Complement Activation and Inhibition in Autoimmune Hemolytic Anemia: Focus on Cold Agglutinin Disease

Seminars in Hematology 55 (2018) 141–149

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Review

Complement Activation and Inhibition in Autoimmune Hemolytic Anemia: Focus on Cold Agglutinin Disease Sigbjørn Berentsen* Department of Research and Innovation, Haugesund Hospital, Helse Fonna HF, Haugesund, Norway

a r t i c l e i n fo

Keywords: Autoimmune hemolytic anemia Cold agglutinin disease Complement therapy Complement inhibitors

a b s t r a c t The classical complement pathway and, to some extent, the terminal pathway, are involved in the immune pathogenesis of autoimmune hemolytic anemia (AIHA). In primary cold agglutinin disease (CAD), secondary cold agglutinin syndrome and paroxysmal cold hemoglobinuria, the hemolytic process is entirely complement dependent. Complement activation also plays an important pathogenetic role in some warm-antibody AIHAs, especially when immunoglobulin M is involved. This review describes the complement-mediated hemolysis in AIHA with a major focus on CAD, in which activation of the classical pathway is essential and particularly relevant for complement-directed therapy. Several complement inhibitors are candidate therapeutic agents in CAD and other AIHAs, and some of these drugs seem very promising. The relevant in vitro findings, early clinical data and future perspectives are reviewed. & 2018 Elsevier Inc. All rights reserved.

Introduction Autoimmune hemolytic anemia (AIHA) is a heterogeneous group of disorders, characterized by autoantibody-initiated destruction of red blood cells (RBCs). AIHAs can be classified according to the properties of the autoantibody as shown in Table 1 [1-5]. Our knowledge of etiology and pathogenesis, including the role of complement for RBC breakdown in subgroups of AIHA, is rapidly growing [6,7]. Although paroxysmal nocturnal hemoglobinuria (PNH) is not an autoimmune disease, lessons learnt from the entirely complement-mediated pathogenesis and the success of therapeutic complement inhibition in PNH have proved useful in understanding and treating AIHA [8,9]. The first examples of clinically effective complement modulation in subtypes of AIHA were published less than 10 years ago [10,11] and, during the same period, several new complement inhibitors have been developed for potential clinical use [12-14]. This review will address the role of complement for antibodyinitiated hemolysis in AIHA as well as its present and future implications for therapeutic use of complement inhibitors. Established, noncomplement directed therapies will be mentioned as far as relevant. Essential characteristic findings in specific types of

The author has received research support from Mundipharma, travel grants from Alexion, and has consulted for True North Therapeutics, Bioverativ, and Apellis. ⁎ Corresponding author. Sigbjørn Berentsen, Department of Research and Innovation, Haugesund Hospital, P.O. Box 2170, NO-5504 Haugesund, Norway. Tel.: þ47 5273 2000. E-mail address: [email protected] https://doi.org/10.1053/j.seminhematol.2018.04.002 0037-1963/$/& 2018 Elsevier Inc. All rights reserved.

AIHA are listed in Table 2. These diagnostic procedures will not be described in detail; comprehensive guidelines can be found elsewhere [4,15]. In cold agglutinin disease (CAD), the pathogenesis has been shown to be entirely dependent on the classical complement pathway [16-20]. The theoretical rationale for therapeutic complement inhibition is particularly strong and the documentation for this treatment modality has been further developed in CAD than in other types of AIHA [9,10,20-23]. The discussion on pathogenesis and potential role of pharmacological complement inhibitors, therefore, will focus especially on CAD.

Immune Hemolysis and the Role of Complement The initial step of the autoimmune hemolytic process is an antigen-antibody reaction resulting in deposition of the autoantibody on the erythrocyte surface, with or without complement fixation [24]. The routine method for detecting immunoglobulins or complement proteins bound to RBCs is the direct antiglobulin test (DAT) [15,25]. DAT is often first performed using a polyspecific antiserum, which will detect any significant deposition of IgG or complement, but not IgA [25,26]. If the polyspecific DAT is positive (or if an IgA-mediated AIHA is suspected in cases of negative DAT), the monospecific or “extended” DAT (using monospecific antibodies) is carried out to further identify the immunoglobulin class or complement protein [2,25]. Figure 1 gives an overview of the hemolytic mechanisms in AIHA. In about half of the cases of warm-antibody AIHA,

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Table 1 Autoimmune hemolytic anemia (AIHA). Warm-antibody types

Cold-antibody types

Atypical AIHA

Primary warm-antibody AIHA Secondary warm-antibody AIHA • Other autoimmune disorders • Chronic lymphocytic leukemia (CLL) • Lymphoproliferative disorders other than CLL • Drug-induced

Primary cold agglutinin disease (CAD) Secondary cold agglutinin syndrome (CAS) • Specific infections (Mycoplasma, Epstein-Barr virus, others) • Aggressive lymphoma • Other malignancies

Mixed warm and cold-antibody type DAT-negative AIHA

Paroxysmal cold hemoglobinuria (PCH)

noncomplement mechanisms are responsible for the hemolysis, as described in Warm–antibody-mediated autoimmune hemolytic anemia. Complement-dependent hemolysis is mediated by the classical pathway and, depending on the type and severity of the AIHA, to some extent by the terminal pathway [7,27-29]. On RBCs opsonized with immunoglobulin, the amount of antigen-antibody complex can be sufficient or not sufficient for binding complement protein complex C1 and, thereby, for initiating the classical pathway [30]. Immunoglobulin M (IgM) is a potent trigger of the classical complement pathway, although usually not found on the RBC surface by DAT [31,32]. Complement activation by IgG is weaker and depends on the subclass. IgG3 activates more efficiently than does IgG1, while IgG2 is a still weaker activator and there is no evidence for complement activation by IgG4 [28,33]. IgA does probably not activate complement, but IgA-deposition can still lead to fulminant hemolysis [26,34]. A probable explanation is involvement of IgM even in cases where IgG or IgA is the only immunoglobulin class detected, since IgM will usually detach from the RBC before it can be demonstrated by DAT [29,32]. Upon initiation of the classical pathway, C1 esterase activates C2 and C4, generating C3 convertase, which cleaves C3 into C3a

and C3b. Especially in heavy complement activation induced by IgM, C3 proteolysis will leave the cell surface coated with large amounts of C3d and other split products, which can be detected using monospecific DAT for C3 fragments [32,35]. Therefore, even though a positive DAT for C3d may result from binding of any Ig class able to activate complement, this finding should raise the suspicion of IgM involvement [22,25,32]. C3b-opsonized erythrocytes are prone to phagocytosis by the mononuclear phagocytic system, previously termed the reticuloendothelial system. This extravascular hemolysis is responsible for a variable part of the red cell destruction [20,24,28]. On the surviving RBCs, C3b is cleaved into inactivated fragments. Because these split products have lower affinity for the C3 receptors expressed on macrophages, C3d opsonized cells have been considered resistant to further phagocytosis [35,36]. However, the possibility that even C3d may contribute to extravascular hemolysis has not been completely ruled out [36]. In addition or alternatively, C3 fixed to the cell surface can initiate the terminal complement cascade by binding C5, which is then split into C5a and C5b. The production of C5b results in formation of the C5b6789 complex, also known as the membrane attack complex, with subsequent intravascular hemolysis. The extent

