P­RI­MER

­­­C­ry­og­lo­bu­li­na­em­ia­­­­­­­­­­­­
Dario Roccatello1*, David Saadoun2,3, Manuel Ramos-​Casals4,5,6, Athanasios G. Tzioufas7,
Fernando C. Fervenza8, Patrice Cacoub9, Anna Linda Zignego10 and Clodoveo Ferri11
Abstract | Cryoglobulinaemia refers to the serum presence of cryoglobulins, which are defined as
immunoglobulins that precipitate at temperatures <37 °C. Type I cryoglobulinaemia consists of
only one isotype or subclass of monoclonal immunoglobulin, whereas type II and type III are
classified as mixed cryoglobulinaemia because they include immunoglobulin G (IgG) and IgM.
Many lymphoproliferative, infectious and autoimmune disorders have been associated with
mixed cryoglobulinaemia; however, hepatitis C virus (HCV) is the aetiologic agent in most
patients. The underlying mechanism of the disorder is B cell lymphoproliferation and
autoantibody production. Mixed cryoglobulinaemia can cause systemic vasculitis, with
manifestations ranging from purpura, arthralgia and weakness to more serious lesions with skin
ulcers, neurological and renal involvement. This Primer focuses on mixed cryoglobulinaemia,
which has a variable course and a prognosis that is primarily influenced by vasculitis-​associated
multiorgan d­­­­­­­­­­­a­­­­­m­­­­­­­­a­­g­­e. I­n a­dd­it­ion, t­he u­nd­er­ly­ing associated disease in itself may cause
considerable mortality and morbidity. Treatment of cryoglobulinaemic vasculitis should be
modulated according to the underlying associated disease and the severity of organ involvement
and relies on antiviral treatment (for HCV infection), immunosuppression and immunotherapy ,
particularly anti-​CD20 B cell depletion therapies.

*e-​mail: dario.roccatello@
unito.it
https://doi.org/10.1038/
s41572-018-0009-4
Cryoglobulinaemia is defined as the presence of cryo­
globulins in the serum, which are immunoglobulins that
reversibly precipitate and form a gel when the tempera­
ture is <37 °C and re-​dissolve if the temperature rises
to >37 °C. According to Brouet’s classification1, three
subtypes of cryoglobulinaemia exist based on immuno­
globulin composition. Type I cryoglobulinaemia com­
prises single monoclonal immunoglobulins (most
commonly immunoglobulin M (IgM), rarely IgG or IgA),
whereas type II and type III are classified as mixed
cryoglobulinaemia because they include two types of
immunoglobulins (normally IgG and IgM). Type II
mixed cryoglobulinaemia comprises a combination of
monoclonal and polyclonal immunoglobulins (usually
IgMκ plus IgGκ or IgGλ), whereas type III mixed
cryoglobulinaemia comprises IgM and IgG, both poly­
clonal (Fig. 1; Table 1). Sensitive methodologies, such
as immunoblotting or 2D polyacrylamide gel electro­
phoresis, or advanced techniques of immunofixation
can be used to reveal the microheterogeneous com­
position of type II mixed cryoglobulins. Often, oligo­
clonal IgM or mixed polyclonal and monoclonal IgM
can be detected together with polyclonal IgG. This par­
ticular serological subset, known as type II–III mixed
cryoglobulina­emia (Fig. 1; Table 1), may be an inter­
mediate evolution from type III to type II mixed cryo­
globulinaemia. Importantly, the cryoglobulins found
in mixed cryoglobulinaemia are autoantibodies with
rheumatoid factor activity (that is, an antibody with the
ability to bind another antibody), which enables them to
form immune complexes; this ability is important in the
pathogenesis of cryoglobulinaemic vasculitis.
The fundamental mechanism contributing to cryo­
globulinaemia is aberrant autoantibody production by
B cells and B cell proliferation. Several underlying dis­
eases can facilitate this by interfering with normal B cell
function. The presence of type I cryoglobulins is always
linked to a B cell lymphoproliferative disorder (Table 1).
By contrast, mixed type II or type III cryoglobulinae­
mias are associated with systemic autoimmune diseases,
lymphoproliferative disorders and chronic infections
(Box 1), with hepatitis C virus (HCV) infection caus­
ing 80–90% of the cases of mixed cryoglobulinaemia2,3.
Autoimmune diseases that are associated with mixed
cryoglobulinaemia are primary Sjögren syndrome,
systemic lupus erythematosus (SLE) and rheumatoid
arthritis. When mixed cryoglobulinaemia is found in
the absence of well-​defined disease and therefore lacks
a known aetiology, the syndrome is designated essential,
mixed cryoglobulinaemia.
Strictly speaking, cryoglobulinaemia refers only to
the presence of cryoglobulins in a patient’s serum; how­
ever, this term is often used to refer to a systemic inflam­
matory syndrome that involves small-​to-medium vessel

