Lupus nephritis (LN) is a dreaded complication of systemic lupus erythematosus (SLE). One in six patients with SLE will have evidence of kidney involvement at the time of diagnosis and at least 40-75% patients will develop nephritis in the course of their SLE.(1)(2) Lupus nephritis continues to significantly affect morbidity and mortality. Both in the U.S. and in Europe, non-Caucasians have increased rates of nephritis.(3)(4) In the U.S., risk factors for progression to end-stage renal disease include: being African American, Hispanic, male, less than 24 years old and having high activity and chronicity on renal biopsy.(5)

The treatment of lupus nephritis has undergone a dramatic change from steroids with azathioprine (as a steroid-sparing agent used in the early 1960s) to immunosuppressive therapy like cyclophosphamide, mycophenolate, azathioprine and rituximab. Despite these advances there continue to be significant morbidity and mortality from this disease, and 10-15% of patients will progress to end-stage renal disease requiring renal replacement therapy. Treatment for lupus nephritis is based on an initial phase of intensive immunosuppressive therapy to halt the disease (the induction period) followed by maintenance therapy to maintain the response.

Classification

The histopathologic classification of lupus nephritis continues to guide therapy. In 2003 the International Society of Nephrology/Renal Pathology Society (ISN/RPS) proposed a classification of lupus nephritis based upon light, immunofluorescent and electron microscopic changes. Class I, or minimal mesangial lupus LN, was applied to normal glomeruli by light microscopy, but mesangial immune deposits by immunofluorescence. Class II is mesangial proliferative LN, mesangial hypercellularity of any degree or mesangial matrix expansion by light microscopy, with mesangial immune deposits. Class III is used to denote active or inactive focal, segmental or global endo- or extracapillary glomerulonephritis involving <50% of all glomeruli, typically with focal subendothelial immune deposits, with or without mesangial alterations.

Within Class III are subclassifications, III-A, for active lesions: focal proliferative LN; III-A/C for active and chronic lesions: focal proliferative and sclerosing LN; and III-C for chronic inactive lesions with glomerular scars.

Class IV or diffuse LN includes active or inactive diffuse, segmental or global endo- or extracapillary glomerulonephritis involving ≥50% of all glomeruli, typically with diffuse subendothelial immune deposits, with or without mesangial alterations. This class is divided into diffuse segmental (IV-S) LN when ≥50% of the involved glomeruli have segmental lesions, and diffuse global (IV-G) LN when ≥50% of the involved glomeruli have global lesions. Again “A” and “C” can be suffixed after class IV to denote active and chronic lesions respectively.

Class V is membranous LN with global or segmental subepithelial immune deposits. Class V lupus nephritis may occur in combination with class III or IV in which case both will be diagnosed. Class VI denotes advanced sclerosing glomerulonephritis where ≥90% of glomeruli are globally sclerosed without residual activity.

The current approach to treating LN is largely guided by histologic findings with consideration to presenting clinical features and degree of renal impairment. In general, classes III and IV need aggressive management to prevent progression to end-stage renal disease.

Treatment Options

Methylprednisolone
Cathcart et al. first proposed the use of high-dose, intravenous methylprednisolone (MP) for severe LN. Seven patients with diffuse proliferative LN were subjected to high-dose IV MP therapy. Following the pulse, five patients with rapidly deteriorating renal functions improved within three days and their serum creatinine levels returned to baseline by one month. All seven patients demonstrated reversal of several immunological abnormalities including decreased serum C3 levels, and reduced number of T lymphocytes in the peripheral blood.(6)

In another study, eight patients with acute exacerbations of LN were treated with nine monthly courses of alternate day IV MP pulse therapy, for three doses of 30 mg/kg/day. They were followed for 24 to 68 months (mean 35.6) post pulse. Stabilization of renal function was observed. Proteinuria declined, ANA titers fell and low serum complement levels improved. Steroids were discontinued by 24 months in four patients, and switched to alternate day therapy in the remaining five patients. No patient required the addition of cytotoxic therapy. No other untoward side effects were observed.(7) These studies showed that pulse MP can be used to treat LN. In fact, pulse MP is the medication of choice whenever there is a life-threatening manifestation of SLE and is used in conjunction with cyclophosphamide (CYP) or mycophenolate mofetil (MMF) to treat LN. However, the use of high-dose intravenous corticosteroids is associated with significant adverse effects that include hypertension, sepsis, osteonecrosis and behavioral changes such as irritability, depression or even frank psychosis.