Table 2 Diagnostic tests in autoimmune hemolytic anemia (AIHA). Type of AIHA

Polyspecific DAT Monospecific DAT positive for

Other diagnostic tests

Warm-antibody

Positive

IgG 7 C3d

Warm-antibody, IgA only Warm-antibody, IgM ( þ IgG) CAD

Negative

IgA

Positive

C3d (7 IgG, 7 IgM)

Positive

C3d (7 weak IgG)

• Cold agglutinin titer • Thermal amplitude if required

CAS

Positive

C3d (7 weak IgG)

• Cold agglutinin titer • Thermal amplitude if required

PCH

Positive or negative for IgG or C3d IgG þ C3d

• Donath-Landsteiner test

Mixed-type

Positive or negative Positive

DAT-negative

Negative

Negative

Clinical exclusion More sensitive tests for RBC-bound Ig or C fragments. Options: • Elution techniques → IAT • Flow cytometry • MS-DAT

• IAT at 37°C (warm agglutinin) • Cold agglutinin titer • Thermal amplitude

C ¼ complement; IAT ¼ indirect antiglobulin test; MS-DAT ¼ mitogen-stimulated DAT.

Confirmatory or further work-up

Search for associated or underlying disorder(s) Search for associated or underlying disorder(s) Search for associated or underlying disorder(s) • Ig classes quantification • Serum electrophoresis • Immune fixation • Bone marrow flow cytometry • Bone marrow biopsy Search for underlying disorder

Search for underlying disorder Search for associated or underlying disorder(s)

Search for associated or underlying disorder(s)

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Macrophage Spherocyte

Ig

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lymphatic leukemia [1,39]. Examples of associated immunological disorders are systemic lupus erythematosus (SLE), rheumatoid arthritis, Sjögren’s syndrome, autoimmune hepatitis, hypothyroidism, inflammatory bowel disease, immune thrombocytopenia, and primary hypogammaglobulinemia [1,3]. An increased risk of thrombosis has been well documented [2].

Ig Ig

Erythrocyte Destruction in Warm-AIHA

Spleen C1

Extravascular lysis

Ig C3b

C2, C4 C3

C3b C5 Liver

C3b Extravascular lysis

C5b6789

Intravascular lysis

C3d Survival

Fig. 1. Destruction of antibody-opsonized red blood cells (RBCs) in warm-antibody autoimmune hemolytic anemia. Four mechanisms are illustrated: (1) Macrophageinflicted membrane damage resulting in formation of spherocytes and phagocytosis (extravascular hemolysis), mainly in the spleen. (2) Extravascular hemolysis in the spleen of opsonized RBCs without previous spherocyte formation. (3) Complement classical pathway activation on IgM- or heavily IgG-coated RBCs, resulting in extravascular lysis of C3b-opsonized cells in the liver and survival of C3d-labeled cells. (4) Activation of the terminal complement pathway, resulting in intravascular hemolysis. C ¼ complement protein. (Color version of the figure available online.)

to which this occurs depends on the type of AIHA, the involved antibody class, and other factors activating the complement system such as any acute phase reaction [22,24,37]. In general, terminal pathway activation with intravascular hemolysis is the main hemolytic mechanism in PCH, relatively prominent in many cases of warm-AIHA, and only a minor mechanism in most patients with CAD [22,29,38]. Part of the explanation for this is the protective effect of the physiological cell surface complement inhibitors CD55 and CD59 which, unlike in PNH, are intact in AIHA [17].

Warm–Antibody-Mediated Autoimmune Hemolytic Anemia

The autoantibodies involved in warm-AIHA are most often of the IgG class. IgA autoantibodies occur in 15%-20% of the patients, either in combination with IgG or, infrequently, alone [2,26]. IgM is probably involved in up to 50% of the patients, leading to complement activation and signaled by a positive DAT for C3d [25,32,42]. Several hemolytic mechanisms are involved in warm-AIHA, as illustrated in Figure 1. One mechanism is membrane damage inflicted by macrophages that express Fc receptors, resulting in formation of spherocytes, which are prone to destruction in the spleen [43]. Second, opsonized cells can undergo phagocytosis by macrophages in the spleen without previous formation of spherocytes [7,24]. Third, RBCs that have bound IgM or are heavily coated with IgG are destroyed by the complement system as described in Section Immune hemolysis and the role of complement, resulting in either (A) intravascular hemolysis mediated by the terminal pathway [24,29], or (B) classical pathwayinitiated, C3b-induced extravascular hemolysis by the mononuclear phagocytic system, mainly in the liver [16,24]. Fourth, several cytokines as well as cytotoxic CD8 þ T-cells and natural killer cells can be involved the hemolytic process [6,29,44]. Established Therapies for Warm-AIHA Initially, most patients with primary warm-antibody AIHA respond well to corticosteroids [2,45], but the probability of staying in complete remission after discontinuation at 15 months has been estimated to less than 40% [46]. Addition of rituximab in the first line has been found to increase the probability of sustained remission [47,48]. Second-line therapies are rituximab (if not given in the first line) or splenectomy. Unspecific immunosuppressants are often used in the third line, but the efficacy is poorly documented. Some patients with acute or subacute, severe warm-AIHA can be very difficult to treat, especially those with IgM involvement and strong complement activation [11,32,49]. IgA involvement and DAT-negative AIHA are associated with more severe anemia and an increased risk of initial therapy failure [2,25,34]. Transfusions require specific precautions described elsewhere [15,45,50]. Thus, the treatment of warm-AIHA reveals several unmet needs.