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Author addresses
1
Center of Research of Immunopathology and Rare Diseases—CMID, Coordinating
Center of the Network for Rare Diseases of Piedmont and Aosta Valley, and Nephrology
and Dialysis Division (ERKnet member), Department of Clinical and Biological Sciences
of the University of Turin, and San Giovanni Bosco Hospital, Turin, Italy.
2
Sorbonne Universités, UPMC Université Paris, Département Hospitalo-​Universitaire
Inflammation-​Immunopathologie-Biotherapie (DHU i2B), Paris, France.
3
AP HP, Groupe Hospitalier Pitié-​Salpêtrière, Département de Médecine Interne et
Immunologie Clinique, National Center for Autoimmune and Systemic Diseases and for
Autoinflammatory Diseases, Paris, France.
4
Department of Autoimmune Diseases, ICMiD, Hospital Clínic, Barcelona, Spain.
5
Laboratory of Autoimmune Diseases Josep Font, IDIBAPS-​CELLEX, Barcelona, Spain.
6
Department of Medicine, University of Barcelona, Barcelona, Spain.
7
Department of Pathophysiology and Joint Academic Rheumatology Program, Medical
School of the National and Kapodistrian University of Athens, Athens, Greece.
8
Division of Nephrology and Hypertension, Mayo Clinic, Rochester, MN, USA.
9
Hôpital La Pitié-​Salpêtrière, Department of Internal Medicine and Clinical Immunology,
Paris, France.
10
Center for Systemic Manifestations of Hepatitis Viruses (MaSVE), Department of
Experimental and Clinical Medicine, University of Florence, Florence, Italy.
11
Rheumatology Unit, Medical School, University of Modena and Reggio Emilia,
Modena, Italy.
vasculitis caused by cryoglobulin-​containing immune
complexes. In this Primer, the terms ‘cryoglobulinae­
mia’ and ‘cryoglobulinaemic vasculitis’ are used to define
the presence of cryoglobulins (with or without clinical
symptoms) and the clinically apparent disorder (pur­
pura, arthralgia and/or arthritis, weakness, skin ulcers,
peripheral neuropathy, nephritis), respectively. Mainly
type II and type II–III, and less often type III, mixed
cryoglobulinaemia may lead to cryoglobulinaemic vas­
culitis. The majority of these cases of cryoglobulinae­
mic vasculitis are related to an HCV infection2,3 and, in
the case of type II mixed cryoglobulinaemic vasculitis,
to low-​grade proliferative B cell lymphomas2,4. Of the
available classification systems for vasculitis, the current
most widely used criteria set is that coined by the Chapel
Hill Consensus Conference5 and reviewed in 2012 (ref.6),
which is mainly based on anatomical distinctions of the
dominant vessels affected. Cryoglobulinaemic vasculitis
is included among the non-​anti-neutrophil cytoplasmic
antibody (ANCA)-associated, small-​vessel vasculitis,
with an immune complex pathogenesis, and, in the case
of HCV-​associated cryoglobulinaemic vasculitis, with a
known aetiology.
This Primer provides an update on the pathogenesis,
diagnosis and management of cryoglobulinaemia, with a
special focus on mixed cryoglobulinaemia and cryoglob­
ulinaemic vasculitis. On the basis of both academic and
clinical perspectives, the Primer aims to offer an outlook
on future research topics with the acknowledgement that
many of the currently established concepts may change
in upcoming years.
Epidemiology
Data of prevalence and incidence of cryoglobulinae­
mia in the general population are scarce because only
a few studies have addressed this issue. Globally, cryo­
globulinaemic vasculitis is considered a rare disease
(<5 cases per 10,000 individuals in the general European
and North American population), although prevalence
has been shown to be higher in some areas such as
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© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
the Mediterranean Basin7. There is an abundance of
data available on the underlying associated diseases,
which will be briefly summarized below in the context
of cryoglobulinaemia.
HCV-​related cryoglobulinaemia
The main aetiology of type II and type III mixed cryo­
globulinaemia is chronic HCV infection, accounting
for 80−90% of cases of mixed cryoglobuliaemia3,8–13.
HCV infection is a common problem, globally affect­
ing more than 184,000,000 people. The prevalence of
HCV infection varies depending on the geographical
region. HCV infection is highly prevalent in central
Asia, eastern Asia, the Middle East and North Africa,
infecting >3.5% of the population in these regions.
Southeast Asia, Central and South America, Australia,
the Caribbean, Oceania and sub-Saharan Africa
show moderate prevalence (1.5–3.5%), whereas in
Europe, prevalence varies from <1% in several western
European countries (for example, the United Kingdom,
Denmark, France, Germany, Sweden and Switzerland)
to 1–2% in the Mediterranean Basin and 2.5–3% in
eastern Europe14. High-​income Asia-​Pacific countries
(Brunei, Japan, South Korea and Singapore) and North
America are the regions with the lowest prevalence of
HCV (<1.5%)14,15.
In two large prospective studies, mixed cryoglobulin­
aemia was found in ~40–60% of patients infected with
HCV but only 5% of individuals with HCV infection
developed cryoglobulinaemic vasculitis16,17. Patients
with chronic HCV infection and cryoglobulinaemia
reportedly have higher incidences of cirrhosis and
higher fibrosis scores than those without cryoglob­
ulinaemia (after adjusting for age, sex and length of
infection) 15,18. However, no association was found
with advanced fibrosis on the basis of type and level of
circulating cryoglobulins.
The reported variation in the proportion of cases of
mixed cryoglobulinaemia and cryoglobulinaemic vas­
culitis that are attributable to HCV infection is mainly
due to population selection and lead time (the length of
time between the detection of cryoglobulinaemia and
its usual clinical presentation) biases and to the fact that
clinical assessment is not standardized, and the labora­
tory test for mixed cryoglobulinaemia is prone to false-​
negative results16,17. Lastly, cryoglobulinaemia seems to
correlate with the duration of HCV infection16.
Non-​HCV-related cryoglobulinaemia
In the 10–20% of cases of mixed cryoglobulinaemia
that are not associated with chronic HCV infection, the
main causes include other infectious diseases, B cell
malignancies and autoimmune diseases19. Specifically,
an association of cryoglobulinaemia with chronic
hepatitis B virus (HBV) infection has been suggested16.
However, only ~2% of cases of cryoglobulinaemic
vasculitis seems to be attributable to HBV infection20.
Occasionally, cryoglobulinaemic vasculitis may be
associated with HIV infection, especially in cases of
co-​infection with HCV21. Non-​HCV-related infectious
mixed cryoglobulinaemia is mainly caused by viruses,
bacterial pathogens or parasites (Box 1).
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Patients with active SLE and rheumatoid arthritis A French study of the underlying B cell disease in
may have circulating serum cryoglobulins; mixed 36 patients with type I cryoglobulinaemia found 13 patients
cryoglobulinaemia has been observed in ~10% of had a nonmalignant monoclonal gammopathy of undeter­
patients with SLE or rheumatoid arthritis; however, mined significance (MGUS) and 23 patients had a hae­
these patients do not usually have the same clinical course matological malignancy (12 patients with Waldenström
as patients with SLE or rheumatoid arthritis who have macroglobulinaemia, 6 patients with low-​grade non-​
cryoglobulinaemic vasculitis22. Approximately 5–20% of Hodgkin lymphoma, 4 patients with multiple myeloma
patients with primary Sjögren syndrome may have type II and 1 patient with chronic lymphocytic leukaemia)28.
cryoglobulinaemia23. Type II mixed cryoglobulinaemia is
currently considered one of the key prognostic factors inMechanisms/pathophysiology
primary Sjögren syndrome as it has been associated with The pathogenesis of cryoglobulinaemia depends on an
extraglandular involvement, the development of systemic only partially understood interaction between a host
vasculitis, B cell lymphoma and poor survival24. predisposition29 and environmental triggers, which
leads to aberrant B cell functions 9. In this model,
Associated haematological disorders pathogenetic cryoglobulinaemic responses depend
The presence of cryoglobulinaemia is associated with the on several factors, including lymphoproliferation
development of haematological disorders. The overall and/or chronic immune stimulation, resulting in the
risk of non-​Hodgkin lymphoma in patients with cryo­ production of monoclonal, oligoclonal or polyclonal
globulinaemia is ~35 times higher than in the general cryoglobulins, and immune complex precipitation fol­
population or 12 times higher if nonaggressive lym­ lowing the binding between cryoglobulins and their
phomas are excluded25. B cell non-​Hodgkin lymphoma target antigens in conjunction with an insufficient and/
includes marginal zone lymphoma, de novo or trans­ or defective clearance of cryo­globulin–target antigen
formed diffuse large B cell lymphoma and follicu­ immune complexes, which accumulate throughout the
lar lymphoma. Serum cryoglobulin levels >0.6 g per body and mediate disease. Animal models showed that
litre, the presence of cryoglobulinaemic vasculitis and cryoglobulins may physically obstruct vessels and/or
hypogammaglobulinaemia are independent variables mediate inflammatory vasculitis after immune com­
associated with the development of B cell non-​Hodgkin plex precipitation30. For many cryoglobulins, aggre­
lymphoma26. Individuals with HCV can develop non-​ gation and pathogenicity appear to depend on several
Hodgkin lymphoma after a long period of infection factors, including pH, weak noncovalent interactions,
(>15 years). Considering patients with B cell non-​ cryoglobulin concentrations and antibody-​specific
Hodgkin lymphoma, the proportion who develop trans­ conditions of temperature31.
formed diffuse large B cell lymphoma is considerably
higher in patients who are HCV-​positive (32%) than in HCV-​related cryoglobulinaemia
patients who are HCV-​negative (6%)27. The mechanism of cryoglobulin pathogenicity is best
As mentioned above, type I cryoglobulinaemia is described for HCV-​associated mixed cryoglobulinae­
always associated with a B cell proliferative disorder. mia. HCV is able to concomitantly infect B cells and
hepatocytes owing to the common expression of the
HCV entry receptor CD81 on the plasma membrane
$EGNN %JTQPKEKPHGEVKQPU
*%8 of both cell types3,20,32,33 (Fig. 2). Active HCV replication
N[ORJQRTQNKHGTCVKXG $EGNNN[ORJQRTQNKHGTCVKXGFKUGCUGU
FKUGCUGU #WVQKOOWPGFKUGCUGU
has been shown in CD19-positive B cells34; HCV RNA
'UUGPVKCNOKZGFET[QINQDWNKPCGOKC and HCV core and non-​structural NS3 proteins can
be detected in CD19-positive, but not CD19-negative,
peripheral blood mononuclear cells34,35. HCV replica­
6[RG+ 6[RG++ 6[RG+++



 tion has also been described in monocytes, peripheral
dendritic cells and macrophages36,37.
+I)
B cell proliferation and HCV infection. B cell prolifer­
+I/ ation is an important mechanism in the pathogenesis
/QPQENQPCN+I /QPQENQPCN+I
+I/κ 2QN[ENQPCN+I/ of cryoglobulinaemia and cryoglobulinaemic vascu­

+I/ +I) +I# RQN[ENQPCN+I  RQN[ENQPCN+I)
litis38–40. A multistep process supports the progression
from a simple serological alteration (cryoglobulinaemia)
/KZGFET[QINQDWNKPCGOKC
to clinical manifestations (cryoglobulinaemic vasculitis)
Fig. 1 | Classification of cryoglobulinaemia based on immune typing. Type I and ultimately to overt malignant B lymphoproliferation
cryoglobulinaemia is associated exclusively with B cell proliferative diseases; the serum of (such as non-​Hodgkin lymphoma)41,42. The discovery of
an individual with type I cryoglobulinaemia contains monoclonal immunoglobulin M (IgM), the HCV envelope affinity to a transmembrane protein,
and more rarely contains IgG, or IgA cryoglobulins. Type II mixed cryoglobulinaemia CD81 (ref.43), provided a cornerstone in the understanding
comprises serum immune complexes formed of monoclonal IgM and polyclonal IgG, and
of the mechanisms of HCV-​induced lymphoprolifer­
type III mixed cryoglobulinaemia comprises serum immune complexes formed of polyclonal
IgM and polyclonal IgG. Both type II and type III mixed cryoglobulinaemias are associated ation18,43. On the surface of B cells, CD81 creates a multi­
with hepatitis C virus (HCV) infection, autoimmune diseases or B cell proliferative diseases. protein complex with CD21 and CD19. When activated
Occasionally , an individual will present with mixed cryoglobulinaemia that has no known by the binding of HCV envelope, this multiprotein
aetiology , which is termed essential, mixed cryoglobulinaemia. In the figure, different complex regulates the polyclonal expansion of B cells.
coloured antibody shapes reflect different antibody clones. In addition, B cells are stimulated by the binding of an

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Table 1 | Types of cryoglobulinaemia, classification and associated diseases

Type of Composition of cryoprecipitates Main associated or underlying diseases
cryoglobulinaemia
Type I Monoclonal IgM (rarely IgG or IgA) Lymphoproliferative diseases, plasma cell dyscrasias,
multiple myeloma, Waldenström macroglobulinaemia,
MGUS, chronic lymphocytic leukaemia, B cell
non-Hodgkin lymphoma and hairy cell leukaemia
Type II Combination of monoclonal HCV (80–90% of cases) and other infections (such as HBV)
(usually IgMκ) and polyclonal
(usually IgGκ and IgGλ)
Type III Polyclonal IgM; polyclonal IgG HCV and other infections; autoimmune diseases
Type II–III Oligoclonal IgM; polyclonal IgG HCV and other infections, autoimmune diseases and
lymphoproliferative diseases
HBV, hepatitis B virus; HCV, hepatitis C virus; Ig, immunoglobulin; MGUS, monoclonal gammopathy of undetermined significance.
Adapted with permission from ref.156, BMJ Publishing Group.

