¹Department of Haematology, Birmingham Heartlands Hospital, Birmingham, UK

²Department of Public Health Laboratory, Birmingham Heartlands Hospital, Birmingham, UK

Correspondence to: Dr D W Milligan, Department of Haematology, Birmingham Heartlands Hospital, Birmingham, B9 5SS, UK

Adenovirus has been recognised as an important pathogen in BMT recipients, especially in patients with GVHD and those receiving T cell-depleted allografts. We report adenovirus infections from an ongoing surveillance study in four patients after a non-myeloablative transplant and their improved outcome following withdrawal of immunosuppression in two patients and donor lymphocyte infusion for relapsed disease in the others. We discuss the control of adenovirus infections following immune manipulations and the feasibility of adoptive immunotherapy for post-transplant adenovirus infections. Bone Marrow Transplantation (2000) 26, 305-307.

adenovirus; immunosuppression; donor lymphocyte infusion

Adenovirus infections have been increasingly recognized as an important cause of morbidity and mortality in allogeneic stem cell transplant recipients.^1,2,3 Most of the serious infections are reported in patients with GVHD or T cell-depleted graft recipients.^1,2,3 Despite its emergence as a life-threatening pathogen treatment of adenovirus infections remains difficult. Anti-viral drugs have been used with variable results.⁴ Immunotherapy has been used with success in other viral infections,⁵ but has not yet been seriously explored in the context of adenovirus infections.

We are currently carrying out a viral surveillance study, examining stool, urine and throat specimens from BMT recipients weekly for 6 months after transplantation. We report the effects of immunological interventions carried out after transplant for control of the primary disease on adenovirus infections in four patients from this surveillance study.

Case reports

Patient 1

A 38-year-old female received an allogeneic stem cell transplant from a matched sibling donor in July 1998 for refractory AML. She was given pre-transplant conditioning with Campath 1H (anti-CD52 antibody), fludarabine, and melphalan with cyclosporin A as GVHD prophylaxis. She engrafted with full donor chimerism on day +16 post transplant. At the same time, adenovirus type 2 (Ad2) was identified from the stool sample on electron microscopy (EM) and culture. Although she was asymptomatic, 14 consecutive stool samples continued to grow adenovirus on culture. Polyomavirus (BK virus) was persistently identified in the urine by EM. Twelve weeks following the transplant she had early evidence of relapsed leukemia. Cyclosporin A was stopped. This was followed 1 week later by donor lymphocyte infusion (DLI) of 3 ´ 10⁷ CD3⁺ cells/kg. Adenovirus was eradicated from the stool 4 weeks after the DLI and six subsequent stool specimens were negative for 5 weeks until she succumbed to progressive leukaemia 23 weeks after transplant without any evidence of GVHD. The polyomavirus was also cleared from the urine 6 weeks after the first DLI.

Patient 2

A 50-year-old female received an allogeneic stem cell transplant from a matched sibling donor in July 1998 because of refractory AML. She was given identical pre-transplant conditioning and GVHD prophylaxis as patient 1. She engrafted on day +14 post transplant with full donor chimerism and developed grade 2 gut GVHD on day +18 following the transplant which promptly responded to steroids, and these were tapered over the next 8 weeks. Fourteen weeks post transplant she developed severe watery diarrhoea. Adenovirus type 2 was cultured from stool samples. Cyclosporin A was stopped at this stage. There was no evidence of GVHD. Clostridium difficile was also isolated from faecal samples. The diarrhoea persisted, though less severely after 10 days of treatment with metronidazole with clearance of clostridium difficile toxin. Adenovirus was isolated in two consecutive stool samples over the next 2 weeks. Five weeks after stopping cyclosporin A, adenovirus was no longer isolated from the stool over the next 3 months surveillance. This corresponded to clinical improvement and complete resolution of diarrhoea.

Patient 3

An 18-year-old male received an allogeneic bone marrow transplant from an unrelated donor (UD) for relapse of acute lymphoblastic leukaemia 6 months following an autologous BMT in January 1999. Conditioning treatment and engraftment kinetics were similar to the previous patients. The immediate post-treatment course was uneventful. Adenovirus (serotype awaited) was cultured from surveillance stool samples 12 weeks post transplant. This was not associated with any symptoms and there was no evidence of adenovirus in other samples. Cyclosporin A was stopped 2 weeks after the first isolation of the virus in view of continued adenovirus infection. The virus was not detectable 3 weeks after the cessation of cyclosporin A. There was no further reactivation of adenovirus over the next 4 months surveillance. He continues to be in remission without any evidence of GVHD.

Patient 4

A 35-year-old male received an allogeneic transplant from a matched sibling for relapsed high grade non-Hodgkin's lymphoma in July 1999. Conditioning treatment and engraftment kinetics were similar to the previous patients. The immediate post-treatment course was uneventful except for CMV reactivation which was treated with ganciclovir. Cyclosporin A was stopped after 60 days following the transplant. Adenovirus was cultured from surveillance stool samples 120 days after the transplant. He was asymptomatic at this stage. He received donor lymphocyte infusion (1 ´ 10⁷ CD3⁺ cells/kg) at the same time for residual lymphoma. Adenovirus was also isolated from the stool for 3 consecutive weeks and from throat samples 2 weeks later. Four weeks after the DLI, adenovirus was not isolated from any further sample for 3 months at the time of this report.

Discussion

Five to 20% of BMT recipients develop adenovirus infections and 1-6.5% develop invasive disease.^1,2 The incidence is higher in T cell-depleted allografts and unrelated allografts with a mortality of up to 50%.³ The major risk factor identified in unmanipulated allograft recipients in these studies has been GVHD. This underscores the role of immune suppression in the genesis of adenovirus infection in allograft recipients.

