# Presentation: 512134

## Epstein-Barr Virus: Cancer and Immunosuppression  

- Jeffrey I. Cohen
- Head, Medical Virology Section
- Laboratory of Clinical Infectious Diseases
- NIH

##  Pathogenesis of EBV Infection

-  Pathogenesis of EBV Infection

-                     Cohen
-                         NEJM  2000

##      Cellular Immune Responses Are Critical 				For Control of EBV

- 		non-HLA specific CTLs
- Late IM:	HLA-restricted CTLs (CD8 and CD4):
-      		Lytic epitopes   - up to 40% of CD8 cells
-      		Latent epitopes - up to  2% of CD8 cells
- Healthy EBV seropositive persons:
-      		Latent epitopes-  4% of CD8 cells
-      		Lytic epitopes- 0.1 to 5% of CD8 cells

-      Cellular Immune Responses Are Critical 				For Control of EBV

##      EBV Transforms B Cells In Vitro and the Cells 	Express Limited Viral and Cellular Proteins

-         Rickinson and Kieff, Fields Virology

-   EBV LCLs

- EBV Latency Proteins

-  Cell Genes Induced

##                     EBV Latency Proteins

-                Cohen NEJM 2000

## Oncogene			   Expression in transgenic mice leads to B cell lymphoma; expression in fibroblasts leads to tumors in nude mice 

- B Cell Proliferation                  Upregulates adhesion molecules, CD23, CD40, IL-6, IL-10, etc. Activates NF-B
- Inhibits apoptosis                         Upregulates Bcl-2, A20, Mcl-1

- 	LMP-1 is the EBV Oncogene

- LMP-1

-  H & E

-   (Kulwichit et al PNAS 1998)

##   LMP-1 Mimics constitutively form of CD40 in B cells

- Thorley-Lawson, Nature Rev Immunol, 2001

##                                                 Liebowitz NEJM 1998

-                                                 Liebowitz NEJM 1998

-    Activation of NF-B in Tumor from Patient with                       Post-Transplant EBV Lymphoproliferative Disease

- Lane 1: EBV- B cell
- Lane 2: EBV+ B cell
- Lane 3: EBV- LPD
- Lane 4: EBV+ LPD

##      Diseases Associated with EBV

- 	Infectious mononucleosis
- 	X-Linked Lymphoproliferative Disease
- 	Chronic active EBV
- 	Hodgkin Disease
- 	Burkitt Lymphoma
- 	Lymphoproliferative disease
- 		_EBV in Other Cells_
- 	Nasopharyngeal carcinoma
- 	Gastric carcinoma
- 	Nasal T/NK cell lymphomas
- 	Peripheral T cell lymphomas
- 	Oral hairy leukoplakia
- 	Smooth muscle tumors in transplant patients

-      Diseases Associated with EBV

## Diseases Driven by Epstein-Barr Virus 

- Infectious mononucleosis
- Chronic Active EBV
- X-linked lymphoproliferative disease
- Lymphoproliferative disease
- Oral hairy leukoplakia
- Hodgkin disease                                EBV            EBV-Driven
- Nasopharyngeal carcinoma             Gene	         Cell
- T cell lymphoma                          Expression      Proliferation
- Burkitt lymphoma

##        Patterns of EBV Latent Infection

- Latency
- _Type_     _EBER_  _EBNA-1_  _EBNA-2_  _EBNA-3_  _LMP-1_  _LMP-2_    _Disease_
- 1	          +         +              -              -            -           -            BL
-           +         +              -              -            +          +            NPC, HD
- 	   +         +              +             +           +           +           IM, LPD
- Other        +         +/-            -               -            -          +/-          Carrier

-        Patterns of EBV Latent Infection

##              Burkitt Lymphoma  

-            20% cases in US – children with abdominal tumors
-            AIDS patients – tumors in lymph nodes
- EBV may be one “hit” but all tumors have c-myc translocations
- Dysregulation of c-myc oncogene
- Only EBV EBNA-1 expressed
- Therapy: Chemotherapy

