BIOPROCESS EQUIPMENT X -RAY STERILIZATION OF SINGLE -USE

2021

X-RAY STERILIZATION OF SINGLE-USE

BIOPROCESS EQUIPMENT PART I – INDUSTRY NEED, REQUIREMENTS AND RISK EVALUATION

© 2021 Bio-Process Systems Alliance. Copying and Distribution Prohibited.

X-RAY STERILIZATION OF SINGLE-USE BIOPROCESS EQUIPMENT: PART I – INDUSTRY NEED, REQUIREMENTS AND RISK EVALUATION

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X-RAY STERILIZATION OF SINGLE-USE BIOPROCESS EQUIPMENT:

PART I – INDUSTRY NEED, REQUIREMENTS AND RISK EVALUATION


AUTHORS and CONTRIBUTORS

James Hathcock, Pall Biotech (subcommittee lead)

Samuel Dorey, Sartorius (subcommittee lead)

Monica Cardona, MilliporeSigma

Sean Lynch, AdvantaPure/NewAge Industries

Nick Troise, PendoTECH

CD Feng, Broadley James

Kirsten Strahlendorf, Sanofi Pasteur (Scientific Advisory Board)

Amit Bhatt, Merck & Co.

Timo Neumann, MilliporeSigma

John Murphy, Merck & Co.

Michael Allard, Venair

Bhuvnesh Sharma, Pall Biotech

Noel Long, Cytiva

Etienne Durant, GSK

Jeffrey Noyes, Steris

Marisa Caliri, Cytiva

Dennis Annarelli, PendoTECH

Rafael Rodriguez, Cytiva

Stephen Hodder, Pall Biotech

Larry Nichols, Steri-tek

Helene Pora, Pall Biotech (sponsor)

CONTRIBUTORS

Jeff Carter, Cytiva

Sade Mokuolu, WMFTG Biopure

John Benson, PendoTECH

Mary Marcus, AdvantaPure/NewAge Industries

Emily S. Alkandry, Saint-Gobain

Max Blomberg, Meissner

Olivia Butterfield, Meissner

Ken Baker, AdvantaPure/NewAge Industries

Mark Petrich, Merck & Co.

Mike Smet, Cytiva

Clive Wingar, Thermo Fisher Scientific

Paul Calverley, Sterigenics

Gabrielle McIninch, Saint-Gobain

Janmeet Anant, MilliporeSigma

Dominic Moore, Sanofi Pasteur

Brian McEvoy, Steris

Acknowledgements

We kindly thank John Logar, Thomas Kroc, and Mark Murphy for their incredible expertise and guidance.

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Table of Contents

1 EXECUTIVE SUMMARY .......................................................................................................................... 4

2 OVERVIEW............................................................................................................................................. 4

3 SUPPLY-CHAIN RISKS ASSOCIATED WITH GAMMA IRRADIATION ........................................................ 5

3.1 Growing Demand for Gamma-Irradiation in a Highly Consolidated Market ................................ 5

3.2 Production and Resupply Challenges with 60Co ............................................................................ 5

3.3 Off-Switch Not Included (Always On, Always Irradiating) ............................................................ 6

3.4 Regulatory, Government, and Security Pressures ........................................................................ 6

3.5 Market Plans to Support Future Irradiation Capacity ................................................................... 6

3.6 Urgency, Timelines, and Need for Action ..................................................................................... 6

4 ALTERNATIVES TO GAMMA IRRADIATION ............................................................................................ 7

4.1 TECHNICAL EVALUATION OF X-RAY AS COMPARED TO GAMMA ................................................. 8

4.1.1 Irradiation Beam Characteristics .................................................................................... 8

4.1.2 Product Impact Characteristics ...................................................................................... 9

4.2 ISO11137 REQUIREMENTS TO TRANSITION FROM GAMMA TO X-RAY ...................................... 10

4.2.1 Requirements on the Irradiation Source. .................................................................... 10

4.2.2 Transferring the sterilization dose. .............................................................................. 10

4.2.3 Transferring the maximum acceptable dose. .............................................................. 11

5 IMPACT OF IRRADIATION TO POLYMERS ............................................................................................ 12

6 RISK-BASED TESTING APPROACH TO QUALIFICATION OF X-RAY ........................................................ 13

6.1 Identifying Materials & Component Tests that Best Assess the Risk ......................................... 14

6.1.1 Connector-Specific Testing Rationale .......................................................................... 20

6.1.2 Container and Film-Specific Testing Rationale ............................................................ 20

6.1.3 Sensor-Specific Testing Rationale ................................................................................ 21

6.1.4 Tubing-Specific Testing Rationale ................................................................................ 21

6.1.5 Filter-Specific Testing Rationale ................................................................................... 22

6.2 Single-use assemblies ................................................................................................................. 23

7 THE PATH FORWARD .......................................................................................................................... 24

7.1 Key Technical Steps for Collective Industry Approach ................................................................ 24

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7.2 Implementation of X-ray as Alternative to Gamma-Irradiation ................................................. 25

Disclaimer .................................................................................................................................................... 25

About BPSA ................................................................................................................................................. 25

8 REFERENCES ........................................................................................................................................ 26

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1 EXECUTIVE SUMMARY A prospective assessment of the contract gamma-irradiation used for sterilization of bioprocess single-use systems,

highlights increasing capacity constraints that may impact delivery and business continuity for a rapidly growing single

use market by 2022. X-ray sterilization, now considered a mature technology, is positioned as a highly similar alternative

to gamma, and contract irradiators are planning much of the future irradiation sterilization capacity in the form of X-ray.

Qualification of alternate sterilization modalities such as X-ray, in addition to addressing industry capacity constraints,

offers increased flexibility to accommodate disruptions or demand spikes and may result in less unwanted effects on

plastics. Technical similarities and differences between X-ray and gamma are reviewed herein, as well as a risk-based

testing strategy for evaluation of x-ray sterilization of single-use systems. The consensus testing strategy indicates the

types of data that may be generated by single-use suppliers on representative single-use materials and is expected to

confirm that existing gamma-irradiation validation packages can be considered applicable to X-ray.

2 OVERVIEW Continued success and rapid growth of single-use technologies in bioprocessing relies critically on a robust irradiation-

sterilization supply chain well-prepared to address growing market demand and unique business continuity challenges.

There is growing global demand for contract irradiation, increasing business and regulatory challenges associated with

cobalt 60 (60Co), limited construction of new gamma-irradiation sites, and the advent of new accelerator technologies.

Coupled with new X-ray service providers entering the market, historically focused on e-beam, a growing biotech single-

use community will likely need to embrace these highly similar, accelerator-based alternatives to gamma irradiation,

such as X-ray.

In addition to reviewing alternatives to gamma-irradiation that can help ensure future business continuity, ISO-11137

prescribed requirements as other recent industry guidances for qualifying e-beam or X-ray as alternative irradiation

modalities are summarized. Whereas as the requirement to demonstrate a minimum sterilizing dose is achieved

through X-ray is relatively straight forward, arguments that X-ray irradiation at the maximum dose impacts single-use

materials in a way that is equivalent or better than gamma, while well-supported by an abundance of heuristic, science-

based rationale by industry experts, is limited by a paucity of publicly available data.

A holistic approach to assessment and qualification of X-ray sterilization entails a fundamental understanding of the

impact of X-ray on single-use materials and components, as well as an overall assessment of the final packaged

assembly. Working as a collective industry group of end-users and suppliers of single-use systems, the BPSA working

team employed the 2015 BPSA quality matrix tool of standard tests performed on single-use components [1], to identify

which tests would most incisively characterize any potential impact of X-ray irradiation as compared to gamma. In

addition to establishing a cross-industry consensus view on the types of testing that will best assess any potential risk,

the working team has identified specific tests that will be performed on representative components and the data shared

with the single-use community. It is expected this risk and data-based assessment of materials and components used in

the biotech single-use industry will support the strongly-touted arguments that X-ray is equivalent or better than

gamma, thereby enabling much of the qualification data already in place for gamma, to be leveraged as fully applicable

to X-ray. Lastly, the path forward including steps required to fully qualify X-ray irradiation and provide appropriate

customer notification timelines is outlined. A successful industry approach to qualifying alternative modes of irradiation

sterilization may strengthen business continuity in the rapidly growing single-use industry, with the end goal of ensuring

innovative patient therapies can be rapidly developed and delivered [2].