Pathogenesis and Associated Diseases Cold Agglutinin Disease Warm-antibody types account for approximately 75% of the cases of AIHA [1-3]. The autoantibodies have a temperature optimum at 37°C and are invariably polyclonal, even when the warm-AIHA complicates a clonal B-cell lymphoproliferative disorder (LPD) [39,40]. A general dysregulation of the immune system with impaired distinction between self and nonself seems essential to pathogenesis. This dysfunction involves an inadequate T-cell mediated regulation of the humoral immune system, polymorphism of the gene for the signal substance CTLA-4 and the role of regulatory T-cells (Treg-cells) [40,41]. It is not surprising, therefore, that a large number of immunological and lymphoproliferative disorders can be associated with warm-AIHA. Secondary AIHA, that is, cases with a demonstrable associated or underlying disease, accounts for more than 50% of w-AIHA, whereas the remaining cases are classified as primary [3,42]. The most frequently occurring associated LPD is chronic

CAD is a Clonal, Lymphoproliferative Disorder We should distinguish between CAD, which is now considered a well-defined clinicopathologic entity and should be called a disease, not syndrome [9,17,22,51-53], and secondary cold agglutinin syndrome (CAS), further described in Section Other cold– antibody-mediated hemolytic anemias [4]. CAD accounts for 15%25% of AIHA with a reported prevalence in Northern Europe of 16 per million and an incidence rate of 1 per million per year [1,2,18]. It affects mainly elderly and middle-aged people but has been reported down to an age of 30 [18,54]. CAD is a specific, clonal B-cell LPD of the bone marrow, which typically can be distinguished from lymphoplasmacytic lymphoma and other low-grade LPDs. This notion has been supported by a recent histopathology study [51] as well as studies of monoclonal

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phase reaction will cause enhanced production of complement proteins; the low levels are replete, and exacerbation of hemolysis may ensue [37]. Direct activation of the complement cascade may also contribute to the exacerbation in these situations. Diagnosis and Clinical Features

Fig. 2. Complement-mediated hemolysis in cold agglutinin disease. Antigen-bound IgM (cold agglutinin) on the red blood cell (RBC) surface binds C1q and initiates the classical pathway. Sequential reactions lead to the formation of C3b. Upon warming to 37°C, IgM detaches from the cell, allowing agglutinated RBCs to separate, whereas C3b remains bound. C3b-coated cells are sequestered by the mononuclear phagocytic system, mainly in the liver. On the surviving RBCs, C3b is cleaved, leaving high numbers of C3d molecules. In some situations, complement activation proceeds beyond the C3b step with cleavage of C5, resulting in activation of the terminal pathway and intravascular hemolysis. C ¼ complement protein. (Color version of the figure available online.)

immunoglobulin found in these patients [18,54,55], flow cytometry [18,56,57], heavy and light chain gene usage [58,59], and molecular biology [60,61]. The concept of CAD as a clonal LPD has been increasingly accepted [9,17,52,62]. The autoantibodies in CAD are monoclonal, produced by the clonal B-cells and termed cold agglutinins (CAs) because they are able to agglutinate RBCs at an optimum temperature of 3-4°C [18,19,51]. The thermal amplitude (TA) is the highest temperature at which the CA will react with its antigen [63,64]. A high TA, overlapping with normal temperatures in the acral parts of the body, will make the CA pathogenic. CAs are of the IgMκ class in more than 90% of the patients and usually have specificity for the I antigen, a carbohydrate antigen which is almost universally present at the RBC surface of humans aged 1-2 years or more [18,19,55,65]. CAs are semi-quantitatively assessed by the titer, which should be expressed as an integer, defined as the inverse of the highest dilution at which they are able to agglutinate RBCs [19]. Hemolysis in CAD is Entirely Complement Dependent Cooling of the blood during passage through acral parts of the circulation allows CA to bind to RBCs and cause agglutination. The antigen-IgM complex binds C1q on the cell surface and initiates the classical complement pathway (Fig. 2) [9,19,20,23]. Upon warming to 37°C in the central circulation, CAs detach from the cells, allowing agglutinated erythrocytes to separate. C3b remains bound and C3b-opsonized cells undergo phagocytosis [16,28]. The classical complement pathway-mediated, extravascular hemolysis is probably the predominant mechanism of erythrocyte destruction in CAD during stable disease [17,20,37,66]. At least in some patients and situations, however, complement activation is thought to proceed beyond the C3 stage, thereby triggering the terminal complement cascade with intravascular hemolysis. The occurrence of hemoglobinuria in about 15% of the patients supports this notion [54], as do the rather frequent finding of hemosiderinuria [53] and the modest but statistically significant beneficial effect of therapy with eculizumab [10,67]. Because of continuous complement consumption, most patients with CAD have low-serum levels of C3 and C4, which is probably rate-limiting for the hemolysis [19,37]. During febrile infections and following major surgery or major trauma, acute

CAD is defined by chronic hemolysis, CA titer Z64 at 4°C and typical findings by the DAT [17,18,68]. The typical DAT pattern is a positive polyspecific test and a monospecific test positive for C3d only. However, DAT can be weakly positive for IgG in addition to C3d in up to 20% of the patients [18,54]. It should be emphasized that not all subjects with a positive DAT for C3d have CAD, CAS, or even AIHA [2]. Furthermore, not all individuals with CA in serum have CAD or CAS, and detectable CA was found in 0.3% in a cohort with nonrelated disorders [69]. These normally occurring CAs are remnants of a phylogenetically ancient, primitive adaptive immune system [70]; they are polyclonal, have low TA, and are present in low titers, usually less than 64 and rarely exceeding 256 [68,71]. For CA titration, serum electrophoresis and other immunoglobulin analyses, it is of critical importance that blood specimens are kept at 37-38°C from sampling until serum has been removed from the clot [17,19]. In some patients it is also necessary to prewarm ethylenediaminetetraacetic (EDTA) blood samples before analyzing hemoglobin (Hb) levels and blood cell counts. Despite this procedure, a spuriously high median corpuscular volume and a false reduction in RBC counts are regularly seen, making estimated hematocrit values unreliable [72]. Additional workup to detect a clonal lymphoproliferation is not required for diagnosis, as this may be a matter of sensitivity, but should be done because the results may be confirmative and have therapeutic implications. Such examinations include quantification of serum Ig classes, capillary or agarose electrophoresis with immunofixation, flow cytometric immune phenotyping in bone marrow aspirate and examination of a bone marrow biopsy sample. At least in cold or cool climates, 90% of the patients have coldinduced circulatory symptoms, ranging from slight acrocyanosis to disabling Raynaud phenomena [18]. In a population-based study of 86 patients with CAD, the median Hb level was 8.9 g/dL and the lower tertile 8.0 g/dL (range: 4.5 g/dL to normal; a few patients have fully compensated hemolysis). Characteristic seasonal variations in hemolysis have been demonstrated in individual patients [73]. Totally, 70% of the patients have experienced complementmediated exacerbation of anemia during febrile infections, and approximately 50% have been transfused [18]. A suspected risk of thromboembolic complications, mostly attributed to the hemolysis itself, has more difficult to demonstrate in CAD than in warm-AIHA [2,74]. In theory, patients might have an additional risk inflicted by the complement activation, as there are numerous points of interaction between the complement system and the coagulation cascade [75]. A registry-based study of individuals with the term CAD repeatedly mentioned in their medical records reported a relative frequency of 1.8 as compared with age-matched controls [76]. Nonpharmacological Management of CAD Patients should avoid low ambient temperatures and use warm clothing, including protection of extremities, ears and face [15,62]. In the ward or outpatient department, they should keep warm and avoid infusion of cold liquids. To prevent exacerbation induced by complement activation, any bacterial infection should be treated [18,37]. Transfusions can safely be given when indicated, provided the necessary precautions are observed, and the compatibility problems typical for warm-AIHA are not encountered in CAD