antigen to the B cell receptor (BCR) on the cell surface, B cells from undergoing apoptosis, leading to an oligo­
resulting in polyclonal expansion44,45. clonal monotypic lymphoproliferation50. Mutations in
Although in vitro studies have shown that specific other oncogenes, such as MYC, and regulators of apop­
anti-​HCV antibodies can stimulate the BCR in B cells, tosis may be the important ‘missing link’ to our under­
it seems that CD81 activation by the HCV envelope standing of lymphomagenesis in the setting of chronic
protein induces naive B cell proliferation independently HCV infection.
from BCR stimulation. CD81-mediated activation of
naive (CD27-positive) B cells with subsequent differen­ Autoantibody production. Chronic stimulation by HCV
tiation to autoantibody-​producing memory cells could infection induces B cells to produce a variety of auto­
play a role in the development of lymphoproliferative antibodies51. This widespread production of autoanti­
disorders. In addition, chronic antigenic stimulation bodies supports the development of a number of immune
may lead to the overexpression of B cells (favouring manifestations associated with HCV infection, which are
certain clones) and support mechanisms of immune variably assembled in what is collectively termed ‘HCV
dysregulation that lead to the development of mixed syndrome’29,52,53. HCV syndrome may include clinical
cryoglobulinaemia and eventually the malignant trans­ manifestations beyond the characteristic picture of
formation sometimes observed in patients who are cryoglobulinaemic vasculitis, such as autoimmune
chronically infected with HCV45–49. thyroiditis, sicca syndrome (dryness of the exocrine
glands, particularly the eyes and mouth), thrombocyto­
B cell transformation. In addition to the induction of B penia, haemolytic anaemia, autoimmune diabetes and
cell proliferation and the reduction of the B cell activation pulmonary fibrosis20,29,38–49.
threshold, HCV infection can result in B cell transforma­ Clonal B cell populations are present in the liver and
tion45–49. Patients with HCV infection have clonal popu­ peripheral blood of patients with a chronic HCV infec­
lations of B cells that are predominantly IgM-​producing tion. Interestingly, clonally restricted B cells demon­
memory B lymphocytes, which express modestly hyper­ strate biased usage of the rheumatoid-​factor-encoding
mutated immunoglobulin genes45–49. Many of the same variable heavy1–69 and variable3–20 immunoglobulin
immunoglobulin idiotypes and restricted gene sequence gene segments, as do B cells isolated from the lymph
rearrangements are seen in both HCV-​positive non-​ nodes of patients with HCV-​associated non-​Hodgkin
Hodgkin lymphoma and cryo­globulinaemic vasculitis, lymphoma39,40,47. In patients with HCV-​related mixed
suggesting a shared pathophysiology. In addition, emerg­ cryoglobulinaemia, lymphoid infiltrates with cells
ing studies regarding the pathogenesis of HCV-​associated expressing oligoclonal or monoclonal IgM with rheu­
lymphomas have revealed evidence that HCV may have matoid factor activity can be detected in several organs,
mutagenic potential. B cells exposed to HCV in vitro had including the portal tracts of the liver, spleen and bone
up to a tenfold increase in mutations in the immuno­ marrow. Thus, mixed cryoglobulinaemia appears to be
globulin heavy chain gene and other sites. Furthermore, a crosslink between classic autoimmune disorders and
increased mutations are seen in HCV-​associated lym­ haematologic neoplasia (that is, B cell lymphoma)20,54–59.
phomas as compared with HCV-​negative lymphomas40. The constant stimulation of B cells induced by viral anti­
Among genetic mutations, the translocation t(14;18) of gens and the increased expression of lymphomagenesis-​
the anti-​apoptotic BCL2 gene (which encodes apopto­ related genes (particularly activation-​induced cytidine
sis regulator BCL-2) is the most common chromosomal deaminase, which is critical for somatic hypermuta­
rearrangement in lymphoid cancers, especially follicu­ tion60) lead to a polyclonal and later monoclonal expan­
lar lymphoma, a subtype of non-​Hodgkin lymphoma. sion of B cells (Fig. 2). These interactions ultimately
Of patients with chronic HCV infection, 35% have induce a lympho­proliferative disorder that may even­
evidence of the common chromosomal rearrangement tually evolve to B cell non-​Hodgkin lymphoma. Indeed,
t(14;18) translocation in their peripheral mononuclear among other haematologic malignancies, a strong asso­
cells39. This HCV-​dependent gene translocation prevents ciation has been found between HCV infection and

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© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.

large B cell lymphoma, marginal zone lymphoma and
lymphoplasmacytic lymphoma61.
In summary, HCV-​induced B cell lymphoprolifer­
ation and autoantibody production are probably the
direct result of the transformation of infected B cells but
also an indirect process arising from chronic antigenic
stimulation of a limited pool of autoreactive B cells.
Immune complex formation. The clinical symptoms of
cryoglobulinaemic vasculitis are caused by the deposi­
tion of immune complexes in small blood vessels and
associated endothelial injury. In HCV-​associated mixed
cryoglobulinaemia, the appearance of immune complexes
formed by circulating HCV particles, anti-​HCV poly­
clonal IgG and monoclonal IgM with rheumatoid factor
activity is sustained by a permanent clone of B cells trig­
gered by chronic HCV infection (see above). These cryo­
precipitable immune complexes are known to escape the
erythrocyte transport system owing to clonally restricted
IgM. The presence of IgM in the cryoprecipitable immune
complexes makes them able to induce complement
activation and consumption but unable to incorporate
complement fragments, including complement C3b
(a complement component, which favours the binding of
the immune complexes to the erythrocyte complement
receptor 1 (CR1))51. These immune complexes (Fig. 2)
remain free to circulate in the blood, not only owing
to escaping the erythrocyte transport system but also
because hepatic and splenic macrophages are unable to
process the immune complexes owing to HCV-​induced
abnormalities in the biogenesis of lyso­somal enzymes62.
In addition, this abnormality was clearly shown in cir­
culating monocytes62. Interestingly, investigations of
renal tissue samples using electron microscopy have
found monocytes that contain engulfed cryoglobulins63;
however, the exact role of these cells is unclear.
In cryoglobulinaemic nephritis, there is a phago­
cyte influx to the glomerulus (small blood vessels of
the kidney). Phagocytes attempt to remove deposited
cryoglobulins; however, they are not able to process
phagocytized cryoglobulins, and the entrapment of
cryoglobulins probably reflects ineffective cryoglobu­
lin clearance62. This mechanism potentially perpetuates
glomerular damage, as shown in a study using a mouse
model of cryoglobulinaemic membrano­proliferative
glomerulonephritis64 in which macrophage ablation
conferred protection from mesangial matrix expan­
sion and collagen accumulation (markers of nephritis)
without affecting cryoglobulin removal. This experi­
mental model suggests that macrophage recruitment
into the glomeruli plays a critical role in the progres­
sion of kidney injury64 instead of cleaning cryoglob­
ulins from the glomeruli; macrophage influx, vessel
infiltration and diapedesis are related to the amplifi­
cation of injury following immune complex deposi­
tion. Altered monocytes’ lysosomal enzymes (possibly
associated with HCV infection), including extracellu­
lar release of pro-​cathepsin D62, and/or the release of
danger-​associated molecular patterns (DAMPs) from
injured resident cells64 could reduce the innate func­
tion of macrophages to clear immune complexes via
immunoglobulin crystal­lizable fragment (Fc) gamma
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receptors. Mesangial matrix expansion and prolifera­
tion of glomerular cells can be sustained by the extra­
cellular activation of released pro-​cathepsin D62 or by
pro-​inflammatory cytokines released from DAMP-​
activated macrophages64. In addition to the classical
pathway of immune complex deposition, clonally
restricted IgM shares strong affinity for the glomeru­
lar matrix components, including fibronectin, opening
the possibility for an ‘in situ’ binding mechanism30 of
immune complexes to components of the kidney. The
pathogenetic role of cryoglobulins in inducing vas­
culitis is therefore related to both the recruitment of
leukocytes in the vessels and the deposition of immune
complexes, with activation of the complement system
and microvascular injury. The low level of complement 4
(C4), a diagnostic hallmark of cryoglobulinaemic
vasculitis, is related to both the complement system
activation with C4 binding to immune complexes and
the genetic polymorphisms of C4 in these patients65.
Non-​HCV cryoglobulinaemia
When compared with HCV-​related cryoglobulinaemia,
data on the pathogenetic mechanisms underlying the
development of mixed cryoglobulinaemia in the con­
text of other disorders (infectious or autoimmune) are
Box 1 | Infectious triggers of MC
Viruses
• Hepatitis C virus
• Hepatitis B virus
• Epstein–Barr virus
• Cytomegalovirus
• Hepatitis A virus
• HIV
• Adenovirus
• Parvovirus B19
Bacteria
• Various species of bacteria that cause endocarditis
• Streptococcus (causing diverse infections in humans)
• Brucella species (causative agents of brucellosis)
• Coxiella burnetii (causative agent of Q fever)
• Mycobacterium leprae (causative agent of leprosy)
• Borrelia burgdorferi (causative agent of Lyme disease)
• Treponema pallidum subspecies pallidum (causative
agent of syphilis)
Parasites
• Plasmodium species (causative agents of malaria)
• Leishmania species (causative agents of leishmaniasis)
• Toxoplasma species (causative agents of toxoplasmosis)
• Schistosoma species (causative agents of schistosomiasis)
• Echinococcus species (causative agents of echinococcosis)
Fungi
• Candida species (causative agents of candidiasis
infections)
• Coccidioides species (causative agents of
coccidioidomycosis)
MC, mixed cryoglobulinaemia.
5
P­r i­m e r
scarce. Type I cryoglobulinaemia sometimes induces
hyperviscosity of the serum that results from a high
concentration of monoclonal cryoglobulins owing to
the concomitant lymphoproliferative disorder.
In summary, a chronic infectious (most often HCV-​
related) B cell stimulation represents the most investi­
gated pathogenetic mechanism for cryoglobulinaemic
vasculitis. The mechanism of HCV-​related cryoglobulin­
aemia development relies on: the synthesis of IgM
with rheumatoid factor activity and the subsequent
formation of cryoprecipitable immunocomplexes; the
abnormal clearance kinetics with tissue deposition
*%8
%&
#PVK%&
$EGNN *GRCVQE[VG
$%4
2NCUOC %&
EGNN
+I/
2QN[ENQPCN+I/
V[RG+++ET[QINQDWNKPCGOKC Detection and characterization of cryoglobulins. The
1NKIQENQPCN+I/
V[RG++CPFQT+++ET[QINQDWNKPCGOKC nosis of cryoglobulinaemia and in particular of cryo­
/QPQENQPCN+I/ +I)
V[RG++ET[QINQDWNKPCGOKC
+%HQTOCVKQP
%4 .GWMQE[VG
FKCRGFGUKU
/CETQRJCIG
4$% 'PFQVJGNKCN
KPLWT[
%[VQMKPGU
2JCIQE[VQUKUCPFFGHGEVKXG
'T[VJTQE[VGVTCPURQTV )NQOGTWNCTFCOCIG RTQEGUUKPIQHET[QRTGEKRKVCVG+%U
U[UVGOGUECRG D[+%FGRQUKVKQP D[*%8KPHGEVGFOCETQRJCIGU
Fig. 2 | Mechanisms of HCV-​related cryoglobulinaemia vasculitis. B cells are targets
of hepatitis C virus (HCV) infection owing to cell surface expression of the CD81
receptor, which also allows hepatocyte infection. HCV-​induced B cell proliferation and
lowering of the B cell activation threshold causes widespread autoantibody production.
An HCV-​dependent gene translocation able to protect cells against apoptosis sustains
the oligoclonal monotypic lymphoproliferation that occurs in mixed cryoglobulinaemia.
Clonal B cells produce immunoglobulin M (IgM) that has rheumatoid activity
(autoantibody activity) towards anti-​HCV IgG; these components bind to each other and
to HCV and form mega-​immune complexes (ICs) that do not bind to the erythrocyte
transport system, therefore remaining free to circulate and saturating the ability of
phagocytes to remove ICs from the blood51. HCV infection blocks lysosomal enzymes in
phagocytes, which makes cells unable to digest cryoglobulins following phagocytosis62.
In the kidney , owing to the affinity of the mesangial matrix to the monoclonal IgM
component, cryoprecipitable ICs deposit in the glomeruli, where cytokine production
favours leukocyte diapedesis and endothelial injury. The anti-​CD20 monoclonal antibody
rituximab acts at the very first step of this cascade, blocking B cell proliferation and, thus,
IgM production, which is critical for both cryoglobulin production and deposition in the
glomeruli54. In the figure, different coloured antibody shapes reflect different antibody
clones. BCR , B cell receptor ; CR1, complement receptor 1; RBC, red blood cell. Adapted
with permission from ref.55, Taylor & Francis Ltd, http://www.tandfonline.com.
6 | Article citation ID: (2018) 4:11 www.nature.com/nrdp
© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
of these immune complexes combined with an inef­
fective cryo­globulin clearance by monocytes and/or
macrophages (which are implicated in perpetuating
glomerular damage); and a subclinical, underlying
lymphoproliferative disorder.
Diagnosis, screening and prevention
Diagnosis and screening
The diagnosis of cryoglobulinaemic vasculitis princi­
pally relies on the laboratory demonstration of serum
cryoglobulinaemia in association with characteristic
clinical signs and symptoms (Fig. 3). The production
of cryoglobulins is most often the consequence of an
underlying disorder that needs an aetiological evalua­
tion depending on the immunochemical determination
of the cryoglobulin components (Fig. 1; Table 1). In clin­
ical practice, cryoglobulin detection should be carried
out in all patients with or without HCV or HBV pre­
senting with typical manifestations of cryoglobulinae­
mic vasculitis, including orthostatic purpura, arthralgias
and/or arthritis, weakness, peripheral neuropathy, sicca
syndrome, urinary abnormalities, rheumatoid factor
seropositivity or low C4 level.
laboratory detection of circulating serum cryoglobulins
is always required for the correct classification and diag­
globulinaemic vasculitis66,67. Although internationally
accepted methodologies for cryoglobulin measurement
are still lacking, simple standardized indications are suf­
ficient for testing serum cryoglobulins66. In standard
serological tests to detect cryoglobulins, false-​negative
results are commonly caused by the cold precipitation
of antibodies, which can occur if the appropriate steps
are not followed. The first steps (blood sampling, clot­
ting and serum separation by centrifugation) must be
carried out shortly after the blood is sampled, and all
these initial steps must always be performed at 37 °C.
The next steps are cryocrit determination (that is,
determining the percentage of packed cryoglobulins
compared with total serum volume after centrifugation
at +4 °C), separation and washing of the cryoprecipitate
(that is, the precipitated cryoglobulins) and cryoglob­
ulin characterization by immunofixation. These steps
must be performed at 4 °C after at least 7 days of cold
serum storage. Serum containing cryoglobulins should
be tested for revers­ibility of the cryoprecipitate by
re-​warming an aliquot at 37 °C for 24 hours. This step
rules out false-​positive results caused by cryofibrinogen
or heparin-​precipitable proteins.
The immunoglobulin components of cryoprecipi­
tate can be identified and classified by immunoelectro­
phoresis or immunofixation at 37 °C to avoid possible
precipitation and/or loss of cryoglobulins. Alternatively,
immunoblotting or 2D polyacrylamide gel electrophor­
esis can be employed to classify serum cryoglobulins.
The characterization of serum cryoglobulins is manda­
tory for a definitive diagnosis of both cryoglobulinae­
mia and cryoglobulinaemic vasculitis66,68. The levels of
cryoglobulins in the serum do not usually correlate with
the severity and/or activity of the cryoglobulinaemic
 P­r i­m e r