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3 SUPPLY-CHAIN RISKS ASSOCIATED WITH GAMMA IRRADIATION Virtually all supplier-sterilized single-use systems (SUS) on the rapidly growing biotech market today are sterilized

through gamma-irradiation by cobalt-60 (60Co), which is associated with a number of unique supply chain risks. These

risks range from the complex production of the 60Co isotope, regulatory restrictions and approvals for handling and

distributing 60Co, a growing overall market demand for ionizing radiation, and fundamental requirements for accurate

long-term planning. The markets for 60Co supply and contract gamma irradiation have been reported to be highly

consolidated [3] [4]. In order to ensure business continuity and mitigate risks, sterilization experts from pharma, medical

device and contract irradiators have advocated exploring multiple sterilization modalities to support sustainable

business continuity plans [2].

3.1 Growing Demand for Gamma-Irradiation in a Highly Consolidated Market The global sterilization market, which is largely dominated by ethylene oxide (EtO, 50%) and gamma-irradiation (40%),

was reported as $4.7 B in 2016 and expected to reach $6.9B by 2021, at a CAGR of 8.8% [5]. For gamma-irradiation,

there are over 200 large-scale gamma irradiators scattered globally utilizing 400- million curies (Ci) of 60Co. As 60Co is

constantly decaying with a radioactive half-life of 5.2 years, each irradiation site must replenish its 60Co at a rate of 12%

per year [4] [6], or 48 MCi/yr globally. Regionally, 60Co utilization is split by the US (51%), EMEA (20%), Asia (15%), and

LATAM (14%); among contract irradiation sites located in North America (24%), Europe (25%), APAC (40%), and LATM

(7%) [6] [7] [4]. In the current scenario, with a high and increasing demand for gamma-irradiation, the need for

accurate long-term planning, and a limited number of service providers in the market, the long-term security of supply

for Biotech SUS providers depends strongly on substantial, long-term commitments to irradiators to ensure their

products receive priority, both for routine processing as well as in the event of any disruption.

3.2 Production and Resupply Challenges with 60Co Production and distribution of 60Co used in contract gamma irradiation of SUS is complex, highly regulated, and strongly

dependent on accurate 2-3 year out market need forecasts. 60Co is produced in nuclear reactors by exposing naturally

occurring and stable cobalt-59 to a neutron flux, where a neutron is added to the nucleus to produce 60Co. As of 2017, 60Co was currently produced in approximately 40 nuclear reactors located in eight countries, with Canada and Russia the

largest producers [6]. However, the vast majority of 60Co generated for use in contract sterilization was produced in

Ontario, Canada. The production in CANDU nuclear reactors is performed using adjuster rods made primarily out of 59Co

instead of the typical stainless steel. After 1 to 3 years, the rods are harvested during a routine shutdown, and

encapsulated in stainless steel pencils to prevent leakage. Production starting today will need to satisfy demand 2-3

years from now, when the 60Co is harvested. Future demand forecasts estimate 4.4 % growth [6], and suggest demand

will double in 15-18 yrs [4] [6].

Current market shortages in availability of 60Co have largely stemmed from decommissioning or refurbishments of

reactors used to produce the isotope. As a result, Nordion, the primary global supplier of 60Co used in contract gamma

irradiation, has reported taking a number of key steps to ensure continued availability for 60Co that include securing IP

enabling 60Co production a broad range of reactors and new multinational supply agreements with Russia, China and

India [8] [9]. However, it remains unclear how much industry capacity can be economically added [4].

Resupply of a contract gamma irradiator’s source 60Co to address the activity lost to radioactive decay is typically

performed once per year and needs to be planned well in advance to address security, logistic and installation

requirements. A well-planned installation process typically requires several days, with 1 full day dedicated to

installation, during which time no processing can take place. Although the resupply process is generally reliable, known

issues have occurred leading to an inability to access the irradiation areas for weeks or months [6]. Other scenarios

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include reports of irradiators requiring 6 weeks to complete maintenance repairs during resupply, while asking SUS

suppliers to reduce shipments by 50% during this period.

3.3 Off-Switch Not Included (Always On, Always Irradiating) As costly 60Co continuously produces photons, regardless of whether it is being used for commercial sterilization

purposes, or sitting unused, there is a high commercial pressure to ensure the right amount is located in the region and

facility needed, and to maximize 60Co utilization 24 hrs/day, 7 days/week. However, for an irradiator who has

successfully-optimized its 60Co utilization and is operating at or near capacity, any disruptions to its operations will

quickly ripple through the supply chain causing delays or capacity restraints in irradiation of goods. Similarly, demand

spikes in the need for biopharma goods, such as observed during the 2020 COVID pandemic, can also stress the supply

chain impacting factors such as single-use availability, contractual-commitments and industry need.

3.4 Regulatory, Government, and Security Pressures The radioactive nature of 60Co and fears that such isotopes – due to accident, oversight or sabotage – could be acquired

and used in radiological dispersal devices (“dirty bomb”). Its use and transport remains highly scrutinized by authorities

[10] [11], with growing pressure from US [11] and European [12] authorities to research and evaluate alternative

technologies for their radioactive sources. One example of the regulatory challenges and approval timelines associated

with construction of new contract gamma irradiation, is the Gammatec facility (Languedoc-Roussillon, France) which

opened in 2013, 7 years following the license application in 2006 [13]. In the US, a 2015 US Appropriations Bill that

would have phased out the use of radioisotopes was proposed and failed. The US Committee on Homeland and National

Security then established an interagency working group on alternatives to high-activity radioactive sources, with the

remit to establish best practices for transitioning to non-radioisotopic technologies [11]. Consequently, whereas it

remains possible to expand irradiation capacity at existing facilities, there are enormous and increasing regulatory

hurdles for construction of any new gamma irradiation sites.

3.5 Market Plans to Support Future Irradiation Capacity The contract irradiation market has and continues to experience significant consolidation. This consolidation appears to

be impacting strongly how and how quickly the industry is allocating future capacity of X-ray vs gamma. Due to business

sensitivities, the market-drivers and how they impact the urgency for qualification of X-ray sterilization are beyond the

scope of this paper.

3.6 Urgency, Timelines, and Need for Action In today’s interconnected global economy that underpins a $300B biologics drug manufacturing industry critical to

public health, business continuity risks such as hurricanes, tsunamis, and global pandemics no longer sound so far-

fetched. Moreover, the current shortage of 60Co, increasing demand for sterilization, and plans by the leading contract

sterilizer to build future capacity with X-ray suggests the gamma-irradiation market is strained and susceptible to risk.

Although hard numbers in terms of total market irradiation capacity, requirements for the rapidly growing SUS industry,

and impact of other irradiation consumers (e.g. medical device, food irradiation) are difficult to exact and predict, an

analysis by a SUS supplier focused on Western Europe has been shared within the BPSA X-ray working group, including

contract sterilizers, with general agreement that it is representative of the larger, global industry trend already being

observed today (Figure 1). Whereas this analysis suggests irradiation capacity will start to have a more significant

impact on market dynamics in 2022, the specific time frames and degree of impact remain educated best guesses based

on limited predictive data. Regardless, the bulk of future capacity appears planned in alternative modalities. Gamma

irradiation will continue to be a cornerstone of overall irradiation market capacity.

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Figure 1: Analysis of gamma-irradiation market demand for SUT vs expected market capacity (Western Europe). Vertical axis indicates estimated biotech consumption of 60Co irradiation capacity. Red arrow indicates expected time in which demand starts to significantly outpace capacity.

