Studies on the Safety and Efficacy of Plasma-derived Products and Their Recombinant Analogs

Basil Golding, M.D.

Office of Tissues and Advanced Therapies
Division or Plasma Protein Therapeutics
Plasma Derivatives Branch

Biosketch

PROFESSIONAL EXPERIENCE:

After completing Medical School at the Witwatersrand University in South Africa in 1968, I spent one year training as an intern in Israel (Tel Hashomer Hospital), four years training in Internal Medicine, one year in Israel (Rambam Hospital) and three years in South Africa (Johannesburg General Hospital). I then took a two-year training elective in Cardiology in Israel (Hadassah Medical Center). I am a diplomat of the South African College of Physicians.

Starting in 1976, I completed a clinical fellowship in allergy immunology and rheumatology at the Oregon Health Sciences Center in Portland, Oregon. I am a diplomat of the American College of Physicians and the American College of Rheumatology. From 1978-82, I was the Acting Director of Rheumatology at the Portland Veterans’ Hospital.

From 1982-85 I was a Senior Staff Fellow at the NIH working under Thomas Waldmann in the Metabolism Branch at NCI.

In 1985 I joined the FDA as a senior staff fellow. I became an FTE in 1988 as a principal investigator, promoted to a lab chief in 1997, and as division director in 2004 until the present.

General Overview

The goal of our research is to enhance the safety of plasma-derived products, such as proteins used to treat clotting disorders, immune system disorders, and certain other diseases.

Our laboratory is pursuing this goal by developing tests that detect contamination of these products by substances released by microorganisms. Although microbes are filtered out of plasma used to make the products, the small molecules they make can escape filtration and trigger inflammation in individuals treated with the products.

Our tests use a variety of cells that carry molecules called toll-like receptors (TLRs) on their surface. Each group of cells carries a specific type of TLR, which detects and binds to a specific protein made by microorganisms. Tests based on this interaction between TLRs and these proteins, called TLR ligands, will help FDA and industry screen plasma-derived products for contamination by microorganisms or their products.

FDA will make any tests that we develop available to industry so they can be used to increase the safety of plasma-derived products.
More recent work from our laboratory investigates the potential effects of viral contamination of our products on activation of endothelial cells (ECs) via dsRNA and triggering of TLR3. We used Poly I:C as a surrogate to investigate the effects of dsRNA on EC function and showed activation via TLR3, loss of tight junction molecules (cladin-5) and increased permeability of EC membranes.

In addition, we regulate protein therapeutics used to treat deficiency diseases. A major challenge to the use of these products is the development of antibodies which negate their effects. Our research is focusing on "next generation" of proteins modified to enhance their potency to determine if they are safe in terms of immunogenicity. Insights gained from this research may facilitate development of products that are less likely to interact with the immune system and therefore less likely to cause adverse events.,

Scientific Overview

Toll like receptors are naturally occurring pathogen-recognition receptors found on cells of the innate immune system in all species, from drosophila to humans. We developed a panel of human cell lines that express TLRs that detect both human pathogens and compounds derived from these pathogens.

Plasma-derived and recombinant protein products undergo sterile filtration of the bulk prior to filling vials. Thus, intact pathogens are removed, but compounds released from pathogens can remain in the final product. One of the commonest adverse effects of immune globulin administration is chills and fever, most likely due to presence of such compounds in the product.

Current methods for detection of bacterial compounds are the rabbit pyrogen assay and the limulus amebocyte assay (LAL test). Both of these are based on reactivity of non-human cells. In contrast, the method we have developed is based on the reactivity of human cells.

We established the utility of the method by showing that a recombinant product associated with serious adverse events in humans contained the contaminant, flagellin. This was derived from E. coli, which had been used as an expression vector. Testing of the product in the final container using routine methods did not detect this contaminant. In contrast, the utilization of our method which consists of a panel of TLR cell lines showed that the product reacted with cells expressing TLR5, the ligand for flagellin. The specificity of this binding was demonstrated by inhibiting it with monoclonal anti-TLR5 antibody. Analysis by Western blot showed that the product contained flagellin, and confirmed that it was E. coli flagellin using mass spectrometry.

