AIHA is anuncommon disorder caused by autoantibodies directed against self-erythrocyte antigens, leading to premature red cell destruction (13). In particular, AIHA is very rare in infancy and childhood (0.2 per 10/year).In these cases it is associated with immune disorders in about 50% of patients (13). AIHA is classified as warm, wAHIA, (IgG autoantibodies) in 90% of pediatric cases (14), cold (IgM autoantibodies) including cold hemagglutinin disease (CAD) and paroxysmal cold hemoglobinuria) or mixed, depending on the thermal range of the autoantibody.

ITP is an autoimmune disorder that induces a premature platelet destruction. ITP is characterized by autoantibodies against platelet glycoproteins, tipically GPIIb/IIIa and GPIb/IX, but ITP pathogenesis is often more complex (12). T cell abnormalities, for example an excessive polarization toward T-helper cell 1 (Th1), Th0, and Th17 cell types, a direct cytotoxic T cell–mediated destruction, and deficiency of T regulatory cells (Tregs) are described (15). A defect in B regulatory cells, megakaryocyte maturation and survival, and myeloid-derived suppressor cells have all also been demonstrated in ITP (12).

ITP is typically self-limited, but, in some cases, thrombocytopenia may be persistent (lasting between 3 and 12 months) or chronic (lasting more than 12 months) (16).

AIN is a benign disease typically with its onset in infants or toddlers. It is caused by autoantibodies directed against a patient's own neutrophils, with subsequent peripheral destruction (17). Spontaneous resolution is frequently observed in most cases within months. Children with AIN rarely suffer from severe or invasive or life-threatening infections despite severe neutropenia (14), because circulating neutrophils, although low in number, can display normal anti-microbial activity and they often increase in number during the acute event (18).

ES is a rare severe autoimmune disorder that affects two or more cell lineages. Initially described as the combination of ITP and AIHA, the recent definition of ES is an autoimmune disorder that affects two or more blood cell lines, occurring together or developing over time. These cytopenias may include ITP, AIHA, or AIN.

Novel treatment approaches are actively being developed for the treatment of AICs.

Among the new drugs, B cell directed therapies for example Ofatumumab (anti-CD20), Alemtuzumab (anti-CD52), and Daratumumab (anti-CD38) have been used with good results in case reports for the treatment of secondary AIHAs (28).

Fostamatinib is an inhibitor of spleen tyrosine kinase (SYK) that reduces macrophage-mediated clearance of RBCs and platelets. Syk plays a central role in FcγR-mediated signal transduction, and for this reason it has been considered a target for inhibition in different autoimmune and malignant conditions (29). The blockage of FcγR signaling through the inhibition of Syk ameliorate platelet destruction, and Fostamatinib is now FDA-approved for the treatment of thrombocytopenia in adult with chronic ITP an insufficient response to a previous treatment and is now in Phase 3 studies in wAIHA (20, 30). Orilanolimab, Rozanolixizumab, and Nipocalimab are monoclonal antibodies that bind and inhibit the neonatal Fc receptor (FcRn). The blockage of FcRn induces an increased clearance of IgG including that of pathogenic IgG autoantibodies (31). Rozanolixizumab has already shown efficacy at increasing platelet counts in patients with persistent/chronic primary ITP (32), while the safety and efficacy of Rozanolixizumab and of the other inhibitors of the FcRn for AHIA are under investigation (30).

Ibrutinib is a Bruton tyrosin kinase (BTK) inhibitor currently used or under investigation in several lymphoproliferative disorders (30). BTK inhibition has a significant effect on both lymphocytes and macrophages. His efficacy is reported in secondary AIHA, while a trial with another BTK inhibitor is currently underway for the treatment of ITP (20).

Belimumab is a human monoclonal antibody that inhibits the binding of soluble B lymphocyte stimulator (BLyS, also known as B-cell activating factor, BAFF) to B cells. Belimumab inhibits the survival of B cells (including autoreactive B cells) and reduces the differentiation of B cells into Ig-producing plasma cells. It is currently approved for the treatment of non-renal systemic lupus erythematosus (33), but it might also be a further solution for refractory AIHA patients (34).