4 ALTERNATIVES TO GAMMA IRRADIATION Current major accepted sterilization practices for the medical device and healthcare industry include EtO (50%), gamma-

irradiation (40%), e-beam (4.5%), and a variety of other modalities (5%) including steam and X-ray [14]. In a recent

response to emissions and closures of EtO sterilization facilities, the FDA issued an innovation challenge to identify new

sterilization methods and technologies [15], from which five applications were accepted focusing on supercritical carbon

dioxide, nitrogen dioxide, vaporized hydrogen peroxide, vaporized hydrogen peroxide-ozone, and accelerator-based

sterilization, such as e-beam and X-ray. Note that although UV radiation has been accepted for disinfection of air,

drinking water and contact lenses, its germicidal effectiveness and use is highly material, organism and application

dependent as it generally does not have sufficient energy to ionize particles in the same way as gamma, X-ray and e-

beam [16] [17].

With regard to ionizing radiation, the definitive standard employed and referenced for sterility validation of single-use

systems, ISO 11137-1:2006 “Sterilization of health care products — Radiation — Part 1” [18], is agnostic with respect to

the modality of irradiation, and treats the requirements for gamma, X-ray or e-beam equally. In this sense, both X-ray

and e-beam offer similar technical and validation strategies to gamma, and avoid complications associated with gas and

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vapor-based sterilization techniques. Both X-ray and e-beam could be used to sterilize single use systems mainly

depending on their configuration and thickness.

E-beam sterilization, initially developed for commercial sterilization of medical devices by J&J in 1956, is a viable, low-

cost alternative to gamma sterilization, and started to gain increasing market share in the 1980’s and 1990’s with

advancements in accelerator power and reliability allowing it to be routinely used for applications such as sutures,

gloves, gowns, face masks, dressings, syringes and surgical staplers [5]. Fundamentally e-beam employs electromagnetic

fields controlled via an accelerator to emit a highly charged stream of electrons that directly impact the product and

DNA of microorganisms. However, unlike the photons emitted via 60Co-generated gamma irradiation, e-beam charged

particles (electrons) possess a charge and very small mass that limits their ability to penetrate product. The poorer

penetration and hence dose uniformity properties of e-beam limit its use to lower density (~0.25g/cm3) products and

box-sizes, as opposed to the larger palettes often used with gamma.

X-ray sterilization technology has rapidly evolved and promises to overcome the hurdles associated with penetration

and dose uniformity, and potentially offer some improvements over gamma. The first commercial dedicated X-ray

facility commenced operations in Hawaii in 2000 for phytosanitary treatment of food products, and a second facility

opened near Philadelphia in 2001, which has been relegated to decontamination of mail [6]. Continued technological

advances in accelerator technology led to the 2010 opening of the Daniken, Switzerland commercial site focused on

sterilization of medical devices and additional X-ray sterilizations sites opening by 2021 near Dallas, Texas;

Northborough, MA; Libertyville, IL, Venlo, Netherlands and Germany. Although this added capacity is currently small

compared to that for existing gamma irradiation, advances in X-ray equipment (IBA, Mevex, and CGN Dasheng), low cost

to entry, and investments by major contract sterilizers position X-ray as the leading technology to supplement market

capacity needs.

4.1 TECHNICAL EVALUATION OF X-RAY AS COMPARED TO GAMMA Both gamma-irradiation and X-ray-irradiation are fundamentally the same in that they both rely on a stream of well-penetrating photons to interact with the product, eliciting Compton-scattering effects whereby scattered electrons generate the killing effects on microorganism DNA [19]. However, the way the initial stream of photons is generated is different [6]. In other words, the key feature demarcating X-rays from gamma-rays is how they originate [20]. Gamma-rays arise from atomic nuclei through isotopic decay while X-rays are produced when high energy electrons decelerate on impact with the nucleus of another molecule [20]. X-ray irradiation equipment are basically electron beam systems where a tantalum (or Tungsten) target is added in front of the e-beam. As the high energy, directed electrons interact with the nucleus of the target, energy is released in the form of a similarly-directed X-ray photon (Bremsstrahlung effect) [5]. The conversion efficiency of the electron to X-ray photon is approximately 12% for a typical 7 MeV electron beam indicating equipment power and cost requirements are substantially higher than e-beam. Although extraordinarily similar, the key differences in X-ray and gamma irradiation in their ability to impact polymers are generally considered in the dimensions below.

4.1.1 Irradiation Beam Characteristics Energy Spectra. Although both gamma and X-ray rely on beams of photons impacting the product material, the energy spectral characteristics of the X-ray and gamma ray are different. Gamma rays from 60Co decay are monoenergetic, having discrete energy peaks at 1.17 and 1.33 MeV. However, scattering within the source and surrounding environment can lead to photons at other energies. In contrast, X-rays generated via Brehmsstrahlung irradiation exhibit a much broader and continuous spectrum of energies, including energies below and above those for gamma [4]. X-rays are

sometimes mistakenly considered to be less energetic than gamma-rays, but their energy bands actually overlap [20]. The

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higher X-ray energies (i.e. > 1.33 MeV) are thought to lead to improved penetration of X-rays (and consequently dose uniformity) as described further below. Directionality. The resulting beam of directed X-ray photons allows the flux to be concentrated in the single direction of

the product, irradiating the product only when in front of the beam and optimizing the photon capture rate [4]. In

contrast, photons generated by 60Co gamma irradiation are isotropic, radiating in all directions. To make best use of the

valuable 60Co source, gamma facilities are designed to position optimal volumes of products around the source, both

vertically and horizontally [6].

Dose Rates. Dose rate, or the amount of irradiation dose absorbed per unit time, is inherently faster with accelerator-

based technologies such as e-beam and X-ray when the product is in front of the beam. Hence a product subjected to X-

ray may receive the target dose approximately 6x faster than when subjected to gamma. Typical reported dose rates of

60 kGy/hr have been reported for X-ray sites (372 kW electron beam power), whereas average dose rates of 10 kGy/h

may be expected for similar gamma-irradiation sites [6]. However, it is worth noting that whereas X-ray exhibits a

constant dose rate when product is exposed to the beam, the dose rate for gamma is an average of both lower and

higher dose rates as the product moves on a conveyer over a period of hours around the gamma facility initially far from

and then closer to the 60Co source.

The higher dose rates, and hence shorter irradiation times, are considered advantageous with regard to material impact

[4], typically associated with key benefits including decreased odor generation, color change, and ozone-induced

oxidation [6].

4.1.2 Product Impact Characteristics Penetration. Both the directionality [21] and broader energy spectra [22] of X-rays are thought to contribute to

improved penetration properties of the product [19] [4] as compared to gamma. The directionality, or more narrow

angular distribution of X-rays (i.e. shooting directly at the target product) enables better penetration of materials as

compared to omnidirectional gamma, because the most intense zone of emitted radiation is perpendicular to the

surface of the target products [21].

Dose Uniformity. The directionality and improved penetration achieved with X-ray also contributes to better

consistency and uniformity of absorbed dose across the product as compared to gamma. From a directionality

perspective, X-ray systems improve uniformity by irradiating product loads consistently as the product moves

continuously through the X-ray beam via multiple passes from both sides and at different elevations [21]. For all areas

of the product to receive the absolute minimum sterilizing dose, some portions of the product will invariably receive

higher doses. The Dose Uniformity Ratio (DUR), defined as the ratio between the maximum dose and minimum dose, is

never less than unity, and characterizes the level of overdosing or wasted energy. Trials comparing X-ray and gamma-

irradiation processes under matched conditions have demonstrated DURs as low as 1.25 can be achieved with X-ray,

which contrasts with a poorer DUR of 1.45 for the same pallet irradiated by a 60Co source [21]. Another example with a

pallet of medical devices requiring a minimum sterilization dose of 25 kGy, indicated a X-ray pallet could be processed

with a maximum dose of no more than 30 kGy, whereas the equivalent gamma process would be expected to receive a

maximum dose of 35 kGy [6]. In the immediate field of complex single-use systems used in biomanufacturing where

gamma irradiation windows easily range from 25 kGy to as much as 50 kGy, the improved DURs associated with X-ray

could lead to either more consistently irradiated SUS at lower levels of irradiation, or slightly larger volumes processed

within the same irradiation pallet.

