Successful Treatment of Epstein-Barr Virus Reactivation-associated Transverse Myelitis Following Herpes Zoster with Intravenous Rituximab in a Cord Blood Transplant Recipient

Abstract

The Epstein-Barr virus (EBV) is associated with many malignancies and autoimmune diseases, including multiple sclerosis. In addition, EBV rarely but occasionally causes central nervous system (CNS) complications. We herein report a case of transverse myelitis (TM) associated with systemic EBV reactivation after herpes zoster infection in a cord blood transplant recipient. Identification of EBV-infected peripheral blood cells revealed a predominance of B cells. Notably, intravenous rituximab ameliorated EBV reactivation and TM. Since the CNS infiltration rate of intravenous rituximab is markedly low, the clinical efficacy of rituximab against TM suggests that EBV reactivation may cause TM via immune-mediated mechanisms.

Introduction

Transverse myelitis (TM) is a pathologically heterogeneous syndrome characterized by acute or subacute paresis, as well as sensory and autonomic dysfunction (bladder, bowel, and sexual) below the affected spinal level (1,2). Parainfectious TM is associated with various antecedent infections, including bacteria, viruses, fungi, and parasites (1). The pathogenesis of parainfectious TM remains unclear, although possible mechanisms include direct microbial central nervous system (CNS) infiltration, pathogen-induced immune responses, and immune-mediated inflammatory responses triggered by distant infection (1). Empirical treatment of TM typically includes immunological therapies, such as high-dose glucocorticoids, plasma exchange, and intravenous cyclophosphamide (1). However, there is limited clinical evidence to support the efficacy of these treatments.

Epstein-Barr virus (EBV) is a human lymphotropic herpes virus present in approximately 90% of the adult population worldwide. It usually exists as a latent infection in the human body (3). EBV infection is associated with many cancers, including nasopharyngeal carcinoma, malignant lymphoma, gastric cancer, and leiomyosarcoma. Under immunosuppressive conditions, EBV can induce severe lymphoproliferative diseases (4). Recently, EBV has also been identified as a significant risk factor for several autoimmune diseases including multiple sclerosis (4,5). In addition, although rare, EBV can lead to CNS complications, such as meningitis, encephalitis, myelitis, acute disseminated encephalomyelitis, and acute cerebellar ataxia in primary infection (infectious mononucleosis) or reactivation, of which myelitis is extremely rare (6,7). However, the pathogenesis and evidence-based treatment of EBV-associated CNS complications remain poorly understood.

We herein report a case of TM associated with EBV reactivation after herpes zoster infection in a cord blood transplant recipient. Interestingly, in this case, intravenous rituximab was effective against both EBV reactivation and TM infection.

Case Report

A 69-year-old woman developed acute myeloid leukemia (M4Eo) with inv (16)(p13.1q22). The patient had no medical history of collagen disease. She achieved complete remission after 1 course of induction therapy with idarubicin and cytarabine (80% dose) according to the Japan Adult Leukemia Study Group AML201 protocol, followed by 4 courses of consolidation therapy.

However, she showed hematological relapse four months later. Despite her advanced age, she requested and received unrelated cord blood transplantation at 70 years old while in hematologic second remission and molecular non-remission (CBFβ::MYH11 mRNA 60 copies/μg DNA) after reinduction therapy with idarubicin and cytarabine. The conditioning regimen consisted of fludarabine (180 mg/m²), busulfan (12.8 mg/kg), and melphalan (80 mg/m²). Graft-versus-host disease (GVHD) prophylaxis included tacrolimus and mycophenolate mofetil at 1,500 mg/day from days -1 to 35. Serological human leukocyte antigen (HLA) (HLA-A, HLA-B, DR) matches were 4/6 in the GVH direction and 5/6 in the HVG direction. The number of cells infused was 2.2×10⁷/kg for nucleated cells and 1.0×10⁵/kg for CD34+ cells. Neutrophil engraftment was achieved on day 15.

On day 38, upper gastrointestinal endoscopy for persistent nausea revealed GVHD, and acute grade II GVHD (gut stage 1, skin stage 0, and liver stage 0) was diagnosed. Oral prednisolone (PSL) was administered at 5 mg/day. Nausea gradually improved, but tacrolimus and low-dose PSL were continued due to persistent mild nausea and subsequent chronic GVHD of localized skin. On day 284, acyclovir (ACV) prophylaxis was discontinued due to nausea. Post-transplant viral monitoring was performed for cytomegalovirus (CMV) only and not for HHV-6 or EBV; no CMV infections were observed.

