The use of Rituximab in Kidney Disease

Rituximab, the monoclonal chimeric anti-CD20 antibody, is an effective B-Cell depleting agent and continues to gather data for its use in a wide range of conditions relevant for the Nephrologist. It was also a pre-season favourite in the recent NephMadness event run by our friends at eAJKD, so I figured a quick recap was timely. As the literature is vast and grows by the week, I will only give a brief review of the current data, much of which is weak consisting of small observational reports.

Lupus Nephritis
The jury is still out. The LUNAR study randomized 144 patients with proliferative LN to Rituximab 1g x 4 or placebo with both groups receiving MMF & steroids. The experimental group had a decrease in anti-dsDNA and complement levels. Remission rates were numerically, but not statistically, better with add-on Rituximab (57% V 46%). While a lack of benefit with additional use of Rituximab was demonstrated, whether it could be an alternative to MMF is not known. In cases of resistant LN, we again have multiple favourable case series but no hard evidence.

Steroid-Resistant Nephrotic Syndrome (SRNS)
Evidence suggests Rituximab may be effective in steroid-dependent or calcineurin inhibitor-dependent patients, allowing withdrawal of one/both agents. An open-label RCT in 54 children with SRNS examined standard therapy (with steroids & calcineurin inhibitors) to Rituximab with lowering doses of usual therapy. The experimental arm had lower proteinuria, less relapse and was more likely to be drug free at 3 months. However, relapse did occur in 18.5% of Rituximab treated patients at the time of recovery of the B-Cell population.
The data does not all demonstrate a benefit however, which brings us to a recent small case series in the NEJM. Another anti-CD20 monoclonal antibody, this time the humanized preparation Ofatumumab, was reported to be an effective treatment in 5 cases of SRNS refractory to Rituximab. Although both are anti-CD20 antibodies, they have different epitope specificities which may explain the outcome in this small series. This reinforces the idea that B-Cell depletion is more complex than some may presume.

Minimal Change Disease
No RCT data exists but observational series suggest a benefit in steroid-dependent, but not resistant cases. A new case series in NDT reports on 16 adult patients with MCD who were steroid-dependent (n=12) or resistant and given 2-4 doses of Rituximab. Overall, 13 had a complete and 2 a partial remission with one non-responder. No serious adverse events were reported but 7 relapsed after 9–28 months.

Membranous Nephropathy (MN)
Guess what? No RCT data exists but limited data suggests it may be a useful agent. (See Nate’s previous post). The largest observational study I could find included 100 patients, 32 of whom had disease resistant to other immunosuppressive agents. Baseline proteinuria of 9g/day had been present for a mean of 2 years. Complete/partial/no remission was achieved in 27/38/35 patients respectively, remissions after a mean of 7 months. Prior immunosuppressant use did not appear to alter outcome. Other smaller studies also report that patients, including those with resistant disease, may respond.
There is evidence that Rituximab may cause a decrease in Anti-M-type Phospholipase A2 Receptor (PLA2R) antibodies. In this study, 25/35 patients with idiopathic MN had Anti-PLA2R antibodies, and these autoantibodies declined or disappeared in 17 (68%) of these patients within 1 year of rituximab treatment. The patients who demonstrated antibody response had much improved rates of complete and partial remission in this small study. Perhaps these autoantibodies may prove to be a useful biomarker for treatment response in MN.

ANCA associated vasculitis (AAV)
As per the RAVE study (& RITUXIVAS), there is now robust evidence that a 4 week course of rituximab is non-inferior to cyclophosphamide in the treatment of AAV, including a finding that it may be superior to conventional immunosuppression in relapsing patients. Note that only 2/3 of patients had renal impairment in RAVE with creatinine clearances of 54-69mls/min in the 2 groups. See my previous post for more detail.

FSGS
Limited evidence suggests it may be beneficial in steroid-dependent but not steroid-resistant cases. We must be wary of publication bias from early series which reported  positive findings, especially as they have often failed to be replicated. It has been used with some success for recurrent FSGS post-transplantation, often together with plasma exchange. There is growing evidence for a direct effect on the podocyte, as well as its known anti-B Cell effect, possibly via binding of podocyte proteins such as SMPDL-3b.

Transplantation
It is beyond the scope of this post to delve into the use of Rituximab in transplantation. It can be used in desensitization protocols, PTLD and treatment of acute antibody-mediated rejection (AMR) as adjuncts to IVIG and plasma exchange (trial data awaited regarding allograft outcome). There is no clear evidence for its use in chronic AMR, which tends to have little response to any agent.