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Oxidative Effects on Polymers. In medical device conferences and white papers from irradiation equipment and service

providers, gamma is frequently touted as much harsher on plastics compared to X-ray and e-beam [6] [21] [23]. The

rationale for diminished oxidative effects is attributed to the high dose rates and shorter exposure times, which

minimize time exposed to ozone [21] and the time for oxygen replenishment required for radical-oxygen reactions,

frequently associated with irradiation [23].

Temperature Effects on Polymers. It is generally regarded that gamma-irradiation is associated with higher ambient

temperatures during processing as compared to X-ray. Although temperatures within the gamma irradiation chamber

vary seasonally, the high density of product around a 60Co source, whereby the product and room equipment are

constantly absorbing irradiation over many hours contributes to higher ambient room temperatures associated with

gamma, as compared to X-ray or e-beam, where absorption occurs only in front of the beam [24]. Comparative data

report temperatures reaching as high as 50C in summer months associated with gamma versus 32.7C for the same

month with X-ray [24]. Temperature increases associated with adsorption of ionizing radiation may also be considered,

but the shorter duration of exposure and lower ambient temperatures associated with X-ray suggest temperatures are

unlikely to approach meaningful transition temperatures within the materials.

4.2 ISO11137 REQUIREMENTS TO TRANSITION FROM GAMMA TO X-RAY The requirements for sterilization of healthcare products via irradiation, whether by gamma, X-ray or e-beam, are

defined in ISO 11137-1. In order to qualify an alternate irradiation modality, such as switching from gamma to X-ray,

three key points need to be evaluated as described by the Panel on Gamma and Electron Irradiation [25]. These include

requirements on the irradiation source, transfer of the sterilization dose, and transfer of the maximum acceptable dose.

4.2.1 Requirements on the Irradiation Source.

ISO 11137-1 (5.1.2) requires the energy levels of the X-ray or e-beams be specified and that, in cases where the level for

X-ray exceeds 5 MeV (7 MeV is typical) or 10 MeV for e-beam, the potential for inducing radioactivity in the product be

assessed. This is also referred to as activation of the irradiated product.

In assessing product radioactivity, the ISO11137 (A5.1) guidance section references a publication by Gregoire et al,

which provides a comprehensive review of materials associated with medical devices and concludes that any imparted

radioactivity in the such devices is negligible and lower than the most conservative regulations [26]. Materials

evaluated by Gregoire using a 7.5 MeV beam with doses up to 50 kGy included the categories below.

materials that have very small potential for becoming radioactive (non-metallic hydrocarbon-based materials,

e.g. polyethylene and polystyrene);

materials that have a potential to be activated at a measurable but low level (e.g. stainless steel and brass); and

materials that have a potential to be activated to comparatively higher levels (e.g. tantalum) requiring detailed

evaluation.

Materials not covered by existing reviews may require further detailed evaluation due to their potential for activity (e.g.

silver and gold) [26].

4.2.2 Transferring the sterilization dose. ISO 11137 Section 8.4.2 addresses transfer of the sterilization dose (and corresponding verification dose) and requires

data indicating that any differences in the operation conditions of the two irradiation sources have no effect on the

microbial effectiveness. To demonstrate that the microbial effectiveness is not altered, a successful dose verification

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experiment is considered sufficient [25] [27]. Such dose verification studies, described in ISO11137-2, are routinely

performed on representative SUS as part of the bioburden assessment and dose audit process for gamma sterilization,

and would need to be similarly performed on representative systems subjected to X-ray. In other words, X-ray

verification dose experiments can be performed with the same irradiation dose used in gamma verification dose

experiments or dose audits.

Data supporting the position that X-ray irradiation dose and gamma-irradiation dose achieve equivalent killing

effectiveness are largely available in existing literature. To answer the question whether there are differences between

the effectiveness of the irradiation modalities, D-values or decimal reduction dose values need to be compared. The D-

value determines the dose that is necessary to kill 90% (or 1 log) of relevant microorganisms [28]. Several studies have

been performed to compare logarithmic survival data or D-values. Tallentire et al. showed that microbicidal

effectiveness’s for gamma, electron and X-ray radiations are equal [29] for the spores of Bacillus pumilus. For other food

borne microorganisms like Escherichia coli, Salmonella typhimurium, Staphylococcus aureus, Listeria monocytogenes no

significant differences in bactericidal efficiency could be observed [30]. Furthermore, decontamination efficiency of X-

ray and gamma irradiation on spices [31] and dried pepper powder [32] were compared with no major differences being

observed. Overall no differences are expected between X-ray and gamma sterilization dose audit experiment studies,

which demonstrate sterility of the single use products.

4.2.3 Transferring the maximum acceptable dose. ISO 11137-1 Section 8.4.1 indicates the maximum acceptable dose for an existing modality (i.e. gamma) can be

transferred through a documented assessment indicating any differences in irradiation conditions do not affect the

validity of the established maximum dose [25]. Guidance associated with clause 8.4.1 pays specific attention to

temperature and dose rate with the remark that higher dose rates, such as with the move from gamma to X-ray, may

lower unwanted effects upon product.

In the biotech industry, there is typically an abundance of historical test data (e.g. extractables, performance

characterization, etc.) performed on SUS and components subjected to the maximum gamma irradiation dose (e.g. 50

kGy +/- 10%) qualified by the SUS supplier. These data packages support justification that SUS irradiated in the range

from the minimum sterilization dose to the maximum dose are well-suited for biopharmaceutical applications in which

they have been qualified. In order to transfer the existing maximum irradiation dose established with gamma to X-ray,

an assessment is required to demonstrate the X-ray photons do not detrimentally impact the materials as compared to

the equivalent levels of photons generated by gamma irradiation. In this way, existing data support packages generated

with gamma at the maximum dose, can be justified as fully relevant and applicable to X-ray sterilization.

Ongoing cross-industry efforts qualifying X-ray sterilization of medical devices, such as that led by Team NABLO of

Pacific-Northwest-National-Labs, have generally indicated X-ray continues to be less detrimental, yielding equivalent or

better results compared to gamma irradiation [33] [24]. These studies, which aim to support increased acceptance of

alternative irradiation modalities to gamma, typically combine a fundamental evaluation of the impact of X-ray on the

basic materials of construction as well as limited scope performance testing of the device. As more data is shared in the

public domain evaluating the impact of X-ray on materials, the expectation is that X-ray will be seen as a gentler or

equivalent alternative to gamma, which can help ensure business continuity and sustainable growth of single use

systems.

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5 IMPACT OF IRRADIATION TO POLYMERS When radiation from a gamma, electron beam, or X-ray source interact with a polymer material, it either directly (e-

beam) or indirectly (X-ray, gamma) yields an abundance of electron interactions within the product material, that are

responsible for the killing effects on microorganisms as well as any impact to the polymer itself [34]. Thus nearly all

physical and chemical changes in polymers are produced by energetic electrons, and no major differences are expected

with respect to effects caused by the different irradiation forms.

The results of these electron interactions can lead to active species such as radicals, which initiate various chemical

reactions. The fundamental processes that result from these interactions are summarized below, where degradation

related to chain scission, and oxidative or free radical effects are generally a primary concern for bioprocess polymers.

[35]

Crosslinking where polymer chains join forming a network

Chain scission where the molecular weight of the polymer is reduced

Oxidation where the polymer molecules react with oxygen via peroxide

Radical formation (oxidation and chain scission often occur simultaneously)

Long-chain branching where polymer chains are joined, but a three-dimensional network is not yet formed

Grafting where a new monomer is polymerized and grafted onto the base polymer chain

In general, many polymers can, based on their intrinsic chemical structure, be grouped into categories indicating how

they respond to ionizing irradiation [34].

