University of Edinburgh: Virology: Professor Dorothy H. Crawford

Professor Dorothy H. Crawford

MBBS, PhD, MD, DSc, FRCPath, FRSE, OBE
Robert Irvine Professor of Medical Microbiology

Contact Details

Address	Centre for Infectious Diseases, Summerhall, Edinburgh EH9 1QH, UK
Telephone	+44 (0)131 650 3142
Fax	+44 (0)131 650 6511
Email	D.Crawford@ed.ac.uk

Research Focus

Epstein-Barr virus (EBV) is a herpesvirus which establishes a persistent life-long infection in over 90% of adults world-wide. Infection is usually asymptomatic but the virus is associated with a variety of diseases, including human tumours (see table).

Conditions associated with EBV infection in humans

1. Clinically silent childhood primary infection	Universal in developing countries
2. Delayed primary infection (teenagers/ young adults)	Seen in developed countries: ~75% are clinically silent, as in childhood ~25% have acute infectious mononucleosis (glandular fever) - usually resolves; may become chronic
3. Primary infection with fatal outcome	Very rare familial x-linked lymphoproliferative disease (Duncan syndrome)
4. B lymphoproliferative disease in the immunocompromised host	In organ graft recipients In AIDS patients
5. Oral hairy leucoplakia	In AIDS patients
6. Endemic Burkitt lymphoma	Common in children where falciparum malaria is hyperendemic. Areas of Equatorial Africa and Papua New Guinea
7. Nasopharyngeal carcinoma	Common in south Chinese and Inuit adults
8. Tumours with suspected links to EBV infection	Hodgkin's disease, some T cell lymphomas, leiomyosarcomas, salivary gland cancers, undifferentiated carcinoma of the stomach

(From: Epstein and Crawford, Gammaherpesviruses: Epstein-Barr Virus. In: Topley and Wilson's Microbiology and Microbial Infections. Volume 1. Eds: BWJ Mahy, L Collier, Arnold 1998).

Our studies focus on EBV infection and persistence in the immunocompromised host, particularly organ transplant recipients, where EBV-associated post transplant lymphoproliferative disease (PTLD) occurs in 1-10% and is fatal in around 50% of cases despite treatment.

Immunotherapy for PTLD

Persistent EBV infection in healthy people is mainly controlled by cytotoxic T cells which recognise cells expressing viral and eliminate them. However when the immune system is suppressed either because of a genetic disorder, an infection like HIV, or drugs, then EBV infected cells in the body are not controlled and the virus can drive them to proliferate uncontrollably forming a tumour. The drugs taken by transplant recipients to prevent rejection of their grafted organ suppress their immune system and often allows reactivation of the persistent EBV infection which may lead to PTLD.

Around 80% of PTLD are EBV associated with the tumour cells containing EBV DNA and expressing viral antigens. These tumour cells are therefore targets for EBV specific cytotoxic T lymphocytes (CTLs) and can only grow where these T cells are lacking. In a recent study we used EBV specific CTL grown in the laboratory to treat PTLD.

PTLD in bone marrow transplant recipients has been prevented and treated successfully using EBV specific CTL derived from the blood of the bone marrow donor (1). But in solid organ transplantation (liver, heart, bowel, lung) the donor is not generally available. We therefore established a bank of EBV-specific CTL lines generated from the blood of 100 HLA-typed healthy blood donors (2). We select CTL lines for infusion into PTLD patients with EBV-positive PTLD on the basis of the best HLA-match and highest specific cytotoxicity in in vitro assays (3).

We have now treated 28 PTLD patients with weekly infusions (1 to 7 doses of 2x106/kgm body weight) of partially HLA-matched, allogeneic, EBV specific CTL. We have seen no CTL related immediate or long-term adverse effects or graft-versus-host disease. At 5 weeks after the last infusion all 28 patients were alive and at 6 months 27 of the 28 (96%) were still living. Fourteen patients (50%) achieved complete response with PTLD tumour regression and improved graft function and clinical status. Three patients had a partial response and 11 showed no response. Thus overall a response (complete or partial) to CTL therapy was recorded in 21 of 28 (75%) patients at 5 weeks and 17 of 27 (63%) patients at 6 months. Significantly more female than male patients responded to CTL therapy.

