SARS‐CoV‐2‐associated warm autoimmune haemolytic anaemia in an immunosuppressed patient: The trend continues

Dear Editor,

Since the onset of the coronavirus disease 2019 (COVID‐19) pandemic, an increasing number of reports of haematologic sequelae secondary to severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection are being described. ¹ , ² Among these is autoimmune haemolytic anaemia (AIHA), particularly in patients with a predilection to this condition, such as those with autoimmune or lymphoproliferative disorders. ³ However, AIHA in the setting of COVID‐19 does not appear restricted to only ‘high‐risk’ individuals, as several case reports have described severe AIHA in patients without underlying conditions, ⁴ , ⁵ supporting the theory that SARS‐CoV‐2 itself may be a stimulus for immune dysregulation and subsequent autoimmune haemolysis. ⁶ However, the mechanisms that induce AIHA in the absence of inflammation remain unclear, though may be related to molecular mimicry between SARS‐CoV‐2 antigens and red blood cell (RBC) membrane antigens. ⁷ Furthermore, the development of AIHA in the setting of immunosuppression wherein autoantibody formation should theoretically be limited has not, to our knowledge, been described in patients with COVID‐19; thus, the risk for AIHA in these individuals is unknown.

A 59‐year‐old female with a 15‐year history of well‐controlled mixed connective tissue disease (MCTD) [manifesting predominantly as myositis and Raynaud's syndrome, managed with chronic immunosuppressive therapy including azathioprine (100 mg daily), hydroxychloroquine (200 mg daily), and intermittent prednisone (4 mg daily)] presented to an outpatient clinic complaining of fever and fatigue. She was diagnosed with COVID‐19 with detection of SARS‐CoV‐2 via reverse‐transcriptase polymerase chain reaction, but denied significant dyspnea, was not hypoxic, and did not require hospitalisation or treatment. She had received four doses of a SARS‐CoV‐2 mRNA vaccine (manufacturer not known), with the fourth dose (second booster dose) administered 2 weeks prior to testing positive for SARS‐CoV‐2. One week after testing positive, her fever had resolved, but she endorsed persistent, worsening fatigue. She also developed new‐onset palpitations and dyspnea on exertion. At a rheumatology appointment 2 weeks later, routine laboratory evaluation revealed a haemoglobin of 5.4 g/dl (baseline 11–11.5 g/dl) and she was admitted to the hospital for further evaluation.

A blood sample was sent for type and screen and the patient's ABO type was determined to be A RhD‐positive. An antibody screen using three reagent RBCs via gel‐based methodology demonstrated panreactivity at the anti‐human globulin (AHG) phase. The patient's plasma was tested against a panel of 11 reagent RBCs, which again showed panreactivity. An auto‐control using standard tube methods was reactive. A direct antiglobulin test (DAT) using a poly‐specific anti‐IgG, ‐C3d reagent via standard tube methodology was reactive (3+). Testing using a monospecific anti‐IgG reagent was reactive (3+), while a monospecific anti‐C3 reagent was non‐reactive. An acid glycine elution was performed and tested against a panel of 11 reagent cells at AHG phase, which demonstrated panreactivity. The patient's blood sample was referred to an immunohematology reference laboratory for autoadsorptions and elutions, which revealed no underlying alloantibody in the patient's plasma at multiple phases of testing, including in saline at immediate spin and room temperature, as well as in low ionic strength saline at 37°C and AHG.

Further laboratory evaluation (Table 1) revealed mild leukopenia with a normal absolute neutrophil count, absolute lymphopenia, normal platelet, and elevated reticulocyte percentage. Haemolysis biomarkers were abnormal, including elevated lactate dehydrogenase and undetectable haptoglobin There was no evidence of an active flare of the patient's underlying MCTD, as her creatine kinase, erythrocyte sedimentation rate, and C‐reactive protein were normal. Flow cytometric analysis demonstrated no evidence of a clonal lymphoproliferative disorder but did reveal an absolute deficiency of CD19+ B cells (3.6% of lymphocytes; reference: 4.0%–26.0%).

TABLE 1.

Patient laboratory results on admission

Test	Patient result	Reference range
Haemoglobin	5.4 g/dl	11.7–15.5 g/dl
White blood cell count	3.1 × 103/μl	4.0–11.0 × 103/μl
Absolute neutrophil count	2.68 × 103/μl	2.0–7.6 × 103/μl
Absolute lymphocyte count	0.14 × 103/μl	0.6–3.7 × 103/μl
Platelet count	268 × 103/μl	150–420 × 103/μl
Reticulocyte percentage	9.6%	0.6%–2.7%
Lactate dehydrogenase	496 U/L	122–241 U/L
Haptoglobin	<10 mg/dl	30–200 mg/dl
Creatine kinase	176 U/L	11–204 U/L
Erythrocyte sedimentation rate	8 mm/h	0–20 mm/h
C‐reactive protein	1.1 mg/L	1.0–3.0 mg/L

The patient had no personal or family history of autoimmune cytopenias or other haematologic conditions, had never received a transfusion, and had a negative DAT 1 year prior. She was initiated on prednisone 60 mg daily and received three units of A Rh‐D positive RBCs, with rituximab therapy planned.

This case illustrates the potential for SARS‐CoV‐2 to induce severe AIHA, even in immunosuppressed individuals with mild COVID‐19 and no evidence of hyperinflammation. A number of prior cases have shown that patients with an underlying predisposition to AIHA are at increased risk following SARS‐CoV‐2 infection, and our group recently demonstrated that SARS‐CoV‐2 may be an independent risk factor for AIHA. ³ However, while our patient may have theoretically been more prone to additional autoimmune conditions given her history of MCTD, ⁸ her immunosuppressed state, with objective evidence of B cell suppression, in conjunction with absence of acute inflammation refutes the hypothesis that immunological overstimulation is a requirement for autoreactivity in COVID‐19.

In general, the development of AIHA and other autoimmune haematologic conditions in immunosuppressed individuals is not well characterised or understood. However, it has previously been shown that AIHA may occur in the setting of immunosuppression for solid organ transplantation, which is thought to be due to the selective reduction in T cells, resulting in a skewed humoral response, which, in conjunction with a depletion of regulatory T cells, permits disinhibition of autoreactive B cells, allowing production of anti‐erythrocyte antibodies. ⁹ This may partially explain the predisposition to AIHA in our patient, as azathioprine ¹⁰ and hydroxychloroquine ¹¹ both exert suppressive effects on cytotoxic and regulatory T cells, potentially allowing for a dysregulated humoral response. However, other authors have shown that hydroxychloroquine in particular is capable of suppressing B cell differentiation and IgG production, ¹² a finding supported by the absolute deficiency of CD19+ B cells in our patient.

While our patient had absolute B‐cell lymphopenia by flow cytometry, we recognise that tissue‐resident B cells are not accounted for in flow cytometric analysis of peripheral blood. ¹³ Thus, immunoglobulin studies would have been useful to further assess the degree of humoral immunosuppression in this patient; however, further studies were unavailable.

Given this unusual presentation of severe, IgG‐mediated haemolytic anaemia in a patient despite both cellular and humoral immunosuppression, additional studies are needed to elucidate the potential mechanism(s) by which SARS‐CoV‐2 elicits autoimmune haemolysis.

AUTHOR CONTRIBUTIONS

Jeremy W. Jacobs wrote the first draft and approved the final version. Savanah D. Gisriel revised the manuscript and approved the final version.

CONFLICT OF INTEREST

The authors declare no conflict of interest.