-              Burkitt Lymphoma

## 		Hodgkin Disease

- EBV+:  60-70% of cases in   		     developing countries
-             35-50% cases in US
- EBV in Reed-Sternberg cells
- Therapy: Chemotherapy, radiation
-                Anti-EBV CTLs effective 		in some cases

- 		Hodgkin Disease

- LMP-1 expression

## EBV-Associated Smooth Muscle Tumors 

- Occur in transplant recipients, AIDS patients, congenitial immunodeficiency
- Pathology: leiomyosarcomas and leiomyomas in various organs (especially transplant) and lymph nodes
- Some tumors regress with reduced immunosuppression

## 	EBV Lymphoproliferative Disease

- Occurs with immunodeficiency    	(AIDS, congenital) or after                	transplantation, RA and MTX
- Symptoms:	                                                	 Infectious Mononucleosis	 	 Mass lesions in organs                       	    (less often lymph nodes)
- Risk Factors:                                                    	 Primary infection                                     	 GVHD with increased 	                      	       immune suppression                             	 T cell depleted bone marrow	 	 CMV

-        Cohen NEJM 2000

## Risk for EBV PTLD

- Primary infection- higher viral loads, no memory T cells to EBV
- CMV infection
- Polymorphisms corresponding to low production of IFN-, TNF-; high levels of IL-10
- Level of intensity of T cell immunosuppression

##        EBV Viral Load is Increased in Patients 

- 	   with Lymphoproliferative Disease

-                       Riddler, Blood 1994

-                Viral Load Used to Monitor Transplant Patients:				Increased EBV load at onset of LPD	                    		Used to initiate preemptive therapy

##   Treatment of EBV Lymphoproliferative Disease

- Reduce immunosuppression-                                                         Early, polymorphic lesions often responsive                         Later monomorphic lesions can have chromosomal changes
- Excise localized lesions
- Radiation therapy (for CNS lesions) or chemotherapy
- Anti-CD20 monoclonal antibody (rituximab)
- Interferon-
- For stem cell transplant recipients: donor lymphocyte infusions or donor EBV-specific cytotoxic T cell infusions
- For solid organ transplant recipients: autologous or HLA-matched, EBV-specific, cytotoxic T cell infusions

## Cutaneous Lymphomas Associated with EBV-infected T cellsNon-immunosuppressed Patients                More often in Asians

- Hydroa vacciniforme: vesciulopapular lesions on face and hands, fever, can progress to T cell lymphoma
- Angiocentric NK/T cell lymphomas:ulcers, vesicles, nodules, papules on nose, checks, lips, extremities, trunk
- EBV subcutaneous T cell lymphoma: plaques, fever, hepatosplenomegaly, pancytopenia, panniculitis, hemophagocytosis

## Cutaneous Lymphomas Associated with EBV-infected B cellsImmunosuppressed Patients

- Cutaneous ulcerated nodules- B cell lymphomas after transplant or in patients with AIDS
- Cutaneous B cell lymphomas in patients with rheumatoid arthritis or polymyositis receiving methotrexate- resolution in some after drug stopped

## EBV LPD More Common at Sites with Chronic Inflammation

- Disease more frequent in transplanted organ              	Higher frequency of EBV+ cells
-       	Antigenic stimulation with B cell proliferation
- 		Cytokine activation in organ
- Reports of EBV+ pyothorax-associated pleural lymphomas at site of pleural inflammation after tuberculosis (Arch Pathol Lab Med. 1996)
- Report of 3 cases of EBV+ large B cell lymphomas in patients with chronic inflammation (osteomyelitis- tumor at site of bone, chronic venous ulcers- tumor at site of ulcer) (J Pathol. 1997 )

## Immunosuppressive Agents Associated with EBV LPD

- Steroids and Azathioprine
- Methotrexate: Patients with RA, Polymyositits
- Antibodies:                                             	ATG:   anti-thymocyte globulin                   	ALG:   anti-lymphocyte globulin                      	OKT3: anti-CD3
- Calcineurin inhibitors: cyclosporine, tacrolimus
- Sirolimus