In general, if a patient does not responded to first-line therapy, a diagnostic reevaluation should be considered, focusing on any previously overlooked cause of secondary AICs. In these subjects, the disease can be characterized by recurrent, chronic or refractory cytopenias affecting more than one blood lineage (concurrently or sequentially) or be associated with adenopathy and/or hepatomegaly and/or splenomegaly or concurrent severe infections (16). In these cases, a further diagnostic evaluation is needed for the identification of the underlying disorder and, potentially, of novel treatment options.

Evaluation of anti-vaccine antibody response, immunoglobulin levels (IgG, IgA, IgM), extended immunophenotype and proliferation test in response to mitogens, in conjunction with molecular tools for IEI diagnosis are essential for identifying a previously undiagnosed IEI (7, 35). Immunological investigations are always recommended, especially in the case of positive family history for IEI or immunedisregulation, very early onset and relapsing or refractory (11) ( Figure 1 ).

Patients with an underlying IEI may often required more second-and third-line immunomodulating therapies and may benefit from a specific target therapy (36) (35).

Central tolerance is one of the function of the thymus, consisting in the elimination of self-reactive T lymphocytes through negative selection, also known as clonal deletion (40). A breakdown of this central tolerance leads to autoimmunity. Aire is a gene expressed in thymic medullary epithelial cells, and it mediates the ectopic induction and presentation of many tissue-specific antigens to maturing T cells (41). This antigen presentation promotes the negative selection of autoreactive thymocytes, as well as self-tolerance. If aire is nonfunctional or absent, autoreactive T cells can escape clonal deletion and may later cause autoimmune disease (42). In addition, aire controls immune tolerance by the stimulation of a population of FOXP3-positive T regulatory cells (Tregs) in the thymus that can suppress autoreactive cells, thus influencing peripheral tolerance (43).

Mutations affecting this gene cause autoimmune polyendocrinopathy, candidiasis, and ectodermal dysplasia (APECED), also called autoimmune polyendocrinopathy syndrome type 1 (APS-1).

Levels of aire are also decreased in patients with RAG deficiency and Omenn syndrome thus contributing to the escape and peripheral expansion of autoreactive T cells (44, 45). A defect in peripheral tolerance is also implicated in the development of autoimmunity (46) that coexists with a marked immune impairment.

The thymic aplasia/hypoplasia characterizes DiGeorge syndrome (DGS) resulting in a decreased expression of aire, which may contribute to the autoimmunity often presents in DGS patients (46).

Impaired negative selection of autoreactive T cells as in APECED, RAG deficiency and DGS causes different autoimmune features, including AIC (44).

Thymocyte escape of central deletion can also occur because of abnormalities of TCR signaling, with subsequent defective activation-induced cell death (AICD) as in ORA1 and STIM1 deficiency, which are both characterized by immunodeficiency and autoimmunity manifestations associated with AIC (47).

Many self-reactive cells escape from the central tolerance mechanisms, but other mechanisms operating in the periphery can delete autoreactive lymphocytes.

Apoptosis, the process of programmed cell death, plays a central role in the destruction of autoreactive T and B cells at central and peripheral tolerance checkpoints. Pathogenic variants in genes involved in the apoptotic pathway (fas, fasl, casp10) result in the uncontrolled proliferation and accumulation of autoreactive (TCRalphabeta-/CD4+/CD8+ double-negative) T cells. The death receptor FAS, his ligand FASL, and the other genes of the FAS/FASL pathway (fadd, casp8, casp10) play a central role in apoptosis both in central and peripheral tolerance. Pathogenic variants in these genes cause autoimmune lymphoproliferative syndrome (ALPS), a primary immunodeficiency characterized by autoimmune disorders with AIC, splenomegaly, lymphadenopathy, and risk of lymphoma (39).

Peripheral tolerance is controlled by a subset of CD4+ CD25+ T cells (Regulatory T cells, Tregs). The development and function of T reg are regulated by Forkhead box protein P3 (foxp3), which affects T regs functional activity. Variants in foxp3 and related components cause a reduction in the number and diversity of Tregs, with severe consequences in the maintenance of peripheral tolerance leading to immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) with a very early onset (47, 48).

Development and functioning of T reg cells depend on IL-2 mediated signaling, and defects in IL-2 signaling, involving STAT5b and CD25, lead to IPEX- like syndromes, with similar autoimmune and autoinflammatory symptoms.

