Several lines of evidence have suggested that proteinuria and glomerular histologic alterations can be mediated by the direct activity of a circulating factor. These data were obtained from ex vivo analysis of glomerular changes after incubation with serum from patients with FSGS, as firstly described by Sharma et al[11] in 1999, as well as from analysis of animal models in which kidneys from a specific line of affected mice showed recovery from FSGS after transplantation into normal mice[12]. The most striking data, however, was obtained from a study of a kidney with FSGS recurrence that had been re-grafted from a patient to another and led to total regression of the disease[13]. However, identification of the responsible factor(s) is still a matter of investigation, although some different molecules are considered likely candidates.

In recent years research interest has focused on the soluble form of the urokinase type plasminogen activator receptor (suPAR). suPAR appears to be able to cause podocyte foot effacement in mice[14], and suPAR levels observed in patients with FSGS are higher than those in patients with other glomerulopathies[15]. Nevertheless, the specific involvement of suPAR in glomerulonephritis has not been confirmed by other studies, which showed increased (plasma) suPAR levels in other pathological situations (i.e., bacterial and viral infections, sepsis, and cancer)[16]. Rather, increased suPAR levels were observed primarily in patients with reduced glomerular filtration rate (GFR), suggesting that an elevation of suPAR levels may merely be an indicator of reduced GFR[17]. Finally, the usefulness of suPAR to distinguish between FSGS and non-FSGS glomerulonephritis has been questioned by Bock et al[18], who showed similar (plasma) suPAR levels among FSGS patients, non-FSGS controls, and healthy volunteers.

Other circulating factors, such as cardiothropin-like cytokine 1 (CLC-1), vasodilator-stimulated phosphoprotein and apolipoprotein A-I, have also been proposed as effectors in the glomerular permeability process, but their clinical and pathological roles remain unknown[19]. Recently, detection of a panel of serum antibodies directed towards podocyte antigens was found to be associated with a high percentage of relapses in FSGS (predictive recurrence value of 92%)[20]. The most prominent of these antigens is CD40; the expression of which is up-regulated in podocytes in FSGS, supporting the hypothesis of a potential direct pathogenetic effect of anti-CD40 antibodies.

Demonstration of the precise permeability factor(s) remains elusive. Yet, recent findings have confirmed the critical role played by podocytes in FSGS development, and different podocyte antigens/cellular pathways have been associated with the disease course and medical treatment response (Figure 1). For example, it has been postulated that the B71 and sphingomyelin-phosphodiesterase-acid-like-3b (SMLPD-3b) proteins (both of which are expressed on the podocyte membrane) may directly interact with the cytoskeleton-inducing foot process effacement in response to a permeability factor[21,22]; interestingly, this effect could be antagonized by some drugs recently adopted in FSGS treatment [abatacept (Orencia^®)/belatacept (Nulojix^®)] for B71 and Rituximab^® for SMLPD-3b, in particular), as outlined below in the therapeutic section.

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Figure 1 Evolution of the therapeutic approaches for focal segmental glomerulosclerosis recurrence and related recent perspectives. The various treatments and their mechanisms are represented by the star shapes. CyA: Cyclosporine; PE: Plasma exchange; MMF: Mycophenolate mofetil; suPAR: Soluble form of the urokinase type plasminogen activator receptor; CYC: Cyclophosphamide; RTX: Rituximab^®; IA: Immuno-adsorption; Synpo: Synaptopodine; SMLPD-3b: Sphingomyelin-phosphodiesterase-acid-like-3b protein. Note: Steroids also regulate the podocyte activity of stabilizing the actin cytoskeleton, preserving glomerular permeselectivity, and directly reducing apoptosis via the PI3K/Akt signaling pathway.

Drug-induced or genetic-related alterations of the podocyte metabolic pathways may also lead to a maladaptive response to cell injury, defining a “pro-FSGS” phenotype, as has been observed in some patients with specific donor APOL1 polymorphisms[23] or in animal models with inhibition of the mTOR/Akt axis[24].

Another step forward in defining this disease may be achieved upon increasing our knowledge of the influence of micro (mi)RNAs on podocyte activity. In a mouse model, Gebeshuber et al[25] observed that transgenic expression of miR-193a (a down-regulator of WT1, itself a crucial effector in podocyte homeostasis) rapidly induces FSGS and observed up-regulated expression of miR-193a in isolated glomeruli from individuals with FSGS, as compared to kidney levels in healthy individuals or individuals with other glomerular diseases.

