SPECIAL FEATURE - Analytical Testing Trends in 2022

An increasing number of clinical trial registrations, more R&D investment, growing demand for biopharma products, a continued focus on safety and quality, and more third-party testers entering the market are key reasons why the US pharmaceutical analytical testing outsourcing market is expected to reach $5.55 billion in the next five years.¹ Globally, the market could reach $12.4 billion by 2028.²

Specifically, bioanalytical testing is anticipated to experience the fastest growth over this period.¹ In fact, this segment led the global market in 2020 due to the high number of clinical trial registrations.²

This annual Drug Development & Delivery report reveals the innovative technologies and techniques that leading outsourcing providers currently offer for both small and large molecules.

Alcami: Full-Service Biologic Drug Development

Alcami offers comprehensive serv­ices to support biologic drug product development from preclinical/early-phase programs through commercial. Its bioanalytical testing services are designed to support drug substance and drug product development, vali­dation, and routine analysis, including associated raw materials, excipients, components, and finished goods. Typ­ical programs include therapeutic pro­teins, peptides, and nucleic acid products. Capabilities consist of cell-based assays with associated cell cul­ture workflows, ELISA, electrophoresis, amino acid analysis, HPLC/UPLC with various detection modes such as UV/PDA, CAD, ELSD, RI, FLD, and MS.

“Alcami utilizes high resolution MS for intact mass analysis. We perform identity testing of post-translational modifications, glycan analysis, and peptide mapping,” explains Katie Schlipp, Vice President, Laboratory Operations, Alcami. “Additionally, these products are commonly ana­lyzed by ion pairing chromatography, ion-exchange chromatography, and PCR-based approaches.”

Alcami also offers a high-throughput method development for efficient screening of solvents and columns. This platform allows Alcami to select optimal parameters quickly and develop robust and QC-friendly methods in much shorter timeframes, says Ms. Schlipp.

There are some specific key mar­ket trends that Alcami is seeing in the industry. One is the need for microbial in-use or admixture compatibility stud­ies. A microbial in-use study is intended to evaluate the growth of a low level of microorganisms inoculated into the diluted product over the hold time to represent inadvertent contam­ination during rehydration or dilution of the product.

These studies determine if the product will support the growth and/or proliferation of this inadvertent contamination during the holding pe­riod prior to patient administration. “Our team of experts can support the experimental design and execution of in-use studies and provide a final sci­entific report for our client’s filing,” she says.

A second trend is an increase in the need for environmental monitor­ing. “The global health crisis triggered by the COVID-19 pandemic has in­creased the demand for sterilized pharmaceutical formulations and sci­entific advancements in cleanroom technology,” says Ms. Schlipp. “Alcami experts can guide customers through the stringent regulatory framework as­sociated with cleanroom space and provide support specifically tailored to the needs of each cleanroom to en­sure that quality, safety, and efficacy are maintained.”

She adds that a successful envi­ronmental monitoring program be­gins with the appropriate risk assessment, qualification, and certifi­cation activities for the space and util­ities that lay the groundwork for routine testing. To this end, Alcami provides support in personnel plating, sampling performed during client manufacturing activities, and expertise in remediation efforts in case of unfor­tunate events such as failure to HVAC systems.

She says: “Environmental services are holistically enhanced by Alcami’s microbiological services, from micro­bial and fungal identification to disin­fection efficacy and biological indicators testing.”

Beyond these capabilities, Alcami is investing in new capabilities, includ­ing analytical ultracentrifugation, vari­able pathlength UV for A260/A280 determinations, and new Tandem Quadrupole and QTOF instruments as part of a 16,000-square foot reno­vation in its Durham, NC, laboratory to be commissioned early this year.

Along with the significant invest­ments in analytical capabilities and expansions, Alcami recently an­nounced the acquisition of Masy BioServices. “GMP storage is a critical need in the pharma industry,” says Ms. Schlipp. “Masy offers secure and tightly controlled GMP temperature storage from -196°C to 70°C, includ­ing all ICH stability conditions, for var­ious materials including vaccines, biopharmaceuticals, cell banks, tis­sues, compounds, and medical de­vices. In addition, pharma support services through Masy include equip­ment calibration, large-scale valida­tion and qualification projects, SenseAnywhere monitoring system, and equipment sales, and rentals.”

