 1x dose

SUMMARY OF SETUP

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ICE-FOG INLET (2 IN)

OUTLET (3 IN)

Nucleation time in probed vials reduced from 118 min to < 2min (see plot)

Nucleation time in wireless probed vials reduced from 152 min to <2 min (not included in plot)

Nucleation temperature range reduced from 9°C to <0.8°C

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NUCLEATION-THERMOCOUPLE

DATA

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CYCLE DURATION COMPARISON

Point of reduced sublimation rate reduced by 12.4 hr. Primary Drying time reduced by 5.4 hr. at same process conditions

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VERISEQ® Ice Fog demonstrated on a 39m2 shelf area freeze dryer

(9 m3) with no silicone jacket cooling with 45,540 vials (35% of 20x

dose)

Nucleation time in probed vials reduced from 152 min to < 2min

(Wireless probes not included in nucleation plot)

Nucleation temperature range in probed vials reduced from 9 °C to

< 0.8 °C

Point of reduced sublimation rate reduced by 10% (12.4 hr.)

Primary Drying time reduced by 5% (5.4 hr.) at same process

conditions

SUMMARY

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Full Scale Test: 195,960 vials in a 56m2 Freeze Dryer at Client Site

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ICE-FOG INLET (3 IN)

OUTLET (3 IN)

Shelf area=56 m2,

Chamber volume=17.5 m3

20+1 shelves Walls cooled with cooling

jacket during ramp 195,960 vials:

1.2 ml fill in 3 ml vials

SUMMARY OF SETUP

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• Nucleation time in probed vials reduced to < 2.5 min • Nucleation temperature range in probed vials reduced to 0.6 °C

Summary of setup

195,960 vials: 1.2 ml fill in 3 ml vials

Shelf load: 5 ˚C Ramp to -7 ˚ C Walls cooled with cooling

jacket during ramp Hold: 90 min

NUCLEATION-THERMOCOUPLE

DATA

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PROCESS HISTORY

Primary Drying time reduced by 812 (19%) minutes, can be further reduced by increasing primary drying temperature

Dotted line indicates original primary drying duration

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Parameter Without Ice Fog With Ice Fog

Loading temperature, ˚C -45 ˚C

5 ˚C

Nucleation temperature, ˚C 0 to -45 ˚C

-6 to -6.6 ˚C

Total variation in nucleation

time between vials

~10 hours (theoretical) 2.5 min

Nucleation completed in < 80 min (variable for vials) 140 min (all during ice fog injection ~ 2.5

min), some with probes inside nucleate early

Primary drying time (time

from Tsh=-9 ˚C to time for

TC to reach shelf temp.)

1001 min 812 min (*not optimized) -19% reduction in

primary drying time, can be reduced further

by increasing shelf temp.

Total time to end

secondary dry

1514 min 1854 min (*not optimized)

Product uniformity 0% (Control) 40% reduction in range of activity values

throughout batch

By using VERISEQ® Nucleation technology, a 40% improvement has been achieved

CYCLE COMPARISON

All times are from time of loading complete (t=0) CYCLE IS NOT OPTIMIZED WITH ICE FOG

Comparison of product uniformity with and without controlled nucleation

Scalability of VERISEQ® Nucleation technology was demonstrated and the following observations were made Overall benefits of VERISEQ® nucleation:

Nucleation time in probed vials reduced from 152 min to < 2min (from 39m2 test) Nucleation temperature range in probed vials reduced from 9 °C to < 0.8 °C (from 39m2

test) Improvement in product uniformity: 40% (from 56m2 test) ~19% reduction in primary drying time even at the same shelf temperature (from 56m2

test) Learning from the scale-up tests:

Ice-fog distribution: robust, not affected by scale of freeze dryer (batch size) The technology is capable of ensuring effective ice fog circulation throughout the

entire chamber volume

Temperature conditions – Critical requirement! The product must be at the right temperature across the entire batch in order to

nucleate!

Duration of ice-fog injection: does not noticeably affect the nucleation temperature and, therefore, uniformity of product attributes Ice fog injection was completed in < 2.5 min (can be less if optimized)

SUMMARY

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Wexler, E., Brower, J., Controlled Nucleation for a Monoclonal Antibody

Formulation – A Case Study, Pharm. Tech. Bioprocessing and Sterile

Manufacturing, pp. 31-34, May 2015.

Wexler, E., Brower, J., New Developments in Controlled Nucleation, Pharm.

Manufacturing, pp. 26-29, October 2015.

Brower, J., Lee, R., Wexler, E., Finley, S., Caldwell, M., & Studer, P. (2015).

New Developments in Controlled Nucleation: Commercializing VERISEQ®

Nucleation Technology. In Lyophilized Biologics and Vaccines (pp. 73-90).

Springer New York.

DeMarco, F., Renzi, E., Lee, R., Chakravarty, P, Ice Fog as a Means to Induce

Uniform Ice Nucleation During Lyophilization, BioPharm International, vol, 25

(1), January 1, 2012

REFERENCES

A STRONG PARTNERSHIP

THE LINDE GROUP

Linde has developed the VERISEQ® Nucleation Technology and the CUMULUS LF Technology (cryogenic cooling of freeze dryer shelves and condenser) in cooperation with IMA Life.

Linde also provides for the pharma industry:

VERISEQ process-grade gases featuring high-purity, traceable N2, O2, CO2, Ar, He & synthetic air

HiQ - specialty gases for calibrating gas monitoring equipment

Process chemicals for applications like for example amination, catalysis or fluorination

Gas applications for ozonolysis, inerting, purging, charging, mixing

Gas applications for Reactor Cooling, cryogenic storage and transport, freezing.

IMA Life

Exclusive rights to market VERISEQ® Nucleation technology.

World leader in the design and manufacture of aseptic processing and freeze-drying equipment.

5 manufacturing sites in Italy, US and China, and 3 service centers in Italy, Netherlands and Japan.

Worldwide networking covering more than 80 countries.

Access to freeze-drying laboratory for development work with 1 laboratory scale, 1 pilot scale freeze-dryer and other analytical tools including tunable diode laser absorption spectroscopy (TDLAS) instrumentation, residual gas analyzer and other re-configurable instrumentation ports.

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CONTACTS

www.ima-pharma.com

Joe Azzarella (Technical Product Manager, Ice Fog) - joseph.azzarella@imalife.com

Arnab Ganguly (Technology Manager) - arnab.ganguly@imalife.com

Frank DeMarco (US Sales, Technical Product Manager, Freeze Drying) - francis.demarco@imalife.com

Geoffrey Sortais (Asia Sales) - Geoffrey.Sortais@imalife.com

Christophe DeStLeger (Europe Sales) - Christophe.DeStLeger@imalife.com

THANK YOU!

www.linde-gas.com/pharma

Eugene Wexler (Product Manager and Global Technical Support) - eugene.wexler@linde.com

Peter Studer (Americas) – peter.studer@linde.com

Rolf Heninger (Europe) – rolf.heninger@linde-gas.com

Arthur Zhang (Asia Pacific) – arthur.zhang@linde.com

Ulrich Hanke (Marketing) – ulrich.hanke@linde.com