• Users Online: 239
  • Print this page
  • Email this page

 Table of Contents  
Year : 2022  |  Volume : 12  |  Issue : 4  |  Page : 183-187

Cold agglutinin autoimmune haemolytic anaemia as an initial presentation of diffuse large B cell lymphoma: A case study

1 Department of Blood Transfusion Medicine, Sir Ganga Ram Hospital, New Delhi, India
2 Department of Clinical Hematology, Sir Ganga Ram Hospital, New Delhi, India
3 Department of Research, Sir Ganga Ram Hospital, New Delhi, India

Date of Submission27-Nov-2021
Date of Decision20-Apr-2022
Date of Acceptance19-Jul-2022
Date of Web Publication30-Aug-2022

Correspondence Address:
Dr. Rashmi Rana
Department of Research, Sir Ganga Ram Hospital, New Delhi -110060
Login to access the Email id

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/cmrp.cmrp_115_21

Rights and Permissions

Cold agglutinin disease, commonly affecting females in the their seventh decade of life, is otherwise a rare entity and has an incidence of one case per million people per year. Cold reactive antibody is associated with autoimmune haemolytic anaemia (AIHA) in approximately 20% of the cases. Although the occurrence of AIHA in patients with non-Hodgkin's lymphomas is well known, aggressive lymphomas such as diffuse large B cell lymphoma (DLBCL) initially presenting as cold agglutinin AIHA is extremely rare. Here, we describe a case of DLBCL presenting few months before the diagnosis as cold antibody-AIHA.xs

Keywords: Autoimmune haemolytic anaemia, B cell lymphoma, cold agglutinin disease

How to cite this article:
Ranjan V, Dhingra G, Gupta N, Khillan K, Rana R. Cold agglutinin autoimmune haemolytic anaemia as an initial presentation of diffuse large B cell lymphoma: A case study. Curr Med Res Pract 2022;12:183-7

How to cite this URL:
Ranjan V, Dhingra G, Gupta N, Khillan K, Rana R. Cold agglutinin autoimmune haemolytic anaemia as an initial presentation of diffuse large B cell lymphoma: A case study. Curr Med Res Pract [serial online] 2022 [cited 2022 Sep 27];12:183-7. Available from: http://www.cmrpjournal.org/text.asp?2022/12/4/183/355198

  Introduction Top

Binding of auto–antibodies optimally to RBC at a specified temperature is one of the ways by which AIHA is classified. Warm autoantibody AIHA is associated with haemolysis mediated by antibodies binding to Red blood cells (RBCs) at 37°C (98.6°F). It constitutes about 80%–90% of adult cases of AIHA.[1],[2],[3]

Cold antibody (CA) AIHA is a rare type of autoimmune haemolytic anaemia (AIHA) with an incidence of 1 per million populations and cold-reactive antibodies optimally exhibiting affinity for RBCs at temperatures below 37°C.[4] Most cold agglutinins in CA-AIHA are specific for antigens of the I/i/Pr system, are immunoglobulin M (IgM) type and have an optimal RBC binding temperature between 0°C and 4°C. Pentameric or hexameric structure of IgM antibody results in its high efficacy for activating the complement system. C3b fragment resulting from the cleavage of C3 complement deposits on erythrocyte surface and binds to the complement receptors on phagocytes leading to erythrocyte destruction in the liver. Haemolysis is not necessarily severe. A positive direct antiglobulin test (DAT) for C3d, a key diagnostic feature of computer aided diagnosis (CAD) and is detectable after the attached C3b on the surviving RBC is cleaved to C3d fragment.[5],[6],[7],[8] CAD has traditionally been classified into a primary and secondary type, with latter most commonly associated with accompanying infection (such as Epstein-Barr virus and mycoplasma pneumoniae) and malignant disease, most often malignant lymphoma. However, the primary type has also been found to be frequently associated with an underlying lymphoproliferative disease in the bone marrow.[9] Váróczy et al. reported that of 421 non-Hodgkin's lymphoma (NHL) patients, 7.6% exhibited an autoimmune disease and only one patient exhibited AIHA.[10] Aggressive lymphomas such as diffuse large B cell lymphoma (DLBCL) initially presenting as CA-AIHA are extremely rare with only few cases reported in literature. Here, we report one such case where the patient presented with CA-AIHA 3 months before the diagnosis of DLBCL.