Another indication is essential mixed cryoglobulinemia. Here, Rituximab has been reported to be beneficial in cases usually associated with Hepatitis C infection. See Gearoid’s previous post.
Overall, B-Cell depletion in general is an exciting treatment strategy for many of the disease processes we deal with. Like much in Nephrology, we lack strong data for if, when and how to use it. We also lack thorough knowledge as to its precise mechanism of action. Suggestions of a direct podocyte effect in glomerular disease and different mechanistic effects of alternate anti-CD20 preparations illustrate how much we have to learn.

Genetic Defects of the Glomerular Basement Membrane

The glomerular basement membrane is a thin layer of extracellular membrane proteins that is an important part of the filtration barrier, particularly glomerular permselectivity, by preventing proteins from crossing into the filtrate. The major proteins in the GBM are laminin, type IV collagen, nidogen and the heparan sulphate proteoglycan agrin. Interestingly, in the past, I was taught that the highly negative charge on agrin played the major role in mediating charge selectivity. However, recent studies have shown that mutations in the gene encoding agrin, leading to a reduction in charge along the GBM, have no effect of glomerular function in mice. Similarly, deletion of agrin has little effect on permselectivity further suggesting that the role of the proteoglycans in selective filtration is minor at best.

There are two conditions associated with genetic defects in glomerular basement membrane proteins:

Syndrome	Gene(s) affected	Protein	Phenotype
Alports Syndrome	COL4A3 COL4A4 COL4A5	Type IV Collagen, α3, α4, α5 subunits	Initial normal formation of GBM but eventually hematuria, proteinuria and eventual ESRD with characteristic splitting of the GBM. COL4A5 mutations are commonest and are X-linked. Other forms are autosomal.
Pierson Syndrome	LAMB2	Laminin β2	Autosomal recessive disorder with variable phenotype depending on the particular mutation. However, ocular abnormalities (microcoria) are present at birth and the majority of affected individuals progress to ESRD within the first few weeks/months of life. Extrarenal manifestations including hypotonia and neurodevelopmental defects have been reported.

Please see this excellent review in Nature Reviews Nephrology for further information.

Still mysterious: the elusive circulating factor for FSGS

Important new findings were recently published in relation to proteinuria and FSGS, which are definitely of interest to our community.

First, the punch line:

There is new evidence for a “circulating factor” in recurrent FSGS in a fascinating case of a re-transplanted kidney (here)

BUT

There is growing evidence that suPAR is a non-specific marker of kidney disease and therefore not likely to be the “circulating factor.”(here)

In fact, it appears that it is non-specifically found in CKD, and correlates with a declining GFR.

Now for some details:

The re-transplanted kidney

A letter to the NEJM editor (here) describes an amazing case of resolution of recurrent FSGS after re-transplantation. 

A 27 year old patient with primary FSGS receiving a kidney from his healthy 24 year old sister developed proteinuria in the nephrotic range (up to 25 g/day!) within 2 days of transplantation, and had no improvement after plasmapheresis and standard immunosuppressive treatment. A renal biopsy confirmed foot process effacement, the first hallmark of recurrent podocyte damage heralding recurrent FSGS. Incredibly, with all appropriate consents and institutional approval, the transplant team removed the allograft from Patient 1 and re-transplanted it into another patient who had ESRD due to diabetes. Within 3-4 days, the proteinuria resolved and a repeat biopsy showed resolution of foot process effacement and re-establishment of a normal podocyte architecture. Eight months later, Patient 2 is reported to be doing very well, with good allograft function and no proteinuria.

This case demonstrates in a remarkable way that recurrent FSGS results from an elusive “factor” rapidly produced by the recipient (with primary FSGS), and that the allograft itself can remain fully functional if removed from the influence of this “factor” and placed in another patient.

suPAR is not suPER specific

What may have seemed to be exciting news in 2011, namely the notion that soluble uPAR may be predictive of recurrent FSGS (here), appears to be unfortunately evolving into yet another unsuccessful attempt to identify the ever elusive circulating factor.

Recent work published in Kidney International by Maas et al. (here) confirms that suPAR is not able to distinguish between idiopathic FSGS, secondary FSGS or minimal change disease. 

This is actually not surprising, because a closer look at the clinical data in Wei et al. (here) reveals that the admittedly arbitrary cut-off for separating primary FSGS from all other glomerular disease (3000 pg/ml) did not hold up when tested among their patient cohorts with idiopathic, recurrent versus non-recurrent FSGS (all had suPAR> 3000 pg/ml, thus suPAR could not predict the recurrent from the non-recurrent cases). 

The second figure in the Maas et al. paper may help explain this conundrum: they show a negative correlation between suPAR and eGFR, meaning that as GFR drops, suPAR levels rise, which essentially means that suPAR is simply a marker of CKD.