The adaptative immune response is strictly controlled by positive and negative regulators.

The positive signal is induced by the T cell receptor (TCR) on naïve T cells through the recognition of the antigen presented by the MHC complex expressed on the antigen-presenting cells (APCs). The second signal is the formation of an immunological synapse (IS), through the binding of the costimulatory T- cell receptor CD28 on the T-cell surface to B7 molecules (CD80 and CD86) on the APCs (i.e dendritic cells, macrophages and B cells). The CD28/B7 interaction is essential for IS creation, as it is required to ensure a complete T-cell activation (49). Cytoskeletal proteins are essential for many cellular functions, including the IS between T cells and APCs, and the regulation of lymphocyte proliferation. Variants in was are responsible of Wiskott-Aldrich syndrome. Was gene econdes WAS protein, an actin-nucleation promoting factor expressed in hematopoietic stem cells. WAS is characterized by the triad of thrombocytopenia with small-size platelets, eczema, and lymphopenia, mainly affecting T cells. The absent function of was is reflected by ineffective T cell proliferation and function, reduced Treg activity (with normal number of Treg), and hyperproliferation of B cells, which subsequent production of autoantibodies (50). Autoimmune manifestation are common, with AIHA, autoimmune neutropenia, peripheral vasculitis, and arthritis considered the most common.

Tregs act as negative regulators of adapative immune response. Defective Tregs development and function associated with autoimmune and lymphoproliferative disease are described in pathogenic variants of cytotoxic T-lymphocyte-associated protein 4 (ctla-4), lipopolysaccharide responsive beige-like anchor protein (lrba), and differentially expressed in FDCP6 homolog (def6) (47).

Ctla-4 is an essential negative immune checkpoint constitutively expressed in Treg cells and it is an inhibitor of T cell activity. Ctla-4 impairment is also observed in biallelic mutations of lrba and def6. Therefore, patients with CTLA-4 insufficiency, LRBA, and DEF6 deficiency have a similar clinical phenotype due to a defective suppressive activity of Tregs, as all three diseases present with reduced expression or defective function of CTLA-4 (49).

CTLA-4 is the CD28 homologue protein that “switches off” the T-cell-dependent response after pathogen elimination. CTLA-4 is mostly intracellularly stored in naïve T cells or constitutively expressed, and up-regulated under stimulus on the surface of Tregs. Upon TCR-dependent activation, when naïve T cells activation through the TCR, CTLA-4-is mobilized to the cell surface, forming homodimers that outcompete CD28 and bind to B7 molecules with higher affinity and avidity, activating cell-intrinsic and cell-extrinsic inhibitory mechanisms.

Lrba and def6 essentially control CTLA-4 recycle. Thereby, pathogenic variants in these genes lead to increased CTLA-4 lysosomal degradation and decreased CTLA-4 recycling.

The reduction of CTLA-4 on the T-cell surface is the common hallmark of ctla-4 insufficiency, lrba, and def6 deficiency, and induces immune homeostasis disruption due to the prolonged T-cell activation and migration, with subsequent multiorgan lymphocytic infiltration and a breakdown of the peripheral immune tolerance. A higher circulation of autoreactive lymphocytes may explain the development of autoimmunity, including AIC (51). Autoimmunity (AIC, enteropathy) is often associated with recurrent and/or severe infections, hypogammaglobulinemia, and lymphoproliferation.

Signal Transducers and Activator of Transcription (STAT) molecules play a central role in different signaling pathways activated by several cytokines and are responsible for the transcription of genes essential in the immune and inflammatory response (52). STAT1 signaling is activated by multiple cytokines including IFN-α and IFN-gamma, while STAT3 mediates responses to IL-6. The presence of Janus kinase (JAK) molecules is required for STAT1 and STAT3 phosphorylation. Impaired or enhanced function of the JAK/STAT-dependent molecular pathways results in immune dysregulation and susceptibility to infections.

In stat1 gain of function (GOF), due to reduced TH17 cells proliferation, patients present with an increased susceptibility to different bacterial and fungal infections, resulting in chronic mucocutaneous candidiasis and pulmonary infections (53). Autoimmunity and immune dysregulation are reported in 30% of patients with stat1 GOF mutations, mainly represented by endocrinopathies, blood cytopenias, and enteropathy (54, 55).

