Beatriz Fernandez-Fernandez

Tenofovir Nephrotoxicity: 2011 Update

Tenofovir is an acyclic nucleotide analogue reverse-transcriptase inhibitor structurally similar to the nephrotoxic drugs adefovir and cidofovir. Tenofovir is widely used to treat HIV infection and approved for treatment of hepatitis B virus. Despite initial cell culture and clinical trials results supporting the renal safety of tenofovir, its clinical use is associated with a low, albeit significant, risk of kidney injury. Proximal tubular cell secretion of tenofovir explains the accumulation of the drug in these mitochondria-rich cells. Tenofovir nephrotoxicity is characterized by proximal tubular cell dysfunction that may be associated with acute kidney injury or chronic kidney disease. Withdrawal of the drug leads to improvement of analytical parameters that may be partial. Understanding the risk factors for nephrotoxicity and regular monitoring of proximal tubular dysfunction and serum creatinine in high-risk patients is required to minimize nephrotoxicity. Newer, structurally similar molecular derivatives that do not accumulate in proximal tubules are under study.

Therapeutic approaches to diabetic nephropathy—beyond the RAS

Despite improvements in glycaemic and blood pressure control, and the efficacy of renin–angiotensin system (RAS) blockade for proteinuria reduction, diabetic nephropathy is the most frequent cause of end-stage renal disease in developed countries. This finding is consistent with the hypothesis that key pathogenetic mechanisms leading to progression of renal disease are not modified or inactivated by current therapeutic approaches. Although extensive research has elucidated molecular signalling mechanisms that are involved in progression of diabetic kidney disease, a number of high-profile clinical trials of potentially nephroprotective agents have failed, highlighting an insufficient understanding of pathogenic pathways. These include trials of paricalcitol in early diabetic kidney disease and bardoxolone methyl in advanced-stage disease. Various strategies based on encouraging data from preclinical studies that showed renoprotective effects of receptor antagonists, neutralizing antibodies, kinase inhibitors, small compounds and peptide-based technologies are currently been tested in randomized controlled trials. Phase II clinical trials are investigating approaches targeting inflammation, fibrosis and signalling pathways. However, only one trial that aims to provide evidence for marketing approval of a potentially renoprotective drug (atrasentan) is underway—further research into the potential nephroprotective effects of novel glucose-lowering agents is required.

Identification of a urine metabolomic signature in patients with advanced-stage chronic kidney disease

The prevalence of chronic kidney disease (CKD) is increasing and frequently progresses to end-stage renal disease. There is an urgent demand to discover novel markers of disease that allow monitoring disease progression and, eventually, response to treatment. To identify such markers, and as a proof of principle, we determined if a metabolite signature corresponding to CKD can be found in urine. In the discovery stage, we analyzed the urine metabolome by NMR of 15 patients with CKD and compared that with the metabolome of 15 healthy individuals and found a classification pattern clearly indicative of CKD. A validation cohort of urine samples from an additional 16 patients with CKD and 15 controls was then analyzed by (Selected Reaction Monitoring) liquid chromatography-triple quadrupole mass spectrometry and indicated that a group of seven urinary metabolites differed between CKD and non-CKD urine samples. This profile consisted of 5-oxoproline, glutamate, guanidoacetate, α-phenylacetylglutamine, taurine, citrate, and trimethylamine N-oxide. Thus, we identified a panel of urine metabolites differentially present in urine that may help identify and monitor patients with CKD.

Osteoprotegerin in Exosome-Like Vesicles from Human Cultured Tubular Cells and Urine

Urinary exosomes have been proposed as potential diagnostic tools. TNF superfamily cytokines and receptors may be present in exosomes and are expressed by proximal tubular cells. We have now studied the expression of selected TNF superfamily proteins in exosome-like vesicles from cultured human proximal tubular cells and human urine and have identified additional proteins in these vesicles by LC-MS/MS proteomics. Human proximal tubular cells constitutively released exosome-like vesicles that did not contain the TNF superfamily cytokines TRAIL or TWEAK. However, exosome-like vesicles contained osteoprotegerin (OPG), a TNF receptor superfamily protein, as assessed by Western blot, ELISA or selected reaction monitoring by nLC-(QQQ)MS/MS. Twenty-one additional proteins were identified in tubular cell exosome-like vesicles, including one (vitamin D binding protein) that had not been previously reported in exosome-like vesicles. Twelve were extracellular matrix proteins, including the basement membrane proteins type IV collagen, nidogen-1, agrin and fibulin-1. Urine from chronic kidney disease patients contained a higher amount of exosomal protein and exosomal OPG than urine from healthy volunteers. Specifically OPG was increased in autosomal dominant polycystic kidney disease urinary exosome-like vesicles and expressed by cystic epithelium in vivo. In conclusion, OPG is present in exosome-like vesicles secreted by proximal tubular epithelial cells and isolated from Chronic Kidney Disease urine.

Fn14 in podocytes and proteinuric kidney disease.

