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  1. Biologics Research Projects

Development of Technologies and Evaluation of Methods Used to Test the Safety of Tissue Intended for Transplantation

Principal Investigator: Shyh-Ching Lo, MD, PhD
Office / Division / Lab: OTAT / DCGT / TVBB

General Overview

Tissue banks distribute more than 1.5 million donated bone and tissue grafts to be used be used in the medical practices each year, according to the American Association of Tissue Banks (AATB), which accredits these facilities. The transplanted tissues usually come from cadaveric donors (e.g., deceased individuals) and include bone, skin, corneas, ligaments, tendons, dura mater (membrane covering the brain), and heart valves. Other tissues obtained from living donors include hematopoietic stem/progenitor cells (cells that give rise to specific types of blood cells) derived from peripheral (vein) and umbilical cord blood, and reproductive tissues such as oocytes (immature eggs) and semen. FDA began regulating the tissue industry in 1993 in order to ensure the safety and effectiveness of human tissues as well as human cellular and tissue-based products (HCT/Ps) that are also drugs, biological products, or devices.

The FDA's donor eligibility rule requires up-to-date and appropriate donor screening and testing. To prevent transmission of viral infectious diseases through tissue transplantation, all human tissues intended for transplantation shall be found negative for viral pathogens Human Immunodeficiency Virus (HIV, type1 and type-2), Hepatitis B virus (HBV) and Hepatitis C virus (HCV) by testing. Additionally, Human T-cell Leukemia Virus-I (HTLV-I) and Human T-cell Leukemia Virus-II (HTLV-II) are considered as Relevant communicable Disease Agents and Diseases (RCDAD) for viable, leukocyte-rich cells and tissues. The screening uses both virus-specific antibody tests and/or virus-specific nucleic acid tests (NAT) approved and licensed by FDA for detecting infections of donors by these 6 viral pathogens.

The agency's Current Good Tissue Practices rule requires methods be put in place to reduce risks during tissue processing. Although transmission of infections is rare, a single human donor has the potential to provide infected tissues for many recipients that can potentially cause life-threatening infections or illnesses for over 100 recipients. Patients whose immune systems are already weakened due to disease or medical treatments are particularly vulnerable to infections transmitted by infected donated tissue. It is important for the FDA to develop and evaluate industry standards for methods designed to ensure tissue sterility and disinfection and to approve tests for emerging infectious diseases that threaten the safety of donated tissues. The agency is also to facilitate development of improved protocols or methods for screening and testing donors for infections, for preventing contamination, and for removing or inactivating organisms that may be present in the allografts.

The agency enhanced its ability to regulate tissues based on sound science by establishing a human tissue microbiology research laboratory in 2009 to study, evaluate and regulate human tissues and tissue-based products for potential risks of transmitting various infectious agents. This laboratory has been developing new methods and molecular technologies for rapid detection and characterization of microbial pathogens including viruses, bacteria, fungi and protozoa in human tissues intended as grafts. Moreover, the laboratory has been developing new scientific capability to increase preparedness against previously unrecognized or emerging infectious pathogens that threaten the safety of human tissue and human cellular and tissue-based products as well as general public health.

Scientific Overview

The Human Tissue Microbiology Laboratory (HTML) in OCTGT has been established to improve the safety of human tissues intended as grafts and HCT/Ps. The mission of HTML is to have better understandings of new scientific trends relevant to regulatory science for the improvement of human tissue safety and prevention of transmitting infectious agents or communicable diseases.

Since both established and newly emerging infectious pathogens pose constant threats to the safety of the public health, it is essential to develop highly sensitive new molecular technologies that enable scientists to detect, identify and characterize individual microbes without the need of the conventional approach of isolation of the microbes by culture. Moreover, the laboratory is developing capabilities that are particularly useful in detecting and identifying previously unknown pathogens. The development of new technologies using microbial genomic approach and bioinformatics could directly support the emergency preparedness mission in improving emergency countermeasure response capability against either naturally occurring or man-made infectious pathogens. The new scientific capabilities and new molecular technologies the laboratory is developing include:

(1) Highly sensitive and specific real-time qPCR: The primer sets for PCR-detection of target microbial pathogens (viruses, bacteria, fungi and protozoa) that threaten the safety of human tissues intended as grafts are designed from microbe-specific signature DNA sequences. The signature sequences are identified from whole genome sequence analysis of the target microbes using bioinformatics tools with various computational algorithms that recognize different patterns of DNA sequences among genomes in different genera and species of microbes. The primer sets designed from selected signature sequences are critically evaluated for their sensitivity and specificity in PCR against the target microbes. A group of validated PCR primer sets can be organized and used to design the real-time qPCR array for target viruses, "high-risk" bacteria and Candida species of fungi.