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r2QUUKDNGRTGENKPKECN %NKPKECN 5GTQNQIKECN UGTWO/%U
 EQPFKVKQPYKVJQTYKVJQWV r2WTRWTC r/KZGF r#DUGPEGQHET[QRTGEKRKVCDNG+%
 4(CPFQTNQY% r9GCMPGUU  ET[QINQDWNKPU r4GRGCVET[QINQDWNKPFGVGEVKQP
r%CTGHWNENKPKECNGXCNWCVKQP r#TVJTCNIKCU r4(  CVFKHHGTGPVVKOGKPVGTXCNUVQ
 QHRQUUKDNGWPFGTN[KPI r.KXGTKPXQNXGOGPV r.QY%  EQPȰTOVJGFKCIPQUKUQH
 KPHGEVKQWU
*%8QT*$8 r4GPCNKPXQNXGOGPV 2CVJQNQIKECN  ET[QINQDWNKPCGOKEXCUEWNKVKU
 CWVQKOOWPGCPFQT r5MKPKPXQNXGOGPV r.GWMQE[VQENCUVKE r%CTGHWNENKPKECNGXCNWCVKQPQH
 JCGOCVQNQIKECNCPFQT r2GTKRJGTCN  XCUEWNKVKU  RQUUKDNGWPFGTN[KPIKPHGEVKQWU
HQT
 PGQRNCUVKEFKUGCUG  PGWTQRCVJ[ r$EGNNGZRCPUKQP  GZCORNG*%8*$8CWVQKOOWPG
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 VTGCVOGPV  PGQRNCUVKEFKUGCUG

Fig. 3 | The clinical, serological and pathological hallmarks of cryoglobulinaemic vasculitis. The presence of serum
mixed cryoglobulins (MCs) may be an isolated serological finding, the expression of a possible underlying disorder or the
hallmark of definite cryoglobulinaemic vasculitis. Cryoglobulinaemic vasculitis is a combination of serological alterations
(mainly MCs with rheumatoid factor (RF) activity and frequent low serum complement C4 (C4)) and typical clinico-​
pathological features (orthostatic purpura, leukocytoclastic vasculitis and multiple organ involvement). HBV, hepatitis B
virus; HCV, hepatitis C virus; IC, immune complex.

vasculitis53,66. An exception is the classical hypervis­
cosity syndrome that may appear in the presence of
very high cryocrit levels; it is characterized by abnor­
mally high serum and/or whole-​blood viscosity due to
increased circulating serum immunoglobulins and typ­
ical clinical symptoms, principally the triad of mucosal
bleeding, visual changes and neurological symptoms66.
Hyperviscosity syndrome typically occurs in type I IgM
cryoglobulinaemia. Importantly, circulating cryoglob­
ulins may interfere with a variety of laboratory tests
and have been associated with spurious quantification
of plasma proteins and erythrocyte sedimentation
rate, pseudo-​leukocytosis, pseudo-​thrombocytosis or
pseudo-​macrocytosis66. If a hyperviscosity syndrome
is suspected, the ‘capillary tube’ (that is, Ostwald tube)
viscometer is commonly used for laboratory testing69.
Viscosity is measured by the time required for a serum
or plasma sample to flow through a tube under the
influence of gravity. Viscous samples flow more slowly;
therefore, viscosity is proportional to time.
If type I cryoglobulinaemia is diagnosed by lab­
oratory testing, searching for an underlying B cell
lympho­prolife­rative disorder, mainly Waldenström
macroglobulinaemia, multiple myeloma or a mono­
clonal gammopathy, is mandatory70. More rarely, other
B cell diseases may be found, such as chronic lymphocytic
leukaemia, hairy cell leukaemia or B cell non-​Hodgkin
lymphoma. Laboratory detection of cryoglobulins com­
posed of IgM is suggestive of Waldenström disease,
whereas cryoglobulinaemia comprising IgG or IgA is
found in multiple myeloma or MGUS. Type II mixed
cryoglobulinaemia can be detected by electrophor­
esis and immuno­electrophoresis that reveal polyclonal
hypergammaglobulinaemia (usually IgG) with a mono­
clonal component (usually IgMκ) (Fig. 1). Type III cryo­
globulinaemia includes only polyclonal immunoglobulin
components. Type II and III cryoglobulinaemias are
more related to cryoglobulinaemic vasculitis than type I.
Diagnosis of cryoglobulinaemic vasculitis. The diagno­
sis of cryoglobulinaemic vasculitis is usually based on
the association of suggestive clinical vasculitis symp­
toms, laboratory detection of serum cryoglobulins and
decreased C4 serum level. Decreased serum levels of
early components of the complement cascade, such as
C4, are almost always seen in mixed cryoglobulinaemia
and are used as an indirect test. Moreover, patients with
serum cryoglobulins and cryoglobulinaemic vasculitis
should be tested for rheumatoid factor as it is almost
invariably found, especially in type II and type III
mixed cryoglobulinaemia70,71.
Figure 3 summarizes the possible clinical presentation
of patients with cryoglobulinaemia with and without overt
cryoglobulinaemic vasculitis. Type I cryoglobulinaemia
is a serological finding that is observable during the
course of various haematological disorders66, whereas
the detection of type II and III mixed cryoglobulinae­
mia represents the main laboratory hallmark of cryo­
globulinaemic vasculitis. However, in clinical practice,
mixed serum cryoglobulins may frequently be recorded