Ascendia Pharmaceuticals: Rescuing Drug Development Programs

Analytical testing is a crucial part of the drug development process for both small and large molecules. And, Muhammad Asif, PhD, executive di­rector, Analytical R&D and Quality Control,  Ascendia Pharmaceuticals, says that Ascendia’s analytical testing has rescued important drug develop­ment programs. For example, Ascen­dia has developed methods that have predicted and helped control the in vivo time-dependent release of an in­jectable drug that precipitates at the injection site and releases slowly. “Such site-specific injections are in­creasingly becoming common and are made in a specific organ, joint or intervertebral space,” Dr. Asif says. “These drugs are intentionally de­signed to precipitate or coagulate to provide maximum effect at the inject site without creating unwanted toxicity by lowering systemic absorption.”

Whether testing liposomal drugs or drugs embedded in a polymer, As­cendia has methods to analyze drug content and release. High-speed chromatography such as Ultra Per­formance Liquid Chromatography (UPLC) and Ultra High Performance Liquid Chromatography (UHPLC) not only cut time, but also the use of or­ganic solvents, explains Dr. Asif. Addi­tionally, higher selectivity offered by such techniques, in many cases, al­lows substitution of a solvent such as acetonitrile, which has a higher toxicity with a less toxic solvent such as methanol. “Actually, Ascendia has a program in place to substitute acetoni­trile with a more environmentally friendly solvent, methanol, wherever feasible.”

Bethyl Laboratories: Custom Antibodies With mIHC Analysis

With 50 years of expertise in on-site manufacturing of antibodies and validation/immuno-analysis services, Bethyl Laboratories (a Fortis Life Sciences brand) has been supporting biopharma/biotech indus­tries and academic researchers for their immunoassays-related. In-house quality control (QC) labs validate the antibodies Bethyl makes for several immuno analytical methods, such as western blot, immunoprecipitation, flow cytometry, and immunohisto­chemistry (IHC). Bethyl’s full-service IHC lab can fix and process tissue for embedding, histological sectioning, and perform immunostaining analysis of a range of analytes. Recently, Bethyl introduced tyramide amplification sys­tem-based fluorescent multiplex IHC (mIHC) to its IHC validation/analytical services.

“mIHC is becoming a key tool in understanding cellular interactions in drug discovery and drug development process e.g., immune cells’ interaction with cancer cells in tumor microenvi­ronment,” says Senior Director of Im­munohistochemistry and Digital Pathology, Dr Mike Spencer, who over­sees the IHC analysis services at Bethyl Laboratories. “As a result, we have in­vested a lot of effort into expanding our mIHC service, improving effi­ciency of target panel-building and optimization, and decreasing turn­around times.”

He describes one of the recent services Bethyl provided to a client that needed to stain tissue microarrays containing control and pathological samples to analyze the status of im­mune infiltrating cells. This was done using a multiplex panel comprised of previously validated antibodies that were translated to the client’s sample type. “We were able to quickly/effec­tively optimize an antibody panel to determine the T-cell numbers and phenotype several T-cell sub-types within the samples,” he explains. “Our ability to efficiently move from opti­mization to immunostaining samples with our mIHC expertise saved months of time, precious samples, and other resources for the customer.”

The Bethyl IHC service lab uses automated whole slide scanners for imaging of the stained tissue sections. This allows for analysis of the entire section as opposed to select regions of interest, providing for a more com­plete picture of markers and their spa­tial relationships, Dr. Spencer says. This is particularly important, as tissue heterogeneity (e.g., in tumors) is one of the leading concerns in immunos­taining analysis. “The automation also allows for consistency and reliability amongst and between samples. With the combination of imaging systems in our lab, we have the capability to image single-stained brightfield im­ages, single-plex fluorescent images, or multiplex images.” These whole slide images are then uploaded to an image database, which the customer uses to view or download their mIHC data for downstream analysis.

Catalent: Innovative Mass Spec Approach Enables Faster Filing

Catalent offers a broad array of small and large molecule analytical solutions, from compendial testing to extractables and leachables and cell-based assays. Specialized capabilities include handling of controlled sub­stances, highly-potent compounds, as well as temperature-, light-, pH-, and oxygen-sensitive APIs.