Our current research focuses on immunogenicity of therapeutic proteins that we regulate. Fc-fusion proteins were designed to increase the half-life of proteins used in replacement therapy. Fc-Factors VIII and IX (Fc-FVIII and Fc-FIX) were approved by our division. A panel of Fc-fusion proteins were studied showing that the Fc domain interacts with Fcg receptors on immune cells and complement (C1q). In a mouse model we showed that Fc-FIX induced a Th1-like response whereas FIX alone triggered a Th2 response. This may explain why FIX has been associated with anaphylaxis.

More recently, we showed that Fc-FVIII binds and activates Natural Killer (NK) cells to secrete IFNg and to release Granzyme B and perforin. In addition, Fc-FVIII was able to induce NK cells to lyse target B cells bearing anti-FVIII receptors. This may explain why Fc-FVIII is more efficient than FVIIII in Immune Tolerance Induction, i.e. regimens used to treat hemophiliacs with high titer inhibitory antibodies.

Important Links

Innovation and Regulatory Science- Scientific Poster: Evidence for a Link between an Immune Globulin Product and Thrombosis

Publications

Ann Rheum Dis 2021 Oct;80(10):1359-61
Pause in immunosuppressive treatment results in improved immune response to SARS-CoV-2 vaccine in autoimmune patient: a case report.
Golding B, Lee Y, Golding H, Khurana S
J Thromb Haemost 2021 Sep;19(9):2102-11
Considerations on activity assay discrepancies in factor VIII and factor IX products.
Ovanesov MV, Jackson JW, Golding B, Lee TK
iScience 2021 Sep 24;24(9):103006
Epitope diversity of SARS-CoV-2 hyperimmune intravenous human immunoglobulins and neutralization of variants of concern.
Tang J, Lee Y, Ravichandran S, Grubbs G, Huang C, Stauft CB, Wang T, Golding B, Golding H, Khurana S
Front Immunol 2021 Jun 28;12:692157
Factor VIII-Fc activates natural killer cells via Fc-mediated interactions with CD16.
Lagasse HAD, Hopkins LB, Jankowski W, Jacquemin MG, Sauna ZE, Golding B
Drugs R D 2021 Mar;21(1):1-8
Dosing considerations for antibodies against COVID-19.
Tegenge MA, Mahmood I, Struble E, Golding B
Antibodies 2020 Jun 19;9(2):E24
Considerations for optimizing dosing of immunoglobulins based on pharmacokinetic evidence.
Mahmood I, Tegenge MA, Golding B
AAPS J 2019 Jul;21(4):62
Fc-Fusion drugs have FcgammaR/C1q binding and signaling properties that may affect their immunogenicity.
Lagasse HAD, Hengel H, Golding B, Sauna ZE
Trends Biotechnol 2018 Oct;36(10):1068-84
Evaluating and mitigating the immunogenicity of therapeutic proteins.
Sauna ZE, Lagasse D, Pedras-Vasconcelos J, Golding B, Rosenberg AS
J Thromb Haemost 2017 Apr;15(4):721-34
Modulating immunogenicity of Factor IX by fusion to an immunoglobulin Fc domain: a study using hemophilia B mouse model.
Levin D, Lagasse HA, Burch E, Strome S, Tan S, Jiang H, Sauna ZE, Golding B
Am J Hematol 2017 Apr;92(4):E44-5
Association of immune globulin intravenous (IGIV) and thromboembolic adverse events (TEEs).
Ovanesov MV, Menis MD, Scott DE, Forshee R, Anderson S, Bryan W, Golding B
PLoS One 2016 Aug 9;11(8):e0160875
Poly(I:C) induces human lung endothelial barrier dysfunction by disrupting tight junction expression of claudin-5.
Huang LY, Stuart C, Takeda K, D'Agnillo F, Golding B
J Pathol 2016 Jan;238(1):85-97
1918 pandemic influenza virus and Streptococcus pneumoniae coinfection results in activation of coagulation and widespread pulmonary thrombosis in mice and humans.
Walters KA, D'Agnillo F, Sheng ZM, Kindrachuk J, Schwartzman LM, Keustner RE, Chertow DS, Golding BT, Taubenberger JK, Kash JC
Trends Biotechnol 2015 Jan;33(1):27-34
Fc fusion as a platform technology: potential for modulating immunogenicity.
Levin D, Golding B, Strome SE, Sauna ZE