Autoimmune pancytopenia occurring late after simultaneous pancreas and kidney transplantation

Abstract

A 35-year-old woman presented with a widespread petechial rash and pancytopenia. She underwent simultaneous pancreas and kidney transplantation for type 1 diabetes 8 years previously followed by a renal transplant 1 year prior to presentation, and was taking tacrolimus as long-term immunosuppression. The full blood count showed haemoglobin 97 g/L, platelet count 2×10⁹/L and neutrophil count 0.22×10⁹/L. Peripheral blood film examination confirmed genuine thrombocytopenia in the absence of any haemolytic or malignant features. Serological testing identified autoantibodies against all three blood lineages, consistent with a diagnosis of autoimmune pancytopenia. Treatment with steroids, intravenous immunoglobulins, romiplostim and mycophenolate mofetil achieved only fleeting remissions. Blood counts eventually normalised following the administration of rituximab and a change from tacrolimus to ciclosporin immunosuppression. Cytopenias are a well-recognised complication of post-transplantation care but we believe this to be the first reported case of autoimmune pancytopenia following solid organ transplantation. In this case report, we discuss the approach to investigation of haematological abnormalities post-transplant and the rationale for, and outcome of, the management of this rare case.

Background

Single and multi-lineage cytopenias are a common complication of solid organ transplant (SOT) occurring in 20%–60% of patients undergoing kidney transplantation.^{1 2} These haematological abnormalities can lead to life-threatening anaemia, bleeding and infection which results in major morbidity and mortality.³ They can occur at any time post-transplant and are challenging to differentiate in patients who often have multiple potential culprits coexisting. This is exemplified by the attempts to document the frequency of tacrolimus-induced anaemia, which vary from 1% to 47% of patients.⁴ Investigation of these cases requires a thorough work-up to ensure uncommon conditions are identified and treated promptly.

We present an unusual case of late-onset autoimmune pancytopenia occurring after kidney and pancreas transplant, but which also serves as a reminder of how to approach the differentiation of cytopenias post-SOT.

Case presentation

A 35-year-old woman presented to the emergency department with a short history of purpura. She reported melaena and heavy menstrual bleeding over the preceding days. Two to three weeks earlier, she developed shingles and received a course of oral acyclovir.

Her medical history was significant for a simultaneous pancreas and kidney transplant, with alemtuzumab conditioning, performed 8 years earlier for type 1 diabetes mellitus with renal failure. One year prior to her current presentation, she had received a second kidney transplant, again with alemtuzumab conditioning, following failure of the original renal allograft.

Her long-standing regular medications were tacrolimus, aspirin, alfacalcidol, ranitidine, simvastatin, folic acid and rifaximin (for small bowel bacterial overgrowth).

Physical examination confirmed the presence of a petechial, non-blanching rash covering the extensor and flexor surfaces of both legs and extending up to the buttocks. Abdominal examination revealed a non-tender, palpable renal graft in the right iliac fossa. There was no fever. Cardiovascular and respiratory examinations were normal. Skin and musculoskeletal examinations were normal with no arthritis, rash or serositis.

Investigations

A full blood count performed 16 days prior to admission was unremarkable (haemoglobin 113 g/L, platelets 190×10⁹/L, neutrophils 2.91×10⁹/L). On presentation she had developed pancytopenia (table 1).

Table 1.

Results of initial investigations

Metric	Result	Local laboratory normal range
Haemoglobin	97	120–156 g/L
Mean cell volume	85	80–99 fL
Platelet count	2	150–370×109/L
White cell count	1.2	3.9–10.2×109/L
White cell differential:
Neutrophil	0.22	1.5–7.7×109/L
Lymphocyte	0.82	1.5–4.5
Basophil	0.01	0–0.1
Eosinophil	0.01	0–0.4
Monocyte	0.08	0.2–0.8
Reticulocyte count	93	25–105×109/L
Haptoglobin	1.7	0.5–2.6 g/L
Prothrombin time	12.9	10.6–12.8 s
Activate partial thromboplastin time	26.3	26.7–35.3 s
D-dimer	295	0–230 ng/mL
Bilirubin	14	0–20 µmol/L
Lactate dehydrogenase	160	120–246 U/L
Creatinine	110	44–97 µmol/L
Folate	>24	>5.38 µg/L
Vitamin B12	289	211–911 ng/L
Ferritin	292	10–291 µg/L
Erythropoietin	101	4–20 IU/L

The direct antiglobulin test (DAT) was positive for monospecific immunoglobulin G (2+) and negative for C3d. Antibody identification confirmed the presence of a warm autoantibody-lacking specificity. Haematinics and biochemical parameters of red cell haemolysis were normal, prompting investigation for alternative causes of anaemia.

