Melissa A. Greenwald, M.D. Office of Cellular, Tissue and Gene Therapies (OCTGT); CBER

1. Issues Related to the Potential Transmission of Trypanosoma cruzi by Human Cells, Tissues and Cellular and Tissue-Based Products (HCT/Ps) Melissa A. Greenwald, M.D. Office of Cellular, Tissue and Gene Therapies (OCTGT); CBER Blood Products Advisory Committee 26 April 2007

2. Overview • Question for the Committee • Background • About HCT/Ps • About Chagas’ disease • Results of Literature Review • Final Comments

3. Question for Committee • Please comment on the current scientific data as it relates to the potential for transmission of Chagas’ disease by HCT/Ps.

4. Background What is an HCT/P? • HCT/Ps are regulated by OCTGT • HCT/Ps encompass a wide variety of products • 21 CFR 1217.3(d) defines HCT/Ps as articles containing or consisting of human cells or tissues that are intended for implantation, transplantation, infusion, or transfer into a human recipient

5. Musculoskeletal tissues Skin Dura Mater Cardiovascular tissues Ocular tissues Reproductive cells and tissues Hematopoietic stem/progenitor cells derived from peripheral and cord blood Other cellular therapies Gene therapies Tissue/device and other combination products Examples of HCT/Ps

6. Not HCT/Ps • Vascularized human organs for transplantation (HRSA oversight); • Whole blood or blood components or blood derivative products; • Secreted or extracted human products, such as milk, collagen, and cell factors

7. Not HCT/Ps (continued…) • Minimally manipulated bone marrow for homologous use and not combined with a drug or a device; • Ancillary products used in the manufacture of HCT/P; • Cells, tissues, and organs derived from animals other than humans; and • In vitro diagnostic products

8. HCT/P Donor Screening & Testing • Donor screening • medical history interview, • physical assessment/examination, and • medical record review • Donor testing • FDA-licensed, cleared or approved donor screening tests • Specifically labeled for cadaveric donors, if applicable and available

9. RCDADs—diseases for which all HCT/P donors are currently screened and/or tested • Human immunodeficiency virus (HIV), types 1 and 2; • Hepatitis B virus (HBV); • Hepatitis C virus (HCV); • Human transmissible spongiform encephalopathy (TSE), including Creutzfeldt-Jakob disease (CJD); and • Treponema pallidum (syphilis) RCDADs are Relevant Communicable Disease Agents or Diseases

10. “New” RCDADs • We have recently issued (final) guidance describing the following to be RCDADs for all HCT/Ps • West Nile Virus • Sepsis • Vaccinia (the virus used in smallpox vaccine) • Today’s discussion will focus on the current scientific data as it relates to the potential for transmission of Chagas’ disease by HCT/Ps and thus, the possible need to test HCT/P donors for T. cruzi

11. Background, continued Chagas’ disease http://www.canr.msu.edu/vanburen/fbenny.htm

12. Transmission of T. cruzi • In addition to transmission by a vector, published literature indicates the following other means of T. cruzi transmission • Vertical (congenitally/mother to infant) • Oral (breast milk or contaminated food) • Entrance via conjunctiva from hand contamination • Blood transfusion • Organ transplantation

13. T. Cruzi lifecycle in humans • Trypomastigotes in blood • Invade tissue/cell • Convert to amastigote form and replicate • Trypomastigotes released into bloodstream to circulate or invade other host cells Tulane.edu

14. Tissue Transmission of T. cruzi • No reports of tissue transmission • Association between the tissue transplant and the development of symptoms may not be recognized • long time between exposure and symptom development in immunocompetent individuals • acute phase usually asymptomatic • chronic (indeterminate) phase asymptomatic

15. Tissue Transmission of T. cruzi (continued…) • May be difficult to recognize a tissue transplant-transmitted infection in an endemic area where there is vector exposure • Tissue allografts generally undergo some processing—some methods may remove or inactivate T. cruzi • Some cellular products are cryopreserved; some tissues are frozen—parasite may survive 2-3 weeks at refrigerator or freezer temperatures, survival beyond that time period is unknown

16. Tissue distribution of T. cruzi in infected individuals • Information is scant • Acute phase • Parasites found in skin lesions at site of transmission • Spread through the bloodstream • Lodge in various tissues, particularly skeletal muscle