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[15,62,77]. The patient and the extremity chosen for transfusion should be kept warm, and the use of an in-line blood warmer is recommended. Because low complement levels are rate-limiting for hemolysis, transfusion of blood products with a high plasma content probably carries a risk of exacerbation [37]. In critical situations, plasmapheresis is an option for “first-aid” [78,79]. Because IgM is found almost exclusively in the blood, this procedure is considered highly efficient, although some conflicting data do exist [79,80]. Exchange of 1-1.5 times the plasma volume with albumin daily or every second day has been recommended [78]. The effect is short-lived and pharmacological therapy should be initiated concomitantly. Splenectomy is not an option in treating CAD, as the extravascular hemolysis mainly takes place in the liver [16,18,77]. Exceptions may exist among the rare patients with CA of the IgG class or with a TA approaching 37°C [77,81]. Surgery performed in hypothermia or cardiopulmonary bypass requires specific precautions because of the cooling itself as well as trauma-induced complement activation. Patients with CAD should have a hematology consultation and TA assessment prior to cardiac surgery, which should preferentially be performed under normothermia [67,82]. Current Pharmacological Therapy for CAD Patients with only mild anemia or fully compensated hemolysis in whom circulatory symptoms are tolerable or absent do not require drug treatment. Descriptive studies have found that 70%80% of the patients had received pharmacologic therapy, often in multiple lines and with few and short-lived remissions [18,54,83]. Most documented therapies have been directed against the pathogenic B-cell clone. The first treatment shown to have an acceptable efficacy was rituximab monotherapy, which is generally well tolerated and the most frequently used therapy for CAD [84,85]. The overall response rate (ORR) is approximately 50%, almost all remissions are partial responses, and the median response duration has been estimated to 11 months [84]. Rituximab in combination with fludarabine has yielded a high ORR at 76% with 21% complete responses (CR) and an estimated median response duration of more than 66 months, however with some toxicity [86]. Rituximab plus bendamustin combination therapy has resulted in 71% ORR, 40% CR, and long response duration (10-percentile not reached after 32 months), with a substantially more favorable toxicity profile than rituximab-fludarabine [87]. Rituximab plus bendamustine may be considered in first line in relatively fit patients. A single cycle of bortezomib monotherapy has been reported to result in 3 CR and 3 partial responses in a cohort of 19 patients [88]. No results have been published regarding the newer, targeted anti-B cell therapies such as ibrutinib, idelalisib, and venetoclax. Chemoimmunotherapy directed against the pathogenic B-cell clone in CAD is characterized by a long time to response (TTR). The median TTR following bendamustine plus rituximab has been reported to be 1.9 months (range: 0.25-12), with a median time to best response of 7.0 months (range: 1.5-30) [87].

Other Cold Antibody-Mediated Hemolytic Anemias Secondary Cold Agglutinin Syndrome A cold hemolytic syndrome similar to CAD is occasionally encountered in patients with aggressive lymphoma [3,4]. CAS has also been associated with a variety of other malignant diseases, although in some cases the association can be questioned [1,4]. CAs in patients with aggressive lymphoma are monoclonal, most often of the IgM class, but the light restriction can be λ as

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well as κ [89]. They are usually anti-I specific, but anti-i specificity has been described [90]. A more acute clinical picture of CAS can complicate Mycoplasma pneumoniae pneumonia, Epstein-Barr virus infection and, rarely, other specific infections. These CAs are polyclonal. In Mycoplasma infection, they are anti-I specific IgM antibodies, whereas Epstein-Barr virus induced CAs have specificity for the i antigen and can be of the IgM or IgG class [4,90]. The further immune pathogenesis is entirely complement-mediated and probably identical to that in primary CAD. The underlying condition should be treated if possible, but otherwise there is no known therapy for secondary CAS. The postulated effect of corticosteroids is poorly documented [4]. Although infection-associated CAS invariably will resolve along with the underlying infection, some patients can remain severely anemic and transfusion dependent for weeks, which identifies an unmet need for novel therapies. Paroxysmal Cold Hemoglobinuria The autoantibodies found in PCH, Donath-Landsteiner antibodies, are biphasic and have specificity for the RBC surface antigen P [38]. The temperature optimum for the antibody-antigen reaction is low, whereas subsequent C1 fixation occurs after warming to normal central body temperature. The classical and terminal complement pathways are activated and the hemolysis is predominantly intravascular [7,38]. PCH is a rare disease, mostly seen in children after virus infections. In adults, PCH used to be associated with tertiary syphilis, but this variant is hardly reported anymore. Nonsyphilitic adult PCH has been described in hematologic malignancies and in cases of unknown etiology [38]. Though postviral PCH is self-remitting, the patients can have severe hemolysis and profound anemia. Corticosteroid therapy has often been used but is poorly documented [7,38,77], and there is an obvious unmet need for effective therapy in these rare patients.

Complement-Directed Therapies Therapeutic Complement Inhibitors A large number of therapeutic and experimental complement modulating substances has been developed [12,13]. As explained earlier, complement inhibitors expected to be effective in AIHA (including CAD) must target the classical or the terminal pathway. Figure 3 shows an overview of such substances and targets. The first complement inhibitor available for clinical use was plasma-derived C1-esterase inhibitor (C1-INH). Although approved only for hereditary angioedema, which is not a complement-mediated disorder, it may seem logical to use this substance in classical complement pathway-dependent diseases as well. Because patients with AIHA produce sufficient amounts of endogenous C1-INH, high doses may be required. In a well-described case report, the administration of C1-INH was shown to immediately stop hemolysis in a patient with a severe, steroid-refractory and transfusion-resistant IgM-driven secondary warm-antibody AIHA, thus offering a bridge for more slow-acting therapy to work [11,91]. Related in vitro experiments confirmed a dose-dependent suppression of C3 deposition on RBCs [11]. Although no prospective study has been done, these observations are considered a proof of principle for classical pathway inhibition in IgM-driven AIHA. Complement modulation in CAD was described in a case report on improvement following therapy with the anti-C5 monoclonal antibody eculizumab [92]. This observation led the authors to perform a prospective clinical trial of eculizumab in 13 patients [10]. Although the increase in hemoglobin levels was marginal and

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IgM C1-INH, ANX005, TNT003, BIVV009 C1 C2, C4

APL-2

C3 ECULIZUMAB C3a

C3b C5a C5 C5b6789

C3b EXTRAVASCULAR LYSIS

C3d

INTRAVASCULAR LYSIS

SURVIVAL

Fig. 3. Complement inhibitors relevant for present or future treatment of autoimmune hemolytic anemia and their targets. (Originally published in Blood by Berentsen et al [22], reused with permission, slightly modified and updated. © Blood, the Journal of the American Society of Hematology.) (Color version of the figure available online.)