Cyclophosphamide
Cyclophosphamide (CYP), an alkylating and cytotoxic agent, depletes both T and B cells, thereby reducing the production of pathogenic autoantibodies. Studies published by Austin et al. and Boumpas et al. were instrumental in making CYP a mainstay for treatment of LN.

Patients with LN enrolled in trials at the NIH between 1969 and 1981 were randomized into one of five treatment protocols: (1) high-dose prednisone; (2) azathioprine + low-dose prednisone; (3) oral CYP + low-dose prednisone; (4) combined oral azathioprine and oral CYP; and (5) intravenous CYP + low-dose oral prednisone. Although this study showed that immunosuppressive therapy preserved renal function, the difference was statistically significant only for the intravenous CYP plus low-dose prednisone arm (p=0.027).(8)

Boumpas et al. later showed that an extended course of pulse CYP is more effective than six months of pulse methylprednisolone in preserving renal function in patients with severe nephritis. In this study, 65 patients with severe LN were randomly assigned to monthly pulse methylprednisolone for six months, monthly pulse cyclophosphamide for six months, or monthly cyclophosphamide for six months followed by quarterly pulse cyclophosphamide for two additional years. Patients treated with pulse methylprednisolone had a higher probability of doubling serum creatinine than those treated with long-course cyclophosphamide (p<0.04). Risk of doubling creatinine was not significantly different between short- and long-course cyclophosphamide. However, patients treated with short-course cyclophosphamide had a higher probability of exacerbations than those treated with long-course cyclophosphamide (p<0.01).

An extended course of pulse cyclophosphamide was found to be more effective than six months of pulse methylprednisolone in preserving renal function in patients with severe LN. Addition of a quarterly maintenance regimen following six monthly pulses of CYP of 1g/m² of body surface reduced the rate of exacerbations.(9) These and several other studies established the NIH protocol for the treatment of LN as a standard of care.(10)(11)(12)(13) The biggest drawback of the NIH protocol was its associated morbidity with complications such as infertility and an increased risk of severe infections.

In response to this drawback, the Euro-Lupus Nephritis Trial (ELNT) was developed to test lower doses of intravenous CYP (500 mg every two weeks for a total of six doses) against the typical NIH protocol for induction. Houssiau et al. found that there was no significant difference between the low-dose and high-dose patients in developing renal failure, recurrent renal flares or in renal remission rates. In this multicenter, prospective clinical trial, 90 SLE patients with proliferative glomerulonephritis were randomly assigned to a high-dose IV CYP regimen (six monthly pulses and two quarterly pulses; doses increased according to the white blood cell count nadir) or a low-dose IV CYP regimen (six fortnightly pulses at a fixed dose of 500 mg), each of which was followed by AZA. Follow-up continued for a median of 41.3 months in the low-dose group and 41 months in the high-dose group. Sixteen percent of those in the low-dose group and 20% of those in the high-dose group experienced treatment failure. These results were not statistically significant. Levels of serum creatinine, albumin, complement component 3 (C3), 24-hour urinary protein and the disease activity scores significantly improved in both groups during the first year of follow-up. Renal remission was achieved in 71% of the low-dose group and 54% of the high-dose group (again not statistically significant). Renal flares were noted in 27% of the low-dose group and 29% of the high-dose group. Although episodes of severe infection were more than twice as frequent in the high-dose group, the difference was not statistically significant.