Cross-linkable polymers: PE, PMA, PCL, PDMS; Polymers with more hydrogen atoms on the side

Radiation degraded polymers: PP, PMMA, PLA, PTFE, POM; polymers with a methyl group (e.g., polypropylene),

di-substitutions (e.g., polymethylmethacrylate) and per-halogen substitutions (e.g., PTFE)

Radiation resistant polymers: PS, PC, PET, aromatic polymers with benzene rings either in the main chain or on

the side

The level of resistance a polymer exhibits to degradation by ionizing radiation generally depends on the base structure

of the polymer as well as additives that may be included to enhance stability [36] [37] [35]. AAMI Technical Report 17

generally serves as the definitive guide on polymer compatibility with various sterilization methods, but in regard to

ionizing radiation, it strongly focused on studies obtained via gamma irradiation and does not discriminate between

modalities such as gamma, X-ray or e-beam. Combined with other published studies [37] [38] [39] and reviews [40] [41]

[42] [43] helps to paint a fuller picture of the general ionizing irradiation compatibility of polymers typically employed in

bioprocessing components (Figure 2).

13




Figure 2: Irradiation compatibility of polymers frequently used in bioprocess components, compiled from multiple sources. Table is intended to be representative of polymers used with bioprocess components, but not exhaustive. (green circle) highly resistant, (yellow triangle) limited resistance, (red diamond) poor resistance at 50 kGy.

Polymers known to be highly resistant to ionizing irradiation effects will generally be expected to warrant less attention,

than those with limited or poor compatibility. Hence data showing that polymers with limited ionizing radiation

compatibility perform equivalent or better when subjected to X-ray as compared to gamma, will be critical to

establishing a sound, fundamental rationale that X-ray is in most all cases less impactful or less harsh to polymers as

compared to gamma.

Conventional approaches for evaluating polymer compatibility with ionizing radiation tend to focus on polymer stress-

strain or physical measurements following exposure. This fundamental assessment of polymer compatibility will be

strengthened through the use of additional techniques such as FTIR, DSC, and coloration that help provide a more

robust characterization of the physicochemical characteristics of the polymer. The use of the spectrometric technique

FTIR-ATR (Fourier Transform Infrared – Attenuated Total Reflectance) allows specifically to examine the polymers

surface [44]. The changes due to irradiation are estimated from the relative increase or decrease in the band intensities

of functional groups present in the polymeric chain. DSC (Differential Scanning Calorimetry) is used to measure and

analyze the reaction of polymers to heat, including properties such as heat capacity, glass transition temperature,

crystallization temperature, and melting temperature [45].

6 RISK-BASED TESTING APPROACH TO QUALIFICATION OF X-RAY Given that the physics associated with X-ray and gamma interaction with materials is comparable, and supporting data

and arguments suggest that X-ray is equivalent or better regarding their impact of materials, a risk-based testing

approach is advocated that seeks to verify through incisive testing of representative components and systems that

existing qualification data for gamma, can directly be applied to X-ray as worst case. Collectively this assessment of the

impact to materials, components and then single-use assemblies represents a holistic approach to risk assessment of X-

ray sterilization.

HD

PE

LDP

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PC

PEE

K

PEI

PET

PS

PSU

PU

E

PV

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Po

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(N

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PB

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s

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Compatiility with Ionizing Radiation - -

Connectors

Containers (bags, bottles, carboys)

Ports on containers

Sensors

Tubing

Filters

TFF devices

Fittings and molded parts

Pumps, check valves

Needles

O-rings, Gaskets, Seals

Packaging

Sin

gle-

Use

Co

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on

ents

14




6.1 Identifying Materials & Component Tests that Best Assess the Risk The 2015 BPSA quality matrix [1], which defines typical standardized tests used to characterize the quality and

performance characteristics of single-use components, is used as a jump off point to identify dimensions of risk that

could be addressed through a standardized testing approach. For each of the standardized testing categories, we

assessed as low (blue), medium (yellow), or high (orange) whether each of the specified test types would be expected to

add significant value in identifying and assessing risks from X-ray irradiation as compared to gamma. In addition,

participating suppliers indicated via filled circles specific tests being planned on representative components, to help

verify no additional risks were being introduced by X-ray.

Table 1: In determining which standard component tests help best address any potential risk from x-ray irradiation as compared to gamma, colored circles are used to indicate low, medium, or high value. Filled circles indicate a component manufacturer has volunteered to generate and share representative data.

In evaluating which tests could most meaningfully verify the absence of any significant risk related to X-ray, efforts were

made to keep the original structure of the 2015 BPSA quality matrix [1], with the only additions related to material

physicochemical characterization. For several test dimensions, such as material color, biological reactivity, and

particulates, the recommendations and thinking rationale are highly consistent for all categories of components and

summarized below the primary table section. More detailed recommendations and rationale specific to individual types

of components can be found in sections 6.1.1 to 6.2, as well as the supplemental appendix.

15




Table 2: Overview of physical testing per components

A. PHYSICAL TESTS

TEST TYPE TEST REFERENCES Connectors

Valves Retainers

Containers &

Film Sensors Tubing Filters

Pressure Burst Test Manufacturer-defined

method, ISO 7241, ASTM D1599, EN 12266, ISO 1402

-

Integrity (Leak) Test Manufacturer-defined

method, ASTM E515 modified, ASTM D4991, ASTM 1003

Tensile (Pull-Off) Test Manufacturer-defined method - - - -

Tear Resistance ASTM D624, ISO 34, ASTM

D1938-14 - - -

O2 and CO2

Permeability ASTM D3985, ASTM F1927,

ISO 15105-2 - - - -

WVTR ASTM F1249, ISO 15106 - - -

Compression Set Test ASTM D395, ISO 815 - - - -

Durometer (Hardness) ASTM D2240, ISO 868 - - - -

Elongation ASTM D412 - - - -

Tensile Strength ASTM D882, ISO 527 - - -

Material Color Manufacturer-defined method

Glass Transition Temperature by DSC

ASTM D3418, ISO11357-2

Material by FTIR-ATR Manufacturer-defined method

The following physical tests from the BPSA 2015 Quality matrix were assessed as low risk, with no additional testing recommended.

(films) Seal Integrity-Peel, Helium leak, puncture, Integrity leak specifically per ISO 9393-2, and helium-leak integrity. (tubing) specific

gravity. “-“ indicates testing not indicated for component type in 2015 BPSA Quality Matrix

Materials assessment. As the higher dose rates associated with X-ray are thought to result in lower unwanted effects on

the product [46], then it can be argued that the material impact profiles and component functionalities following

gamma irradiation may generally be regarded as a worst-case. To verify that materials are equivalently or less impacted

by X-ray, fundamental physicochemical characterization of representative materials will be performed for all component

types following irradiation (i.e. time zero assessment). FTIR can be employed to assess fundamental information on the

modification of polymers due to irradiation. Any changes due to irradiation can be estimated by the relative increase or

decrease in band intensities of functional groups present in the polymeric chain [47]. Differential scanning calorimetry

(DSC) can also be used to determine any changes in the transition temperatures and heat capacity [48]. It is expected

that such testing will verify that the impact of gamma and X-ray irradiation are equivalent. Packaging materials must

be evaluated as well.

16




Material Color & Visual Inspection. With all modalities of ionizing radiation, dose-dependent color changes to the

material (i.e. yellowing) are one of first observations reported. Although largely cosmetic, these observations do reflect

the impact of ionizing radiation on the material and are expected to be assessed for representative materials subjected

to X-ray as compared and gamma. Most observations to date indicate yellowing of X-ray irradiated components is

equivalent or less than gamma.