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P­r i­m e r
C D
E F
G H
Fig. 4 | Renal pathology in cryoglobulinaemic vasculitis. a | Light microscopy ( × 20)
extracapillary proliferation in the kidney , with features of fibrinoid necrosis (arrow) and
interruption of the capsule. b | Light microscopy ( × 40) showing a membranoproliferative
pattern with double contour appearance of the glomerular basement membrane
(arrow). c | Light microscopy ( × 20). Periodic acid–Schiff (PAS)-positive endoluminal
pseudothrombi (corresponding to cryoglobulin precipitates) (arrow) and reduplication
with cellular interposition of the glomerular basement membranes. d | Light microscopy
(x10). PAS stain shows diffuse membranoproliferative glomerulonephritis characterized
by duplication of glomerular basement membrane, interposition by mesangial cells
and especially mononuclear leukocytes and/or macrophages, subendothelial and
mesangial deposition of immune reactants, mesangial expansion and proliferation
with intracapillary leukocyte accumulation and endoluminal hyaline thrombi can
also be observed. e | Electron microscopy showing electron-​dense deposits
corresponding to cryoglobulin precipitates in the subendothelial space (white arrow).
Electron-​dense material with structured shape can also be observed in the capillary
lumen (yellow arrow). f | Immunofluorescence studies showing immunoglobulin M
with segmental accumulation (arrow) corresponding to the thrombi observed with
light microscopy.
(particularly in low amounts) as an isolated laboratory
finding in individuals without any typical symptoms of
cryoglobulinaemic vasculitis (see below)66. The diag­
nosis of cryoglobulinaemic vasculitis (Fig. 3) must be
based on serological findings combined with the clas­
sic clinical triad of purpura, arthralgia and weakness
and multiple organ involvement, which may manifest
as glomerulonephritis, peripheral neuropathy, haema­
tological alterations, endocrine alterations and hepatic
alterations; specific diagnostic procedures (for example,
biopsy of renal, liver or bone marrow tissue and elec­
tromyography) may be appropriate29,66,72. For example,
the presence of urinary abnormalities such as micro­
haematuria and/or proteinuria is helpful to diagnose
8 | Article citation ID: (2018) 4:11 www.nature.com/nrdp
© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
cryoglobulinaemic vasculitis with renal involvement,
but confirmation requires renal biopsy29 (Fig. 4). Isolated
cryoglobulins may be detected in the early stages of the
disease or during clinical remission of the cryoglobulin­
aemic vasculitis. In addition, clinically overt cryoglobu­
linaemic vasculitis without detectable cryoglobulinaemia
may be rarely observed (Box 2). This condition some­
times occurs transiently during the natural history
of the disease and is correlated with the great variability
in the percentage of cryoprecipitable immune complexes
(that is, the amount of cryoglobulins in the serum of any
given patient at a point in time)66. If cryoglobulinaemic
vasculitis is strongly suspected and serum cryoglobulins
cannot be detected by routine methods, a precipitation
technique in hypoionic medium that induces an increase
in cryoprecipitation could represent an additional tool to
detect very low levels of cryoglobulins73.
Differential diagnosis. Patients with suspected cryo­
globulinaemic vasculitis can present with symptoms
that overlap with a variety of conditions; therefore, the
differen­tial diagnosis between cryoglobulinaemic vas­
culitis and non-​organ-specific autoimmune disorders is
often necessary for the correct management of disease.
HCV-​related and non-​HCV-related cryoglobulinaemias
frequently show clinico-​pathological features that over­
lap with a number of other autoimmune and/or neoplas­
tic disorders, particularly primary Sjögren syndrome,
rheumatoid arthritis, membranoproliferative glomeru­
lonephritis, thyroiditis and B cell lymphomas52,53,74. Thus,
careful evaluation of patients is necessary for the cor­
rect classification and diagnosis of the individual clinical
entity (Fig. 5).
Patients with cryoglobulinaemic vasculitis usu­
ally show symptoms of a mild, generally non-​erosive
arthritis53,74. However, some patients with HCV-​associated
cryoglobulinaemic vasculitis may develop symmet­
rical, erosive polyarthritis suggesting an overlapping
cryoglobuli­naemic vasculitis — rheumatoid arthritis syn­
drome. In these cases, the detection of serum anti-​cyclic
citrullinated peptide antibodies (that is, markers of classi­
cal seropositive rheumatoid arthritis) can be employed for
the differential diagnosis of the two conditions20.
The relationship between sicca syndrome, primary
Sjögren syndrome and cryoglobulinaemic vasculitis,
particularly in patients with a chronic HCV infection,
is another intriguing issue52,74. Dryness of the exocrine
glands, particularly of the eyes and mouth, is referred
to as ‘sicca syndrome’ when there is no evidence of
autoimmune-​positive serum tests (antinuclear anti­
bodies and antibodies against extractable nuclear
antigens) found together with a lymphocyte infiltration
of the involved glands. Quite a number of patients with
cryoglobulinaemic vasculitis have the symptoms of
sicca syndrome, but only a few cases of cryoglobu­
linaemic vasculitis meet the current criteria for the
official classification of primary Sjögren syndrome75.
Importantly, cryoglobulinaemic vasculitis, sicca
syndrome and primary Sjögren syndrome may share
several symptoms, which complicates the diagnosis:
xerostomia (dry mouth) and/or xerophthalmia (dry
eyes), arthralgias, purpura, serum rheumatoid factor

and cryoglobu­lins as well as an increased risk of devel­
oping a B cell lymphoma52,53,74. Thus, careful patient clin­
ical and laboratory assessment may be needed in order
to make a correct diagnosis. Specific histopathological
alterations of the salivary glands and seropositivity for
specific autoantibodies (mainly the antibodies against
extractable nuclear antigens anti-​Ro/SSA and/or anti-​
La/SSB) are hallmarks of primary Sjögren syndrome.
Unlikely primary Sjögren syndrome, cryoglobulinae­
mic vasculitis with or without isolated sicca syndrome
is characterized by HCV infection in the vast majority
of patients23,67 (Fig. 5). Differential diagnosis may prove to
be more difficult in patients who are HCV-​negative with
symptoms of both primary Sjögren syndrome and cryo­
globulinaemic vasculitis. These patients would be best
classified as having an overlap syndrome52,74. Overlap
syndrome is often characterized by low serum levels
of anti-​Ro/SSA and/or anti-​La/SSB along with a high
prevalence of mixed cryoglobulinaemia, low serum C4,
systemic autoimmune manifestations and complicating
lymphomas, which may be responsible for more severe
outcomes. It is a matter of current discussion if, in clin­
ical practice, the cryoglobulinaemic vasculitis–primary
Sjögren syndrome overlap syndrome can be considered
a separate entity with relevant implications on patient
monitoring, treatment and prognosis52,74.
Management
Cryoglobulinaemic vasculitis can be successfully treated
with a combination of antiviral treatment for underlying
HCV infection and/or rituximab (anti-​CD20 monoclo­
nal antibody that triggers cell death of B cells). However,
cryoglobulinaemic vasculitis remains a challenging dis­
ease to manage owing to severe organ-​specific involve­
ments and occasionally life-​threatening presentations76.
Studies reporting on the effects of treatment showed
mortality for cryoglobulinaemic vasculitis of 8–15%20,77.
In a cohort of 151 patients with HCV-​associated cryo­
globulinaemic vasculitis, the survival at 1, 3, 5 and
10 years was 96%, 86%, 75% and 63%, respectively78.
The most common causes of death in cryoglobulinae­
mic vasculitis are infection, HCV-​related end-​stage liver
disease, cardiovascular disease and more rarely vasculitis
and lymphoma and/or neoplasia. Baseline factors asso­
ciated with poor prognosis are the presence of severe
liver fibrosis associated with HCV infection, involve­
ment of the nervous system and involvement of the kid­
neys and/or the heart. In a multivariate analysis, severe
Box 2 | Understanding the presence or absence of serum cryoglobulins
The presence of serum mixed cryoglobulins may be an isolated serological finding,
the expression of a possible underlying disorder or the hallmark of definite
cryoglobulinaemic vasculitis. An incomplete cryoglobulinaemic vasculitis (that is, the
typical clinico-​pathological features of the disease in the absence of detectable serum
mixed cryoglobulins) can be observed during the natural course of the disease.
In particular, isolated serological alterations may be often detected in the early stages
of the disease or during clinical remission of cryoglobulinaemic vasculitis. The absence of
serum cryoglobulins in patients with overt cryoglobulinaemic vasculitis may be a
transient biological phenomenon owing to the wide variability in the percentage of
cryoprecipitable immune complexes at the time of measurement or, less frequently,
to the switching from ‘benign’ B cell lymphoproliferation to malignant lymphoma.
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P­r i­m e r
liver fibrosis (HR 10.8) and the severity of vasculitis
(HR 2.49) were statistically significantly associated with
poor prognosis78. Furthermore, in a cohort of patients
with non-​HCV-cryoglobulinaemic vasculitis, 18 out of
133 (13.5%) died, primarily owing to sepsis. In addition,
another multivariate analysis in patients with non-​HCV-
related cryoglobulinaemic vasculitis found that age
>60 years (OR 1.06) and renal involvement (OR 5.20)
were independently associated with death79.
The natural history of cryoglobulinaemic vasculitis
is not predictable and strongly depends on concomitant
diseases and complications as well as response to treat­
ment. Death usually occurs after a prolonged course of
vasculitis, often lasting years. Morbidity due specifically
to cryoglobulinaemic vasculitis may also be impor­
tant. The use of antiviral agents against HCV infection
(especially sofosbuvir-​based therapy) is associated with
better prognosis in patients with HCV-​associated cryo­
globulinaemic vasculitis80. The careful monitoring of
life-​threatening complications (mainly nephropathy,
widespread vasculitis and B cell lymphoma or other
malignancies) should be carried out in all patients with
cryoglobulinaemic vasculitis.
Renal manifestations
Prognosis. When kidney involvement is suspected (for
example, a patient presents with urinary abnormali­
ties, with or without decreased glomerular filtration
rate), renal biopsy is recommended 29 (Fig. 4). Renal
involvement is one of the most harmful complications
of HCV-​associated cryoglobulinaemic vasculitis and
may severely affect the clinical outcome of a patient.
In 5% of patients with renal involvement, acute oliguric
kidney failure is the first indicator of kidney disease.
Hypertension is common, affecting more than 50% of
patients with cryoglobulinaemic vasculitis, and may be
severe. The severity of hypertension may mirror the
severity of the kidney disease81. In many patients, kidney
disease shows an indolent (slow) course, and kid­
ney failure requiring chronic dialysis is rare (<10%).
Patients with cryoglobulinaemic nephritis have poor
prognosis, mainly because of a high incidence of
comorbidities such as infectious diseases, end-​stage
liver disease and cardio­vascular diseases. The overall
10-year survival after a diagnosis of cryoglobulinae­
mic vasculitis ranges from 50–90% in the case of renal
involvement20,29,81. In a cohort of 146 patients with cryo­
globulinaemic nephritis, cardiovascular disease was the
cause of death in >60% of patients29. Older age, higher
serum creatinine levels and increased proteinuria
at diagnosis are associated with the development of
kidney failure and death29.
Treatment. Patients with renal involvement should be
treated according to two broad principles. First, immuno­
suppressive therapy should be provided as initial therapy
in those patients identified as having a rapidly progres­
sive, organ-​threatening or life-​threatening course, regard­
less of the aetiology of the cryoglobulinaemic vasculitis.
This immunosuppressive therapy usually involves a short
course of glucocorticoids (to suppress inflammation)82–84
and/or treatment with rituximab (to deplete B cells)54,85–95
9
P­r i­m e r