Stat3 GOF mutations result in a combined immune defect (56), featured by an increased risk of severe infections, and with a high incidence of autoimmunity (cytopenia, enteropathy, endocrinopathy, arthritis), lymphoproliferation, hypogammaglobulinemia and hepato-splenomegaly.

Among humoral defects, autoimmunity is a common feature in patients with Common Variable Immunodeficiency (CVID, including those with taci defect, baff-r defect, icos, NF-kB1 and NF-kB2 deficiency), selective IgA Deficiency (sIgAD), and hyper-IgM syndrome (HIGM). Autoimmune manifestations, such as arthritis, type 1 diabetes, intestinal bowel diseases, and AIHA are reported in 15% of patients with X-linked agammaglobulinemia (XLA) (48).

CVID is characterized by hypogammaglobulinemia (low IgG and IgA, with or without low IgM levels) with poor antibody response to vaccines or low switched-memory B cells, according to ESID criteria (57). Non-infectious manifestations, including autoimmune, lymphoproliferative disorders, and malignant diseases are present in about 30-50% of CVID patients (58). Autoimmune diseases are described in 20–30% of CVID patients; ITP, AIHA, and ES are the most frequent and can constitute the first manifestation of the immune defect, even in the absence of infection history. Development of autoimmunity can be related to several mechanisms including an altered reaction of the germinal center resulting in abnormal class switch, or uncontrolled B cell proliferation (59). Other mechanisms may include the defective suppressive function of B-regs on activated T cells (60); or hyperactivated T cell phenotypes (61).

CVID is mainly a polygenic disease, but a monogenic cause can be identified in about 15-30% of cases (62). Defects in the common receptor of taci, of the B cell-activating factor (baff) and april (a proliferation-inducing ligand) encoded by the tnfrsf13b gene, have been linked to CVID. CVID patients with CVID-associated variants of tnfrsf13b especially if heterozygous, have a higher risk for autoimmune complications and lymphoid hyperplasia potentially due to lack of normal mechanisms required to establish tolerance. Pathogenic variants in baff-r, impairing B-cell maturation, and in NF-kB1 and NF-kB2, transcription factors essential for B-cell maturation, survival, differentiation, class switching, and self-tolerance, are also associated with autoimmunity.

The HyperIgM (HIGM) syndromes include a group of IEIs caused by defective class-switch recombination, resulting in low levels of IgG, IgA, and IgE with normal or increased levels of IgM (63). HIGM syndromes are heterogeneous diseases, with X-linked, autosomal recessive, and autosomal dominant inheritance. Variants in the gene encoding CD40 ligand (CD40L), a protein expressed on activated T cells, are the most common cause of X-linked HIGM. Autosomal recessive forms of HIGM are associated to biallelic mutations in aid and ung.

Often, but not always, HIGM syndromes are characterized by the association of immunodeficiency and autoimmune diseases, depending on the genetic background. In the X-linked form, for example, self-reactive B-cells cannot be eliminated due to the defective CD40-CD40L mediated interaction, that is also responsible for the reduction of Tregs, while in the recessive form, AID deficiency could hesitate in impaired regulation of self-reactive B cells (59).

Activated phosphoinositide 3-kinase delta syndrome (APDS) is caused by heterozygous gain of function mutations in pi3kδ catalytic p110δ (pik3cd) or regulatory p85α (pik3r1) subunits leading to APDS1 and APDS2, respectively (64). One of the major downstream effectors of PI3K is mTOR, which plays a central role for cell growth and survival and in TH1 and TFH cell differentiation (65). APDS is characterized by the proliferation of effector cells, while naïve, and central memory T-cell subsets remain quiescent, with CD8 senescence, and imbalance between immature and mature cells in the T compartment, likely contributing to autoimmunity, lymphoproliferation, and immunodeficiency seen in this syndrome (55). AICs are the most frequently reported (76.2%) autoimmune manifestations (64).

ALPS is characterized by hyperactivation of PI3K/Akt/mTOR signaling pathway. Sirolimus, an mTOR inhibitor, is used with great success in children with ALPS, reducing lymphoproliferation. The treatment is described as particularly effective in ALPS patients who have autoimmune cytopenias (67).