(2) Genomic sequencing, metagenomics and microbiome study capability using various high-throughput platforms of ultra-deep, non-biased sequencing technology to examine tissue or tissue-based products for the potential hidden microbes. Genomic sequencing and genomic characterization of microbes provide an effective way for studying disease outbreaks and verifying linkage of microbial transmissions through tissue grafts between donors and recipients. Metagenomics and microbiome study are to analyze in large scale all the members of the microbial communities present in the examined tissue sample. These newly available technologies will be uniquely powerful in the detection and characterization of those previously unknown or difficult-to-culture microbes, as well as those microbial agents that transmit an infectious disease with a long incubation time.

(3) Bioinformatic capabilities for HTS data analysis: The HTML is proactively developing capabilities of analyzing massive amount of data using various open sources and commercially available software programs for filtering out human sequences, identifying and characterizing microbial sequences from any given samples examined. The capability of big data analysis is essential for detections, characterization and taxonomical analysis of previously known or newly emerging microbes infecting or contaminating the human tissues to be used in transplantation. The capability has also become increasingly important in fulfilling the regulatory mission of effectively reviewing the newly submitted pre-INDS and INDs.


Genom Data 2017 Jun;12:84-8
Intraspecies comparative genomics of three strains of Orientia tsutsugamushi with different antibiotic sensitivity.
Liao HM, Chao CC, Lei H, Li B, Tsai S, Hung GC, Ching WM, Lo SC

Genome Announc 2016 Aug 18;4(4)
Genomic sequencing of Orientia tsutsugamushi strain Karp, an assembly comparable to the genome size of the strain Ikeda.
Liao HM, Chao CC, Lei H, Li B, Tsai S, Hung GC, Ching WM, Lo SC

Monoclon Antib Immunodiagn Immunother 2016 Jun;35(3):125-34
Developing peptide mimotopes of capsular polysaccharides and lipopolysaccharides protective antigens of pathogenic Burkholderia bacteria.
Guo P, Zhang J, Tsai S, Li B, Lo SC

Microbiol Insights 2016 Apr 18;9:21-8
Development of Candida-specific real-time PCR assays for the detection and identification of eight medically important Candida species.
Zhang J, Hung GC, Nagamine K, Li B, Tsai S, Lo SC

Int J Clin Exp Pathol 2015 Nov 1;8(11):13834-52
Mixed group of Rhizobiales microbes in lung and blood of a patient with fatal pulmonary illness.
Lo SC, Hung GC, Li B, Lei H, Li T, Nagamine K, Tsai S, Zucker MJ, Olesnicky L

Genome Announc 2015 Nov 12;3(6):e01297-15
A novel member of Chitinophagaceae isolated from a human peritoneal tumor.
Lo AS, Merrell DS, Lei H, Sardi A, McAvoy T, Testerman TL

Sci Rep 2015 Nov 23;5:16706
Epstein-Barr virus from Burkitt lymphoma biopsies from Africa and South America share novel LMP-1 promoter and gene variations.
Lei H, Li T, Li B, Tsai S, Biggar RJ, Nkrumah F, Neequaye J, Gutierrez M, Epelman S, Mbulaiteye SM, Bhatia K, Lo SC

Microbiol Insights 2015 Jul 23;8:7-14
DNA sequence signatures for rapid detection of six target bacterial pathogens using PCR assays.
Nagamine K, Hung GC, Li B, Lo SC

Genome Announc 2015 Feb 26;3(1):e01595-14
Draft Genome Sequence of Pantoea sp. Strain MBLJ3, Isolated in a Laboratory Environmental Control Study.
Zhang J, Hung GC, Lei H, Li T, Li B, Tsai S, Lo SC

Genom Data 2014 Dec;2:123-6
Genome sequencing and annotation of Afipia septicemium strain OHSU_II
Yang P, Hung GC, Lei H, Li T, Li B, Tsai S, Lo SC

PLoS One 2013 Dec 18;8(12):e82673
Isolation and Characterization of Two Novel Bacteria Afipia cberi and Mesorhizobium hominis from Blood of a Patient Afflicted with Fatal Pulmonary Illness.
Lo SC, Li B, Hung GC, Lei H, Li T, Zhang J, Nagamine K, Tsai S, Zucker MJ, Olesnicky L

BMC Genomics 2013 Nov 19;14:804
Identification and characterization of EBV genomes in spontaneously immortalized human peripheral blood B lymphocytes by NGS technology.
Lei H, Li T, Hung GC, Li B, Tsai S, Lo SC