Future work will no doubt continue to address these issues, but the apparent lack of specificity of suPAR for FSGS casts serious doubt on its proposed role as the circulating factor.

So, the search is still on!!!

The Plot Thickens: Role of Plasmin in Edema Formation in Nephrotic Syndrome

I was at the ASN Kidney Week in Philadelphia last week and I had the chance to check out some abstracts and discuss with the authors. Three abstracts called my attention right away. I uploaded a post a few months ago related to the mechanisms of edema in nephrotic syndrome (NS) : in patients with NS, plasminogen is filtered from plasma and activated in distal nephron by enzyme urokinase forming plasmin. Plasmin can then proteolytically activate ENaC by cleavage of the γ-subunit, leading to sodium retention and edema. These are the abstracts:

1. Abstract: [FR-PO1777] Urinary Content of Plasmin(ogen) and Activation of ENaC Current by Urine Resides during Remission of idiopathic Nephrotic Syndrome. Buhl et al.

The same group from University of Southern Denmark who published the original plasmin study came back again and presented more evidence for their hypothesis. They took spot urine samples from 20 children with active idiopathic NS and compared them to urine samples obtained after remission in the same patients. Urine samples were analyzed for plasmin and plasminogen concentrations and urinary protease activity. Urine plasmin and plasminogen concentrations (normalized to urine creatinine concentration) and urine protease activity were found to be significantly higher in the active phase of NS in comparison to the remission phase. Not only that, the urine samples obtained in the active phase were able to evoked stronger ENaC currents than the urine samples obtained in remission phase.

2. Abstract: [FR-PO1776] Preeclampsia Is Associated with Significant Urinary Excretion of Plasmin(ogen) and the Ability of Urine To Activate ENaC In Vitro. Buhl et al.

The same group above also did another study where urine samples from 16 preeclamptic patients and 17 normotensive, non-proteinuric pregnant women (control) matched on age and gestational age were compared. Urine was analyzed for plasminogen and proteolytic activity. ENaC currents after exposure to urine was monitored in M1 cells by whole cell patch clamp. Urine plasminogen concentration (normalized to urine creatinine concentration) and proteolytic activity were increased in the urine of preeclamptic patients but not in controls. What is more, a significant positive correlation was found in the preeclamptic group between urinary plasmin(ogen) and diastolic blood pressure. The ability of the urine samples from preeclamptic patients to evoke ENaC current was abolished by amiloride to a lower level than the controls, suggesting that there might be small amounts of plasmin (ogen) present in the urine under normal conditions. The authors speculated that this might have a natural anticoagulant effect in the urine.

3. Abstract: [FR-PO1779] Nephron Expression and Distribution of the Plasminogen Receptor, PLG-RKT, and Colocalization with ENaC and uPAR, in Murine Kidney. Nangia et al.

Dr. Parmer’s group at UCSD has identified the presence of a novel Plasminogen Receptor (PLG-RKT). This PLG-RKT, apparently colocalizes with urokinase, and ENaC on the apical surface of the distal nephron, and all of these are present in an orientation to promote Plasminogen activation and ENaC processing. They actually found that urokinase was also present in the proximal tubule but in a less prominent way than in the distal nephron. The significance of the latter is unknown. Therefore, the machinery for sodium retention is present even under normal conditions.

I believe these studies give more support to the Plasmin hypothesis.

Are we finally getting thE kNaCk of edema in nephrosis?

Mechanisms of edema formation in nephrotic syndrome (NS) are controversial. The traditional underfill hypothesis has come under heavy fire, and there are now many clinical and experimental observations that contradict it. The overfill hypothesis has now captured the zeitgeist. It postulates that sodium retention in many patients with NS is caused by an intrinsic renal defect in sodium excretion, which in turn causes an expansion of plasma volume. However, the exact mechanism of sodium retention has not been clearly elucidated and many renal sodium transporters have been implicated.

One recent paper by Svenningsen et al suggests that the epithelial sodium channel (ENaC) might be strongly implicated. Most of our understanding of the molecular mechanisms of sodium retention in NS is derived from the puromycin aminonucleoside (PAN) mouse model, which resembles minimal change disease. They found that urine of PAN nephrotic rats increased ENaC currents and amiloride abolished them. Proteolytic cleavage has a major role in regulating the activity of these channels by increasing their open probability. Specific proteases have been shown to activate ENaC by cleaving different channel subunits at specific sites within their extracellular domains. They found that plasminogen and/or plasmin were the serine proteases responsible for ENaC activation in nephrotic urine. The urine of nephrotic rats contained both substances, but the plasma from these animals only contained plasminogen, suggesting that plasmin was formed in the urine in situ and was not filtered out of the plasma. Svenningsen et al also observed that cortical collecting duct cells of nephrotic rats had urokinase activity which was directly responsible for the conversion of filtered plasminogen into plasmin. Another important observation was that amiloride not only blocks ENaC but also inhibits urokinase. Significantly, Svenningsen et al were able to reproduce all of the previously described experiments with urine from patients with NS.