According to Jeff Schwartzen­hauer, Analytical Group Leader – Product Development, Catalent, the company has extensive experience and the capabilities to perform ana­lytical testing on a variety of dosage modalities including mRNA, mAbs, cell and gene therapy products, and several dosage forms, including solid oral dosages (capsules, tablets, soft­gels), inhalation products, and injecta­bles. Catalent also has expertise in characterization, spanning techniques from residue level analysis up to higher order structure (HOS) using state-of-the-art instrumentation across spectroscopy, biophysical, bioassay, and mass spectrometry.

Additionally, Catalent offers in-house identification of visible particu­lates using modern Fourier transform infrared (FTIR) spectroscopy and Raman microscopes. “Our team of career forensic scientists provides a level of expertise that can rapidly assist in investigations and avoid third-party outsourced testing, which significantly reduces delays,” says Todd Stone, PhD, Director, Analytical Develop­ment, Catalent Biologics.

With regard to biologics, Catalent deploys two advanced mass spec­trometry (MS) analyses in support of drug substance process development and HOS. “Coverage of host cell pro­tein (HCP) is typically performed using 2D gel assays, but suffers from short­comings preventing a true measure of the HCP species involved,” says Dr. Stone. “Catalent uses an MS ap­proach, which provides a more thor­ough assessment, with depth of information on coverage, identity of HCP species, and potential quantita­tion that outpaces the traditional gel-based method, while shortening the timeline for analysis. For HOS, Catal­ent uses a covalent label to decorate molecules, followed by high-resolution MS to examine structural modifica­tions because of stresses typically en­countered in drug substance or drug product processes.”

As an example, a client with a fast-track biotherapeutic required evi­dence that a standard HCP kit was sufficient to detect and quantify clear­ance in its drug substance manufac­turing process, describes Dr. Stone. Proceeding with the standard gel-based approach through a third-party lab exceeded their timeline for filing. “However, using the innovative MS-based HCP coverage approach devel­oped and described above, Catalent was able to provide the supporting in­formation to enable the client filing in much less time, and meet the filing deadline,” he says.

Providing information relies on technology. Arvind Ramakrishnan, Di­rector, Lab Automation, Catalent, says technology and analytics offer the ability to access raw data in near-real-time. “Having early access to data could potentially de-risk projects that could go out of specifications, which in turn, saves cost and time,” he says. “Replacing outdated equipment in the lab with smart devices helps partners share data streams with sponsors in near-real-time. The resultant data streams could then be harnessed using platform technologies to not only inform sponsors on status but also positively intervene and salvage runs that are going out of specifica­tion. Catalent is actively working on proof-of-concept (PoC) ideas to in­crease the digital maturity of external projects.”

He adds that automation and an­alytics approaches using low-code technologies could help generate in silico predictive models, even before running of the actual samples. “Hav­ing models to predict outcomes before actual sample runs could help compa­nies avoid failure costs,” Mr. Ramakr­ishnan says. “Catalent is working with technology partners to help meet this objective through PoC simulation work on “Lab of the Future” platforms.”

DDL, Inc.: Analytical Testing of Injection Devices

When it comes to testing, devices like prefilled syringes, autoinjectors, and pen injectors have specific FDA requirements. For instance, the bulk of the requirements surrounding the physical and mechanical performance of a prefilled syringe to measure at­tributes such as leakage, break loose, extrusion force, and burst resistance is covered by the ISO 11040 series of standards. Many of these same me­chanical and performance measure­ments for auto and pen injectors are outlined in the ISO 11608 series of standards.

Another key aspect of ISO 11040 for glass Luer syringes is the subject of connectivity. The manufacturing process produces a slippery and slightly irregular Luer taper, which can have issues forming a secure connec­tion with some components. The ISO standard addresses this issue by rec­ommending that certain Luer tests from ISO 80369-7 are conducted to demonstrate adequate connectivity to the same components, which will be attached to the syringe in the actual use situation. ISO 80369-7 replaces ISO 594.