This study showed that in SLE patients with proliferative LN, a remission-inducing regimen of low-dose IV CYP (cumulative dose 3 g) followed by AZA achieved clinical results comparable to those obtained with a high-dose regimen.(14) The 10-year follow up data for survival and kidney function from the ELNT was published in 2010. It showed that death, sustained doubling of serum creatinine and end-stage renal disease rates did not differ between the low-dose CYP vs. the high-dose CYP group (5/44 (11%) vs. 2/46 (4%), 6/44 (14%) vs. 5/46 (11%) and 2/44 (5%) vs. 4/46 (9%), respectively) nor did mean serum creatinine, 24 h proteinuria and damage score at last follow-up. The data confirmed that a low-dose IV CYP regimen followed by AZA (Euro-Lupus regimen) achieved good long-term clinical results.(15)

Mycophenolate Mofetil
Mycophenolate mofetil (MMF), the prodrug of mycophenolic acid, inhibits inosine monophosphate dehydrogenase, which then suppresses DNA synthesis and the proliferation of T and B cells. Initially used to prevent transplant rejection, investigators began to evaluate the drug’s use in LN. Meta-analysis of several small trials showed that MMF was comparable to CYP in the treatment of proliferative LN.(16)(17) Ginzler et al. conducted a 24-week randomized, open-label, noninferiority trial comparing oral MMF (initial dose 1000 mg per day, increased to 3000 mg per day) with monthly IV CYP (0.5 g/m² of body-surface area, increased to 1.0 g/m²) as induction therapy for active LN. The primary end point was complete remission at 24 weeks. A secondary end point was partial remission at 24 weeks. Of 140 patients recruited, 71 were randomly assigned to receive MMF and 69 were randomly assigned to receive CYP. At 12 weeks, 56 patients receiving MMF and 42 receiving CYP had satisfactory early responses. In the intention-to-treat analysis, 16 of the 71 patients (22.5%) receiving MMF and 4 of the 69 patients receiving CYP (5.8 %) had complete remission, for an absolute difference of 16.7 percentage points (95% confidence interval, 5.6 to 27.9 percentage points; p=0.005), meeting the pre-specified criteria for noninferiority and demonstrating the superiority of mycophenolate mofetil to cyclophosphamide. Partial remission occurred in 21 of the 71 patients (29.6%) and 17 of the 69 patients (24.6%), respectively (p=0.51). Three patients assigned to CYP died, two during protocol therapy. Fewer severe infections and hospitalizations but more diarrhea occurred among those receiving MMF. In this 24-week trial, MMF was more effective than IV CYP in inducing remission of LN and had a more favorable safety profile.

The Aspreva Lupus Management Study (ALMS), a large multicenter, randomized clinical trial, was developed to determine if MMF was in fact better than CYP therapy. The researchers found no difference in response rates when comparing MMF to CYP -- 104 (56.2%) of 185 patients responded to MMF compared with 98 (53.0%) of 185 to CYP. Secondary end points were also similar between treatment groups. However, MMF was noted to be better in Hispanic and African American patients.(18) Induction therapy with MMF is associated with fewer side effects than with CYP. MMF may be particularly suitable as induction therapy in women of childbearing age where infertility from CYP treatment is a major concern. It also may work particularly well in African American and Hispanic patients, two groups with a large burden of LN.

Azathioprine
Azathioprine (AZA) is used as a steroid-sparing agent in patients with active SLE and for maintenance therapy after LN induction therapy. It is a purine analog and inhibits nucleic acid synthesis thereby affecting both cellular and humoral immune function. The mycophenolate mofetil versus azathiprine for maintenance therapy of lupus nephritis (MAINTAIN) trial showed that AZA was not inferior to MMF in maintaining remission. In this investigator-initiated trial, 105 patients with proliferative LN were randomized to AZA (target dose: 2 mg/kg/day) or MMF (target dose: 2 g/day) after induction therapy. Mean follow-up of the intent-to-treat population was 48 months. The baseline clinical, biological and pathological characteristics of patients allocated to AZA or MMF did not differ. Renal flares were observed in 13 (25%) AZA-treated and 10 (19%) MMF-treated patients. Time to renal flare, to severe systemic flare, to benign flare and to renal remission did not statistically differ.