In addition to the probably pivotal role of podocytes in the disease process, it is also likely that T and B cells of the immune system contribute to FSGS development. A Th2 phenotype is commonly observed in patients with idiopathic nephrotic syndrome (NS)[26], and overexpression of IL-13, a characteristic Th2 cytokine, is associated with significant proteinuria in Wistar rats[27]. An indirect confirmation of B cell involvement derives from evidence showing a selective Rituximab^®-induced depletion is correlated to disease remission[28]. This association has recently been questioned, however, so the role of B cells in FSGS pathogenesis is still not well defined.

The adoption of plasma exchange (PE) for treatment of FSGS recurrence has been based on the hypothesis of the presence of circulating factor(s) that could be removed in order to treat or prevent the disease. Despite research into this causative factor remaining in a status of “cold case”, PE is still a cornerstone in FSGS recurrence treatment, since the 1985 report of its first positive application by Zimmerman[30]. A systematic review by Ponticelli[4] showed that PE promotes partial or complete remission in 70% of children and 63% of adults with FSGS recurrence. Most of the analyzed studies, however, are limited by their retrospective or uncontrolled design.

Adoption of PE in a pre-emptive protocol to reduce FSGS recurrence has been described by Gohh et al[31] in one of the few prospective studies in the literature. Ten transplanted patients with FSGS and at high risk of recurrence (both children and adults, including 5 transplants from living donors and 5 from deceased donors) were treated with a course of 8 PE sessions in the peri-operative period. Seven of the patients (including all 4 who received first grafts and 3 out of 6 who had prior recurrence) were free of recurrence at the end of follow-up (range of 238-1258 d). The use of pre-emptive PE in a high risk pediatric patient who underwent a second living kidney transplantation (the first kidney was lost due to recurrence) was more recently described by Chikamoto et al[32]. The patient had also received a 2-wk course of Rituximab^® (375 mg/m²; 2 doses), methylprednisolone (1 mg/kg per day), tacrolimus (10 ng/mL) and mycophenolate mofetil (MMF) (600 mg/m² per day) before transplantation; at 12 d before transplantation, 4 PE sessions were performed. No sign of recurrence was found in protocol biopsies at 8 mo after transplantation.

Canaud et al[33] described positive outcome (complete remission at 3 mo after diagnosis) for 10 patients with FSGS recurrence that had been treated with a 9-mo course of intravenous cyclosporine (CyA; C₀ levels at 200-400 ng/mL), followed by oral CyA (C₂ levels at 1200-1400 ng/mL), high dose oral steroids (1 mg/kg per day for the first 4 wk, then progressively tapered) and a course of PE sessions. The only patient who experienced recurrence of proteinuria after post-transplant year 1, concurrent to PE frequency reduction, had been successfully treated with Rituximab^® (2 doses) and PE sessions bimonthly, obtaining a complete proteinuria remission in the 34 ± 6.7 mo of follow-up.

A positive effect is also described for plasma absorption in some papers[34-37], but further studies are needed to define the potential additive benefit in comparison with PE.

KDIGO guidelines suggest for FSGS on native kidneys a 4-wk to 16-wk course of prednisone (1 mg/kg per day, with a maximum of 80 mg and a slow tapering in the 6 mo after remission)[38]. Glucocorticoids may act to stabilize the actin cytoskeleton, thereby preserving glomerular permeselectivity[39] and directly reducing podocyte apoptosis via the PI3K/Akt signal pathway[40]. Efficacy of steroid treatment in recurrent/de novo FSGS has never been evaluated in a randomized trial; on the other hand, considering its pivotal therapeutic role in FSGS on native kidneys, many different regimens have included steroids in post-transplantation FSGS treatment.

Apart from the paper by Canaud et al[33], who described a combined treatment of CyA in association with high dose steroids and PE, Shishido et al[41] also reported a favorable outcome (7/10 complete remission) for pediatric patients with FSGS recurrence in response to a combined treatment with methylprednisolone pulses (20 mg/kg after diagnosis on 3 consecutive days in weeks 1, 3 and 5) and an increase in CyA target levels (area under the curve_0–4 4500-5500 ng/h per milliliter for the first month, 4000 ng/h per milliliter for the next 2 mo, and 3000 ng/h per milliliter thereafter).