We have also treated 4 non-transplant EBV-positive lymphoma patients (ulcerative colitis, chronic lymphocytic leukaemia, haemophagocytic syndrome and primary immunodeficiency) and CTL infusions were well tolerated in all. EBV-positive lymphoma in the brain of one patient with primary immunodeficiency regressed completely after 7 infusions of allogeneic CTL and the patient remains well and tumour-free after a bone marrow transplant to correct her immunodeficiency (4).

Our CTL bank bypasses the need to grow CTL for individual patients and provides immediate access to fully characterised CTL lines for a large number of patients with EBV-positive lymphomas. We have shown that partially HLA matched allogeneic CTL are safe and effective. This approach can now be utilised to treat other infectious and malignant diseases.

References

Rooney et al. Lancet 1995; 345: 9-13.
Wilkie et al Immunotherapy 2004; 27: 309-316
Haque et al Lancet 2002; 360: 436-442
Wynne et al Lancet Oncology 2005; 6:344-346

Recent Publications

Haque T, Wilkie GM, Taylor C, Amlot PL, Murad P, Iley A, Dombagoda D, Britton KM, Swerdlow AJ & Crawford DH. Treatment of Epstein-Barr virus-positive post transplant lymphoproliferative disease using partially HLA-matched allogeneic cytotoxic T cells. Lancet 360 (9331), 436-442 (2002).

Williams H, Macsween K, McAulay K, Higgins C, Harrison N, Swerdlow A, Britton K & Crawford DH. Analysis of immune activation and clinical events in acute infectious mononucleosis. J Inf Dis 190 (1), 63-71 (2004).

Anthony IC, Crawford DH & Bell JE. Effects of Human Immunodeficiency Virus Encephalitis and drug abuse on the B lymphocyte population of the brain. J Neurovirology 10 (3), 181- 88 (2004).

Wilkie GM, Taylor C, Jones MM, Burns DM, Turner M, Kilpatrick D, Amlot PL, Crawford DH & Haque T. Establishment and characterisation of a bank of cytotoxic T lymphocytes for immunotherapy of EBV-associated diseases. J Immunotherapy 27 (4), 309-16 (2004).

Wynne RF, Arkwright PD, Haque T, Gharib MI, Wilkie G, Morton-Jones M, Crawford DH. Treatment of Epstein-Barr virus-associated primary CNS B-cell lymphoma with allogeneic T-cell immunotherapy and stem-cell transplantation. Lancet Oncology 6, 344-346 (2005).

Conacher M, Callard R, McAulay K, Chapel, H, Webster D, Kumararatne D, Chandra, A, Spickett G, Hopwood, PA, Crawford, DH. Epstein-Barr virus can establish infection in the absence of a classical memory B-cell population. J Virol 79, 11128-11134 (2005).

Thomas R, Macsween KF, McAulay K, Clutterbuck D, Anderson R, Reid S, Higgins CD, Swerdlow AJ, Harrison N, Williams H, Crawford DH. Evidence of shared Epstein-Barr virus isolates between sexual partners, and low level EBV in genital secretions. J Med Virol 78, 1204-1209 (2006).

Crawford DH, Macsween KF, Higgins CD, Thomas R, McAulay K, Williams H, Harrison N, reid S, Conacher M, Douglas J, Swerdlow SJ. A cohort study among university students: identification of risk factors for Epstein-Barr virus seroconversion and infectious mononucleosis. CID 43, 276-282 (2006).

Haque H, Johannessen I, Dombagoda D, Sengupta C, Burns DM, Bird P, Hale G, Miele-Vergani G, Crawford DH. A mouse monoclonal antibody against Epstein-Barr virus envelope glycoprotein 350 prevents infection both in vitro and in vivo. JID 194, 584-587 (2006).