Over a 3-year period, 24 h proteinuria, serum creatinine, serum albumin, serum C3, hemoglobin and global disease activity scores improved similarly in both groups. Doubling of serum creatinine occurred in four AZA-treated and three MMF-treated patients. Adverse events did not differ between the groups except for cytopenias, which were statistically more frequent in the AZA group (p=0.03).(19) More recently Houssiau et al. showed that repeat kidney biopsies in 30 patients who were initially enrolled in the MAINTAIN trial failed to detect differences between AZA and MMF maintenance therapy for LN. Thirty patients (16 AZA and 14 MMF) underwent repeat renal biopsy at two years (+/- 6 months). More patients had normal renal biopsies or showed Classes I/II/V LN at follow-up compared to baseline and conversely, fewer patients had Class IV LN at follow-up. No differences could be detected between the groups.(20)

Results from a long-term follow-up study of a randomized controlled trial of azathioprine plus pulse IV methylprednisolone (MP) versus CYP in patients with proliferative LN showed that induction treatment with CYP was superior to AZA+MP in preventing renal relapses, but other parameters for renal function did not differ. In this study, 87 patients with biopsy-proven proliferative LN were treated with either AZA/MP (n=37) or IV CYP (n=50), both with oral prednisone. After two years, renal biopsy was repeated, and all patients continued with AZA/oral prednisone.

After a median follow-up of 9.6 years, no significant differences between AZA/MP versus IV CYP groups were found in sustained doubling of serum creatinine [n=6 (16%) vs. n=4 (8%); p=0.313], end-stage renal disease [n=2 (5%) vs. n=2 (4%); p=1.000] or mortality [n=6 (16%) vs. n=5 (10%); p=0.388]. Renal relapses occurred more often in the AZA/MP group [n=14 (38%) vs. n=5 (10%); p=0.002, HR:4.5]. Serum creatinine, proteinuria and immunosuppressive treatment regimens at the last follow-up were comparable. Induction treatment with IV CYP was superior to AZA/MP in preventing renal relapses, but given that other parameters for renal function did not differ, it could be concluded that AZA/MP can therefore serve as an alternative in patients with proliferative LN who wish to avoid the potential gonadal toxicity of CYP.(21)

In another study looking at MMF versus AZA as maintenance therapy for LN, MMF was found to be superior to AZA with respect to the study’s primary end point, time to treatment failure, time to renal flare and time to rescue therapy. A total of 227 patients were randomly assigned to maintenance treatment (116 to mycophenolate mofetil and 111 to azathioprine). Observed rates of treatment failure were 16.4% (19 of 116 patients) in the MMF group and 32.4% (36 of 111) in the AZA group. MMF was significantly superior to AZA for the key secondary end points of the broader definition of treatment failure [rate, 42.2% (49 of 116 patients) in the MMF group vs. 56.8% (63 of 111) in the AZA group; hazard ratio, 0.66; p=0.03)]. Serious adverse events occurred in 33.3% of patients in the AZA group and in 23.5% of those in the MMF group (p=0.11), and the rate of withdrawal due to adverse events was higher with AZA than with MMF (39.6% vs. 25.2%, p=0.02).(22)

Based on these studies, AZA can be considered an option for therapy, particularly during pregnancy when other medications are contraindicated. Nevertheless, CYP and MMF continue to be the preferred treatment modalities for induction of remission in lupus nephritis and clearly MMF appears to be superior to AZA for maintenance therapy.

Tacrolimus
Tacrolimus was discovered in the 1980s and is useful in preventing transplant rejection. It is a T-cell–specific calcineurin inhibitor that complexes with immunophilin FK506 binding protein 12, and inhibits the phosphatase activity of calcineurin, resulting in decreased interleukin-2 transcription and inhibition of T-cell activation.(23) In addition, tacrolimus is also capable of inhibiting the production of tumor necrosis factor-α and interferon-γ by activated T cells.(24) More recently, there have been several reports of tacrolimus being used in the treatment of LN. However most studies that support tacrolimus use are from China and Japan and it is not known if results from those studies can be extrapolated to African Americans, Hispanics or Caucasians with LN.