there was no significant improvement in quality of life, the study showed a significant reduction in transfusion requirements and lactate dehydrogenase levels. A recent case report demonstrated prophylactic effect of eculizumab in a CAD patient who underwent heart surgery after previously having suffered from several exacerbations induced by surgery or febrile infections [67]. Taken together, the eculizumab data indicate that C5 inhibition has a modest, beneficial effect in some patients. This conclusion should encourage investigations of more proximal complement modulation targeting the classical pathway, which seems more important in CAD. The most comprehensive studies to date on complement inhibition in CAD are in vitro experiments with the anti-C1s mouse monoclonal antibody TNT003 [20], followed by a phase 1B clinical trial studying its humanized counterpart BIVV009, formerly known as TNT009 [21]. The in vitro study tested the effects of TNT003 on CA-induced complement activity against human erythrocytes. Using CA samples from 40 patients with CAD, the authors found that the monoclonal antibody efficiently inhibited CA-induced deposition of C3 fragments on RBCs and prevented erythrophagocytosis by a phagocytic cell line [20]. TNT003 also inhibited the CA-induced, classical pathway-driven generation of anaphylotoxins C4a, C3a, and C5a. Interestingly, only one patient sample out of 40 was able to induce membrane attack complex-mediated hemolysis. Figure 4 illustrates some essential results of these experiments. In the phase 1B trial, 6 patients with typical primary CAD received BIVV009 intravenously [21,93]. A test dose of 10 mg/kg was followed by a full dose of 60 mg/kg 1-4 days later. Three additional doses of 60 mg/kg were administered at 1-week interval, and a weekly fixed dose of 5.5 g was given for maintenance. The patients received vaccination against meningococci, pneumococci and Haemophilus. BIVV009 was well tolerated and immediately stopped hemolysis in all of these 6 severely anemic patients, increasing Hb levels by a mean of 4.3 g/dL and eliminating transfusion requirements while on therapy. Infectious complications did not occur. BIVV009 has now entered phase 3 trials (ClinicalTrials.gov, NCT03347396 and NCT03347422).

Fig. 4. TNT003 prevents complement C3 deposition and phagocytosis of RBCs exposed to CAD patient plasma in the presence of normal human serum as a source of complement. Representative assay using a single patient sample. C3/C5, complement protein 3 and 5, respectively; NHS ¼ normal human serum; IC ¼ internal control. Columns below blue bar represent measurements after addition of cold agglutinin (patient plasma). (Originally published in Blood by Shi et al [19], reproduced with permission. © Blood, the Journal of the American Society of Hematology.) (Color version of the figure available online.)

ANX005, a humanized monoclonal antibody that targets C1q, has also been shown in vitro to prevent C4 and C3 fragment deposition on erythrocytes exposed to CAD sera, resulting in a dose-dependent reduction in hemolysis [94]. The compstatin family is a group of cyclic peptides that inhibit complement activation by binding C3, interfering with convertase formation and C3 proteolysis [95]. Because this step is essential in the classical as well as the alternative and lectin pathways, targeting C3 will allow total blockade of the complement system. Experiments with one of these substances, Cp40, found a dosedependent inhibition of hemolysis and prevention of C3 deposition on PNH erythrocytes in an in vitro system [96]. Preclinical in vivo studies indicated favorable pharmacokinetics following subcutaneous administration [96]. There are no published data on Cp40 in a CAD-related setting. APL-2 is a PEGylated compstatin analog designed for subcutaneous injection [13,97]. Systemic administration of APL-2 was investigated in two phase 1 trials comprising 51 healthy volunteers altogether [97]. Participants received vaccination against meningococci, pneumococci and Haemophilus. Monitoring of complement activity showed high efficacy of the study drug in inhibiting the classical and alternative pathways, and the results of pharmacodynamic assessments were favorable. There were few adverse events and no serious adverse events, although the maximum duration of drug administration was only 28 days. A phase 2 clinical trial of APL-2 in CAD has recently been posted at ClinicalTrials.gov (NCT03226678). Complement Inhibition in AIHA—Future Perspectives The therapeutic use of complement inhibitors in AIHA including CAD is at its very beginning, and only one of these drugs has reached phase 3 trials. Still, the existing data do allow some perspectives. In contrast to B-cell directed therapies for CAD, which are completed after administration of a few cycles, complement-directed therapies will probably have to be continued infinitely to maintain their effect. Probably, most complement inhibitors will also be very expensive. Of the B-cell directed therapies, rituximab plus bendamustine has resulted in high response rates, frequent CRs and long response duration, as described in this review. It should also be noted that because RBC agglutination is not complement mediated, complement

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inhibition cannot be supposed to improve the circulatory symptoms in CAD. On the other hand, chemoimmunotherapy will continue to fail in at least 25% of patients with CAD; relapses will eventually occur in all responders; some patients experience unacceptable toxicity; and others have contraindications or are reluctant to receive cytotoxic drugs. Therefore, a significant proportion of patients with CAD will be candidates for therapy with a complement inhibitor provided that further clinical trials confirm the high efficacy and favorable safety profile. As discussed earlier, chemoimmunotherapy for CAD is characterized by a long TTR, at least in a proportion of the patients [87]. In contrast, the onset of action seems very rapid during therapy with BIVV009 [21], and this may turn out to be the case with other proximal complement inhibitors as well. CAD patients who have profound hemolytic anemia and those with severe exacerbations may profit from this short TTR, which will offer a bridge to the more slowly acting B-cell directed therapies. Furthermore, complement inhibition may become useful as prophylaxis during cardiac or other major surgery [67]. Novel complement inhibitors should also be explored in IgMdriven warm-AIHA, secondary CAS and PCH. In the 2 latter, however, the low incidence will probably make prospective trials unrealistic. A case observation of therapy with eculizumab for PCH did not report any improvement [98], but this single case is not necessarily representative and more upstream inhibition might show better efficacy. Complement inhibition may carry a risk of severe infection. Long-term studies on C5 inhibition in PNH have found this risk negligible provided the patients are vaccinated against encapsulated bacteria or receive prophylactic penicillin [99]. In theory, the risk of infection may be higher with C3 inhibitors (which are able to block all complement activity) than with inhibitors of C1 (which will leave the alternative and lectin pathways intact). According to the limited amount of available clinical data, however, both C1 and C3 inhibition seem safe with regard to infection, but this issue will have to be carefully addressed in future clinical trials. Because hereditary deficiencies in proximal classical pathway components are associated with SLE [100], a risk of developing SLE might be suspected in patients treated with C1 inhibitors. Until now, clinical data have not supported this concern.