These findings suggest a novel mechanism of edema in NS: plasminogen present in plasma is filtered through the defective glomerular barrier of NS and it is then converted into plasmin by the action of urokinase present in the cortical collecting duct. Plasmin then activates ENaC by proteolysis, resulting in sodium retention with the subsequent appearance of edema. More studies are needed, but this could be an important breakthrough.

Helbert Rondon, MD, FASN, FACP

Thrombosis and the nephrotic syndrome

Should patients with nephrotic syndrome (NS) be anticoagulated? Is there any level of proteinuria or albumin at which we should be especially concerned?

The rates of venous thromboembolism (VTE) have ranged from 8-44% in observational studies and it is estimated that patients with NS are 8 times more likely to develop a clot than the general population and that this risk is highest in the first 6 months after diagnosis (140 times the general population or 9.8%). The risk appears to be higher in patients with severe NS; in one study of 89 patients with NS and an albumin of less than 2.0, 31% of patients were found to have PE on pulmonary angiogram.

There are a number of potential mechanisms for the hypercoagulable state seen in patients with NS:

- Increased platelet aggregation: There are a number of potential mechanisms for this including increased production of thromboxane A2 (arachidonic acid is normally albumin-bound and hypoalbuminemia leads to increased availability) and high LDL levels (which appears to independently increase platelet aggregation). There also seems to be an increase in interactions between platelets and the vascular endothelium with increased levels of circulating markers of platelet activation

- Defects in the clotting system: Loss of antithrombin in the urine (serum antithrombin levels correlate with albumin levels in patients with NS). Elevated protein C and protein S have also been noted in patients with NS

- Impaired fibrinolysis: Plasminogen levels are decreased in NS and correlate with proteinuria. Albumin is itself is a cofactor for plasminogen binding to fibrin and thus its lack may contribute to impaired fibrinolysis.

- Etiology of the Nephrotic Syndrome: It appears that patients with membranous nephropathy, MPGN and minimal change disease are more at risk of VTE than patients with other causes of NS. The reason for this is uncertain and the risk is particularly high with membranous nephropathy.

Click to enlarge

Unfortunately, there are no tests that can accurately predict the risk for VTE in patients with NS. It has been suggested prophylactic anticoagulation should be considered in patients with an albumin of 2.0 although this is based on opinion rather than evidence. There is some suggestion that treatment of the hyperlipidemia associated with NS can reduce platelet aggregation and similarly, prophylactic aspirin should be considered although, again, there are no trials proving its efficacy. Overall, although the risk of of VTE in patients with NS is 8 times the general population, this is still less than 1% per year and given that the risk of major bleeding with warfarin is greater than this, it is hard to justify routinely anticoagulating all patients. Perhaps if we had a better measure of risk we could better stratify these patients and select those who would benefit most from anticoagulation early.

Here is a recent review of the topic the picture above is from this review

Cyclosporine in Membranous Nephropathy: A non-immune modulatory effect to maintain remission?

This is the story of a 64 year old male mason with past medical history notable for hypertension and hyperlipidemia who was referred to me for evaluation of proteinuria.

The patient noted onset of lower extremity edema nine months prior to the clinic visit. At the time, he was admitted to the hospital due to increasing complaints of lower extremity edema and shortness of breath, and received the presumptive diagnosis of congestive heart failure. He had an echocardiogram showing preserved ejection fraction and no valvular abnormalities. A chest CTA was negative for pulmonary embolism and imaging of the lower extremities was negative for deep venous thromboses. His serum albumin at the time was 2.5 g/dl down from his baseline of 4.0 g/dl a few months prior. No urinalysis data was available from that admission. He was treated with furosemide to which he responded with resolution of his shortness of breath, but with no effect on his notable 3+ pitting edema. He was discharged from the hospital with cardiology follow up as an outpatient.

Teaching point: not all edema is heart failure! A simple urinalysis would have been revealing.

Now back to the case...