CyA is commonly applied for the treatment of several immune-mediated diseases and as a second-line therapy for steroid-resistant/dependent FSGS on native kidneys[38]. Conversely, CyA does not appear to prevent post-transplant FSGS recurrence when given as a part of the initial immunosuppressive regimen[42,43]; although, this potential has not been evaluated in more recent studies. Standard oral doses of CyA have not been associated with reduced incidence of recurrent FSGS. Nonetheless, higher intravenous doses have been associated with remission of proteinuria for the first time since reported by Ingulli et al[44] 25 years ago.

Overall, limited evidence has supported the administration of high dose CyA to achieve remission of FSGS recurrence with a persistent effect[45,46]. Salomon et al[45] reported a remission of recurrent proteinuria in 14/17 (82%) of children following administration of intravenous CyA (mean period of 21 d; range of 250-350 ng/mL); after 4 years, 11/17 (64%) patients had achieved sustained remission. In a second series, remission was induced in 13/16 patients (81%), which also included PE sessions for 4 of the cases; CyA doses were from 6 to 25 mg/kg per day[46]. At the latest follow-up (range of 10 mo to 12 years), 11/13 (84%) patients had a functioning allograft. It is noteworthy to mention that in this study, as in the studies by Canaud et al[33] and Chikamoto et al[32], the CyA treatment was combined with PE sessions.

The mechanism by which CyA might decrease proteinuria has been elucidated recently. Briefly, CyA has been shown to act by means of a direct effect on the cytoskeleton via dephosphorylation of synaptopodin, a crucial stabilizer of podocyte actin cytoskeleton, rather than through an immunosuppressive activity such as inhibition of T cells[47,48]. According to these clinical evidence, it was postulated that the anti-proteinuric effect had been observed only with high dose CyA because the hypercholestorelemic state induced by NS limits the CyA active fraction[49].

Currently, the option of CyA therapy in FSGS is more frequently used in combined-therapy regimens. The long-term safety/efficacy ratio of such a therapy, however, remains to be confirmed by study, which is of particular importance in light of the severe toxicities associated with high dose CyA.

Cyclophosphamide (CYC) is an alkalizing agent that inhibits cell DNA duplication, leading to cell death. It is active both on resting and dividing lymphocytes[50]. Anecdotal experiences with CYC therapy (2 mg/kg per day) reported achievement of partial or complete remission in patients with FSGS on native kidneys and also in steroid-dependent patients; however, no benefit was found in steroid-resistant patients[51,52].

In FSGS recurrence, Kershaw et al[53] treated 3 pediatric patients with CYC (1-2 mg/kg per day, adjusted for white blood cell count) for 8-12 wk and obtained two complete remissions and one partial; the patient with the longest follow-up (125 mo) experienced two additional relapses, each of which were treated successfully with pulse intravenous steroids. A more recent report described a series of 6 patients with FSGS recurrence all of whom were treated with a combination of CYC and PE (10 sessions over 2 wk, followed by 1 session per week for 2 mo), with complete remission being achieved in 3 of the patients and partial remission in the other 3[54]. A second case series described 11 pediatric patients with FSGS recurrence who were treated with a 2-mo course of CYC (2 mg/kg per day) and PE sessions, with initial remission being achieved in 9/11 and with 7/9 being free of disease at the last follow-up (32 ± 15 mo)[55].

MMF inhibits the inosine monophosphate dehydrogenase-mediated reduction of T and B lymphocyte proliferation. Gbadegesin et al[56] suggested MMF for treatment of steroid-dependent/resistant FSGS on native kidneys. Subsequently, a randomized controlled trial including 138 patients (both children and adults) with primary FSGS compared CyA and MMF plus dexamethasone, but no difference was observed in complete or partial remission rates after 52 wk of follow-up and both groups showed poor outcome (remission in 46% vs 33%, respectively)[57]. At the present time, as reported by Lau et al[58], no randomized controlled trial has yet to demonstrate the efficacy of MMF in association with other therapies or as a single agent in FSGS treatment on native or transplanted kidneys.

Renin angiotensin system (RAS) blockers have an important role in blood pressure control, but they also have anti-proteinuric and systemic anti-inflammatory effects[59]. RAS inhibition represents an important therapeutic strategy in proteinuric glomerular disease as FSGS, for either recurrent or de novo types.