The rapidly growing injectable market has also brought the need for more accurate Container Closure Integrity (CCI) testing to meet USP-NF<1207> deterministic CCI re­quirements, says Chris Murphy, Mar­keting Manager, DDL, Inc. The four primary deterministic tests include:

• Helium Mass Spectrometry – best suited for evaluating the inherent integrity of a package system.
• High Voltage Leak Detection – a standard approach for assessing container closure integrity of a non­porous package system.
• Vacuum Decay Testing – applica­ble to any package containing headspace, including, but not lim­ited to, parenteral vial packages, screw-capped bottles, autoinjec­tors, and flexible bags or pouches.
• Headspace Analysis – Assessment of package headspace via laser-based analysis techniques provides a quantitative, nondestructive measure of oxygen, carbon diox­ide, water vapor, or internal pres­sure in a nonporous, rigid or non-rigid package’s headspace.

Additional testing, which accounts for drug/device interactions, will likely need to be performed to bring a pre­filled syringe product to market in­cludes:

• extractables and leachables;
• USP particulate matter identification and determination;
• ISO 11607 package validation; and
• stability studies to determine prod­uct shelf-life.

Metrics Contract Services: Sensitive Detection for Challenging Compounds

More and more APIs are entering the clinical phase with no chro­mophore or possessing a very weak chromophore, says Jerry “Jr.” Mizell, Senior Director of Analytical Services at Metrics Contract Services. “The need for methods that have adequate sensitivity for assay and impurities could pose a substantial issue,” he ex­plains. “Having other means of detec­tion on an HPLC system, such as charged aerosol detection (CAD), re­fractive index (RI), and mass spec­troscopy (MS) will be critical going forward to be successful with chal­lenging compounds.”

As an example, he says Metrics was recently challenged by a clinical project with a very simple API structure containing no chromophore and highly reactive when exposed to water and several other organic solvents. In addition, one of the known impurities and a process intermediate were mu­tagenic and had to be controlled and analyzed at very low levels (ppm). Several approaches had to be taken for assay and impurity analysis includ­ing gas chromatography, RI, and MS. Diluent selection and sample prepa­ration were challenging due to the re­active nature of the active. “But, all challenges were overcome with suc­cessful method development and val­idation for all methods.”

Mr. Mizell adds that MS is also a valuable tool for a CDMO to possess as the FDA’s requirements for ni­trosamines is a requirement that all NDA filings must meet. “The same can be said for elemental impurities as it behooves a CDMO to possess ICP-OES and ICP-MS instrumenta­tion,” he says.

Salubrent: GCHS & GC/MS Capabilities Handle Complex Projects

“Salubrent Pharma Solutions is a recently formed CDMO focused on supporting the shift toward biologics and personalized medicines, where small batch fill and finish services, combined with continuous batch-fed API processes and just-in-time/direct-to-patient delivery, will ensure patients are never without the life-saving ther­apies they require,” says Anand Pad­manabhan, Director of Analytical Development at Salubrent. “To this end, Salubrent has built its analytical services lab to meet the needs of this changing paradigm.”

In addition to its broader offering of small-molecule analytical testing methods, Salubrent has assembled a team of scientists, and specific instru­mentation, focused on large-molecule analytical method development. One specific challenge developing these therapies is the identification of impu­rities in drug substances and finished goods. Mr. Padmanabhan views gas chromatography (GC) as a vital ana­lytical technique in this effort because of its ability to separate the organic volatile compounds of a sample mix­ture (typically drug substances) and detect them, thus determining their presence or absence and/or how much is present. “GC is also helpful in the determination of potency, dissolu­tion rate, cleaning verification, etc. in instances where compounds cannot be detected using HPLC UV,” he adds.

Analysis for actives that do not have a UV chromophore cannot be detected using HPLC alone. “In this area, GC is a useful tool, and by adding mass spectrometry (MS), this instrument becomes even more powerful in the identification of impu­rities,” according to Mr. Padmanab­han. “Overall, a compound is identified via GC-MS not only by com­paring its retention time to a standard (GC), but also by using its mass spec­trum, making GC in combination with MS an extremely powerful analytical tool.”