(Fig. 2). Cyclophosphamide (a drug used in chemotherapy) plasma in patients at risk of bleeding (for example, fol­
can be used to suppress the immune system; however, lowing kidney biopsy). Owing to the high molecular
rituximab is preferred as it is less toxic. In some patients, mass of cryoglobulins, a double filtration technique can
e\specially those with hyperviscosity syndrome, plas­ be very effective to remove cryoglobulins from plasma
mapheresis may be used to reduce the concentration of during plasma exchange. Plasma exchange does not pre­
serum cryoglobulins96–98. vent the formation of new cryoglobulins, and as such it
Patients with rapidly progressive glomerulonephritis should be combined with rituximab to eliminate B cell
should receive one to three doses of intravenous methyl­ clones that produce cryoglobulins92–94.
prednisolone, followed by oral prednisone. The dose is Second, all patients should receive therapy directed
quickly tapered until it is discontinued. This rapid taper against the underlying aetiology of the mixed cryoglob­
is an attempt to reduce infection complications asso­ ulinaemia. Patients with chronic HCV infection should
ciated with immunosuppression82–84. In cases in which receive antiviral therapy (Box 3), whereas patients with
rituximab therapy is unavailable, fails to produce a an underlying lymphoproliferative or autoimmune
clinical response or is not tolerated, cyclophosphamide disorder should receive appropriate disease-​specific
therapy can be used as an alternative. therapy. In patients with HCV, the choice of specific
If plasma exchange is considered, it should be used antiviral drugs will depend upon the degree of kidney
early in the course of treatment and performed daily function. Direct-​acting antiviral, interferon-​free regi­
for 10 to 14 sessions, or 3 exchanges per week for mens are now the therapy of choice on the basis of the
2–3 weeks. The plasma replacement fluid can be a solu­ improved safety profiles and efficacy of these drugs and
tion of albumin, which should be warmed to prevent the high rates of clinical response (Box 3). The duration
precipitation of circulating cryoglobulins, or fresh frozen of antiviral therapy will depend on the HCV genotype,
the degree of liver fibrosis and hepatic function, prior
response to treatment, the viral load (in some cases)
HCV infection
and tolerance of treatment.
HCV-related sicca HCV-related Antiviral therapy can be delayed for 1–4 months in
syndrome arthritis patients with severe renal disease who require initial
• Mild salivary gland Cryoglobulinaemic • Often non-erosive immunosuppressive therapy. However, in patients with
involvement vasculitis oligo-arthritis
• HCV+ • Mixed • HCV+
cryoglobulinaemic vasculitis associated with infection
• Anti-Ro/SSA cryoglobulin+ • RF± with HIV or HBV, antiviral therapy should always be ini­
and/or La/SSB– • HCV+ • Anti-CCP– tiated before or concurrently with immunosuppressive
• RF± • Low serum C4 • Cryoglobulin± therapy, as these patients are at high risk of enhanced
• Anti-CCP– • Anti-CCP– • Normal C3
viral replication as a result of rituximab or cyclophos­
• Cryoglobulin± • Multiple organ and/or C4
• Normal C3 and/or C4 involvement phamide therapy93,94. In particular, rituximab should not
be used in patients with active hepatitis flares, as fatal
B-NHL fulminant hepatitis has been reported99,100.
Primary SS Rheumatoid The frequency of clinical and laboratory moni­
• Salivary gland • RF+ arthritis toring of patients will depend on the activity of
involvement • Anti-CCP– • Erosive • Anti-CCP+
• Anti-Ro/SSA • Cryoglobulin± polyarthritis • Cryoglobulin± cryoglobulinaemia-​associated disease, the treatment
and/or La/SSB+ • Normal C3 • HCV– • Normal C3 regimen, comorbidities and tolerance of therapy. In
• HCV– and/or C4 • RF± and/or C4 general, patients should have a complete blood count,
electrolyte panel, serum creatinine, liver function test­
Fig. 5 | Clinical overlap between HCV-​related conditions and autoimmune diseases. ing and blood glucose tests weekly during the first
There is a frequent clinical overlap between some possible hepatitis C virus (HCV)-related 2 weeks of antiviral therapy and monthly thereafter.
diseases and certain autoimmune conditions; thus, a differential diagnosis is very difficult, However, patients with cirrhosis and portal hyperten­
requiring a careful clinico-​laboratory assessment of patients suspected of these conditions. sion or a poor degree of kidney function will require
In particular, cryoglobulinaemic vasculitis, primary Sjögren syndrome (primary SS) and more frequent monitoring owing to the higher risk of
rheumatoid arthritis show a clinico-​pathological overlap regardless of concomitant HCV
side effects. All patients with cryoglobulinaemic vas­
infection; moreover, HCV infection may be associated with sicca syndrome and/or arthritis.
These conditions may be correctly classified by taking into account the following culitis who are receiving immunosuppressive therapy
considerations: primary SS shows a typical histopathological pattern of salivary gland should receive prophylaxis against opportunistic infec­
involvement and specific autoantibodies (anti-​Ro/SSA and/or anti-​La/SSB), which are tions, such as Pneumocystis pneumonia, as well as age-​
rarely found in patients with cryoglobulinaemic vasculitis; conversely , cutaneous appropriate immunizations, ideally several weeks before
leukocytoclastic vasculitis, visceral organ involvement (glomerulonephritis and/or the initiation of immunosuppression.
hepatitis), low serum complement C4 (C4) and HCV infection are typically found in
cryoglobulinaemic patients. The erosive symmetrical polyarthritis and serum anti-​cyclic Extrarenal manifestations
citrullinated peptide antibodies (anti-​CCP) of classical rheumatoid arthritis are absent in Treatment should be modulated according to the pre­
HCV-​related cryoglobulinaemia, which is usually characterized by mild, non-​erosive joint dominant aetiopathogenic mechanism — that is, vascu­
involvement. The serum rheumatoid factor (RF), detectable in both HCV-​positive subjects
litis (most common) or hyperviscosity (a rare syndrome
and other autoimmune diseases, is a less helpful diagnostic finding. Finally , ‘indolent’ B cell
neoplasias are a frequent underlying disorder of cryoglobulinaemic vasculitis; however, seen in type I cryoglobulinaemia) and disease severity76.
overt B cell non-​Hodgkin lymphoma (B-​NHL) may complicate cryoglobulinaemic vasculitis. The best example of how therapeutic approaches should
Malignant B-​NHL may be suspected on the basis of clinical symptoms, sudden variation of be aetiologically driven is the essential role that the new
typical laboratory parameters (cryoglobulin disappearance and abnormally high serum direct-​acting antiviral HCV therapies (Box 3) currently
C4 levels) and pathological and/or instrumental investigations. play in the treatment of extrahepatic disease101.

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© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.