Conclusion In CAD, CAS, and PCH, the autoantibody-initiated hemolysis is entirely mediated by the classical complement pathway. The pathogenetic mechanisms in warm-antibody AIHA are more heterogeneous, but complement activation plays a substantial role even in this subgroup, especially when the immune pathogenesis is IgM-driven. Although therapy for CAD has improved greatly during the last 15 years, these achievements have revealed several unmet needs. Pharmacologic inhibition of the classical pathway in AIHA is still investigational but promising, and the potential of such therapy has been best elucidated in CAD. The C1s inhibitor BIVV009 has entered clinical phase 3 trials.

Acknowledgments Some of the author’s original works referenced in this review have been made possible through grants from Helse Vest RHF and Helse Fonna HF (public hospital trusts), to whom I am very grateful. I also thank all investigators who have participated in the Norwegian and Nordic CAD studies as well as international colleagues who have provided valuable input.

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References [1] 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. [2] Barcellini W, Fattizzo B, Zaninoni A, et al. Clinical heterogeneity and predictors of outcome in primary autoimmune hemolytic anemia: a GIMEMA study of 308 patients. Blood 2014;124:2930–6. [3] Michel M. Classification and therapeutic approaches in autoimmune hemolytic anemia: an update. Expert Rev Hematol 2011;4:607–18. [4] Berentsen S, Tjonnfjord GE. Diagnosis and treatment of cold agglutinin mediated autoimmune hemolytic anemia. Blood Rev 2012;26:107–15. [5] Mayer B, Yurek S, Kiesewetter H, Salama A. Mixed-type autoimmune hemolytic anemia: differential diagnosis and a critical review of reported cases. Transfusion 2008;48:2229–34. [6] Garratty G. The James Blundell Award Lecture 2007: do we really understand immune red cell destruction? Transfus Med 2008;18:321–34. [7] Berentsen S, Sundic T. Red blood cell destruction in autoimmune hemolytic anemia: role of complement and potential new targets for therapy. Biomed Res Int 2015;2015:363278. [8] Hill A, Rother RP, Arnold L, et al. Eculizumab prevents intravascular hemolysis in patients with paroxysmal nocturnal hemoglobinuria and unmasks low-level extravascular hemolysis occurring through C3 opsonization. Haematologica 2010;95:567–73. [9] Baines AC, Brodsky RA. Complementopathies. Blood Rev 2017;31:213–23. [10] Röth A, Bommer M, Hüttmann A, et al. Complement inhibition with eculizumab in patients with cold agglutinin disease (CAD): results from a prospective phase II trial (DECADE Trial). Blood 2015;126:274. [11] Wouters D, Stephan F, Strengers P, et al. C1-esterase inhibitor concentrate rescues erythrocytes from complement-mediated destruction in autoimmune hemolytic anemia. Blood 2013;121:1242–4. [12] Risitano AM. Current and future pharmacologic complement inhibitors. Hematol Oncol Clin North Am 2015;29:561–82. [13] Ricklin D, Barratt-Due A, Mollnes TE. Complement in clinical medicine: clinical trials, case reports and therapy monitoring. Mol Immunol 2017;89: 10–21. [14] Ricklin D, Mastellos DC, Reis ES, Lambris JD. The renaissance of complement therapeutics. Nat Rev Nephrol 2018;14:26–47. [15] Hill QA, Stamps R, Massey E, et al. The diagnosis and management of primary autoimmune haemolytic anaemia. Br J Haematol 2017;176: 395–411. [16] Jaffe CJ, Atkinson JP, Frank MM. The role of complement in the clearance of cold agglutinin-sensitized erythrocytes in man. J Clin Invest 1976;58:942–9. [17] Berentsen S. Cold agglutinin disease. Hematology Am Soc Hematol Educ Program 2016;2016:226–31. [18] Berentsen S, Ulvestad E, Langholm R, et al. Primary chronic cold agglutinin disease: a population based clinical study of 86 patients. Haematologica 2006;91:460–6. [19] Ulvestad E, Berentsen S, Bo K, Shammas FV. Clinical immunology of chronic cold agglutinin disease. Eur J Haematol 1999;63:259–66. [20] Shi J, Rose EL, Singh A, et al. TNT003, an inhibitor of the serine protease C1s, prevents complement activation induced by cold agglutinins. Blood 2014; 123:4015–22. [21] Jaeger U, D’Sa S, Schoergenhofer C, et al. Long term efficacy, safety and PK/PD profile of the anti-C1s antibody (BIVV009) in primary cold agglutinin disease patients. Blood 2017;130:703. [22] Berentsen S, Randen U, Tjønnfjord GE. Cold agglutinin-mediated autoimmune hemolytic anemia. Hematol Oncol Clin North Am 2015;29:455–71. [23] Berentsen S. Complement, cold agglutinins, and therapy. Blood 2014;123: 4010–2. [24] Dacie J. Auto-immune haemolytic anaemia (AIHA): pathogenesis. In: Dacie J, editor. The Haemolytic Anaemias, Vol. 3. London: Churchill Livingstone; 1992. p. 392–451. [25] Barcellini W. Pitfalls in the diagnosis of autoimmune haemolytic anaemia. Blood Transfus 2015;13:3–5. [26] Janvier D, Sellami F, Missud F, et al. Severe autoimmune hemolytic anemia caused by a warm IgA autoantibody directed against the third loop of band 3 (RBC anion-exchange protein 1). Transfusion 2002;42:1547–52. [27] Meulenbroek EM, Wouters D, Zeerleder S. Methods for quantitative detection of antibody-induced complement activation on red blood cells. J Vis Exp 2014:e51161. [28] Schreiber AD, Frank MM. Role of antibody and complement in the immune clearance and destruction of erythrocytes. I. In vivo effects of IgG and IgM complement-fixing sites. J Clin Invest 1972;51:575–82. [29] Barcellini W. New Insights in the Pathogenesis of Autoimmune Hemolytic Anemia. Transfus Med Hemother 2015;42:287–93. [30] Kurlander RJ, Rosse WF, Logue GL. Quantitative influence of antibody and complement coating of red cells on monocyte-mediated cell lysis. J Clin Invest 1978;61:1309–19. [31] Arndt PA, Leger RM, Garratty G. Serologic findings in autoimmune hemolytic anemia associated with immunoglobulin M warm autoantibodies. Transfusion 2009;49:235–42. [32] Meulenbroek EM, de Haas M, Brouwer C, et al. Complement deposition in autoimmune hemolytic anemia is a footprint for difficult-to-detect IgM autoantibodies. Haematologica 2015;100:1407–14.