Table 3: Overview of functional testing per components

B. FUNCTIONAL TESTS


Valves Retainers

Containers &


Water Flow Rate and Pressure Drop

ISO 7241-2, ISO 3968, Manufacturer-defined

method -

IEC60534-2-3, DIN EN 1267 - - -

Accelerated Aging (Shelf Life)

Manufacturer-defined method, ASTM F1980

Particulate Matter USP <788>, ANSI/AAMI BF7,

BPSA recommandations, EP2.9.19

Kink Resistance/ Bend Radius

Manufacturer defined method - - - -

Filter Integrity Test Manufacturer defined method - - - -

Bacterial Retention Test (Sterilizing Grade Filters)

ASTM F838 - - - -

Bacterial Challenge/ Soiling Test

Manufacturer-defined method, ANSI/AAMI BF7

- - - -

The following functional tests from the BPSA 2015 Quality matrix were assessed as low risk, with no additional testing recommended.

For all components, Packaging Testing/ Transportation & Shipping Integrity are not recommended; for the films, Break at Cold

Temperature Test, Low Temperature Brittleness, Dart Drop, Gelbo, Haze and Transmittance, Plastic Containers Qualification of

Parenteral / Opthalmics testing are not recommended; for the filters, Solute Rejection testing is not recommended.

Shelf life. Shelf life, which indicates the length of time a component may be stored while remaining fit for use, is not

expected to be impacted by X-ray as compared to gamma irradiation [18]. Although the process of irradiation may

impact shelf life, it is expected that the thermal and mechanical materials analysis described above, will verify that any

detrimental impact of X-ray on polymers is equivalent or better to that expected from gamma. The needs and

magnitude of a full-term shelf life study can be evaluated through a risk assessment based on existing information (e.g.

gamma shelf life data) and time zero materials data comparing X-ray and gamma. If results demonstrate meaningful

differences in materials properties at time zero then further evaluation of shelf life requirements may be warranted.

17




Particulates are predominately dependent on the manufacturing environment and conditions, and not expected to be

significantly impacted by irradiation. Furthermore, because gamma irradiation may be regarded as a worse case with

regard to material impact, particulates attributable to X-ray are considered to be low risk [46]. Successful verification

that the material properties are equivalent following X-ray or gamma, will help confirm the absence of any potential risk

of particulate generation related to X-ray. If any of these tests indicate any potential risks for particulate matter, then

representative samples will be tested by participating suppliers in a preliminary study.

Table 4: Overview of biological testing per components

C. BIOLOGICAL TESTS


Valves Retainers

Containers & Film

Sensors Tubing Filters

Biological Reactivity - In Vitro

USP <87>, ISO 10993-5

Biological Reactivity - In Vivo

USP <88>, ISO 10993-1,6,10,11

Bacterial endotoxin and hemolysis testing were assessed as low risk, with no additional testing recommended for all components.

Biological reactivity. As the materials of construction, manufacturing process and environment, and use of ionizing

radiation remain unchanged, there is no expected change to biological reactivity compliances (e.g. USP <87> and <88>),

and existing compliances are expected to remain fully valid. To further substantiate this assessment, limited scope USP

<87>/ISO10993-5 testing may be performed on some representative single-use materials.

Endotoxin levels are primarily associated with raw materials handling strategy and environmental manufacturing’s

conditions, which are identical for X-ray and gamma irradiation. Hence no impact is expected, and testing is not

planned.

18




Table 5: Overview of chemical testing per components

D. CHEMICAL TESTS


Valves Retainers

Containers & Film


Chemical Process Compatibility

Manufacturer defined method typically aligned with ASTM

D543-14 and/or risk assessment

Extractables Manufacturer defined method, <665> - Moderate risk, <665>

high risk, BPOG

Physicochemical Container Test

USP <661>a

EP/ Physicochemical EP 3.1.xb - -

Conductivity Test USP <645> - - - -

pH Shift Test USP <791> - - - -

Total Organic Carbon (TOC)

USP <643> - - - -

USP <381> Elastomeric closures for injections testing were assessed as low risk, with no additional testing recommended for

connectors, valves, and tubing. a The current version of USP <661> is taken as reference in the present protocol. b The purpose of the

EP 3.1 test series is to analyze Materials used in the Manufacture of Pharmaceutical Containers. Raw materials are considered. We

extended to the EP 3.1 series not to include silicone testing as it was referenced in the BPSA 2015 matrix.

Chemical Process Compatibility is generally assessed based on the materials of construction and process contact

conditions including process fluid formulation, temperature, and exposure duration. As these parameters remain

identical between X-ray and gamma- irradiation, no additional risks are expected, or testing recommended.

Extractables are typically generated by suppliers following reasonable worst-case application conditions, including

following exposure to heat or ionizing radiation. As the expectation, data, and scientific arguments to date indicate X-ray

irradiation yields a less deleterious impact on the materials, limited scope verification testing on representative

components using the USP <665> moderate-risk component testing protocol may be appropriate.

19




Table 6: Overview of regulatory testing

E. REGULATORY TESTS


Valves Retainers

Containers &


Animal Origin Free

EMA 410/01

TSE BSE Statement

EMA 410/01, EC 1774

REACH EC/1907/2006

RoHS 2002/95/EC

Food Contact 21 CFR 177 (2600, 2400, 1550,

2510)

Resin and Material compliances. Many compliances including REACH, TSE/BSA and EP physicochemical compliance

(section 3.1) are largely based on the materials of construction and resin formulation, which are not impacted by the

modality or ionizing radiation. These compliances are expected to remain valid, and no additional testing is

recommended.

Table 7: Overview of sterilization and sanitization testing per components

F. STERILIZATION AND SANITIZATION TESTS


Valves Retainers

Containers & Film


Sterilization Process Compatibility

Manufacturer defined method

Irradiation Validation ANSI/AAMI/ISO 11137, AAMI

TR33, CEN ISO/TS 13004

The following functional tests from the BPSA 2015 Quality matrix were assessed as low risk, with no additional testing recommended.

For the filters, sanitization testing is not recommended.

Sterilization Process Compatibility relates to confirmation of manufacturers’ specified performance claims following

sterilization. Functional performance characteristics and suggested testing are described under physical and functional

tests, with further details and rationales provided in sections 6.1.1 to 6.2.

Irradiation Validation per ISO 1113, AAMI TR33, CEN ISO/TS 13004 are key requirements for all components as

described in Section 3.2. It is not necessary to perform the complete dose verification, but is appropriate and sufficient

to perform a dose audit when changing from gamma to X-ray [18] [49]. The key focus of this section (6.1) is on the

impact assessment of the maximum irradiating dose.

20




6.1.1 Connector-Specific Testing Rationale Materials of Construction. Common materials of construction include polysulfone, polyetherimide and polycarbonate

for the connector body. Seals and O-rings generally consist of silicone and there can be a stainless-steel spring.

Physical Testing. As pressure burst testing addresses what is considered a high risk, it may be prudent to ensure that the

device burst pressure is comparable regardless of sterilization modality, gamma irradiation vs X-ray irradiation. Pressure

burst tests can be performed per respective manufacturer defined method. Successful pressure burst testing is expected

to mitigate risk of hydrostatic leaks from the connector. Impact to integrity (leak) test and tensile (pull-off) tests are

considered moderate risk and can be performed per respective manufacturer defined method.

6.1.2 Container and Film-Specific Testing Rationale Materials of Construction. Common materials for containers and films include for instance polyolefins and linked

copolymers ((L)LPDE, EVA, EVOH, PA, polyesters) used in construction of the webs, as well HPDE, EVA, PVC used in

construction of the different components directly sealed to the bag chamber.

Physical tests. Key evaluation consists on highlighting chemical changes in the polymers and chain entanglement

though gas permeability measurements (oxygen, carbon dioxide, water vapor). The film strength or physical resistance

will be evaluated with tensile tests. Chamber integrity, seal integrity-peel check, pressure burst test and integrity (leak)

are performed before irradiation in routine process and no impact of irradiation have been observed on seals. Another

key evaluation involves physical change investigation, which could be evaluated by polymer molecular weight

distribution (by size exclusion chromatography for instance) and the surface energy (analyzed through the traditional

contact angle approach for instance).