Despite some reports having suggested RAS blockers as effective therapeutics for this disease[60,61], the association of these drugs with other therapies limits a final judgment on their real effect as a single drug. Freiberger et al[62] reported a favorable outcome after the use of a triple RAS blockage [angiotensin-converting enzyme (ACE) inhibitor, angiotensin receptor (blocker) antagonist (ARB), and renin inhibitor] in a transplanted patient with FSGS recurrence; since PE and Rituximab^® treatment produced no apparent benefits in the patient previously, a late response to this treatment may not be excluded “a priori”.

It is noteworthy that a close monitoring of serum creatinine and potassium levels is essential in all subjects treated with RAS blockers, especially when all these drugs are prescribed together and even more so when renal function is suboptimal.

The potential effect of galactose on glomerular permeability and proteinuria was firstly hypothesized by Savin et al[63], stating that sucrose binds with high affinity and inactivates the supposed “permeability factor”, thereby facilitating its plasma clearance.

Jhaveri et al[64] described a patient with severe recurrent FSGS (massive proteinuria of 37 g/d at day 2 after transplantation) who had been previously treated with PE, intravenous immunoglobulin and Rituximab^®, and achieved complete remission of proteinuria after receipt of a high galactose diet and supplemental oral galactose (0.2 g/kg, two times per day). As for other case series mentioned before, the role played by galactose in disease remission vs the role of previous treatment is indistinguishable. More recently, Robson et al[65] also reported a favorable outcome (1 complete and 1 partial response) in 2 patients with FSGS recurrence treated with high galactose diet. Sgambat et al[66] reported in a recent case series a reduction in permeability factor activity in 7 pediatric patients with steroid-resistant NS (2/7 with recurrent FSGS) treated with high galactose diet (0.2 g/kg, twice daily), without any significant improvement in proteinuria values.

The tumor necrosis factor-alpha (TNF-α) signaling pathway is involved in the development of both NS and FSGS, as evidenced by elevated levels of TNF-α detected in plasma and urine obtained from patients with FSGS[67] and increased glomerular permeability to TNF-α observed in vitro[68].

At the present time, very few cases of FSGS have been treated with anti-TNF-α agents. Leroy et al[69] reported a favorable outcome (complete remission) for a 15-year-old patient with recurrent FSGS that was presumably resistant to other treatments (increased immunosuppressant dose, PE, intravenous immunoglobulin, high dose steroids, CyA, and CYC) after administration of an anti-TNF-α blocker (firstly infliximab, then etanercept). Bitzan et al[70] showed that plasmapheresis effluent or fresh plasma (obtained from a child with recurrent FSGS and from two children with primary FSGS) caused cytoskeleton disturbance on podocyte culture. In detail, the plasma from the patient with FSGS recurrence activated β3 integrin and dispersed focal adhesion complexes, and this effect was reversed by pre-incubation with antibodies against TNF-α or either of the two TNF-α receptors. Following this study’s observation, the patient who was plasma resistant was treated firstly with Etanercept and then with Infliximab, which ultimately led to partial remission of the disease.

Rituximab^® is a chimeric monoclonal antibody that recognizes CD20 antigen on B lymphocytes. This agent has several unlabeled applications in the field of kidney transplantation; it has been successfully applied to reduce anti-donor ABO or HLA antibodies[71] and to treat acute humoral rejection of the graft[72], post-transplant lymphoproliferative diseases[73], and also some recurrent/de novo glomerulonephritis.

Rituximab^® treatment also has a long history of interest in its potential as a therapeutic option for idiopathic NS before and after transplantation. However, after the initial reports about its favorable use in FSGS recurrence were published in 2006 and 2007[28,74,75], conflicting results were reported by other studies in the literature. Currently, Rituximab^® may be adopted as a preventive therapeutic approach to reduce FSGS recurrence rate, or as a treatment of FSGS recurrence.

The use of Rituximab^® as a prevention strategy derives from two retrospective studies[21,76]. In the first, Fornoni et al[21] investigated 27 kidney transplanted patients at high risk for FSGS recurrence and showed that use of Rituximab^® in the perioperative period (375 mg/m² within 24 h after the kidney transplantation) was associated with a lower incidence of post-transplant proteinuria and with stabilization of GFR at the 12 mo follow-up. This study also demonstrated for the first time that Rituximab^® operates in a B cell-independent manner; sera obtained from FSGS recurrent patients caused a down-regulation of SMLPD-3b, a protein involved in regulation of podocyte cytoskeleton, and this phenomenon was prevented by pre-treatment with Rituximab^® through direct binding.