PLoS One 2013 Oct 14;8(10):e76142
Isolation of novel Afipia septicemium and identification of previously unknown bacteria Bradyrhizobium sp. OHSU_III from blood of patients with poorly defined illnesses.
Lo SC, Hung GC, Li B, Lei H, Li T, Nagamine K, Zhang J, Tsai S, Bryant R

Exp Biol Med 2012 Dec 1;237(12):1413-23
Differentially expressed genes and pathways induced by organophosphates in human neuroblastoma cells.
Li T, Zhao H, Hung GC, Han J, Tsai S, Li B, Zhang J, Puri RK, Lo SC

MBio 2012 Sep 18;3(5):1-7
A multicenter blinded analysis indicates no association between chronic fatigue syndrome/myalgic encephalomyelitis and either xenotropic murine leukemia virus-related virus or polytropic murine leukemia virus.
Alter HJ, Mikovits JA, Switzer WM, Ruscetti FW, Lo SC, Klimas N, Komaroff AL, Montoya JG, Bateman L, Levine S, Peterson D, Levin B, Hanson MR, Genfi A, Bhat M, Zheng H, Wang R, Li B, Hung GC, Lee LL, Sameroff S, Heneine W, Coffin J, Hornig M, Lipkin WI

J Clin Microbiol 2012 Aug;50(8):2770-3
Identification of DNA signatures suitable for use in development of real-time PCR assays by whole-genome sequence approaches: use of Streptococcus pyogenes in a pilot study.
Hung GC, Nagamine K, Li B, Lo SC

PLoS One 2012;7(8):e43246
Development of real-time PCR array for simultaneous detection of eight human blood-borne viral pathogens.
Pripuzova N, Wang R, Tsai S, Li B, Hung GC, Ptak RG, Lo SC

Science 2011 Nov 11;334(6057):814-7
Failure to confirm XMRV/MLVs in the blood of patients with chronic fatigue syndrome: a multi-laboratory study.
Simmons G, Glynn SA, Komaroff AL, Mikovits JA, Tobler LH, Hackett J Jr, Tang N, Switzer WM, Heneine W, Hewlett IK, Zhao J, Lo SC, Alter HJ, Linnen JM, Gao K, Coffin JM, Kearney MF, Ruscetti FW, Pfost MA, Bethel J, Kleinman S, Holmberg JA, Busch MP, Blood XMRV Scientific Research Working Group (SRWG)

Clin Vaccine Immunol 2011 May;18(5):825-34
In Vitro and In Vivo Studies of Monoclonal Antibodies with Prominent Bactericidal Activity against Burkholderia pseudomallei and Burkholderia mallei.
Zhang S, Feng SH, Li B, Kim HY, Rodriguez J, Tsai S, Lo SC

PLoS One 2011 May 9;6(5):e19867
Production and Characterization of Chimeric Monoclonal Antibodies against Burkholderia pseudomallei and B. mallei Using the DHFR Expression System.
Kim HY, Tsai S, Lo SC, Wear DJ, Izadjoo MJ

Oncotarget 2011 May;2(5):352-5
Mycoplasmas and human prostate cancer: an exciting but cautionary note.
Lo SC, Tsai S

Transfusion 2011 Mar;51(3):643-53
The Blood Xenotropic Murine Leukemia Virus-Related Virus Scientific Research Working Group: mission, progress, and plans.
Simmons G, Glynn SA, Holmberg JA, Coffin JM, Hewlett IK, Lo SC, Mikovits JA, Switzer WM, Linnen JM, Busch MP, Blood XMRV Scientific Research Working Group

J Virol 2011 Mar;85(6):2620-30
Kaposi's sarcoma-associated herpesvirus ORF57 promotes escape of viral and human interleukin-6 from microRNA-mediated suppression.
Kang JG, Pripuzova N, Majerciak V, Kruhlak M, Le SY, Zheng ZM

Proc Natl Acad Sci U S A 2010 Sep 7;107(36):15874-9
Detection of MLV-related virus gene sequences in blood of patients with chronic fatigue syndrome and healthy blood donors.
Lo SC, Pripuzova N, Li B, Komaroff AL, Hung GC, Wang R, Alter HJ

J Biol Chem 2010 Jul 23;285(30):23359-70
The TOR complex 1 is distributed in endosomes and in retrograde vesicles that form from the vacuole membrane and plays an important role in the vacuole import and degradation pathway.
Brown CR, Hung GC, Dunton D, Chiang HL

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