Functional Tests. No changes are expected following X-ray irradiation as compared to gamma for the cold temperature

break point and the low temperature brittleness, if no change of the glass transition temperature (Tg) or other specific

thermal features occur. This will be reviewed once the thermal properties are obtained as described in the materials

analysis. Gamma is considered as a worse case compared to X-rays with regards to deleterious or unwanted effects on

plastics [18]. No plastics change/degradation is expected under conditions of normal that would prevent re-evaluation

of haze and transmittance properties if warranted.

As post-gamma shelf life data exist, and time zero post-X-ray data are expected to show equivalency to gamma, then no

new risks to shelf life are expected. No increases in particulates is expected after gamma or X-rays irradiation if materials

are not degraded in relation with physical testing results. This will be reviewed once material testing results become

available.

Packaging material attributes, such thermal properties and chemical fingerprints may require evaluation (e.g. DSC, FTIR).

If, as expected, packaging materials show no degradation with regard to material and physical testing following X-ray,

then the packaging integrity and sterility results for gamma can be expected to fully apply to X-ray. If significant

degradation is observed following X-ray as compared to gamma, then reevaluation may be warranted.

Chemical tests. No impact to chemical compatibility between materials and the process is expected. Initial compatibility

assessments typically based on formation, contact duration, temperature and base polymer composition, will remain

unchanged. The extractables degradation profile is expected to be equivalent or better to gamma and will be evaluated

as per the current evaluation plan. No additional physicochemical container tests are indicated as extractables testing is

expected to provide more incisive information.

21




6.1.3 Sensor-Specific Testing Rationale Functional Tests: The most important testing required for sensors is functionality testing. Changes to sensor

performance post X-ray sterilization are considered to be high risk, especially for single use components with active

electronics. A risk analysis is recommended to evaluate the extent of testing required for different single use sensors.

Functionality testing will vary depending on sensor type. In general, tests should either compare pre- and post-

irradiation performance, or test against an acceptance criterion based on the sensor’s specifications. If risk analysis

demonstrates low risk, then a rational referencing post gamma irradiation results may be used instead. Other

functionality testing, such as shelf life and particulate matter, are considered low risk for sensors and do not need to be

performed at this time. Post-gamma shelf life data can be used as a replacement as material change is expected to be

less significant with X-ray irradiation because of the lower energy per photon. Likewise, post gamma particulate matter

test results can be used to qualify a sensor as irradiation is not anticipated to produce particulates and any effects of X-

ray are anticipated to be less than gamma.

Physical Tests: Physical testing, specifically leak and burst testing should also be conducted on sensors as there is a

moderate risk. Integrity (leak) testing should be considered for all sensor types. Although the risk for a leak post X-ray

sterilization is considered to be relatively low based on existing post gamma data, testing is advised as a precaution. The

integrity test can be performed at the relevant operating pressure for the sensor. For burst testing, a risk analysis can be

conducted to determine if a sensor is at risk for bursting based on its typical usage. Depending on the risk analysis, post

gamma burst test results may be used as an alternative rational, if available. If a sensor manufacturer uses a similar

design or materials for multiple sensors, then the burst test results of the highest risk sensor may be used as a

justification for all other related sensor types if the risk analysis provides this rational.

Biological and Chemical Tests: A risk analysis is also recommended for biological and chemical testing of sensors. Most

sensors are expected to present low overall risk and not require any additional testing. Materials that comprise single

use sensors are expected to have already met all biological and chemical testing requirements post gamma irradiation,

which is assumed to have a more significant effect than X-ray irradiation. In cases where post gamma data does not

exist, justifications can be made using the test results of other components, which use the same material. Depending on

the risk assessment, a rationale to not conduct testing may also be built around the small surface area to volume ratio of

the sensor. If a sensor only marginally contributes to the overall surface area of a single use assembly, then it likely

presents low risk to the overall biological reactivity, extractables, bacterial endotoxins, chemical compatibility, etc. If this

data is also not available, or the risk analysis deems a sensor to be high risk, then important tests, such as USP<88> or

extractables may be required.

6.1.4 Tubing-Specific Testing Rationale Summary: Several BPSA Quality Test Matrix tests will be applied to various grades of tubing to evaluate the impact of an

alternative sterilization method to gamma irradiation. X-ray sterilization may affect both silicone and thermoplastic

elastomers differently than gamma irradiation, and the effects will be compared. Areas of high interest to evaluate

include the changes in cross link density, material degradation, and physical properties of the material. These changes

may affect product performance tests.

Each test, unless noted, will be performed on unreinforced silicone tubing, unreinforced thermoplastic elastomer tubing,

and braid-reinforced silicone tubing, sized 3/8” ID x 5/8” OD. Where noted, some tests will be performed on silicone

tubing overmolded with liquid silicone rubber. Samples will be split between X-ray sterilization at 50 kGy and gamma

irradiation at 50 kGy, and the results will be compared.

22




Physical Test Program: Several physical tests will be explored.

A burst pressure test can be performed on all tubing samples because the method of irradiation could affect the level of

cross linking and/or lead to degradation of the material itself. This could lead to a deterioration in tensile properties,

reduction in tear strength and changes in modulus. Any such changes would be expected to result differences in burst

test performance.

An integrity leak test can be performed for silicone over molded tubing samples because any change in compound

stiffness or elasticity could affect sealing performance. Additionally, any change in the level of cross-linking present in

the elastomeric component of a TPE or the silicone could affect stiffness/elasticity. Changes in elasticity and crosslink

network or the occurrence degradation would result in changes in material relaxation properties, which would have a

direct effect on the ability of the material to for a seal.

Tear strength may be evaluated on representative samples because a reduction in tear strength would be of importance

in terms of damage resistance, therefore tear strength test would be a primary indicator of the effect of sterilization. It is

particularly relevant in respect of silicone elastomers since the tear strength of these materials is generally lower than

that of other elastomers.

Compression set can be evaluated on representative samples because it will measure any differences in relaxation

properties, indicating likely changes in crosslinking, elasticity and stiffness.

Durometer, elongation and tensile strength can be evaluated on representative samples because changes in elasticity

and modulus could be the result of degradation or changes to state of cure.

DSC (Differential Scanning Calorimetry) and TGA (thermogravimetric analysis) will also be performed on samples as

described in the materials assessment. TGA polymer degradation temperatures can indicate degradation. DSC may

indicate changes in state of cure.

The physical appearance and any noted changes of all samples will also be documented.

Functional Test Program: Several functional tests will be explored. Adhesion testing will be performed on over-molded

samples to evaluate if the bond in over-molded junctions between tubing and LSR that would be affected if state of cure

is changed by sterilization method. Fatigue testing will be performed on all tubing samples and may show differences

related to sterilization technique if material modulus/elasticity is altered.

Sterilization and Sanitation Test Program: Several sterilization tests will be explored. Sterilization process compatibility

(confirmation of manufacturer's specified performance claims after sterilization process), irradiation validation

(qualification of the sterilization of healthcare products by irradiation), and moist heat sterilization validation will be

evaluated using representative sample types, per ISO 11137 and other applicable industry standards.

6.1.5 Filter-Specific Testing Rationale Sterilizing-grade, bioburden reduction, clarification and virus-removal filters are commonly used in SUT bioprocesses,

with sterilizing grade filters being used in high-risk, downstream applications near the formulation and filling stage.

Materials of Construction. Common materials for bioburden and sterilizing grade filters include polypropylene, silicone

gasket materials, and PET, used in construction of the filter capsules hardware, as well functionalized PES, PVDF or nylon

membranes used in construction of the retentive membrane.

23




Physical tests. As the physical integrity of filter capsules, often used under significant driving pressures, is key to

operator safety, pressure burst tests will be performed in accordance with a manufacturer-defined method aligned to

ASTM. Introduction of hydrostatic leaks (ASTM 1003) or loss to filter capsule integrity is not expected to be a

significant risk related to the mode of irradiation, but will be verified as part of the burst-test process. Junctions between

filters and connected tubing in single-use assemblies will be assessed for integrity, including leak and pull-strength

testing.