148

S. Berentsen / Seminars in Hematology 55 (2018) 141–149

[33] Li Z, Shao Z, Xu Y, et al. Subclasses of warm autoantibody IgG in patients with autoimmune hemolytic anemia and their clinical implications. Chin Med J (Engl) 1999;112:805–8. [34] Bardill B, Mengis C, Tschopp M, Wuillemin WA. Severe IgA-mediated autoimmune haemolytic anaemia in a 48-yr-old woman. Eur J Haematol 2003;70:60–3. [35] Schreiber AD, Frank MM. Role of antibody and complement in the immune clearance and destruction of erythrocytes. II. Molecular nature of IgG and IgM complement-fixing sites and effects of their interaction with serum. J Clin Invest 1972;51:583–9. [36] Lin Z, Schmidt CQ, Koutsogiannaki S, et al. Complement C3dg-mediated erythrophagocytosis: implications for paroxysmal nocturnal hemoglobinuria. Blood 2015;126:891–4. [37] Ulvestad E, Berentsen S, Mollnes TE. Acute phase haemolysis in chronic cold agglutinin disease. Scand J Immunol 2001;54:239–42. [38] Shanbhag S, Spivak J. Paroxysmal cold hemoglobinuria. Hematol Oncol Clin North Am 2015;29:473–8. [39] Diehl LF, Ketchum LH. Autoimmune disease and chronic lymphocytic leukemia: autoimmune hemolytic anemia, pure red cell aplasia, and autoimmune thrombocytopenia. Semin Oncol 1998;25:80–97. [40] Fagiolo E. Immunological tolerance loss vs. erythrocyte self antigens and cytokine network disregulation in autoimmune hemolytic anaemia. Autoimmun Rev 2004;3:53–9. [41] Ward FJ, Hall AM, Cairns LS, et al. Clonal regulatory T cells specific for a red blood cell autoantigen in human autoimmune hemolytic anemia. Blood 2008;111:680–7. [42] Barcellini W. Immune hemolysis: diagnosis and treatment recommendations. Semin Hematol 2015;52:304–12. [43] LoBuglio AF, Cotran RS, Jandl JH. Red cells coated with immunoglobulin G: binding and sphering by mononuclear cells in man. Science 1967;158: 1582–5. [44] Gilsanz F, De La Serna J, Molto L, Alvarez-Mon M. Hemolytic anemia in chronic large granular lymphocytic leukemia of natural killer cells: cytotoxicity of natural killer cells against autologous red cells is associated with hemolysis. Transfusion 1996;36:463–6. [45] Lechner K, Jager U. How I treat autoimmune hemolytic anemias in adults. Blood 2010;116:1831–8. [46] Kulpa J, Skrabs C, Simanek R, et al. Probability of remaining in unsustained complete remission after steroid therapy withdrawal in patients with primary warm-antibody reactive autoimmune hemolytic anemia. Wien Klin Wochenschr 2016;128:234–7. [47] Birgens H, Frederiksen H, Hasselbalch HC, et al. A phase III randomized trial comparing glucocorticoid monotherapy versus glucocorticoid and rituximab in patients with autoimmune haemolytic anaemia. Br J Haematol 2013;163:393–9. [48] Michel M, Terriou L, Roudot-Thoraval F, et al. A randomized and double-blind controlled trial evaluating the safety and efficacy of rituximab for warm auto-immune hemolytic anemia in adults (the RAIHA study). Am J Hematol 2017;92:23–7. [49] Barcellini W. The clinical dilemma and management of red cell autoantibodies. Expert Rev Hematol 2016;9:325–7. [50] Barcellini W. Current treatment strategies in autoimmune hemolytic disorders. Expert Rev Hematol 2015;8:681–91. [51] Randen U, Trøen G, Tierens A, et al. Primary cold agglutinin-associated lymphoproliferative disease: a B-cell lymphoma of the bone marrow distinct from lymphoplasmacytic lymphoma. Haematologica 2014;99: 497–504. [52] Jaeger U. Hot therapy for cold agglutinin disease. Blood 2017;130:392–3. [53] Stone MJ, Berentsen S. Hyperviscosity syndrome, cold agglutinin hemolytic anemia, and cryoglobulinemia. In: Leblond V, Treon SP, Dimopoulos M, editors. Waldenström’s Macroglobulinemia. Switzerland: Springer International Publishing; 2017. p. 171–83. [54] Swiecicki PL, Hegerova LT, Gertz MA. Cold agglutinin disease. Blood 2013;122:1114–21. [55] Harboe M, van Furth R, Schubothe H, Lind K, Evans RS. Exclusive occurrence of K chains in isolated cold haemagglutinins. Scand J Haematol 1965;2: 259–66. [56] Berentsen S, Bo K, Shammas FV, Myking AO, Ulvestad E. Chronic cold agglutinin disease of the “idiopathic” type is a premalignant or low-grade malignant lymphoproliferative disease. APMIS 1997;105: 354–62. [57] de Tute R, Rawstron A, Evans P, Owen R. Cold agglutinin disease is a phenotypically distinct clonal B-cell disorder. 15 International Myeloma Workshop, Rome, Italy: Sept 23-26, 2015. Clin Lymphoma Myeloma Leuk 2015;15:e184. [58] Pascual V, Victor K, Spellerberg M, et al. VH restriction among human cold agglutinins. The VH4-21 gene segment is required to encode anti-I and anti-i specificities. J Immunol 1992;149:2337–44. [59] Malecka A, Trøen G, Tierens A, et al. Immunoglobulin heavy and light chain gene features are correlated with primary cold agglutinin disease onset and activity. Haematologica 2016;101:e361–4. [60] Malecka A, Trøen G, Tierens A, et al. Frequent somatic mutations of KMT2D (MLL2) and CARD11 genes in primary cold agglutinin disease. Br J Haematol 2017; E-pub ahead of print. https://doi.org/10.1111/bjh.15109. [61] Arthold C, Skrabs C, Mitterbauer-Hohendanner G, et al. Cold antibody autoimmune hemolytic anemia and lymphoproliferative disorders:

[62] [63] [64] [65] [66] [67]

[68]

[69]

[70] [71] [72] [73] [74]

[75] [76]

[77]

[78]

[79]

[80]

[81]

[82]

[83]

[84]

[85]

[86]

[87]

[88]

[89] [90]

