Regulatory Challenges of Recent Low Endotoxin Recovery Regulatory Queries: Pfizer’s Strategy to Evaluate and Implement Global Changes to Meet Evolving Expectations

The frequency and scope of inquiries related to Low Endotoxin Recovery (LER) from health authorities for biotherapeutic product submissions have dramatically increased over the past year. LER studies are required by the Center of Drug Evaluation and Research (CDER) – Food and Drug Administration (FDA). The Parenteral Drug association (PDA) Technical Report (TR) No. 82, Low Endotoxin Recovery, is used as a guidance for performing LER studies for Biologics License Application (BLA) submissions to CDER, although some recent queries diverge from PDA TR 82. In September 2023, the European Medicines Agency (EMA) published Questions and Answers on biological medicinal products, containing the question, “When should Low Endotoxin Recovery (LER, also known as “endotoxin masking) be investigated?”. The provided answer to this question has been updated to refer to TR82 as a guidance for these studies. Due to the increase in queries, an assessment of the queries in a trackable database was completed. This presentation will highlight the major themes observed in these queries, particularly those that had tangible impacts on our LER strategy.

Presented by Jody Peraino MS, Principal Scientist, Pfizer

Jody Peraino is a Principal Scientist in in Analytical Research and Development in the Biotherapeutics Pharmaceutical Sciences organization at Pfizer. She is an internal endotoxin Subject Matter Expert and leads the Low Endotoxin Recovery Expert Working group at Pfizer. She is also responsible for developing, troubleshooting, and validating microbiology methodologies. Jody holds a B.S. in Biology from the University of Missouri – St. Louis, a M.S. in Quality Assurance/Regulatory Affairs from Temple University and has over 18 years of industry experience.

Demystifying Low Endotoxin Recovery (LER)

This presentation will demonstrate that Control Standard Endotoxin (CSE) and Reference Standard Endotoxin (RSE), while adequate as calibrators in water, are not representative of the endotoxin likely to contaminate a parenteral product. Water from the upstream purification system is the most probable source of contamination and is the recommended analyte in Low Endotoxin Recovery (LER) studies. LER is described as the inability to recover a known concentration of CSE or RSE over time. It is a two‑part reaction involving both a chelator and a surfactant. The chelator removes divalent cations, causing lipopolysaccharide (LPS) aggregates to disassemble. Because Lipid A is hydrophobic, polysorbate encapsulates the LPS in a lipid micelle. In this form, endotoxin cannot react with Factor C of the Limulus Amebocyte Lysate. FDA and EMA require all new biologic drugs submitted for approval to be assessed for LER. Organisms used to produce CSE/RSE are grown under high‑nutrient conditions containing divalent cations, which allow “salt bridges” between LPS molecules. In contrast, organisms in purified water systems adapt to low‑cation environments via the PhoP/PhoQ system, stabilizing the outer membrane without divalent cations. Their endotoxin remains intact in the presence of chelators and is therefore the material most relevant to contamination risk.

Presented by Allen L. Burgenson, Global Subject Matter Expert, Testing Solutions at Lonza

Allen L. Burgenson has over 40 years of experience as a Microbiologist in industries regulated by the FDA, including foods, drugs, biologics, medical devices, and cosmetics. He has worked in R&D, QC, QA, regulatory affairs, and now as an SME for endotoxin detection. Allen is involved in several scientific organizations, including the immediate past chair of the Horseshoe Crab Advisory Panel for the Atlantic States Marine Fisheries Commission (ASMFC), a member of the Horseshoe Crab Working Group of the International Union for the conservation of Nature (IUCN), and the immediate past president of the Capital Area Chapter of the Parenteral Drug Association (PDA). He is a co-author of three PDA technical reports, including, TR-50 and 51 regarding mycoplasma detection and filtration and TR-82 regarding Low Endotoxin Recovery (LER). Allen also contributed to USP Informational Chapter <1228.5> on Endotoxin Indicators. Allen has a BA in Microbiology from Rutgers University and an MS in Biotechnology Management from the University of Maryland

Followed by a live Question and Answer session

Spnsored by Lonza

Lonza is the preferred global partner for the pharmaceutical, biotech, and nutrition markets. We focus on enabling treatments that prevent illness and support healthier lifestyles. We optimize scientific innovation and manufacturing technology to enable our customers to serve their patients and consumers. Our comprehensive range of testing solutions supports your efforts in pyrogen and endotoxin testing of raw materials, in-process samples, and manufactured products.

No matter where you are in your process, Lonza’s testing products optimized with our world-class software and hardware solutions and supported by our experts will help streamline your workflows and meet regulatory requirements for injectable drugs and implantable medical devices, including for new modalities used in modern vaccines and biologics. We can help answer your questions about endotoxin testing, whether you use qualitative gel clot, quantitative turbidimetric, or quantitative chromogenic LAL-based assays, or want sustainable recombinant factor C endotoxin testing or sustainable pyrogen testing.

Our BioScience Division provides a large portfolio of testing solutions to support the pharmaceutical and medical device industries, including classical LAL assays (Pyrogent® gel clot, Pyrogent-5000® kinetic turbidimetric, and KineticQCL® kinetic chromogenic assays) and sustainable recombinant Factor C (PyroGene®) assays for endotoxin detection.  We provide monocyte activation assays to detect pyrogenic substances as a sustainable alternative to the Rabbit Pyrogen Test.  We also supply the industry-leading MODA software for laboratory data that eliminates paper from the process, as well as the PyroTec^TM PRO Automated Endotoxin Testing solution, which is the industry leader in automated endotoxin detection.