Audard et al[76] observed the absence of a clinical FSGS recurrence (not biopsy proven) in 4 patients who received Rituximab^® (375 mg/m²) in their induction protocol for a second kidney transplant (first kidney lost due to a recurrent disease). Nevertheless, the short follow-up (12-54 mo), the difference in Rituximab^® schedule (a single administration in 2/4 patients and 2 doses in the other 2 patients), and PE adoption in 2/4 patients partially limit the significance of this uncontrolled study.

To date, Rituximab^® has been widely used, alone and in combination protocols, as a treatment for recurrent FSGS in cases of incomplete remission, PE dependence, or as a first-line therapy in specific patient subsets. Despite successful results having been obtained[77-86], other studies have shown a transient or even absent response to Rituximab^®[62,87-94] (Table 1).

Abatacept is a biologic agent, specifically the CTLA4-Ig recombinant fusion protein derived from the extracellular portion of CTLA4-Ig and genetically fixated to the high and constant portion of the IgG1 immunoglobulin. Its effect is exerted by interfering with lymphocyte co-stimulation[95,96] upon binding to the APC protein ligands B71 (CD80) or B72 (CD86) and displacing their T cell counterpart or CD28[97]. In some experimental models of organ transplantation, the systemic administration of CTLA4-Ig effectively dampened the immune response, preventing experimental acute and chronic rejection and resulting in prolonged graft survival and tolerance[98-100]. On the basis of these findings, different biological T cell co-stimulation blockers became the subject of clinical trials. A high affinity variant of CTLA4-Ig, named LEA29Y (belatacept, Nulojix^®; Bristol-Myers Squibb Pharma, Uxbridge, United Kingdom), has been developed and was awarded approval by the Federal Drug Administration (FDA) in 2011 for prophylactic use for organ rejection in adult kidney recipients[101].

Abatacept was approved by the FDA in 2005 for the treatment of rheumatoid arthritis and active juvenile idiopathic arthritis[102], and quite recently has been proposed as a new treatment strategy for FSGS recurrence. Yu et al[103] reported a positive outcome in 4 patients (2 children) affected by early and Rituximab^®-resistant FSGS recurrence and in 1 patient with glucocorticoid-resistant primary FSGS on native kidneys. All these patients received abatacept, at a dose between 250 mg/d and 500 mg/d, the most commonly used dose for rheumatoid arthritis treatment. Before using abatacept, PE sessions were also performed in all 4 patients with FSGS recurrence, while the patient with primary disease on native kidneys received an immunosuppressive treatment composed of prednisone and CyA, with tacrolimus applied as a second line therapy. All patients achieved and maintained a significant proteinuria regression after 10-48 mo of follow-up. The authors suggested that this response was directly correlated with the B71-positive immuno-stained podocytes found in the kidney-biopsy specimens, because B71 may be expressed on the podocyte surface in some proteinuric conditions such as FSGS, thereby altering cytoskeleton organization, a condition that is known to be abrogated by abatacept.

Nevertheless, other studies of patients with FSGS recurrence have shown a slight/absent response after treatment with CTLA4-Ig[104-107], despite the fact that in some of these cases belatacept (able to bind B71 with an higher affinity than abatacept) was adopted.

The use of bone marrow mesenchymal stem cells (BM-MSCs) has been reported to reduce kidney injury in different experimental models of kidney disease[108-111]. Ma et al[111] showed in a well-established murine model of FSGS (adriamycin nephropathy) that human umbilical mesenchymal stem cells (HuMSCs) may improve kidney fibrosis and modulate the inflammatory response. Recently, BM-MSCs have been demonstrated as effective treatment for a wide range of immuno-mediated diseases[112-114].

Belingheri et al[115] reported successful application of their innovative approach with BM-MSCs in a 13-year-old kidney transplanted patient who had developed an immediate biopsy proven FSGS recurrence after renal transplantation and who was non-responsive to PE and Rituximab^® (2 doses). The patient had received allogeneic BM-MSCs infusions (6 doses, according to the most commonly adopted protocol for treatment of graft vs host disease) at months 7, 10 and 14 after transplantation and at month 5 after Rituximab^® administration. Remission of proteinuria was achieved after three BM-MSCs infusions, and at the last follow-up (22 mo) both renal function and proteinuria values were stable. The treatment appeared as well tolerated, and no adverse events were noted.