Chen et al. conducted a multicenter, noninferiority randomized controlled trial comparing tacrolimus to CYP as induction therapy. Eighty-one patients with biopsy proven class III, IV and V LN were randomized to either prednisone and tacrolimus (n=42) or prednisone and IV CYP (n=39) for six months. Tacrolimus was started at 0.05 mg/kg/d and titrated to achieve a trough blood concentration of 5-10 ng/ml. Intravenous cyclophosphamide was initiated at 750 mg/m² of body surface area, then adjusted to 500-1000 mg/m² every 4 weeks for a total of six pulse treatments. After the 6-month induction therapy, the tacrolimus group achieved higher cumulative probabilities of complete remission and response (52.4% vs. 38.5% and 90.5% vs. 82.1%, respectively) than the IV CYP group, but differences were not statistically significant (log-rank test, p=0.2 and p=0.7, respectively). Proteinuria was significantly decreased in tacrolimus vs. IV CYP treated patients after the first month of treatment, even with adjustment for baseline proteinuria (protein excretion, 1.76 vs. 2.40 g/d; p=0.02 for the log-transformed analysis). After treatment, serum creatinine levels and estimated glomerular filtration rates were not significantly different between treatment groups. The authors of this study concluded that tacrolimus in conjunction with prednisone as induction therapy was at least as efficacious as intravenous CYP and prednisone in producing complete remission of LN and had a more favorable safety profile.(25)

A systematic review of clinical trials on tacrolimus in LN showed that tacrolimus might be effective as an induction and maintenance therapy for LN or as a treatment for LN with persistent proteinuria.(26) However, randomized controlled trials are needed to compare tacrolimus with standard regimens for the treatment of LN. Another drawback of tacrolimus use is the frequency of side effects. Hypertension, headaches, hyperglycemia, tremors, cytopenias and worsening renal functions are a few of the commonly seen side effects and need to be kept in mind while following any patient on this drug. In addition, levels of tacrolimus need to be monitored and dose of tacrolimus adjusted to keep tacrolimus concentration between 5-15 ng/ml.

B Cell Targeted Therapies

Anti-CD20 Therapy
Rituximab (RTX) is a chimeric antibody directed against CD20, a cell surface protein expressed on certain B cell subsets but not plasma cells. The mechanisms by which it induces B cell depletion is likely through the induction of apoptosis as well as cell mediated toxicity.(27) Initially used in non-Hodgkin’s lymphoma, off-label use in lupus has shown potential efficacy in SLE and LN.(28)(29)

Looney et al. conducted one of the first trials investigating the role of RTX in SLE. Eighteen SLE patients were recruited into a phase I/II dose escalation trial, where seven of the 18 patients had LN: three with class III, three with class IV, and one not classified. Of the 10 patients who successfully depleted their B cells, all experienced some disease improvement. The authors do not comment on the overall efficacy in the nephritis patients but do mention that one patient with class IV nephritis went into remission, with resolution of proliferative changes on repeat biopsy.(30)

An open label trial was then conducted looking at the combination of CYP and RTX in the treatment of refractory SLE. Thirty-two patients who had previously failed immunosuppressive therapy for lupus were included in this study. Of these 32, 24 had received CYP prior to inclusion in this study. All but three patients received a combination protocol of RTX and CYP. (Although 21 of the 32 patients had LN, the authors do not comment on these patients specifically.) Overall, 12 remained disease free after one cycle of B cell depletion therapy. Global British Isles Lupus Assessment Group (BILAG) scores were also significantly improved at six months. Therefore we assume that the nephritis patients did well after RTX and CYP therapy, given the entire group did well post treatment. However, we do not know if they responded to the same magnitude as the non-nephritis patients.(31) Several small trials that have shown that rituximab is efficacious in treating lupus as well as LN.(32)(33)(34)

The LUpus Nephritis Assessment with Rituximab trial (LUNAR) was conducted to specifically look at the role of RTX therapy in the treatment of class III/IV LN. This Phase III randomized, double-blind, placebo-controlled, multi-center study included 144 patients with Class III or IV LN. Study participants were randomized 1:1 to receive RTX or placebo in two infusions, 15 days apart, on top of standard immunosuppressive therapy with MMF and steroids. Although there were more renal responders in the RTX group (57% v 45.9%), the difference was not statistically significant. Though underpowered to detect a statistically significant difference, the African American group had a higher proportion of patients who responded to RTX versus placebo (70% v 45%). While there may be some benefit to the addition of RTX in African Americans, the medication overall does not seem to grant any additional benefit to current regimens.