GC/MS analysis has endless ap­plications in material testing, identifi­cation, and certification. Identification of organic volatile impurities is one of the main applications. In the pharma­ceutical industry, GC/MS is used in re­search and development, production, and quality control. In medicinal chemistry, GC/MS is used in the syn­thesis and characterization of com­pounds and in pharmaceutical biotechnology.

“Salubrent has the GCHS and GC/MS instruments analysis capabili­ties and expertise necessary to handle these types of complex projects,” says Mr. Padmanabhan, adding that he views Salubrent’s ELISA testing, Malvern Mastersizer particle size analysis, and HPLC with PDA impurity and assay potency quantitation capa­bilities as key to supporting the indus­try’s increasing focus on biologics and personalized therapies.

SGS: Sophisticated Instrumentation for ID Testing

SGS has three centers of excel­lence in North America that offer an­alytical testing for biologics – located in Pennsylvania, Illinois, and Ontario, Canada – providing services at every stage of a product’s lifecycle, from early-phase cell bank safety assess­ment and product characterization to later phase method development and final phase GMP product release.

For the last seven years, SGS’s chemistry laboratory in Lincolnshire, IL, has extensively used Pinnacle PCX, a post-column derivatization system from Pickering Laboratories that per­forms analysis of amino acids for in­dividual raw materials as well as the evaluation of small peptides. This in­strument replaced the thin layer chro­matography (TLC) test used in the past to monitor ninhydrin positive sub­stances, explains Natalia Belikova, PhD, Analytical Services Director, SGS. “High Performance Liquid Chro­matography instrument (HPLC) tech­nology is more specific than TLC, has better sensitivity, is faster, and costs less. This instrumentation was also successfully used for identification tests for small peptides, when identification is based on molar ratios of known amino acids and presence/absence of other known amino acids.”

Additionally, the SGS laboratory has an X-ray powder diffractogram D2-phaser (XRPD) from Bruker that is used extensively for the identification of different known polymorphic forms of small molecules. “It also allows us to evaluate API purity,” she says. “Sev­eral clients have asked us to run con­firmatory testing that polymorphic structure of active pharmaceuticals in­gredients does not change when an API is incorporated into the final drug product during the manufacturing process. This methodology is useful when clients ask us to evaluate if sta­bility storage (under International Conference on Harmonization or ac­celerated studies) affects the polymor­phic form of active ingredient.”

Triclinic Labs, Inc.: Chemical Analysis & Solid-State Development

Triclinic Labs provides both cGMP and non-cGMP materials characteri­zation of organic and inorganic mol­ecules. The lab has a DEA-Controlled Substance Registration for Schedule I-V compounds and offers a variety of techniques, including PXRD, NMR, TGA, DSC, DVS, SEM, EDX, IR, FT-IR, Raman (Dispersive, Low Frequency), optical, digital, hot-stage, and polar­ized microscopy, optical rotation, con­tact angle, particle size and quantity analysis, HPLC, GC with headspace, ICP-MS, and a variety of other tech­niques for identifying chemical com­position, contaminants, and unknown substances.

Determination of crystalline in amorphous mixtures is a requirement of the FDA to demonstrate control in chemical manufacturing (CMC). Aeri Park, PhD, Chief Scientific Officer, Tri­clinic Labs, explains that Triclinic has developed innumerable quantitative methods to determine the presence or absence of crystalline polymorphs in solid mixtures at extremely low levels of detection, and well below the FDA’s typical 5% w/w request. These meth­ods are then used to release materials for pharmaceutical drug product manufacturing.

References

1. United States $5.55 Billion Pharmaceutical Analytical Test­ing Outsourcing Market – Size, Share & Trends Forecast to 2027, Research and Markets, Feb. 1, 2021, https://www.globenewswire.com/en/news-release/2021/02/01/2167066/28124/en/United-States-5-55-Billion-Pharmaceutical-Analytical-Testing-Outsourcing-Market-Size-Share-Trends-Forecast-to-2027.html.
2. Global Pharmaceutical Analytical Testing Outsourcing Market Analysis Report 2021-2028, ResearchAndMarkets.com, Oct. 12, 2021, https://www.businesswire.com/ news/home/20211012006008/en/Global-Pharmaceutical-Analytical-Testing-Outsourcing-Market-Analysis-Report-2021-2028.