Box 3 | The emerging role of direct antiviral agents
Interferon-​based antiviral therapy showed positive effects on several
hepatitis C virus (HCV) infection extrahepatic manifestations131–134;
however, these drugs have toxic side effects and limited efficacy135.
The introduction of direct antiviral agents (DAAs) that directly target
non-structural viral proteins involved in HCV replication has led to a
breakthrough in the therapeutic approach to HCV infection. HCV
non-structural protein 3 (NS3) inhibitors target the viral protease and
were introduced in 2011. These small-​molecule drugs block the catalytic
site of NS3, preventing cleavage of the viral poly-​protein and therefore
preventing HCV replication. This first generation of DAAs (telaprevir
and boceprevir) were administered with interferon-​based therapy and
ribavirin (a synthetic guanosine nucleoside that interferes with the
synthesis of viral mRNA); after this, new generations of DAAs were
introduced, including NS5A and NS5B inhibitors that allowed
interferon-free treatment (see the list below). Current therapy is based
on various interferon-​free DAA combinations with short therapy duration
(usually 8–24 weeks) and minimal side effects136 as well as antiviral
efficacy approaching 100%136.
Interferon-​free DAA treatments were characterized by low rates of
adverse events and a high clinical response rate, together with an
improvement of antiviral effectiveness80,137–142. Interferon and ribavirin
have limitations in patients with renal failure; thus, the availability of
interferon-​free and ribavirin-​free therapies allows the early, combined
use of antiviral therapy in HCV-​related cryoglobulinaemic vasculitis
with renal involvement143. Limited, although positive, data exist on the
effect of interferon-​free regimens134,138,139,144. A recent study evaluated
the effectiveness and tolerance of an all-​oral interferon-​free and
Haematological cryoglobulinaemia. The main therapeu­
tic goal must be the cure of the underlying haematologi­
cal disease, which are predominantly B cell neoplasms.
The most common neoplasias are multiple myeloma
(predominantly associated with type I cryoglobulinae­
mia and hyperviscosity) and B cell lymphoma (more
frequently related to type II or III mixed cryoglobulin­
aemia and cryoglobulinaemic vasculitis). Treatment of
the underlying monoclonal B cell proliferative disorder
has been associated with improvement and/or stabiliza­
tion of cryoglobulinaemic symptoms in most patients
with type I cryoglobulinaemia102, although a decrease
of serum cryoglobulins was achieved in only half the
patients. Alkylating agents such as cyclophosphamide,
bortezomib (a proteasome inhibitor) and rituximab-​
based regimens are the main therapeutic options70,102,103.
Patients presenting with symptomatic hyperviscos­
ity will require urgent therapeutic intervention using
plasma exchange or plasmapheresis (see above) to
remove cryoglobulins from the circulation.
HCV-​related cryoglobulinaemic vasculitis. In HCV
patients, pathogenic damage is related to vasculitis.
A case-​by-case stratification according to the severity
of vasculitic involvement (mild, moderate, severe or
life-​t hreatening presentations) is recommended 104.
Patients with mild vasculitic involvement might be
treated with the new antiviral agents alone (Box  3),
whereas those with moderate to severe vasculitis may
require a short-​term course of glucocorticoids in order
to subdue vasculitic-​induced damage quickly as a bridge to
antiviral therapies2,101. Intravenous pulses of methylpred­
nisolone and immunosuppressants should be used in
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ribavirin-​free regimen with sofosbuvir plus daclatasvir in patients with
cryoglobulinaemic vasculitis. The incidence of complete remission from
cryoglobulinaemic vasculitis at 3, 6 and 12 months was 85%, 90% and
90%, respectively. A sustained virological response at week 12 after
treatment was achieved in all cases. The disappearance of cryoglobulin
was shown in 50% of cases80, suggesting that cryoglobulin-​producing
B cell clones may persist even after HCV suppression.
NS3–NS4A inhibitors (protease inhibitors)
• Paritaprevir–ritonavir
• Grazoprevir
• Glecaprevir
• Voxilaprevir
• Telaprevir
• Boceprevir
NS5A inhibitors
• Velpatasvir
• Ombitasvir
• Elbasvir
• Pibrentasvir
• Daclatasvir
NS5B inhibitors
• Sofosbuvir
• Dasabuvir
life-​threatening presentations, including multiple and/or
extensive cutaneous ulcers (Fig. 6), severe neuropathy,
central nervous system involvement, alveolar haemor­
rhage or gastrointestinal ischaemia, in combination with
plasma exchange105. The most promising glucocorticoid-​
free, non-​antiviral therapeutic approach to HCV-​related
cryoglobulinaemia is based on the use of B cell-​depleting
agents such as rituximab106. B cell-​depleting agents show
far more evidence of efficacy compared to other options
(glucocorticoid and chemotherapeutic agents), not only
regarding the number of treated patients (>500 patients)
but also regarding the excellent results of small con­
trolled trials, even in patients with complications such
as cirrhosis76.
When separately taking into account each clinical
manifestation of cryoglobulinaemic vasculitis, no ran­
domized clinical trial is available investigating the best
treatment on the basis of organ involvement. However,
B cell depletion has been seen to be an effective tool for
the management of peripheral neuropathy associated
with cryoglobulinaemic vasculitis, with improvement of
symptoms and electrophysiological improvement within
6 months after treatment95,107 as well as improvement of
the symptoms and signs of glomerulonephritis54,55,108
and of skin involvement91,108. Overall, specific organ-​by-
organ data confirm the efficacy of rituximab in 85% of
patients with cutaneous ulcers, in 79% of patients with
arthralgia, in 77% of patients with weakness and in 67%
of patients with peripheral neuropathy91.
Some patients with a history of HCV infection that
was successfully treated with antiviral therapy, and
undetect­able viral load, may present with the clinical
manifestations of cryoglobulinaemia; this is likely the
11
P­r i­m e r

%NKPKECN 2CVJQNQIKECN for inducing clinical remission, based on high-​dose

glucocorticoid and cyclophosphamide, are derived
from therapeutic strategies employed in other sys­
temic vasculitides. Azathioprine (an immunosuppres­
sive medication belonging to the purine analogue and
antimetabolite family of medication) and mycophenolate
mofetil (an immunosuppressant, which is a reversible,
noncompetitive inhibitor of inosine-5ʹ-monophosphate
dehydrogenase, an enzyme essential to the de novo
synthesis of guanosine-5ʹ-monophosphate) are mainly
used as remission maintenance agents. However, accord­
ing to recent data112,113, the use of glucocorticoid and
rituximab (instead of cyclophosphamide) may now
be considered the first-​line therapeutic approach in
patients with severe non-​infectious cryoglobulinaemic
vasculitis.

Quality of life
Only one small study has specifically analysed health-​
related quality of life (HRQoL) in patients with cryo­
globulinaemia. The study evaluated 15 patients with
severe HCV-​related cryoglobulinaemic vasculitis using
Fig. 6 | Skin ulcers in severe cryoglobulinaemic vasculitis. Left panels: severe vasculitic the Medical Outcomes Study 36-item Short Form
ulcerative lesions located at the ankle and the leg. Right panels: skin histology Health Survey (SF-36)114. Lower scores were observed
(hematoxylin and eosin (H&E) staining (× 20)). Cryoglobulinaemic vasculitis with fibrinoid for all the physical and mental domains in this group
necrosis of the capillary walls with pyknotic nuclei and nuclear debris of granulocytes than in the control group. However, these scores sig­
and mononuclear cell infiltrates.
nificantly improved following treatment with rituxi­
mab. At present, no studies have been carried out in
result of expanded B cell clones or underlying lymphoma patients with non-​HCV cryoglobulinaemia, and the
that have developed as result of the HCV infection. main HRQoL studies that may be applicable to cryo­
These patients should be treated with rituximab. globulinaemia involved unselected patients with
chronic HCV infection. A recent meta-​analysis of the
Non-​HCV infectious cryoglobulinaemic vasculitis. Non-​ prevalence, patient quality of life and financial burden
HCV infectious cryoglobulinaemic vasculitis is a fairly of chronic HCV infection in the United States115 clearly
rare clinical condition that has been reported in several supports the hypothesis that HCV infection has nega­
international studies19,109,110. Patients are treated with tive effects on the overall physical and mental health of
the most appropriate specific anti-​infectious therapy as patients. A large number of patients with chronic HCV
first-​line therapy, in association with short-​term gluco­ infection present with chronic pain (including fibro­
corticoid regimens, plasma exchange and/or rituximab myalgia) and/or chronic fatigue, which are considered
in the most severe vasculitic presentations. Patients who the main culprits of the reported reduced HRQoL116,117.
are refractory to treatment or who relapse are mainly These chronic symptoms are closely related to the onset
affected by an underlying HBV infection, which is the of sleep disorders, depression and other neurocognitive
main non-​HCV infectious aetiology of cryoglobulinae­ alterations, which are independent of the severity of
mia19. The therapeutic goal for HBV patients should be the liver disease, viral genotypes or HCV replication
eradication of HBV DNA (a marker of active chronic rates118. These chronic manifestations of HCV infec­
infection) using antiviral agents (entecavir, interferon-​ tion typically occur in the absence of signs of structural
based agents, adefovir or lamivudine)110. The successful brain damage observed using neurological imaging
use of rituximab combined with antiviral agents has also techniques, although a few studies have reported some
been reported19. However, relapses are frequent, with a metabolic or microstructural changes118. Other features
reported mortality of about 25% in patients with chronic may worsen the psychological status of patients with
HBV infection and cryoglobulinaemic vasculitis110. HCV infection, including low socio-​economic status,
HBV or HIV co-infections, discrimination and/or lim­
Autoimmune cryoglobulinaemia. Cryoglobulinaemia ited access to adequate health care74. The use of the new
that is unrelated to underlying haematological or interferon-free and ribavirin-​free direct-​acting anti­
infectious processes should be considered a primary viral regimens for HCV infection has resulted in a better
vasculitic disease and is associated in some cases with experience for patients and increased work productiv­
other systemic autoimmune diseases (mainly primary ity following treatment119. Personalized environmental
Sjögren syndrome)109,111. The therapeutic approach is and individual psychological assessment of patients
mostly based on the use of conventional immunosup­ with HCV is mandatory in order to establish a com­
pression and/or B cell depletion in an effort to suppress prehensive counselling approach to manage the main
the clonal B cell expansion that is responsible for the HRQoL items74. No data are available on the restricted
synthesis of cryoglobulins. Traditional approaches subset of non-​HCV-related cryoglobulinaemia.