Despite the failure of RTX for lupus nephritis to show a statistically significant efficacy in randomized clinical trials, data from the French Autoimmunity and Rituximab registry shows that overall 80 of 113 patients in the cohort responded to RTX therapy. Of those patients with renal disease, 74% showed at least a partial, if not a complete response.(28)

While the use of RTX as first-line treatment or in patients with a mild form of the disease is not recommended, its off-label use in severe, refractory SLE appears to be sufficiently positive to warrant its use.

Belimumab
B-cell activating factor, BAFF, a TNF-like cytokine, supports survival and differentiation of B cells. BAFF, also known as B-lymphocyte stimulator (BLyS), exerts its affects by interacting with three receptors on B cells: BAFF-receptor (BAFF-R); transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI); and B-cell maturation antigen (BCMA). BAFF-R can be activated by BAFF only, while TACI and BCMA can be activated by both BAFF and another member of the TNF-protein family called APRIL.(35) Binding of BAFF to the BAFF-R promotes B cell survival and development, allowing for progression past the T1 stage into follicular and marginal zone cells.(36) BAFF, as well as BAFF-R and TACI, have been implicated in murine models of SLE and nephritis. Researchers have demonstrated that early inhibition of BAFF-R in murine models improved mortality, decreased renal inflammation and decreased glomerular immune complex deposition.(37) Compared to normal controls, BAFF levels appear to be elevated in patients with SLE.(38)

Given the role of BAFF in SLE/LN, belimumab was designed as a recombinant, fully human antibody, with a high affinity for soluble BAFF. In a phase II, 52-week, placebo-controlled trial, 449 SLE patients were randomized to several doses of belimumab versus placebo. Although patients had moderate disease activity at entry, active nephritis patients were excluded from this trial. The medication was well tolerated; however, this study failed to meet its primary end points of reduction of disease activity scores and decreased time to flare at 24 weeks. However a significant percentage of these patients (38.5%) were ANA negative at enrollment. In subgroup analyses, seropositive patients performed significantly better on belimumab than on placebo with regard to disease activity, physician’s global assessment (PGA) and Short Form-36 (SF-36) physical component score.(39)

Two multi-national phase III trials were conducted in ANA positive, SLE patients with a similar study design to the phase II study. One trial followed patients for 52 weeks (BLISS-52) and the other for 76 weeks (BLISS-76). In both studies, all patients were seropositive for anti-nuclear antibodies and had moderate to severe disease activity. Patients were randomized to either standard of care + placebo, standard of care + belimumab 1 mg/kg, or standard of care + belimumab 10 mg/kg. Again, patients with active nephritis were excluded, and less than 15% of the patients randomized had a history of nephritis. The primary end point was the response rate at 52 weeks, as measured by the SLE Responder Index (SRI). When assessed with SRI, significantly more people showed a response in the belimumab 1 mg/kg and 10 mg/kg groups than in the placebo group at week 52.(40)

Despite the modest improvement in disease activity, belimumab is the first medication for SLE approved by the Food and Drug Administration (FDA) in 50 years. It remains to be seen, though, what role this drug will play in the treatment of LN given that active nephritis patients were excluded from these studies.

Conclusion

Lupus nephritis causes significant morbidity and mortality. While current regimens (summarized in Table 1 below) have reduced the burden of disease, there continues to be a large number of patients who progress to end-stage renal disease despite treatment with aggressive immunosuppression.

CYP remains the drug of choice for induction therapy in LN. However MMF, given its safety profile, is a valuable alternative, especially in African American and Hispanic patients. For maintenance therapy, AZA is another effective alternative to MMF, particularly for pregnant women where MMF is contraindicated. Tacrolimus could be the ‘new kid on the block,’ though randomized controlled trials are needed to compare it with the standard therapy being used for LN.

Given that B cells play such an integral role in nephritis through the generation of pathogenic autoantibodies, B cell depleting therapy would seem to be a logical target for the treatment of LN. However, rituximab and belimumab have both failed to show increased efficacy in SLE/LN and SLE respectively. But all is not lost -- in patients who do not respond to standard immunosuppressive therapy, rituximab and now belimumab could be tried as a “rescue therapy.” And given the results of the LUNAR study, which showed a trend toward benefit using rituximab in African American and Hispanic patients, rituximab could be considered as an option in this sub group pending further studies.