During this time period, the patient’s lower extremity edema and exertional dyspnea had not improved in spite of escalating doses of furosemide. He was admitted to the hospital again two months prior to the renal clinic visit, this time for hypertensive emergency treated with intravenous labetalol followed by escalation in his anti-hypertensive regimen which included beta blocker, calcium channel blocker and a small amount of ACE inhibitor. A urinalysis sent at this time showed 3+ protein. He was also subsequently noted to have a serum albumin of 1.3 g/dl. Of note, his renal function was normal at his baseline of 1.0 mg/dl on presentation to renal clinic. Quantification of his proteinuria revealed 13 gms/day.

The patient denied a family history of kidney disease or any known past episodes of kidney problems. He denied fever, headache, chest pain, visual problems, rashes, joint pain, muscle pain, abdominal pain, nausea, vomiting, dysuria, hematuria, or hemoptysis. He had noticed foamy urine over the past few months. He also complained of occasional dyspnea on exertion, which was markedly different from his baseline a year prior. Renal ultrasound was notable for kidneys measuring 14 cm bilaterally with no hydronephrosis and no masses. Ultrasound evaluation of the renal arteries revealed no renal artery stenosis.

He underwent a renal biopsy which revealed membranous glomerulonephritis.

The patient was initially managed with angiotensin converting enzyme inhibitor, angiotensin receptor blocker and diuretics. After a six month period -- fifteen months since the onset of symptoms -- during which there was no sign of spontaneous remission, we initiated treatment with Cyclophosphamide and high dose Prednisone.

The patient developed severe leukopenia, so Cyclophosphamide was discontinued. While we have some experience with Rituximab at our institution, this patient had normal renal function and therefore appeared to be a good candidate for a trial of Cyclosporine. We therefore initiated treatment with Cyclosporine at a dose of 3 mg/kg/day.

He responded well with reduction of his proteinuria to less than 2 gms/day. However, due to an episode of VZV/shingles, the Cyclosporine dose had to be decreased to 1 mg/kg/day, which resulted in trough levels less than 40 ng/ml. In spite of Cyclosporine levels well below these known to have a significant immunomodulatory effect, the patient achieved a surprising partial remission. It is of course conceivable that his remission was spontaneous, but this is not supported by the fact that he had been persistently nephrotic for 1.5 years prior to initiation of Cyclosporine treatment. Furthermore, the fact that the patient continues to be in remission 3 years later suggests that Cyclosporine may continue to exert a beneficial effect.

It is not surprising that Cyclosporine can be successfully used to achieve remission in patients with IMN. However, past experience has shown that many patients with IMN have recurrence of proteinuria once Cyclosporine is removed. On the other hand, the chronic use of Cyclosporine is known to result in a decline of renal function, and nephrologists have been reluctant to use it for a prolonged period of time in the nontransplant setting. It would therefore seem advantageous to devise therapeutic strategies for the use of low dose Cyclosporine to prevent recurrence of nephrosis.

Intriguingly, a 2008 study by the Mundel group suggests that Cyclosporine may have a new target: the glomerular podocyte. The authors present data in mouse models of acquired proteinuria, and specifically in mice that have been genetically modified to express calcineurin in a constitutive fashion. Interestingly, these mice develop severe albuminuria. Through a series of experiments, the authors subsequently determine that Cyclosporine confers an anti-proteinuric benefit through its protective effect on a critical, podocyte-specific molecule called synaptopodin. The preservation of synaptopodin ensures that the podocyte cytoskeleton remains intact, preventing the disruption of the glomerular filtration barrier.

These findings challenge our understanding of Cyclosporine as an immune modulatory agent for proteinuric kidney disease, and our long held belief that Cyclosporine puts patients in remission through its effects on T-cell activity. These data necessitate a closer look at the effects of traditional “immune modulatory” agents on the podocyte cytoskeleton.

Extrapolating from these data, is it possible to maintain podocyte health – and keep nephrotic patients in remission – by using Cyclosporine, or preferably targeted agents with similar podocyte-specific effects?

My patient enjoys a partial remission (currently proteinuria is less than 0.8 gm/day) on what would be considered "subtherapeutic" levels of Cyclosporine. However, if we begin to think of Cyclosporine as a podocyte-modulating agent, perhaps we also need to redefine what levels are “therapeutic.” This case suggests that lower doses may allow us to use Cyclosporine to maintain patients in remission with less concern for its nephrotoxicity. We have now employed this approach in a few other patients with IMN, with encouraging results thus far. Moreover, the use of low dose Prograf in some of our patients with FSGS has also proven to be effective in maintaining remission. Future clinical studies on these questions will be revealing. In the meantime, we are hard at work in the lab to identify new podocyte-specific anti-proteinuric agents.

The actin cytoskeleton of the podocyte

Peter Mundel, a well-known researcher of podocyte biology, gave our Renal Grand Rounds today.  Here's what I took away from this morning's talk:

-regulation of the podocyte's actin cytoskeleton is postulated to be the final common pathway in most instances of nephrotic syndrome/proteinuria.