17. Tissue Distribution of T. cruzi in infected individuals (continued…) • Chronic asymptomatic phase • muscle (especially cardiac muscle) • nerve • digestive tract • Little investigation of distribution within other tissues during this phase (tendency to look at tissues that have known clinical manifestations) • Parasites have been demonstrated in the affected tissues of individuals who die with cardiomyopathy, megaesophagus, or megacolon

18. Review of T. cruzi literature provided to the committee http://www.ctegd.uga.edu/misc_pages/parasites.html

19. Mouse Studies • Mice inoculated with T. cruzi demonstrated parasites in skeletal muscle, heart, bladder, peripheral nerve, liver, spleen, adrenal gland, brain, and adipose tissue • Parasite load decreased 100-fold over 10 months, still demonstrated visible parasites in skeletal muscle and bladder • Mice subcutaneously inoculated with T. cruzi demonstrated PCR positivity for T. cruzi DNA in ocular tissue and surrounding structures

20. Mouse Studies (continued…) • Experimentally infected mice demonstrated pseudocysts filled with amastigotes in <1% of evaluated tissue sections; but IHC methods demonstrated T. cruzi antigens in 11% of inflammatory infiltrates (visualization of amastigotes may be insensitive) • Experimentally infected mice demonstrated T. cruzi in sternum chondroblasts, osteoblasts, macrophages and fibroblasts (osteocyte and chondrocyte invasion was rare)

21. Human Placenta Study • Human placentas (collected post-partum) were perfused and inoculated with a bolus containing T. cruzi trypomastigotes • Specimens collected immediately following perfusion; after 24 and 48 hours of incubation • T. cruzi DNA was identified in cells within all postinoculation placental tissue specimens

22. Clinical data • Individuals with chronic Chagas’ disease who underwent endomyocardial biopsy demonstrated • T. cruzi antigenic deposits by immunohistochemical techniques • T. cruzi DNA by PCR • Histopathology evaluation—necrosis, inflammatory infiltrates, and fibrosis

23. Clinical Data (continued…) • Study demonstrated a correlation between the presence of T. cruzi antigen and the severity of myocardial inflammatory process • T. cruzi DNA by PCR has been demonstrated in esophageal tissues of persons who died of esophageal Chagas’ disease

24. Final Comments Science Daily.com Credits: WHO/TDR/Stammers

25. How do HCT/Ps transmit infection? • Infectious disease transmission by HCT/Ps is complex—much is unknown • In cases of known transmission of other infectious disease agents (e.g., HIV, HBV, and HCV), it has been difficult to determine whether transmission occurs because of • agent in the tissue, or • agent in blood that is in the tissue

26. How do HCT/Ps transmit infection? (continued…) • Infectious dose of T. cruzi is not clearly defined in the literature, generally believed to be low • What activates the organism to mobilize from the intracellular amastigote stage into bloodborne trypomastigotes is also unknown, but has been demonstrated to occur in persons who were infected via organ transplantation

27. Summary • T. cruzi is found in blood and various tissues • Much is unknown about potential transmission from tissue allografts • Necessary to make public health decisions based on best available information

28. References • Anez N, et al. Myocardial Parasite Persistence in Chronic Chagasic Patients. Am J Trop Med, 1999;60(5):726-732 • Bellotti, et al. In vivo detection of Trypanosoma cruzi antigens in hearts of patients with chronic Chagas’ heart disease. American Heart Journal, 1996 Feb;131(2):301-307 • Buckner FS, et al. Detection of Live Trypanosoma cruzi of Infected Mice by Using Histochemical Stain for β-Galactosidase. Infection and Immunity, 1999 Jan;67(1):403-409

29. References, cont. • Herrera L, et al. Cornea as a tissue reservoir of Trypanosoma cruzi. Parasitol Res, 2006 Nov • Morocoima A, et al. Trypanosoma cruzi: experimental parasitism of bone and cartilage. Parasitol Res, 2006;99:663-668 • Shippey SH, et al. Use of the placental perfusion model to evaluate transplacental passage of Trypanosoma cruzi. Am J Ob Gyn, 2005;192:586-591

30. References, cont. • Vago AR, et al. PCR detection of Trypanosoma cruzi in oesophageal tissues of patients with chronic digestive Chagas’ disease. Lancet, 1996 Sep;348:891-892 • Younes-Chennoufi AB, et al. Persistence of Trypanosoma cruzi in the inflammatory lesions of chronically infected mice. Transactions of the Royal Society of Tropical Medicine and Hygiene, 1988;82:77-83