Serological testing identified human leucocyte antigen class I, GIIb/IIIa-specific platelet antibodies and granulocyte-specific antibodies, which lacked human neutrophil antigen specificity. One and 24-hour platelet increments showed no response to platelet transfusion. A low-level paroxysmal nocturnal haemoglobinuria (PNH) clone (1.5% of neutrophils) was detectable by immunophenotyping.

Serial bone marrow biopsies were performed. Morphological examination demonstrated a hypercellular marrow with increased megakaryocyte numbers, severe erythroid hypoplasia and severe myeloid hyperplasia with maturation arrest (figure 1). Haemophagocytosis was noted on one occasion but this occurred in the absence of clinical haemophagocytic syndrome. Dysplasia was not present. Immunophenotyping and cytogenetic analysis was normal. Peripheral blood and bone marrow chimerism was 0% donor from the recent renal transplant.

Figure 1.

Bone marrow examination at diagnosis. (A) Aspirate prepared with modified Wright's stain, demonstrates granulocyte maturation arrest, predominantly at the promyelocyte stage with absence of mature neutrophils. (B) Trephine prepared with H&E (×20) demonstrates marrow hypercellularity. (C) Trephine with CD68P immunostaining highlights macrophage activation with haemophagocytosis. (D) Trephine with glycophorin A immunostaining demonstrates severe erythroid hypoplasia.

Tests for all infections listed in box 1 were negative.

Box 1. Differential diagnosis.

Infection

• For example, CMV, EBV, parvovirus B19, HHV6, HIV, hepatitis B and C, leishmaniosis.

Drugs

• For example, tacrolimus, ciclosporin, alemtuzumab, ATG, valganciclovir, ganciclovir, co-trimoxazole.

Immune dysregulation

• Autoimmune cytopenia, for example, ITP.

• MAS, HLH.

• PTLD (may be EBV-driven).

Haematinic deficiency

• Vitamin B₁₂, folate.

Primary haematological conditions

• Leukaemia, lymphoma, aplastic anaemia, myelodysplasia.

Bone marrow infiltration

• Solid organ malignancy, for example, prostate, breast.

Alloimmune processes

• Graft versus host disease.

• Alloimmune haemolytic anaemia, for example, passenger lymphocyte syndrome.

Thrombotic microangiopathies

• For example, TTP, HUS.

• ATG, antithymocyte globulin; CMV, cytomegalovirus; EBV, Epstein-Barr virus; HLH, haemophagocytic lymphohistiocytosis; HHV6, human herpes virus 6; HUS, haemolytic uraemic syndrome; ITP, immune thrombocytopenic purpura; MAS, macrophage activation syndrome; PTLD, post-transplant lymphoproliferative disorder; TTP, thrombotic thrombocytopenic purpura.

Differential diagnosis

Our diagnosis of autoimmune pancytopenia was made after the exclusion of the differentials listed in box 1.⁴ Positive evidence for the diagnosis was provided by the demonstration of autoantibodies against all three blood cell lineages using serological tests and the presence of a PNH clone.

Treatment

Prednisolone at a dose of 1 mg/kg/day was started and three units of platelets were transfused. Granulocyte colony-stimulating factor (GCSF) was started in view of the neutropenia. After 48 hours there was no response in the platelet count (3×10⁹/L) to steroids or transfusion and so intravenous immunoglobulin 1 g/kg was administered (figure 2).

Figure 2.

Weekly blood counts since presentation accompanied by key interventions. IVIg, intravenous immunoglobulin; MMF, mycophenolate mofetil.

Three weeks later the platelet count remained unresponsive and the patient started the thrombopoietin agonist romiplostim. The platelet count rose to 133×10⁹/L after three doses.

The response was short-lived and after 2 weeks she once again became pancytopenic (haemoglobin 72 g/L, platelets 6×10⁹/L, neutrophil 0.99×10⁹/L), this despite GCSF 300 µg/kg daily and also romiplostim 10 µg/kg weekly.

At this point she became transiently reticulocytopenic and received 15 units of packed red cells over a 3-month period. A repeat bone marrow biopsy demonstrated red cell aplasia alongside myeloid hyperplasia with granulocyte maturation arrest.

Eight weeks after her initial presentation, mycophenolate mofetil (MMF) 500 mg two times per day was commenced as a steroid-sparing immunosuppressant, in addition to her long-term tacrolimus.