-in some ways it is better to think of the podocyte as a modified smooth muscle cell rather than a modified epithelial cell based on its intricate network of actin & myosin which mediate contractility.

-the medication cyclosporine (used in the treatment of some refractory forms of nephrotic syndrome) acts NOT on modulation of immune cell function as previously assumed, but rather on direct effects on the podocyte actin cytoskeleton, as detailed in a prior post.

-a key event in generating abnormal, protein-leaking podocytes is turning them from a stationary cell to a motile one.  This key transition is mediated by three distinct Rho GTPase proteins, already established to mediate motility in several cell types. In general, Cdc42 and Rac1 promote podocyte motility, whereas RhoA promotes podocyte stabilization. It is possible that targeting these Rho GTPase pathways could lead to novel therapies for podocytopathies, and this is being investigated.  

Check out a good 2009 Kidney International review of the topic by Mundel and Reiser.

Potential Use of Rituxan in Membranous Nephropathy

Take-home points from Mayo Clinic's Fernando Fervenza's excellent Renal Grand Rounds this morning, in which he addresses the potential use of rituximab as a treatment for membranous nephropathy:

-the "rule of thirds" that we learn about membranous nephropathy (e.g., 1/3 of patients will spontaneously remit, 1/3 of patients will stay the same, and 1/3 of patients will progress to ESRD) is not entirely accurate in that patients with an extremely high degree of proteinuria (e.g., more than 8-10 grams per day) have a very high rate of progression of kidney disease.  This becomes critical in interpreting the results of clinical trials; you really have to look at the baseline proteinuria characteristics of the cohort.

-in non-randomized controlled trials, investigators are finding that dosing Rituxan either using a 1gm iv at D+1 and D+15 protocol OR using a 375 mg/m2 every 4 weeks for 4 doses total leads to at least a partial remission in between 60-75% of membranous nephropathy patients, a rate which is seemingly greater than one would expect for spontaneous remissions. One such paper by Fervenza et al is shown here.  

-a randomized, controlled trial comparing Rituxan to another therapy (e.g., Cytoxan-prednisone or a calcineurin inhibitor) would really be necessary to fully recommend using this strategy in the treatment of membranous nephropathy.  However, it is not clear where the money for performing such a trial would or should come from--industry or government.  

-it may be necessary to look at longer end-points to fully assess the effectiveness of Rituxan.  Since Rituxan targets pre-B cells but not plasma cells, you presumably have to wait until the plasma cells die off for it to have full effect.  Furthermore, monitoring proteinuria (the current standard for evaluating clinical response) is not perfect since it may reflect chronic podocyte damage (which may take months to years to fully heal, if it ever does) rather than immunologically-mediated proteinuria.  

-it looks as if levels of the antibodies against the phospholipase A2 receptor which explains a good chunk of patients with membranous nephropathy correlates generally well with patients who respond to Rituxan; that is, patients who successfully achieve a remission with Rituxan tend to show a complete disappearance of this antibody as assessed by Western blog.  Although this does not prove causality, it does point to a potential means of monitoring patients for a potential response.  

Interferon effects on the kidney

Interferons are cytokines which play a central role in the inflammatory response, and commercially prepared interferons have proven useful in the treatment of several diseases. For instance, interferon-alpha (often used in conjunction with ribavirin) is often used in the treatment of hepatitis C, and has also proven useful in the treatment of certain cancers, such as malignant melanoma. Interferon-beta has also been very successfully used in the treatment of multiple sclerosis. With the increasing use of interferons, however, has come the realization that they can have renal side effects in some patients.

A variety of mechanisms of injury have been reported, though most attention has focused on the ability of interferon therapy to cause proteinuria and nephrotic syndrome. This has been noted most commonly with interferon-alpha therapy, though in many of the patients with hepatitis B or C it may be difficult to be certain whether or not the development of nephrotic syndrome comes from the interferon therapy or a direct hepatitis-mediated renal injury such as MPGN. There have also been some recent case reports suggesting that interferon-beta can also cause a minimal change nephrotic syndrome in patients treated for multiple sclerosis and malignant melanoma.

Other mechanisms of renal injury reported with interferon use include acute tubular necrosis, acute interstitial nephritis, and even hemolytic-uremic syndrome. Occasionally, tubuloreticular structures as seen on electron microscopy of a kidney biopsy can be a clue as to the diagnosis of interferon-induced renal injury; these may also be seen in HIV-associated nephropathy.