  Case Report Top

A 55-year-old male presented in June 2015 to his local physician with complaints of weakness and easy fatigability for 1 month. There was no history of fever, bleeding and weight loss. Complete haemogram showed haemoglobin 6.4 g/dl, white blood cell (WBC) count 8100/μl with normal differential count and reticulocyte count 3.5%. Peripheral smear showed polychromasia, spherocytes and occasional nucleated RBCs (nRBCs). Biochemical investigations showed indirect hyperbilirubinaemia (total 3.85 mg/dl, indirect 2.41 mg/dl) and normal liver enzymes. DAT was positive. Bone marrow examination showed erythroid hyperplasia. There was no history of previous blood transfusion. Prednisolone was started, but there was no improvement in haemolytic anaemia even after 1 month of steroid therapy. At the same time, he noticed progressive swelling of the face and neck more on the right side along with watering from right eye which was attributed to steroid side effect. He then presented to our hospital with above complaints. On examination, the patient had pallor, mild icterus and there were firm immobile non-tender swelling of right submandibular region and parotid region suggestive of lymphadenopathy. The patient also had right periorbital puffiness with continuous watering from right eye. His complete haemogram showed severe anaemia (Hb 5.2 g/dl) and peripheral smear revealed marked red cell agglutination, 15 nRBC/100 WBC, polychromasia and spherocytes. Reticulocyte count was raised to 10.6% (corrected 3.6%.). Biochemical investigation showed indirect hyperbilirubinaemia (total 3.6 mg/dl, indirect 2.6 mg/dl) and increased lactate dehydrogenase 398 U/L (normal range 90–180 U/L). A blood sample and request was received in hospitals transfusion laboratory for DAT and indirect antiglobulin test (IAT) to find out the presence of auto- and/or allo-antibodies. Simultaneously, a request of two units of packed RBC for cross match was also received on urgent basis. Due to difficulty in finding the confirmatory blood group of patients two units of 'O' Rh (D) negative best matched units were provided on emergency basis. An initial blood grouping showed forward blood group as AB and revere blood group as O. However, later on, his blood group was reported as A Rh (D) positive after resolving the blood group discrepancy by washing the patient's RBCs with warm (37°C) normal saline, treatment of IgM coated patients RBCs with dithiothriol (DTT) and auto-adsorption of patient's serum [Table 1]. Patients red cells were subjected to polyspecific and monospecific DAT with autocontrol through column agglutination gel technology (BioRad, Cressier s/Morat, Switzerland) using Liss–Coombs cards and DC-screening immunoglobulin G (IgG), IgM, immunoglobulin A (IgA), C3c, C3d, Ctl card, respectively. Patients sera were subjected to antibody screening IAT through gel technology using the reagent three-cell panel (BioRad). The patient was found positive for DAT (polyspecific [4+]) with positive autocontrol (3+) and negative for IAT. A positive reaction with IgM and C3d was observed in monospecific DAT evaluation. Subsequent blood test for thermal amplitude and cold agglutinin titre revealed a thermal range of ≥30°C and a titre of 1:1024 obtained after incubating the O cells with the patient's serum at 4°C, RT and 30°C for 20 min.
Table 1: Cell and serum grouping reaction pattern

Click here to view

Computed tomography (CT) of para-nasal sinuses and neck was done which showed enhancing soft tissue lesion in right ethmoid, maxillary and nasal cavity. There were enlarged LN in right pre-vertebral, upper and middle deep cervical and submandibular region [Figure 1] and [Figure 2].
Figure 1: CT PNS showing enhancing soft tissue lesion in right ethmoid, maxillary and nasal cavity. CT: Computed tomography, PNS: Para-nasal sinus

Click here to view
Figure 2: CT neck showing enlarged LN in right upper and middle deep cervical and submandibular region. CT: Computed tomography, LN: Lymph node

Click here to view

Trucut biopsy from cervical node showed sheets of intermediate to large lymphoid cells with large areas of necrosis. The cells had large hyperchromatic nuclei and scanty cytoplasm. Immunohistochemistry (IHC) revealed that these large cells were CD 20 positive and negative for CD 3 and CD5 [Figure 3] and [Figure 4]. Ki-67 labelling index was 70–80%. Further IHC showed positivity for MUM1 and negative for BCL 6 and CD10 suggestive of non-germinal center phenotype DLBCL shown in [Figure 5] and [Figure 6]. Positron emission tomography (PET) CT showed uptake in right cervical and mandibular, bilateral axillary and para-aortic lymph nodes [Figure 7]. Bone marrow examination revealed erythroid hyperplasia. Hence, the diagnosis of DLBCL, stage III with cold autoimmune haemolytic anaemia was made. The patient was transfused 2 units of 'O' Rh (D) negative best matched PRBC through inline warmer without any complication. The patient was then treated with 6 cycles of RCHOP chemotherapy (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisolone) with rapid clinical improvement. A total of 6 cycles of RCHOP chemotherapy was given which was completed by January 2016. At the end of treatment, PET-CT revealed complete remission and the patient continued to be in clinical remission after 4 years of completion of treatment.
Figure 3: CD 20 Positive