On day 669, the patient developed herpes zoster along the left T7 dermatome, which was successfully treated with intravenous ACV. Two weeks after the onset of herpes zoster, she was admitted to our hospital because of progressive gait disturbance, numbness in the left lower leg, and marked allodynia in the resolved herpes zoster area. The clinical course is shown in Fig. 1, with the first day of hospitalization set as day 1. Vital signs and general physical examination results were unremarkable. A neurological examination revealed mild somnolence (Glasgow Coma Scale E3V4M6), normal cranial nerves, no neck stiffness, left-dominant proximal lower extremity paresis, paresthesia at the left Th7 to L2 level, dysesthesia at the same level, voiding dysfunction, and urinary retention. Deep tendon reflexes showed increases in the bilateral biceps, bilateral triceps, brachioradialis, and patella and a normal Achilles reflex. The Babinski's sign was bilaterally negative. Although cervical to lumbar magnetic resonance imaging (MRI) showed no abnormal intramedullary signals or cord compression lesions, the neurological findings were suggestive of TM. Head MRI also showed no remarkable findings. The laboratory findings on admission are presented in Table. The results showed nothing of note regarding the complete blood count, an elevated fibrinogen level, mild hyperferritinemia, a mildly decreased IgG level, and no CMV antigenemia. On day 6, EBV polymerase chain reaction (PCR) in the blood showed a high value of 1.8×10⁴ copies/10⁶ white blood cells. On day 9, a flow cytometric analysis of peripheral blood mononuclear cells (PBMCs) showed that 49.0% of lymphocytes were positive for CD3 with a CD4:CD8 ratio of 1:1, 34.7% of lymphocytes were positive for CD2 with CD3-CD16+ CD56+, and 10.7% of lymphocytes were positive for CD19 with CD20 and no light chain restriction, indicating no B-cell monoclonality. Analyses of EBV genomes by Southern blotting and T-cell receptor rearrangement in PBMCs were not performed.

Figure 1.

Clinical course.

Table.

Laboratory Findings on Admission.

Peripheral blood															Cerebrospinal fluid
WBC		6,500	/µL		Alb		4.0	g/dL		Ferritin		676	ng/mL		Appearance		Clear
Seg		66.0	%		AST		17	U/L		IgG		826	mg/dL		pH		7.5
Eos		3.0	%		ALT		7	U/L		IgA		236	mg/dL		Cells		100	/µL
Baso		2.0	%		LDH		228	U/L		IgM		63	mg/dL		Lym		94	/µL
Mono		9.0	%		ALP		158	U/L							Neutro		2	/µL
Lym		20.0	%		γ-GTP		25	U/L		Anti-VCA IgM		0.7 (±)			Mono		4	/µL
RBC		446	×104/µL		T-Bil		0.5	mg/dL		Anti-VCA IgG		9.3 (+)			Protein		127	mg/dL
Hb		14.0	g/dL		BUN		16.6	mg/dL		Anti-EBNA IgG		0.3 (-)			Glucose		60	mg/dL
PLT		26.2	×104/µL		Cr		0.65	mg/dL							LDH		30	U/L
Reticulocytes		11.6	×104/µL		Na		133	mmol/L		CMV pp65 antigen		Negative
					K		4.7	mmol/L		(C7-HRP)					Culture		Negative
PT-INR		0.92			Cl		101	mmol/L
APTT		28.2	s		UA		4.4	mg/dL							HSV-DNA		Negative
Fibrinogen		438	mg/dL		Glucose		127	mg/dL							VZV-DNA		Negative
D-dimer		0.8	μg/mL		CRP		0.03	mg/dL							EBV-DNA		1.2×103	copies/mL

WBC: white blood cell count, Seg: segmented neutrophil, Eos: eosinophil, Baso: basophil, Mono: monocyte, Lym: lymphocyte, RBC: red blood cell count, Hb: hemoglobin, PLT: platelet count, PT-INR: prothrombin time-international normalized ratio, APTT: activated partial thromboplastin time, Alb: albumin, AST: aspartate transaminase, ALT: alanine transaminase, LDH: lactate dehydrogenase, ALP: alkaline phosphatase, γ-GTP: gamma glutamyl transpeptidase, T-Bil: total bilirubin, BUN: blood urea nitrogen, Cr: creatinine, CRP: C-reactive protein, IgG: immunoglobulin G, IgA: immunoglobulin A, IgM: immunoglobulin M, VCA: Epstein-Barr virus viral capsid antigen, EBNA: Epstein Barr virus nuclear antigen, CMV: cytomegalovirus, Neutro: neutrophil, HSV: herpes simplex virus, VZV: varicella zoster virus