Transplant Glomerulopathy

As acute rejection rates continue to fall, causes of late allograft loss, such as transplant glomerulopathy (TG), become increasingly important. TG should spring to mind when you encounter a renal transplant recipient who develops heavy proteinuria and progressive allograft failure, usually late post-transplant. The clinical presentation overlaps with that of chronic allograft nephropathy, although proteinuria tends to be greater in TG and patients are likely to have a history of donor-specific anti-HLA antibodies. The pathogenesis is believed to relate to the presence of these donor-specific antibodies, which are often often anti-HLA Class II. These may wax and wane in concentration and, as a result, may not be detected on a single assay but repeat testing is usually successful. C4d staining is typically negative.

Various immunosupressive regimens have been tried, but none are known to be effective. Progressive graft failure and return to dialysis is the usual outcome. Finally, the increasing use of protocol biopsy informs us that ultrastructural changes that predate the TG lesion develop within the first months post-transplant and in apparently well-functioning kidneys.

Secondary Causes of Membranous Nephropathy

Although the majority of membranous nephropathy falls under the "idiopathic" category, there are a variety of causes of secondary membranous nephropathy. It is quite important to eliminate these entities as possible diagnoses, as the treatment for secondary membranous nephropathy relies primarily on treatment of the underlying cause of disease whereas idiopathic membranous nephropathy often involves the use of steroids and/or cytotoxic agents.

Broadly speaking, the secondary causes of membranous nephropathy can be broken down into 4 general categories:

1. Infection-associated Membranous Nephropathy: hepatitis B and hepatitis C are both associated with membranous nephropathy. Because diseases such as malaria, schistosomiasis, TB, and leprosy are likewise associated with membranous nephropathy, secondary nephrotic syndrome is likely much more common in developing countries subject to these tropical diseases. Syphilis is also on the list.

2. Disease-associated Membranous Nephropathy: it is well-known that MN may be see in lupus ("WHO Type V Lupus Nephritis") either alone or in combination with other lupus-related pathologies. A variety of the other conditions associated with MN are also autoimmune diseases (e.g., rheumatoid arthritis, Sjogren's syndrome). Diabetes mellitus and membranous nephropathy have also been linked, as has sickle cell disease.

3. Drug-induced Membranous Nephropathy: the classic offenders are gold, penicillamine, NSAIDs, and captopril, though there are case reports for many others.

4. Malignancy-associated Membranous Nephropathy: although there appears to be a increased relative risk of solid tumors and lymphomas in patients with membranous nephropathy as compared to the general population, this association is still somewhat poorly defined and remains controversial. Many would simply recommend age-appropriate cancer screening (e.g., mammogram, screening colonoscopy, etc) in the patient with newly-diagnosed nephrotic syndrome without any other evidence for malignancy rather than a large-scale imaging workup.

The Proteinuria Controversy

One of the largest controversies in the field of proteinuria/nephrotic syndrome research derives from a 2007 Kidney International paper published by Russo et al. Briefly stated, the authors suggest the paradigm-shifting idea that the glomerulus filters massive amounts of albumin, and that nephrotic syndrome is a defect in tubular reabsorption of albumin. This flies in the face of decades worth of research on proteinuria, which based on a combination of micropuncture and other physiologic experiments in mice and man has led to the conventional model in which albumin is prevented from entering Bowman's space by the charge-selectivity of the glomerular filtration barrier, and nephrotic syndrome results from a breakdown of this barrier. What are we to make of such a debate? Who is right and who is wrong? The following is a (hopefully unbiased) list of the pros and cons of each side of the argument.

Pro: Normal glomeruli filter nephrotic levels of albumin.

The major piece of data in support of this hypothesis comes from the Russo et al paper in which the authors use a relatively new imaging technique, intravital 2-photon microscopy, which enables in vivo imaging of the kidney using injected fluorescent compounds at a resolution previously unachievable. Essentially, the crucial experiment involved injection of a fluorescently-labeled albumin into the vasculature of rats; by quantifying the degree of fluorescence in the plasma compared to Bowman's space, the authors were able to calculate a "sieving coefficient" for albumin. The sieving coefficient they arrived at was about .02--which is orders of magnitude higher than the previous value obtained via the micropuncture method, about .0006. This implies that nephrotic levels of proteinuria are being filtered from normal glomeruli, and the authors postulate a proximal tubular-based mechanism of rapid albumin reclamation. The nature of such a mechanism is unclear, but the authors suggest that vesicles of intact albumin are transcytosed through proximal tubular cells, providing evidence of this by showing an electron micrograph of endogenous albumin within proximal tubular cells. Finally, in a 2008 JASN article by Dr. Wayne Comper, the author sites various methodologic problems with some of the initial experiments used to demonstrate the "charge selectivity" model of the glomerulus. A recent follow-up paper in JASN by Russo et al furthermore suggests that it is an impaired tubular uptake of filtered albumin which accounts for the changes seen in early diabetic nephropathy.