[91]

a retrospective study of 20 patients including clinical, hematological, and molecular findings. Wien Klin Wochenschr 2014;126:376–82. Berentsen S. How I manage patients with cold agglutinin disease. Br J Haematol 2018;181:320–30. Schubothe H. The cold hemagglutinin disease. Semin Hematol 1966;3:27–47. Rosse WF, Adams JP. The variability of hemolysis in the cold agglutinin syndrome. Blood 1980;56:409–16. Issitt PD. I blood group system and its relationship to disease. J Med Lab Technol 1968;25:1–6. Berentsen S. Role of Complement in Autoimmune Hemolytic Anemia. Transfus Med Hemother 2015;42:303–10. Tjønnfjord E, Vengen OA, Berentsen S, Tjønnfjord GE. Prophylactic use of eculizumab during surgery in chronic cold agglutinin disease. BMJ Case Rep 2017, https://doi.org/10.1136/bcr-2016-219066. Dacie J. Auto-immune haemolytic anaemia (AIHA): cold-antibody syndromes I: idiopathic types: clinical presentation and haematological and serological findings. In: Dacie J, editor. The Haemolytic Anaemias, Vol. 3. London: Churchill Livingstone; 1992. p. 210–39. Jain MD, Cabrerizo-Sanchez R, Karkouti K, et al. Seek and you shall find–but then what do you do? Cold agglutinins in cardiopulmonary bypass and a single-center experience with cold agglutinin screening before cardiac surgery. Transfus Med Rev 2013;27:65–73. Litman GW. Sharks and the origins of vertebrate immunity. Sci Am 1996;275:67–71. Bendix BJ, Tauscher CD, Bryant SC, Stubbs JR, Jacob EK. Defining a reference range for cold agglutinin titers. Transfusion 2014;54:1294–7. Bessman JD, Banks D. Spurious macrocytosis, a common clue to erythrocyte cold agglutinins. Am J Clin Pathol 1980;74:797–800. Lyckholm LJ, Edmond MB. Images in clinical medicine. Seasonal hemolysis due to cold-agglutinin syndrome. N Engl J Med 1996;334:437. Chen EC, Loftus PD, Weber SC, et al. Autoimmune hemolytic anemia confers risk of thromboembolism that is not attributable to usual thrombosis risk factors: a longitudinal, retrospective cohort study using the “Stride” database. Blood 2016;128:1258. Hill A, Kelly RJ, Hillmen P. Thrombosis in paroxysmal nocturnal hemoglobinuria. Blood 2013;121:4985–96 [quiz 5105]. Broome C, Cunningham JM, Mullins M, et al. Incidence of thromboembolic events is increased in a retrospective analysis of a large cold agglutinin disease (CAD) cohort. Blood 2017;130:928. Dacie J. Treatment and prognosis of cold-antibody AIHA. In: Dacie J, editor. The Haemolytic Anaemias, Vol. 3. London: Churchill Livingstone; 1992. p. 502–20. Szczepiorkowski ZM, Winters JL, Bandarenko N, et al. Guidelines on the use of therapeutic apheresis in clinical practice—evidence-based approach from the Apheresis Applications Committee of the American Society for Apheresis. J Clin Apher 2010;25:83–177. Schwartz J, Padmanabhan A, Aqui N, et al. Guidelines on the use of therapeutic apheresis in clinical practice-evidence-based approach from the Writing Committee of the American Society for Apheresis: the Seventh Special Issue. J Clin Apher 2016;31:149–62. von Baeyer H. Plasmapheresis in immune hematology: review of clinical outcome data with respect to evidence-based medicine and clinical experience. Ther Apher Dial 2003;7:127–40. Silberstein LE, Berkman EM, Schreiber AD. Cold hemagglutinin disease associated with IgG cold-reactive antibody. Ann Intern Med 1987;106: 238–42. Barbara DW, Mauermann WJ, Neal JR, et al. Cold agglutinins in patients undergoing cardiac surgery requiring cardiopulmonary bypass. J Thorac Cardiovasc Surg 2013;146:668–80. Berentsen S, Beiske K, Tjønnfjord GE. Primary chronic cold agglutinin disease: an update on pathogenesis, clinical features and therapy. Hematology 2007;12:361–70. Berentsen S, Ulvestad E, Gjertsen BT, et al. Rituximab for primary chronic cold agglutinin disease: a prospective study of 37 courses of therapy in 27 patients. Blood 2004;103:2925–8. Schöllkopf C, Kjeldsen L, Bjerrum OW, et al. Rituximab in chronic cold agglutinin disease: a prospective study of 20 patients. Leuk Lymphoma 2006;47:253–60. Berentsen S, Randen U, Vagan AM, et al. High response rate and durable remissions following fludarabine and rituximab combination therapy for chronic cold agglutinin disease. Blood 2010;116:3180–4. Berentsen S, Randen U, Oksman M, et al. Bendamustine plus rituximab for chronic cold agglutinin disease: results of a Nordic prospective multicenter trial. Blood 2017;130:537–41. Rossi G, Paolini F, Fattizzo B, et al. Short course of bortezomib in anemic patients with refractory or relapsed cold agglutinin disease. A phase II prospective study by the Gimema Group. Blood 2017;130:2199. Crisp D, Pruzanski W. B-cell neoplasms with homogeneous cold-reacting antibodies (cold agglutinins). Am J Med 1982;72:915–22. Dacie J. Auto-immune haemolytic anaemia (AIHA): cold-antibody syndromes II: immunochemistry and specificity of the antibodies; serum complement in autoimmune haemolytic anaemia. In: Dacie J, editor. The Haemolytic Anaemias, Vol. 3. London: Churchill Livingstone; 1992. p. 240–95. Wouters D, Zeerleder S. Complement inhibitors to treat IgM-mediated autoimmune hemolysis. Haematologica 2015;100:1388–95.

S. Berentsen / Seminars in Hematology 55 (2018) 141–149

[92] Röth A, Hüttmann A, Rother RP, Dührsen U, Philipp T. Long-term efficacy of the complement inhibitor eculizumab in cold agglutinin disease. Blood 2009;113:3885–6. [93] Derhaschnig U, Gilbert J, Jager U, et al. Combined integrated protocol/basket trial design for a first-in-human trial. Orphanet J Rare Dis 2016;11:134. [94] Gertz MA, Qiu H, Kendall L, et al. ANX005, an Inhibitory antibody against C1q, blocks complement activation triggered by cold agglutinins in human disease. Blood 2016;128:1265. [95] Mastellos DC, Yancopoulou D, Kokkinos P, et al. Compstatin: a C3-targeted complement inhibitor reaching its prime for bedside intervention. Eur J Clin Invest 2015;45:423–40. [96] Risitano AM, Ricklin D, Huang Y, et al. Peptide inhibitors of C3 activation as a novel strategy of complement inhibition for the treatment of paroxysmal nocturnal hemoglobinuria. Blood 2014;123:2094–101.

149

[97] Grossi FV, Bedwell P, Deschatelets P, et al. APL-2, a complement C3 inhibitor for the potential treatment of paroxysmal nocturnal hemoglobinuria (PNH): phase I data from two completed studies in healthy volunteers. Blood 2016;128:1251. [98] Gregory GP, Opat S, Quach H, Shortt J, Tran H. Failure of eculizumab to correct paroxysmal cold hemoglobinuria. Ann Hematol 2011;90: 989–90. [99] Kelly RJ, Hill A, Arnold LM, et al. Long-term treatment with eculizumab in paroxysmal nocturnal hemoglobinuria: sustained efficacy and improved survival. Blood 2011;117:6786–92. [100] Aggarwal R, Sestak AL, D’Souza A, et al. Complete complement deficiency in a large cohort of familial systemic lupus erythematosus. Lupus 2010;19: 52–7.