A cerebrospinal fluid (CSF) analysis showed 100 cells/mm² (55% lymphocytes, 1% macrophages, and 44% atypical lymphocytes), elevated protein (127 mg/dL), and normal glucose (60 mg/dL). CSF PCR was positive for EBV at 1.2×10³ copies/mL and negative for varicella zoster virus (VZV) and herpes simplex virus (HSV). CSF PCR for CMV and human herpesvirus-6 was not performed. A flow cytometric analysis of CSF cells showed that 89.6% of lymphocytes were positive for CD3 with a CD4:CD8 ratio of 2:1, while 3.0% of lymphocytes were positive for CD10 with CD19, CD20, and no light-chain restriction. An analysis of immunoglobulin and T-cell receptor gene rearrangement in the CSF was not performed because of insufficient sample volume. CSF cytology showed predominantly medium-sized lymphocytes, partially with irregularly shaped nuclei, positivity for CD3 and granzyme B, and negativity for CD5, L-26, CD56, and EBV-encoded RNA on in situ hybridization (EBER-ISH) immunostaining, leading to the diagnosis of reactive T-cell proliferation in the CSF.

After admission, she complained of progressive nausea; suspecting exacerbation of gastrointestinal GVHD, the dose of PSL was increased from 3 to 5 mg/day, but there was no improvement. On day 20, endoscopy revealed two elevated pyloric portions of the stomach, which pathologically showed pyloric mucosa with inflamed fibromusculosis and a partially hyperplastic foveolar epithelium. Sparse EBER-ISH-positive cells were identified in the biopsy specimen by immunostaining, but no neoplastic changes were observed. Whole-body computed tomography (CT) showed no lymphadenopathy or hepatosplenomegaly. Based on these findings, systemic reactivation of EBV was diagnosed, including a pathologically documented EBV infection in the stomach.

On day 20, the following procedure (Fig. 2) was performed to identify the origin of the EBV-infected PBMCs as previously described (8). First, PBMCs were isolated by centrifugation. Next, CD19+, CD4+, CD8+, CD56+, and CD14+ cells were serially isolated from PBMCs using a magnetic cell separator. Third, DNA extraction was performed, and quantification of EBV-DNA was conducted using real-time PCR for each fraction. Finally, the amount of EBV-DNA in each fraction of PBMCs showed the highest DNA values in CD19+ B cells, followed by CD16+ CD56- natural killer cells. In addition, contemporary flow cytometry of PBMCs showed an increase in CD19+ and CD16+ CD56- cells. Most CD16+ cells did not express HLA-DR, indicating that activation did not occur (Fig. 3). These findings suggest that the predominant EBV-infected cell fraction in PBMCs is B cells.

Figure 2.

An identification analysis of EBV-infected peripheral blood cells fractions. (a) The amount of EBV-DNA in blood cells and plasma showed high values. (b) The amount of EBV-DNA in each fraction of peripheral blood cells showed the highest DNA values in CD19+ B cells (main population), followed by others, including CD16+ CD56- natural killer cells. This suggests that the predominant EBV-infected cell fraction is B cells. PBMC: peripheral blood mononuclear cells, w/o: without, ND: not detectable

Figure 3.

Flow cytometry of peripheral blood mononuclear cells. Flow cytometry showed increases in CD19+ cells (red circle) and CD16+ CD56- cells (solid blue circle). Most of the CD16+ cells did not express HLA-DR, indicating that activation had not occurred (dashed blue circle).

Immediately upon admission, ACV was administered because the neurological symptoms were suspected to be due to CNS involvement associated with herpes zoster reinfection. However, the patient's symptoms did not improve. Because whether or not the symptoms of TM were related to the systemic reactivation of EBV with reactive T-cell proliferation in the CNS was unclear, we preferred B-cell elimination therapy over conventional TM treatment, including high-dose corticosteroids. After receiving intravenous rituximab on day 27, the patient showed rapid improvement in somnolence and nausea, as well as gradual improvement in CSF findings and symptoms of TM. Two months later, the TM symptoms resolved completely. EBV-DNA was undetectable in the CSF on day 71 and in the blood on day 104. The patient was discharged on day 122 and has survived for four years without recurrence of neurological symptoms.