Click here to view
Figure 4: CD3 Negative

Click here to view
Figure 5: Positive MUM1

Click here to view
Figure 6: CD 10 Negative

Click here to view
Figure 7: PET-CT at the time of diagnosis showing increased uptake in right cervical and mandibular, bilateral axillary and para-aortic lymph nodes. PET: Positron emission tomography, CT: Computed tomography

Click here to view

  Discussion Top

Most common type of lymphoma, accounting for 30% of all NHLs is DLBCL. Bone marrow involvement or blood losses due to gastrointestinal involvement are the two most common reasons for anaemia associated with DLBCL. Anaemia in these patients due to immune mediated haemolysis is a rare occurrence. CAD can present with severe anaemia as 50% of patients require a red cell transfusion.[7],[8] Anaemia only improves transiently with RBC transfusions, which should only be considered for severe and acute cases. An in-line blood warmer and slow rate of transfusion are the preferred modalities to reduce increased risk of transfusion reaction. In our case two units of best matched 'O' Rh (D) negative red cell units were transfused through inline blood warmer without any complication.

CAs are IgM type and are produced by B cells, predominantly at the lymphoplasmacytic cell stage. IgM can also be produced by a smaller compartment of plasma cells that are long-lived and not targeted by chemo-immunotherapy. The CA-producing cells are monoclonal in CAD as well as in CAS secondary to lymphoma, but polyclonal in CAS secondary to infection. There is a potential for the generation of auto-reactive lymphocytes due to the process of rearrangement of Ig T-cell receptor gene segment in the DNA of precursor lymphocytes. The failure in preventive breakdown mechanism which prevents the expansion and promotes the elimination of these two types of autoreactive cells may lead to several autoimmune manifestations. Hence, patients with tumours of the immune system such as B or T cell NHL are at risk of developing antibodies against haematopoietic cells or other tissues.[11],[12],[13] AIHA is a rare complication of NHL and is usually caused by warm auto-antibodies regardless of stage of the disease.[14] Cold agglutinin disease makes only a minor group of AIHA.

Cold agglutinin disease occurs due to circulating IgM antibodies (rarely IgA or IgG) which bind RBC at low temperature generally below 30°C leading to agglutination of RBC and frequent fixation of complement and accounts for approximately 20% of all AIHA cases.[2],[4] Cold AIHA is a complication in 1.1%–4.8% of all NHL.[15] Although DLBCL is the most common type of NHL, only few cases presenting as cold agglutinin disease have been reported in literature with uncertain prognostic significance of this association. In a study by Sallah et al., it was demonstrated by the fact that 1 of the 14 patients who achieved response and 6 of 7 NHL patients who were alive at the conclusion of their analysis had warm haemolytic anaemia.[14]

Even in asymptomatic cases, despite supportive approaches, drug therapy is indicated. Steroids are generally less effective in cold agglutinin AIHA as compared with warm AIHA with response rate as low as of 15%–30%.[16] However, in case of some subgroups of patients, particularly those with IgG cold agglutinin steroid can be effective.[17] RBC destruction in CAD is primarily known to occur in liver hence splenectomy is not a recommended treatment for CAD.[5],[8],[18] There are case reports of NHL where patients with cold AIHA have been successfully treated with combination chemo-immunotherapy,[18],[19],[20] while as reported by Sallah et al., 2 out of three patients with NHL and cold agglutinin disease were refractory to CHOP chemotherapy.[14] Rituximab, the B cell targeting antibody, is the gold standard to decrease autoantibody production in patients with CAD. Our case reports that the patient had good response to therapy and is in remission post 6 cycles of chemo-immunotherapy with normal haemoglobin. It appears that treatment with rituximab-based chemo-immunotherapy regimen has improved prognosis in these patients.

  Conclusion Top

Cold antibody AIHA can be a rare presenting feature of underlying aggressive NHLs. There is a lack of information in the literature in terms of prognostic implications of such phenomenon on clinical course of the disease. In many cases, lymphoma can precede or follow the development of haemolytic anaemia. Therefore, an astute clinician dealing with cold agglutinin, AIHA should always consider underlying lymph proliferative disorder in addition to infectious causes. To better address issues relevant to the treatment and prognosis of cold autoimmune haemolytic anaemia in patients with NHL, large-scale prospective trials are required.