Along with symptomatic therapies (anti-infective and anti-inflammatory treatment, first-line immunosuppressive treatment against autoimmune cytopenias and inflammatory parenchymal lung disease), many patients benefit from rapamycin (mTOR) inhibitors, and soluble CTLA-4-Ig (Abatacept, Belatacept), functioning in vivo as inhibitors of T-cell activation, mimicking CTLA4 function (68). In particular, the efficacy of Abatacept in auto-immune cytopenias caused by CTLA-4 haploinsufficiency has been well characterized (69). It is important to underline that the possibility of viral infection/reactivation during the treatment with Abatacept or Belatacept may be a limitation for the use of these drugs. For this reason, hematopoietic stem cells transplantation may be considered a possible definitive therapy (39).

The phenotypic overlap between CTLA4 and LRBA deficient subjects and their common cellular pathway establish a rational basis for the treatment with Abatacept. Abatacept mimics cellular CTLA4 function, rendered missing by LRBA deficiency, and negative regulate the immune responses by blockading or capturing CD80/86 molecules (70).

However, as in CTLA4 haploinsufficiency, the only curative therapy is HSCT: earlier studies showed that most patients with LRBA deficiency who have undergone HSCT achieve complete immune reconstitution (71).

Autoimmune manifestations (blood cytopenias, juvenile diabetes, hypothyroidism) are common in STAT1 GOF patients and they are often refractory to conventional treatment and also to the treatment with multiple immunosuppressants (72).

Although HSCT is the only curative treatment, the molecular understanding of JAK-STAT pathway has allowed the use of target pharmacologic inhibitors in patients with STAT1 GOF. Currently 5 different JAK inhibitors are available: Tofacitinib (JAK1 and JAK3 inhibitor), Ruxolitinib and Baricitinib (JAK1 and JAK2 inhibitors), Filgotinib (selective JAK1 inhibitor), and Decernotinib (selective JAK3 inhibitor) (66).

Jakinibs are also used for the treatment of STAT3 GOF patients, as an alternative to the anti-IL6 receptor (IL6R) mAb Tocilizumab. Indeed, IL6 and other cytokines, such as type I, II, and III interferons, IL-10, and IL-21, activate STAT3-mediated intracellular signaling. The combination of IL6 blockade and Jakinib therapy has proved to be an effective strategy for the treatment of immune dysregulation in STAT3 GOF patients (72).

Standard treatments for APDS include antimicrobial prophylaxis and immunoglobulin replacement as anti-infective prevention (73). Different immunosuppressive regimens (steroids, Rituximab, and calcineurin and rapamycin inhibitors) in different combinations can be used to control the symptoms of lymphoproliferation and autoimmunity (74). Most of APDS clinical symptoms were improved after HSCT, but an elevated rate of post-HSCT viral reactivation and engraftment failure have been reported (75, 76). Selective PI3K delta inhibitors constitute a targeted therapy based on the molecular mechanisms of APDS. Drugs such as leniolisib and nemiralisib have been successfully used, resulting in the reduction of lymphoproliferation, and improvement of clinical manifestations, including autoimmune cytopenia episodes (77, 78).

Several trials with new drugs (B-cell directed monoclonal antibodies; B-cell receptor inhibitors; IgG-mediated phagocytosis inhibitors) are ongoing or being planned. Given their mechanism of action, a potential use in AIC with underlying immunodeficiency may be envisioned. However, no specific information regarding IEI is currently available.

Allogeneic hematopoietic stem cell transplantation constitutes the only life-saving and curative treatment for some severe IEIs (ie: SCID) who currently do not benefit from any target therapy.

As mentioned above, also non-SCID patients may benefit of HSCT, requiring a complete conditioning regim to achieve engraftment.

Other management options, for example enzyme replacement, and gene therapy may provide an alternative approach to HSCT in specific IEIs.

Gene therapy is a relatively new approach to IEIs, based on the infusion of autologous hematopoietic stem cell (HSCs) transplantation to deliver stem cells with added or edited versions of the gene of interest (79). Gene therapies with retroviral and lentiviral vectors are being developed for different IEIs and may be find an application in many immunedeficiencies.