A fleeting partial remission was obtained following the initiation of MMF (haemoglobin 102 g/L, platelets 174×10⁹/L, neutrophils 0.84×10⁹/L). A fortnight later the platelet count fell to 27×10⁹/L and rituximab was added to her regimen at a dose of 375 mg/kg weekly for 4 weeks. Her transplant immunosuppressant was changed from tacrolimus to ciclosporin. In view of the persistent neutropenia, antimicrobial prophylaxis was started (posaconazole and nebulised pentamidine).

A second remission was obtained 6 weeks after the fourth dose of rituximab (haemoglobin 92 g/L, platelets 123×10⁹/L, neutrophils 3.2×10⁹/L). She returned to work for 7 weeks before abruptly relapsing for a second time (haemoglobin 115 g/L, platelets 54×10⁹/L, neutrophil 0.28×10⁹/L). Romiplostim was restarted and a further four doses of rituximab were administered.

Outcome and follow-up

After 9 months she achieved her third remission (haemoglobin 126 g/L, platelets 248×10⁹/L, neutrophils 4.0×10⁹/L), which persists at the time of writing. Repeat serological testing found a negative DAT. Platelet and granulocyte antibodies were no longer detectable.

Treatment was complicated by superficial thrombophlebitis, which was managed with short courses of therapeutic-dose low molecular weight heparin followed by prophylactic dose rivaroxaban for symptomatic relief.

While thrombocytopenic she developed retinal haemorrhages and macular oedema. Both are recognised as uncommon adverse effects of romiplostim.⁵ She received platelet transfusions for the prevention of further retinal bleeding, totalling 24 units for over 6 months.

Discussion

To our knowledge this is the first reported case of autoimmune pancytopenia occurring after SOT. Review of the literature identified only two related cases. A 13-year-old boy with pancytopenia whose T-lymphocytes were found to inhibit the growth of autologous myeloid progenitors when tested in vitro, and a more recent case of autoimmune pancytopenia in a patient after immunochemotherapy for lymphoma.^{6 7}

The timing of the shingles and acyclovir treatment is noteworthy as both have been identified as potential triggers of autoimmune disease.^{8 9} However the infection resolved swiftly on standard doses of acyclovir and so exposure to both was transient.

The high incidence of PNH clones in diseases such as aplastic anaemia is thought to be the result of selection pressure from the immune process, and so represents an attempt by blood cells to escape the offending autoantibody.¹⁰ Although in this case the low-level clone did not translate into haemolysis, its presence lends additional weight to our diagnosis.

We believe our patient’s case was B-cell mediated. This is supported by the triple positive immune serology and clinical response to rituximab that followed. Immune thrombocytopenia (ITP) occurs in 2% of patients receiving alemtuzumab and these cases remit with rituximab therapy.¹¹

With this in mind we relied on approaches used for the treatment of ITP.¹² Tacrolimus is known to be associated with the onset of ITP, and cessation of the drug has been shown to induce remission.^{13 14} Despite this well-documented association, the mechanism is not fully understood and, notably, attempts to correlate tacrolimus use with haematopoietic maturation arrest of the sort seen in this case have failed.^{1 15}

Haemolysis was not present and the patient’s anaemia derived from erythroid hypoplasia in the bone marrow. It has long been posited that autoantibodies directed at erythroid precursors may be responsible for autoimmune anaemias in the absence of peripheral haemolysis.¹⁶

Patient’s perspective.

I am no stranger to complex medical interventions, having undergone two previous transplants. However, I found renal failure an easier process to get a handle on, as I was able to map my progress with a known end point. By comparison, this illness arrived out of the blue, with no rhyme or reason, and since the start it has involved unexpected ups and downs.

During my temporary remission, I was able to return to work and found it even harder to tackle the relapse, which was a knock to my confidence. The steroids negatively affected my mood and cognitive function. I feel that many people do not appreciate the detrimental effects on a person’s ability to think and feel while on high doses of steroids.

These difficult experiences were countered by my supportive family, social and work environments. I enjoy my independence and felt uncomfortable having to rely so heavily on family, while at the same time feeling fortunate to have them. The haematology day unit were able to provide the comfort and reassurance of knowing a team that I was able to form a connection with.

Learning points.

• Extremely rare condition which responded to multi-modal therapy.

• Prompt investigation and treatment is required to prevent major morbidity and mortality.

• The treatment of immune-mediated cytopenias requires a multi-pronged approach to (a) cease production of the offending antibody, (b) stimulate bone marrow cell production and (c) provide adequate supportive care.