Declaration of patient consent

The authors certify that they have obtained all appropriate patient consent forms. In the form, the patient has given his consent for his images and other clinical information to be reported in the journal. The patient understands that his name and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

  References Top

Dacie JV. The Haemolytic Anaemias. The Autoimmune Haemolytic Anaemias. 3rd ed. New York: Churchill Livingstone; 1992.  Back to cited text no. 1
Petz LD, Garratty G. Acquired Immune Hemolytic Anemias. Philadelphia: Churchill Livingstone; 2004.  Back to cited text no. 2
Sokol RJ, Hewitt S, Stamps BK. Autoimmune haemolysis: An 18-year study of 865 cases referred to a regional transfusion centre. Br Med J (Clin Res Ed) 1981;282:2023-7.  Back to cited text no. 3
Berentsen S, Ulvestad E, Langholm R, Beiske K, Hjorth-Hansen H, Ghanima W, et al. Primary chronic cold agglutinin disease: A population based clinical study of 86 patients. Haematologica 2006;91:460-6.  Back to cited text no. 4
Berentsen S, Tjønnfjord GE. Diagnosis and treatment of cold agglutinin mediated autoimmune hemolytic anemia. Blood Rev 2012;26:107-15.  Back to cited text no. 5
Berentsen S. Role of complement in autoimmune hemolytic anemia. Transfus Med Hemother 2015;42:303-10.  Back to cited text no. 6
Berentsen S. How I manage cold agglutinin disease. Br J Haematol 2011;153:309-17.  Back to cited text no. 7
Swiecicki PL, Hegerova LT, Gertz MA. Cold agglutinin disease. Blood 2013;122:1114-21.  Back to cited text no. 8
Ulvestad E, Berentsen S, Bø K, Shammas FV. Clinical immunology of chronic cold agglutinin disease. Eur J Haematol 1999;63:259-66.  Back to cited text no. 9
Váróczy L, Gergely L, Zeher M, Szegedi G, Illés A. Malignant lymphoma-associated autoimmune diseases - A descriptive epidemiological study. Rheumatol Int 2002;22:233-7.  Back to cited text no. 10
Grønbaek K, D'Amore F, Schmidt K. Autoimmune phenomena in non-Hodgkin's lymphoma. Leuk Lymphoma 1995;18:311-6.  Back to cited text no. 11
Polliack A, Lugassy G. Autoimmunity and auto-immune syn-dromes associated with and preceding the development of lymphoproliferative disorders. Leukemia 1992;6:152-4.  Back to cited text no. 12
Borche L, Lim A, Binet JL, Dighiero G. Evidence that chronic lymphocytic leukemia B lymphocytes are frequently committed to production of natural autoantibodies. Blood 1990;76:562-9.  Back to cited text no. 13
Sallah S, Sigounas G, Vos P, Wan JY, Nguyen NP. Autoimmune hemolytic anemia in patients with non-Hodgkin's lymphoma: Characteristics and significance. Ann Oncol 2000;11:1571-7.  Back to cited text no. 14
Economopulos T, Stathakis N, Constatindou M, Papageorugiou E, Anastassiou C, Raptis S, et al. Cold aggulutinin disease in non-Hodgkin's lymphoma. Eur J Haematol 1995;55:69-71.  Back to cited text no. 15
McNicholl FP. Clinical syndromes associated with cold agglutinins. Transfus Sci 2000;22:125-33.  Back to cited text no. 16
Petz LD. Cold antibody autoimmune hemolytic anemias. Blood Rev 2008;22:1-15.  Back to cited text no. 17
Kosugi S, Watanabe M, Hoshikawa M. Primary bone marrow lymphoma presenting with cold-type autoimmune hemolytic anemia. Indian J Hematol Blood Transfus 2014;30:271-4.  Back to cited text no. 18
Eskazan AE, Akmurad H, Ongoren S, Ozer O, Ferhanoglu B. Primary gastrointestinal diffuse large B cell lymphoma presenting with cold agglutinin disease. Case Rep Gastroenterol 2011;5:262-6.  Back to cited text no. 19
Sumi M, Ichikawa N, Shimizu I, Yotsumoto M, Ueno M, Kobayashi H. [Primary diffuse large B-cell lymphoma of the bone marrow complicated with autoimmune hemolytic anemia and erythroid hypoplasia]. Rinsho Ketsueki 2007;48:571-5.  Back to cited text no. 20


  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7]

  [Table 1]


Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

  In this article
Case Report
Article Figures
Article Tables

 Article Access Statistics
    PDF Downloaded15    
    Comments [Add]    

Recommend this journal