A family with cytotoxic T-lymphocyte-associated protein 4 haploinsufficiency presenting with aplastic anaemia

Background

Today, whole-exome sequencing is easily accessible, and more and more diseases are linked with different genetic variants. This is especially true for autoimmune diseases. Aplastic anaemia is a rare disease, and usually presents sporadically without any family history. We report, three family members all diagnosed with aplastic anaemia, which led to the investigation of an underlying genetic cause. To the best of our knowledge, only one patient with aplastic anaemia and CTLA4 haploinsufficiency has been treated with allogenic stem cell transplantation.1 The three family members had very different presentations and response to treatment, which we wish to highlight with this case report. They were initially treated with standard immunosuppressive therapy for aplastic anaemia, and after the CTLA4 haploinsufficiency was unravelled, we changed treatment to sirolimus and abatacept in two of the patients with favourable response in one.

Case presentation

Our index patient (A.III.1), an identical male twin, came to medical attention aged fifteen due to severe pancytopenia, and was admitted to hospital due to epistaxis and fatigue. A diagnosis of severe aplastic anaemia was made. The patient was referred to allogenic stem cell transplantation with his monozygotic twin (A.III.2) as donor. Bone marrow studies of A.III.2 showed a hypocellular bone marrow with hypoplastic erythropoiesis and almost absent thrombopoiesis, which excluded him as donor. Fanconi anaemia was excluded in both twins. Their parents were non-consanguineous.

The twins’ father (A.II.1) was diagnosed with autoimmune haemolytic anaemia 40 years old. He also had signs of liver disease diagnosed as primary sclerosing cholangitis and pulmonary infiltrates of unknown aetiology. In retrospect the pulmonary infiltrations may have been granulomatous-lymphocytic interstitial lung disease now recognised as a manifestation of CTLA4 haploinsufficiency. The haemolytic anaemia and pulmonary infiltrates responded to treatment with corticosteroids. He later developed immune mediated thrombocytopaenia. By the age of 60 years he became transfusion dependent and was diagnosed with severe aplastic anaemia. A case report of the twins’ father, before he was diagnosed with aplastic anaemia, has been published previously.2

Investigations

After the third family member was diagnosed with aplastic anaemia, whole-exome sequencing was performed and revealed a heterozygous variant in CTLA4 (NM_005214.4(CTLA4): c.567+6T>G) in all three. CTLA4 codes for cytotoxic T-lymphocyte associated protein 4, and mRNA analysis of peripheral blood monocytic cells showed that this variant results in exclusion of exon 3 in the CTLA4 mRNA, leading to frameshift. Seventy-two amino acids in the C-terminal end are replaced by 21 amino acids with a different sequence. The result of this variant is loss of function in CTLA4, and it is the likely pathogenic cause of disease in this family.

Treatment

A.III.1: After he was diagnosed with aplastic anaemia, he started treatment with antithymocyte globulin (ATG) combined with cyclosporine. He had an initial response to this treatment, but pancytopenia recurred, and he was given a second course of ATG and continuation of cyclosporine with a sustained good partial response. However, he suffered from Molluscum contagiosum and severe Verruca vulgaris. Several attempts to taper cyclosporine were unsuccessful, and addition of eltrombopag did not make a difference.

When the CTLA4 haploinsufficiency was diagnosed, treatment was changed from cyclosporine to sirolimus and thereafter adding abatacept. There was no response to this treatment. His bone marrow failure gradually worsened with high transfusion demands, and he suffered from several serious infections including Herpes simplex viraemia, pneumonia with Aspergillus fumigatus, sepsis with Corynebacterium jeikeium and Staphylococcus haemolyticus.

While being treated in the intensive care unit for respiratory failure due to pneumonia, he underwent allogenic stem cell transplantation with a matched unrelated donor. Immunosuppression after transplantation was sirolimus. The initial period after transplantation was complicated by Epstein-Barr virus reactivation, Cytomegalovirus reactivation, recurrent infections and acute graft versus host disease (GvHD), all responding to standard treatment.

However, he had persistent cytopenia, and graft failure due to donor specific antibodies was suspected.3 4 Treatment with rituximab and bortezomib was unsuccessful.5 Due to poor graft function, we decided to give him a CD34 +stem cell boost 7,5 months after the initial transplant.

There was concern that he would not tolerate a flare up of GvHD, and he was given a single dose of cyclophosphamide 50 mg/kg on day +3 after the stem cell boost. On day +7, he developed septicaemia and multiorgan failure from which he succumbed.

A.III.2: On diagnosis, he had non-severe aplastic anaemia and was initially observed without treatment. After a few months of observation, he started treatment with ATG and cyclosporine due to increasing pancytopenia, with favourable response. He has been cyclosporine dependant for more than 17 years without any major complications.

A.II.1: When aplastic anaemia was diagnosed, ATG combined with cyclosporine was initiated. The response was poor, and subsequent addition of eltrombopag did not make a difference. When CTLA4 haploinsufficiency was unravelled, cyclosporine and eltrombopag was substituted for sirolimus and abatacept. He became transfusion independent, and venesection to relieve transfusion related haemosiderosis could be undertaken. After 1 year, he suddenly died of septicaemia.

Outcome and follow-up

Both twins became fathers, and presymptomatic testing of their children showed that A.IV.2 had inherited the CTLA4 variant, while A.IV.1 had not. The child with the CTLA4 variant is currently healthy. A.II.2 had three more children (A.IV.3, A.IV.4 and A.IV.5) who are currently not yet tested for the CTLA4 variant. A.IV.3 died within less than 24 hours of birth of unknown cause.

We also did an inquiry about other family members and medical history. There was no other family member with symptoms or medical history indicating CTLA4 haploinsufficiency. The variant has not been detected in other family members who have been tested (A.II.1, A.II.3 and A.II.4). Of note is that the twins’ father had an aunt who was diagnosed with aplastic anaemia at the age of 80 years. She received transfusions for about 1.5 years before she experienced a spontaneous remission and died 91 years old (figure 1).

Figure 1

Pedigree family A with CTLA4 haploinsufficiency. Squares: male subject; circles; female subjects; black filled symbols: subjects with aplastic anaemia and CTLA4 variant; black circle: CTLA4 variant carrier without disease; crossed-out symbols; deceased subjects. The figure is created by the author TSS for this case presentation, and has not been used before.