Functional Tests. The pressure versus flow performance characteristics of sterilizing-grade filters, as well as their ability

to render a product free of bacteria are identified as key risks requiring verification testing. Pressure vs flow

characteristics will be evaluated using manufacturer-defined methods. Bacterial retention capability will be assessed

post-irradiation by testing filters post-irradiation by a manufacturer-defined filter integrity test as well as a bacterial

challenge test aligned to ASTM F838. Both the filter integrity test and the bacterial challenge test are standard QC

release test performed on non-irradiated filters as part of the filter manufacturing process.

Chemical tests. Extractables testing such as that prescribed by USP <665> provides a robust characterization of

chemical entities that could potentially migrate from the irradiated material into the drug manufacturing process. As X-

ray irradiation is expected to generate equivalent or better (i.e. less degradation) profiles as compared to gamma,

limited scope verification testing in 50% ethanol/water will be performed in accordance with the USP <665> PF

requirement for medium risk components. This specific solvent generally produces a large number of peaks

characteristic of filter materials and is regarded as providing the most incisive profile of the industry consensus USP

<665> and BioPhorum proposed solvents. Although nonspecific chemical tests such as conductivity, pH shift, and TOC

are regarded as low risk and less insightful than compound-specific extractables profiling, they may be included as these

are typically performed as manufacturing QC release tests for non-irradiated filters.

6.2 Single-use assemblies Whereas the sections above focus on tests typically applied at a material or component level, irradiation is typically

applied to an assembled single-use system, which has been packaged to maintain a sterile barrier around the system

and accommodate transportation requirements. Hence a holistic approach to X-ray qualification for single-use systems

may require a general, fundamental assessment of the impact to materials and components coupled with an overall

impact assessment to the final assembly and packaging.

Table 8: Overview of assembly testing

Single Use-Assembly

TEST TYPE TEST REFERENCES Representative final system

System Integrity Manufacturer-defined methods for junctions or system integrity

Packaging Integrity ASTM D4169, ASTM D4725, DIN ISO 2872, ISTA 2A

Transportation Validation

24




Single-Use System Integrity. SUS integrity, both in terms of an ability to maintain a sterile barrier as well as in terms of

loss of drug substance, has been a key concern that has been seen strong improvements in recent years. Although

radiation in general, or in this case a move from gamma to X-ray, would not be expected to damage or directly

compromise the sterile barrier, a risk assessment focusing on the strength and integrity of junctions, such as between

flexible tubing and a rigid barbed fitting may be warranted. Industry data demonstrating that the strength and

viscoelastic properties of flexible materials remain equivalent or better to gamma follow X-ray, will likely mitigate much

of this concern. In addition, some suppliers have manufacturing defined methods for validating junctions between

single-use components under stress, and such studies would help further demonstrate the suitability for use following X-

ray.

Packaging. The single-use system packaging, in addition to being compatible with irradiation, must protect sterilized

items against microbial contamination during storage, transport and all steps up until the intended use. For the purpose

of transitioning from gamma to X-ray, the suitability of selected packaging materials has been already validated

following gamma irradiation, and hence the general compatibility of the materials are deemed compatible with ionizing

radiation. General physicochemical features such as thermal and tensile properties of the packaging materials can be

evaluated following X-ray irradiation to confirm the material properties are equivalent or better to gamma. Once

confirmed, integrity of the packaging sterile barrier can be expected to remain in the same or better condition

throughout the existing product shelf life claim. Given concerns and criticality around the sterile packaging barrier,

testing of representative packaging systems to verify sterility as a function of shelf life may help further support this

rationale.

Transportation Validation. Following a careful review of the X-ray irradiation process with the contract irradiators, it is

largely expected that no changes to the packaging will be required, and that existing transportation validation studies

can be fully applied to X-ray-treated single-use systems. If considering e-beam as an alternative, where e-beam is much

more suitable to box or tote-size irradiator loading as compared to pallets, this may warrant a further review of whether

packaging changes may be warranted, as well as a revisit to the transportation validation studies. For further guidance

related to transit testing, please see the BPSA transit testing guidance for single use assemblies [50].

7 THE PATH FORWARD Successful evaluation and qualification of X-ray sterilization for single-use systems may mitigate key business continuity

concerns linked to the market experiencing rapidly growing demand for sterilized SUS, and overall growing overall

market demand for contract sterilization. Whereas the existing gamma-irradiation market is not disappearing and will

continue to support current capacity needs, much of the future capacity planning is being rapidly built solely through X-

ray and e-beam. As a biotech single-use industry looking to qualify possible alternatives to gamma sterilization to

mitigate the impact of availability and lead times critical to the delivery of life changing therapies, a voluntary industry

approach among suppliers, end users, and regulators is expected to yield the greatest benefit.

7.1 Key Technical Steps for Collective Industry Approach As part of an industry-approach, testing that addresses the key risks identified in Section 6 may be performed on

representative single-use components or systems and the results shared as a representative, cross-industry dataset. The

expectation is that resulting data may serve to verify that X-ray yields an equivalent or gentler impact on materials

typically associated with bioprocessing, and that existing datasets establishing compatibility with the maximum gamma

irradiation dose, can be used as worst-case data for an equivalent maximum X-ray irradiation dose. Moreover, the

fundamental material assessments will help support and extend this rationale to a more generalized approach

demonstrating X-ray compatibility of polymer families, such as polypropylene, silicone, PES and so forth.

25




The ISO 11137 requirement to assess inducement of radioactivity in X-rayed materials will also be evaluated based on

literature, and any additional testing deemed critical in conjunction with the irradiation service provider.

The ISO11137 requirement to ensure sterility at the minimum or target dose will also be verified through an assessment

of bioburden and sterility testing following irradiation at the verification dose, similar to the routine process performed

on representative gamma-irradiated materials in conjunction with the quarterly dose audit process.

During qualification with an X-ray facility, dose mapping studies may need to be reviewed or repeated to verify the

minimum dose is achieved throughout the pallet and the maximum dose is not exceeded.

7.2 Implementation of X-ray as Alternative to Gamma-Irradiation Per BPSA and Biophorum change notification requirements, a change to the irradiation modality is considered a major

change, requiring a formal notification at least 12 months in advance of the change. The formal notification would be

expected to include relevant validation reports supporting the change qualification as well as detailed guidance

clarifying how the change will be implemented.

Disclaimer

This document is not intended to, nor should it be used to support a cause of action, create a presumption of a breach of legal duty, or form a basis for civil liability. Nothing expressed or implied in this informational document is intended, or shall be construed, to confer upon or give any person or entity any rights or remedies under or by reason of this informational document.

Determination of whether and/or how to use all or any portion of this document is to be made in your sole and absolute discretion. Use of this document is voluntary.

BPSA shall not be responsible or liable for any inaccuracies in the document or the information presented. All warranties express or implied are disclaimed and waived.

Manufacturers, suppliers and end users should consult with their own legal and technical advisors relative to their SUT use and participation. No part of this document constitutes legal advice.

About BPSA

The Bio-Process Systems Alliance (BPSA) was formed in 2005 as an industry-led corporate member trade association dedicated to encouraging and accelerating the adoption of single-use manufacturing technologies used in the production of biopharmaceuticals and vaccines. BPSA facilitates education, sharing of best practices, development of consensus guides and business-to-business networking opportunities among its member company employees.

For more information about BPSA, visit www.bpsalliance.org.

Visit https://bpsalliance.org/technical-guides/ for the full catalog of BPSA guidance documents.

BPSA educational webinars can be found here.

http://www.bpsalliance.org/

https://bpsalliance.org/technical-guides/

https://bpsalliance.org/bpsa-webinar-speaker-series/

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BIOPROCESS EQUIPMENT X -RAY STERILIZATION OF SINGLE -USE