Con: The case against the "tubular proteinuria" model.

Not surprisingly, this newer model has been met with much resistance. In an article by Christensen et al forcefully entitled, "Controversies in nephrology: renal albumin handling, facts, and artifacts!", the authors describe their opposition to the idea that the glomerulus exhibits such large permeability to albumin. First, they point out several methodologic concerns with the technique of 2-photon microscopy, suggesting that the low fluorescent signal they observed is subject to misinterpretation; perhaps some of the "filtered albumin" seen in Bowman's space is fluorescent bleed-through from nearby blood vessels. The authors also point out a very logical question: if massive amounts of protein are retrieved by proximal tubular cells, why haven't we seen evidence of this throughout decades of research in this field? Many veterans of the field have simply not observed proximal tubular cells chock-full of vesicles containing endogenous albumin, and suggest that the published electron micrographs by Russo et al could be fixation artifact. Furthermore, the authors point out that mice lacking megalin function--thought to be a major player in protein uptake in proximal tubular cells--show only a mild degree of proteinuria, not nearly enough to be consistent with the massive amounts of albumin purported to be filtered by the glomerulus. Finally, virtually all of the mutations identified in patients with congenital nephrotic syndrome target genes known to be important in podocyte function: nephrin, podocin, alpha-actinin 4, etc, which would seemingly point to the podocyte, rather than the proximal tubular cell, as the primary player in regulating proteinuria.

So: Who's right and who's wrong? I don't think we know yet, but for now my money is on the conventional explanation--in general, it seems that the "tubular etiology of nephrotic syndrome" is supported predominantly by a new technique, the limitations of which are not yet fully known, and in order to lend it further credence, alternative techniques which support this theory would be necessary. If it does turn out to be true, however, this would really represent a major paradigm shift in nephrology.

Exciting NEJM Membranous Nephropathy Article

A landmark paper on membranous nephropathy was published by Beck et al in the most recent issue of NEJM. Ultimately we will have to see if the finding is reproducible, but based on the data presented this represents a major breakthrough in understanding the pathogenesis of membranous nephropathy and nephrotic syndrome.

There has long been strong evidence in support of a circulating factor which causes podocyte injury and resultant nephrotic syndrome. Additional evidence from the neutral endopeptidase story further suggested that an autoantibody directed again some podocyte antigen might be the culprit. In this issue, the researchers demonstrate that in 70% of patients with idiopathic membranous nephropathy (but 0% of patients with secondary membranous nephropathy or other forms of proteinuric kidney disease such as FSGS or diabetic nephropathy) contain an autoantibody against the podocyte antigen phospholipase A2 receptor. Furthermore, the autoantibody's presence appears to correlate with disease activity, suggesting a possible pathogenic role.

The work has a number of implications. First, it suggests that membranous nephropathy is indeed a separate disease than FSGS and other distinct forms of nephrotic syndrome. The common final pathway for proteinuria is the same (podocyte injury), but the ways in which to get there is likely different. Second, it suggests that detection of serum antibodies against phospholipase A2 receptor may be a useful part of the diagnostic workup for nephrotic syndrome--perhaps even making biopsies unnecessary--and perhaps could be used to follow disease activity in response to various therapeutic maneuvers. That is, this test may well become the "ANCA" of membranous nephropathy.

Congenital analbuminemia

Plasma oncotic pressure is important to prevent edema, right? And you would think that albumin, which plays a critical role in the binding of multiple plasma proteins and ionized calcium, is essential to life, right?

The rare, autosomal recessive disorder congenital analbuminemia would seem to suggest otherwise. Although it is extremely rare (estimated at less than one in a million), it is able to teach us important physiologic lessons. The disease is caused by mutations in the human serum albumin (HSA) gene which result in a markedly decreased serum albumin concentration (estimated at between 1/100th to 1/1000th of the normal serum albumin concentration), individuals with the disorder generally do well, usually with only mild edema that may even go unrecognized in childhood. Individuals do not have proteinuria but often have hyperlipidemia. It is postulated that oncotic pressure is maintained in part by an increased hepatic synthesis of other plasma proteins to compensate.

The generally mild phenotype is fairly surprising in my opinion because of the many physiologic roles ascribed to albumin in biology. It also hints that perhaps something other than the hypoalbuminemia and subsequent loss of oncotic pressure observed in patients with nephrotic syndrome is responsible for their edema.