Discussion

The patient's TM symptoms were associated with reactivation of EBV, mainly in B cells, after a herpes zoster episode. A CSF analysis revealed reactive T cell proliferation. CSF PCR was positive for EBV and negative for VZV and HSV. Elimination of EBV-infected B cells with rituximab simultaneously improved TM symptoms. Based on these findings, the patient was diagnosed with TM associated with EBV reactivation. To our knowledge, this is the first report of EBV reactivation-associated TM that was successfully treated with intravenous rituximab monotherapy.

In the present case, herpes zoster may have triggered EBV reactivation. Previous studies reported that other CNS infections could trigger EBV reactivation in immunocompromised patients (9). Two case reports described VZV infections associated with EBV reactivation (10,11). However, some reports have discussed the need for careful clinical determination of whether EBV is the true cause of or merely a bystander in CNS infections, as EBV can occasionally be detected along with other CNS infections in immunocompromised patients (9,12). In the present case, VZV was not detected in the CSF, suggesting that myelitis occurred as a result of EBV reactivation triggered by herpes zoster in an immunocompromised state due to hematopoietic stem cell transplantation.

Rituximab therapy may be effective in cases of EBV-associated CNS involvement. Previous reports have described successful treatment with rituximab for CNS vasculitis-associated EBV lymphoproliferative disorder and steroid-refractory EBV-associated acute disseminated encephalomyelitis (ADEM) (13,14). It has also been reported that the CSF penetration rate of intravenous rituximab was approximately 0.1% (15), and serial administration of intravenous rituximab did not significantly increase CSF concentrations (16). The limited CNS penetration of intravenous rituximab suggests that, in the present case, rituximab did not act directly in CSF. Instead, it indirectly ameliorates TM by eliminating systemic EBV-infected B cells. This observation suggests that TM associated with EBV reactivation may occur via immunological mechanisms.

The association between EBV infection and autoimmune reactions in the CNS, particularly in the etiology of multiple sclerosis (MS), has been discussed in several studies. Various mechanisms have been considered, including the involvement of cellular and humoral immunity in the cross-reactivity between EBV and autoantigens, immortalization of autoreactive B-cell clones by EBV infection, and induction of autoreactive cytotoxic T cells in the CNS to counteract the direct invasion of the CNS by EBV (4,5). Indeed, previous reports have shown that B-cell depletion therapy with rituximab reduces inflammatory brain lesions and the rate of clinical relapse in patients with refractory MS (5). In the present case, EBV DNA was elevated in the CSF, but no EBER-ISH-positive or lymphoma cells were identified, suggesting an increase in reactive T cells. However, it was difficult to determine whether these T cells were induced by an immune response to direct EBV invasion in the CNS or by an autoimmune response triggered by EBV-infected B cells systemically in the body. Both hypotheses suggest that rituximab reduces EBV-infected B cells that induce autoreactive T cells or reduces the EBV viral load by depleting B cells, resulting in the clinical efficacy of rituximab in this case.

In the present case, differential diagnoses for TM other than parainfectious causes included inflammatory demyelinating diseases, such as ADEM, MS, and neuromyelitis optica spectrum disorder (NMOSD), and collagen diseases, such as systemic lupus erythematosus and Sjögren's syndrome. The patient had mild somnolence but no meningism on normal head MRI, suggesting that it was not ADEM. Although anti-aquaporin 4 antibody and oligoclonal bands in the CSF were not measured in this case, the clinical course was monophasic, ruling out MS and NMOSD, which typically have polyphasic courses. The patient also had no history of any collagen disease. Since the efficacy of rituximab against recurrent TM has been previously reported (17), the efficacy of rituximab against TM in this case does not definitively prove an association between TM and EBV reactivation. However, based on the close temporal relationship between treatment and effectiveness, rituximab monotherapy against EBV reactivation likely improved TM in this case, supporting the autoimmune hypothesis of TM. This finding may help clarify the pathogenesis of TM in the future. Clarifying this point will require the further accumulation of cases.

In conclusion, we successfully treated a case of EBV reactivation-associated TM with intravenous rituximab following herpes zoster infection in a cord blood transplant recipient. The most notable aspect of this case was the unique clinical course, in which TM improved with rituximab alone, without conventional treatment for TM, such as high-dose steroids. This suggests that EBV reactivation may induce TM via immunological mechanisms.

The authors state that they have no Conflict of Interest (COI).