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© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.

Outlook
Cryoglobulinaemic vasculitis as a distinct multiorgan
disorder was first described in 1966 (ref.120). The dis­
covery of the striking association with HCV infection
25 years later3,10,121,122 profoundly affected both our
understanding of the disease’s pathogenesis and thera­
peutic strategies. Although a minority of patients were
still classified as having non-​HCV-relat\ed cryoglobu­
linaemia, identifying the main triggering factor would
represent a definite advancement. However, several
etio­pathogenetic issues remain to be clarified. First, it is
still unclear whether HCV infection represents a simple
triggering factor or whether it also contributes to the
self-​perpetuating mechanism of the disease. Moreover,
the natural course of type II or type III mixed cryoglobu­
linaemia may develop with different clinical phenotypes
among patients, suggesting a multifactorial and multi­
step process. Thus, the role of possible genetic and/or
environmental cofactors remains under investigation.
The actual role of B cell lymphoproliferation that
represents an underlying mechanism of the disease is
another important issue. In particular, in patients with
HCV-​related mixed cryoglobulinaemia, the persistence
of benign B lymphocyte proliferation may explain the
paradoxical effects observed after HCV eradication
in some individuals who show unremitting immuno­
logical abnormalities and clinical symptoms. Thus,
we may assume that in some individuals, possibly in
those with a longer duration of HCV infection, the
alterations to the immune system have passed a point
of no return. Recognizing this as a biological condi­
tion may be decisive for the overall management of the
Box 4 | DAAs for the management of haematological malignancies
In patients with hepatitis C virus (HCV)-related haematological malignancies, direct
antiviral agents (DAAs; Box 3) may be a suitable treatment option for HCV infection.
In patients with lymphoma, older interferon-​based antiviral therapies can induce a
remission of the haematological disease in patients with low-​grade forms, especially
marginal zone lymphoma42,145,146. Recent data suggest an anti-​lymphoma activity of
interferon-​free DAA regimens147. Antiviral therapy should be considered the first-​line
approach in HCV-​associated low-​grade lymphomas90,145. For HCV-​associated aggressive
lymphomas, such as transformed diffuse large B cell lymphoma, immediate
conventional antiviral therapy is required148; however, when interferon-​based antiviral
therapy is administered in combination with chemotherapy for treatment of lymphoma,
it may substantially increase haematological toxicity149. Recent data suggest that
interferon-​free antiviral therapy should be performed after completion of, or in
combination with, immunotherapy and chemotherapy, following close monitoring for
potential drug–drug interactions150–152. Further prospective studies are needed.
However, no particular overlapping toxicities are expected.
The efficacy of DAAs was recently evaluated in 46 patients with HCV-​positive
indolent B cell non-​Hodgkin lymphoma, consisting mostly of marginal zone lymphoma
(n = 37)153. Sustained virological and haematological responses (including physical
examination, biochemical evaluation, CT scan and bone marrow biopsy) were obtained
in 98% and 67%, respectively. Estimated 1-year progression-​free and overall survival
were 75% and 98%, respectively. Patients with HCV-​positive diffuse large B cell
lymphoma usually have a high International Prognostic Index (a clinical tool developed
by oncologists to aid in predicting the prognosis of patients with aggressive non-​
Hodgkin lymphoma; high risk (4–5 points) corresponds to a 5-year survival of 26%)154,
and treatment, as for their HCV-​negative counterparts, consists of anthracycline-​based
chemotherapy coupled with rituximab. However, sustained viral response after antiviral
therapy has been associated with better overall survival in patients with HCV-​related
marginal zone lymphoma155 and in patients with diffuse large B cell lymphoma.
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P­r i­m e r
patients, especially with regard to the expectations of
antiviral treatment.
Patients with cryoglobulinaemia with no evidence
of causative factors are still classified as having essen­
tial mixed cryoglobulinaemia, which shows a variable
prevalence among patient populations in different coun­
tries. Essential mixed cryoglobulinaemia encompasses a
heterogeneous group of individuals and represents
a very challenging disease variant with regard to both
aetiopathogenetic and therapeutic concerns4.
Outstanding research questions
The mechanisms involved in the development of mixed
cryoglobulinaemia-​related lymphomas should be
investigated more deeply. Shedding light on the general
mechanisms of lymphomagenesis could also be of spe­
cial interest for haematologists. Identifying the factors
involved in non-​HCV-associated mixed cryoglobulinae­
mia remains a research challenge. Presently, the similari­
ties between the clinical presentation of non-​HCV and
HCV-​related cryoglobulinaemic vasculitis70,123 suggest
that these alternative triggers are also viral in nature.
Interferon-​free, direct-​acting antiviral-​based antiviral
therapy represented a paradigm shift, especially in the
field of HCV extrahepatic manifestations. Broadening
the use of antiviral therapy without immunomodula­
tory and/or antiproliferative properties will allow for the
definitive evaluation of the effect of HCV eradication on
cryoglobulinaemia, which was in the past confounded
by the immunoregulatory actions of interferon55,90.
Direct-​acting antiviral-​based antiviral therapy likely
does not actually affect the development of the immune-​
mediated injury once cryoglobulinaemic vasculitis is
established108. Direct-​acting antivirals (Box 4) might
be effective in mild cases of HCV-​related cryoglobu­
linaemic vasculitis80, but the use of direct-​acting anti­
virals alone as a therapeutic tool to treat HCV-​related
severe cryoglobulinaemic vasculitis requires further
validation studies. Furthermore, the uncertainty of the
pharmacokinetics and safety of direct-​acting antivirals
in the presence of renal impairment implies caution
when using them in nephritic patients. However, due to
the absence of the interferon-​related risk of triggering
autoimmune-​related side effects, direct-​acting antiviral-​
based therapy can be recommended to eradicate HCV,
a potential lymphoma-​causing agent that is known to be a
B cell activator.
Immunosuppression has been traditionally considered
as the first-​line intervention in severe cryoglobu­linaemic
vasculitis, especially when renal involvement is present.
Rituximab (used with either lymphoma or rheumatoid
arthritis treatment regimens) is the most widely used
biologic agent for the management of cryoglobu­linaemic
vasculitis. The B cell-​depleting agent has a proven safer
profile than conventional immunosuppressants124, and
its use has substantially changed the natural history of
HCV-​associated cryoglobulinaemic vasculitis by indu­
cing long-​term remission124,125, especially with intensive
regimens (for example, the ‘4 + 2 regimen’)54,108,126. In addi­
tion to having an immunomodulatory effect, rituximab
also plays an important role by depleting CD19-positive
B cells, which are known to be HCV reservoirs34.
13
P­r i­m e r

It may be envisaged that a combination of direct-​ considered. Finally, an underlying infection-​related

acting antivirals and rituximab may result in improve­
ments in the treatment of cryoglobulinaemic vasculitis
and HCV eradication108. The optimal combination pro­
tocol of rituximab plus new interferon-​free anti-​HCV
treatments needs to be defined. Currently, severe cryo­
globulinaemic vasculitis, especially with renal involve­
ment, should first be treated with a rituximab-​based
protocol. Direct-​acting antiviral administration may be
administered concomitantly or may be delayed pending
on individual circumstance, especially in the absence of
a very high viral load108,126.
Further studies are needed to identify rescue treat­
ments for the few patients with cryoglobulinaemic vas­
culitis who are rituximab-​refractory and/or intolerant.
Refractory cryoglobulinaemic vasculitis can be defined
as a clinical picture that does not improve within
4–6 weeks after the initial treatment, or that provides
a <50% improvement after 12 weeks, or that shows
chronic persistent manifestations after >12 weeks of
treatment. First, other conditions that mimic cryo­
globulinaemic vasculitis should be ruled out. Second,
the patient’s adherence to medical prescriptions and
adequacy of administered therapy should be critically
or cancer-​related comorbidity should be considered.
Some alternative approaches have been proposed for
treatment of refractory cryoglobulinaemic vasculitis,
including abatacept (a soluble fusion protein that con­
sists of the extracellular domain of human cytotoxic
T lymphocyte-​associated antigen 4 linked to the mod­
ified Fc portion of human IgG1)108,127 or tocilizumab
(a humanized mono­clonal antibody against the IL-6
receptor)128. Among others, newly available biologi­
cal agents, such as B cell-​activating factor (BAFF; also
known as tumor necrosis factor ligand superfamily
member 13B)-blocking agents129 and IL-2 agonists130,
also seem to be promising. The role of plasma exchange
and double filtration cascade to remove cryoglobulins
from the serum can still be taken into consideration in
the most severe or refractory cases68, especially in the
presence of high cryoglobulin concentrations before
rituximab administration. Again, the combination of
these new agents with direct-​acting antivirals could
favour control of the vasculitic process and eradicate
the viral trigger.
Published online xx xx xxxx

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Author contributions
Introduction (D.R.); Epidemiology (A.G.T.); Mechanisms/
pathophysiology (D.R. and P.C.); Diagnosis, screening and
prevention (C.F. and D.S.); Management (F.C.F., M.R.-C.
and A.L.Z.); Quality of life (M.R.-C.); Outlook (D.R.); Overview
of Primer (D.R.).
Competing interests
The authors declare no competing interests.
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional
claims in published maps and institutional affiliations.
Reviewer information
Nature Reviews Disease Primers thanks F. Dammacco,
Y. Shoenfeld, L. Quartuccio and the other anonymous referee(s)
for their contribution to the peer review of this work.