## Methotrexate, but not other Immunosuppressants, 			Induces EBV Lytic Replication 		

-                      Feng et al JNCI 2004

## Calcineurin Inhibitors and PTLD: Cyclosporine, Tacrolimus

- Inhibit generation of cytotoxic activity
- Induce expression of IL-6 and TGF- that supports B cell activation and proliferation
- Enhance survival of EBV-transformed cells in vitro by protecting from Fas-mediated apoptosis
- Lower doses of cyclosporine allow T cell responses to EBV in vitro and are associated with lower rates of lymphoma than higher doses
- In children tacrolimus is associated with a higher risk of LPD than cyclosporine in some, but not all studies.

## Risk of PTLD in Pediatric Liver Transplant Recipients for Primary Tacrolimus Therapy

- Cacciarelli et at Pediatric Transplantation 2001

## Kaposi’s Sarcoma at the Site of Topical Tacrolimus

- Cho et al. J. Am Acad Dermatol. 50:149-50, 2004

- 28 yo AIDS patient on HAART  (CD 143) with psoriasis and seborrheic dermatitis treated with topical tacrolimus 0.1% ointment to axilla, groin, head for 1 month
- Developed KS at these sites and in lungs while on tacrolimus

## Lymphoma at Site of ATG or ALG Injections

- _Age_	 _Transplant_  _AT/LG_ _Sites of Lymphoma_	    _Ref_
-     kidney        horse   buttock, nodes		       1
-     kidney        horse   buttock, nodes, liver        2
- 32	    heart          rabbit   thigh, brain, lung, nodes  3
-     heart          rabbit   thigh, chest wall,              3
- 					   abdominal nodes

- 1. Deodhar et al N Engl J Med 280:1104-6, 1969
- 2. Cotton et al. Transplantation 16:154-7, 1973;                   	follow-up Herrera et al. Mil Med. 146:652-4, 1981
- 3. Weintraub and Warnke Transplantation 33:347, 1982

## Lymphoma at Site of ALG(Cotton et al 1973; Herrera et al 1981)

- 47 y.o. renal transplant recipient
-  thoracic duct canulation before and 3 wks after transplant to deplete lymphocytes; prednisone, azothioprine
- Horse ALG i.m. in buttocks post transplant on x 14 d,            3 x/wk x 1 yr
- 6 months after last ALG nodule at site >reticulum cell sarcoma (no EBV studies), immunosuppression reduced, radiation to site;
- One year later draining lymph nodes had histiocytic lymphoma, radiation (no EBV studies)
- 2 years later died of bacteremia-lymphoma in liver

## Lymphoma at Site of ALG(Deodhar et al 1969)

- 32 y.o. renal transplant recipient on azathioprine and prednisone
- Rejection 7 months after transplant: treated with actinomycin C and graft irradiation
- Horse ALG i.m. in buttocks: 6 weeks later nodule at site, enlarge over 10 months; excised-reticulum cell sarcoma with lymph node involvement (no EBV studies); died of OI

## Lymphoma at Site of ALG(Weintraub and Warnke 1982)

- 7 patients with NHL/182 heart transplants, 2 developed lymphoma at site of ATG
- 32 yo cardiac transplant recipient underwent two allogeneic heart transplants
- 	Developed high grade immunoblastic lymphoma in thigh at site of rabbit ATG, later in brain and lung
- 18 yo cardiac transplant recipient underwent two allogeneic heart transplants
- 	Developed large noncleaved cell lymphoma in thigh at site of rabbit ATG, later chest wall and abdomen

## Summary: EBV LPD in Persons Receiving Immunosuppressants

- Most early, polymorphic lesions are EBV driven, and may respond to reduction in immunosuppression
- Later monomorphic lesions may have chromosomal changes and often require chemotherapy
- More common with primary EBV infection
- May have genetic component (cytokine polymorphisms)
- More common at site of chronic inflammation
- Some occur at sites of local immunosuppression:                  	ATG or ALG injection sites – all patients on other 	immunosuppression