Anti-viral therapies for adenovirus infections have been of little benefit in fulminant disease or in severely immunosuppressed patients.⁴ Donor lymphocyte infusion has been used effectively to treat relapsed leukaemia and EBV-associated lymphoproliferative disorders.⁵ Hromas et al⁶ reported the only successful treatment of adenovirus infection with DLI. The patient was a recipient of a T cell-depleted graft who developed severe adenovirus-associated haemorrhagic cystitis not responsive to anti-viral drugs or immunoglobulin. He improved following infusion of 1 ´ 10⁶/kg CD3⁺ cells without development of GVHD. The first patient we described with persistent adenovirus infection for 18 weeks ceased to excrete the virus 4 weeks after omission of cyclosporin A and donor lymphocyte infusion, although the treatment was primarily directed at the relapsed leukaemia. The experience was similar with patient 4. Miyamoto et al⁷ reported adenovirus-associated cystitis following a CD34-selected autograft, which was effectively treated by infusion of unmanipulated cryopreserved PBSC.

In the remaining patients we described, withdrawal of immunosuppression alone was followed by control of the localised adenovirus infections without development of GVHD. Complete cessation of GVHD prophylaxis is often used to control early relapse of leukaemia and EBV-associated lymphoproliferative disorders.⁵ In another report, severe adenovirus-associated cystitis in four T cell-depleted allograft recipients did not improve with a delayed T cell add-back while still on cyclosporin A,⁸ emphasising the importance of withdrawal of immunosuppression as the basic requisite to generate CTLs.

In this ongoing surveillance study, we have detected eight adenovirus infections in 50 allogeneic transplant recipients, compared to none from 50 recipients of autologous transplant (unpublished data). Three of the other four allograft recipients with adenovirus infections were severely immunosuppressed. Although they were not symptomatic at the time of isolation of adenovirus from the stool specimens these patients succumbed to fulminant infections within 2-3 weeks of the first isolation of the adenovirus despite treatment with anti-virals.⁴

Our results are entirely based on clinical observations which could have been coincidental. However, the fact that reduction of immunosuppression or adoptive transfer of immunity might have a favourable outcome in post-BMT adenovirus infections is not unexpected, considering the containment of these infections in normal hosts by cellular immune response which is long-lasting.⁹ Flomenberg et al⁹ demonstrated specific CTL response to Ad2 in 29/30 healthy adults, viral structural proteins being important targets for CD4⁺ T cells, whereas in a later study CD8⁺ T cell responses were shown to be directed at the early regulatory proteins as well.¹⁰ Proliferative response was directed against the Ad35 antigens in patients without evidence of prior infection with Ad35. Smith et al¹¹ reported similar results in an in vitro system where CTLs raised against Ad5 could lyse Ad11 infected cells. These results suggest that adenovirus-specific CTLs might recognise conserved epitopes across various serotypes, which might have important implications in the development of adoptive immunotherapy for adenovirus infections.

Despite encouraging experimental studies, clinical reports on immunotherapy in adenovirus infections are sparse. Our observations may generate further interest in the study of adenovirus-specific immune reconstitution following BMT and the generation of adenovirus-specific CTLs.

1 Shields AF, Hackman RC, Fife KH et al. Adenovirus infections in patients undergoing bone marrow transplantation. New Engl J Med 1985; 312: 529-533, MEDLINE

2 Flomenberg P, Babbitt J, Drobyski WR et al. Increased incidence of adenovirus disease in bone marrow transplant recipients. J Infect Dis 1994; 169: 775-781, MEDLINE

3 Blanke C, Clark C, Broun R et al. Evolving pathogens in allogeneic bone marrow transplantation: increased fatal adenoviral infections. Am J Med 1995; 99: 326-328, MEDLINE

4 Chakrabarti S, Collingham KE, Fegan C, Milligan DW. Fulminant adenovirus hepatitis following unrelated bone marrow transplantation: failure of intravenous ribavirin therapy. Bone Marrow Transplant 1999; 23: 1209-1211, MEDLINE

5 Rooney CM, Smith CA, Ng CYC et al. Infusion of cytotoxic T cells for the prevention and treatment of Epstein-Barr virus-induced lymphoma in allogeneic transplant recipients. Blood 1998; 92: 1549-1555, MEDLINE

6 Hromas R, Cornetta K, Srour E et al. Donor leukocyte infusion as therapy of life-threatening adenoviral infections after T-cell-depleted bone marrow transplantation. Blood 1994; 84: 1689-1690, MEDLINE

7 Miyamoto T, Gondo H, Miyoshi Y et al. Early viral complications following CD34-selected autologous peripheral blood stem cell transplantation for non-Hodgkin's lymphoma. Br J Haematol 1998; 100: 348-350, MEDLINE

8 Childs R, Sanchez C, Engler H et al. High incidence of adeno- and polyomavirus-induced hemorrhagic cystitis in bone marrow allotransplantation for hematological malignancy following T cell depletion and cyclosporine. Bone Marrow Transplant 1998; 22: 889-893, MEDLINE

9 Flomenberg P, Piaskowski V, Truitt RL, Casper JT. Characterisation of human T cell responses to adenovirus. J Infect Dis 1995; 171: 1090-1096, MEDLINE

10 Flomenberg P, Piaskowski V, Truitt RL, Casper JT. Human adenovirus specific CD8+ T-cell responses are not inhibited by E3-19K in the presence of gamma interferon. J Virol 1996; 70: 6314-6322, MEDLINE

11 Smith CA, Woodruff LS, Kitchingman GR, Rooney CM. Adenovirus-pulsed dendritic cells stimulate human virus-specific T-cell responses in vitro. J Virol 1996; 70: 6733-6740, MEDLINE