Non-proliferative proteinuric diseases are the most common primary glomerular disorders causing end-stage renal disease. These disorders may associate low level glomerular inflammation and podocyte expression of inflammatory mediators. However, the factors regulating podocyte expression of inflammatory mediators in vivo in non-immune disorders are poorly understood. We have now explored the regulation and role of TWEAK receptor Fn14 in mediating glomerular inflammation in cultured podocytes and in experimental and human non-immune proteinuria. Transcriptomics disclosed Fn14 and MCP-1 mRNA upregulation in glomeruli from patients with focal segmental glomerulosclerosis, as well as a correlation between the expression of both transcripts. Increased glomerular Fn14 and MCP-1 mRNA was confirmed in a second focal segmental glomerulosclerosis cohort and was also observed in membranous nephropathy. In human non-proliferative proteinuric kidney diseases podocytes displayed Fn14 and MCP-1 expression and NFκB activation. Podocyte Fn14 was increased in murine protein overload-induced proteinuria. In Fn14 knock-out mice with protein overload-induced proteinuria, glomerular and periglomerular macrophage infiltrates were reduced, as were MCP-1 mRNA and podocyte MCP-1 staining and podocyte numbers preserved as compared to wild-type counterparts. Adenovirus-mediated overexpression of TWEAK increased periglomerular macrophage infiltration in mice without prior kidney injury. In cultured podocytes inflammatory cytokines increased Fn14 mRNA and protein levels. TWEAK activated NFκB and increased MCP-1 mRNA and protein, an effect prevented by the NFκB inhibitor parthenolide. In conclusion, Fn14 activation results in NFκB-mediated pro-inflammatory effects on podocytes that may be relevant for the pathogenesis of non-proliferative proteinuric kidney disease of non-immune origin.

TWEAK/Fn14 and non-canonical NF-kappaB signaling in kidney disease

The incidence of acute kidney injury (AKI) and chronic kidney disease (CKD) is increasing. However, there is no effective therapy for AKI and current approaches only slow down, but do not prevent progression of CKD. TWEAK is a TNF superfamily cytokine. A solid base of preclinical data suggests a role of therapies targeting the TWEAK or its receptor Fn14 in AKI and CKD. In particular TWEAK/Fn14 targeting may preserve renal function and decrease cell death, inflammation, proteinuria, and fibrosis in mouse animal models. Furthermore there is clinical evidence for a role of TWEAK in human kidney injury including increased tissue and/or urinary levels of TWEAK and parenchymal renal cell expression of the receptor Fn14. In this regard, clinical trials of TWEAK targeting are ongoing in lupus nephritis. Nuclear factor-kappa B (NF-κB) activation plays a key role in TWEAK-elicited inflammatory responses. Activation of the non-canonical NF-κB pathway is a critical difference between TWEAK and TNF. TWEAK activation of the non-canonical NF-κB pathways promotes inflammatory responses in tubular cells. However, there is an incomplete understanding of the role of non-canonical NF-κB activation in kidney disease and on its contribution to TWEAK actions in vivo.

Targeting inflammation in diabetic kidney disease: early clinical trials

ABSTRACT Introduction: The age-standardized death rate from diabetic kidney disease increased by 106% from 1990 to 2013, indicating that novel therapeutic approaches are needed, in addition to the renin-angiotensin system (RAS) blockers currently in use. Clinical trial results of anti-fibrotic therapy have been disappointing. However, promising anti-inflammatory drugs are currently on phase 1 and 2 randomized controlled trials. Areas covered: The authors review the preclinical, phase 1 and 2 clinical trial information of drugs tested for diabetic kidney disease that directly target inflammation as a main or key mode of action. Agents mainly targeting other pathways, such as endothelin receptor or mineralocorticoid receptor blockers and vitamin D receptor activators are not discussed. Expert opinion: Agents targeting inflammation have shown promising results in the treatment of diabetic kidney disease when added on top of RAS blockade. The success of pentoxifylline in open label trials supports the concept of targeting inflammation. In early clinical trials, the pentoxifylline derivative CTP-499, the CCR2 inhibitor CCX140-B, the CCL2 inhibitor emapticap pegol and the JAK1/JAK2 inhibitor baricitinib were the most promising drugs for diabetic kidney disease. The termination of trials testing the anti-IL-1β antibody gevokizumab in 2015 will postpone the evaluation of therapies targeting inflammatory cytokines.

Cardiovascular risk biomarkers in CKD: the inflammation link and the road less traveled

End-stage renal disease patients suffer a syndrome of accelerated aging characterized by a 10- to 100-fold increase in cardiovascular and all-cause mortality when compared to age-matched controls. No specific therapeutic interventions have been shown to improve this dismal outcome. Inflammation, chronic kidney disease-mineral and bone disorder (CKD-MBD) and other biomarkers predict outcome in observational studies. However, we lack clinical trials that address the role of these biomarkers in risk stratification and therapeutic decision making. Biomarkers may also provide insights into the pathophysiology of disease and identify novel therapeutic targets. Inflammation emerges as a prime potential target for intervention. Thus, CKD-MBD biomarkers, asymmetrical dimethyl arginine and tri-iodothyronine have a link to inflammation. Interleukin-6 (IL-6) is one of the inflammation biomarkers with highest predictive value for outcome in ESRD. Biologicals targeting IL-6 are approved for the treatment of chronic inflammatory conditions such as rheumatoid arthritis. Furthermore, trials are underway to test IL-6 targeting potential to decrease cardiovascular injury in non-CKD patients. In this regard, targeting IL-1 was recently shown to decrease systemic inflammation in hemodialysis patients. The success of these trials will likely influence future studies on biomarker targeting in CKD.

Horizon 2020 in Diabetic Kidney Disease: The Clinical Trial Pipeline for Add-On Therapies on Top of Renin Angiotensin System Blockade

Diabetic kidney disease is the most frequent cause of end-stage renal disease. This implies failure of current therapeutic approaches based on renin-angiotensin system (RAS) blockade. Recent phase 3 clinical trials of paricalcitol in early diabetic kidney disease and bardoxolone methyl in advanced diabetic kidney disease failed to meet the primary endpoint or terminated on safety concerns, respectively. However, various novel strategies are undergoing phase 2 and 3 randomized controlled trials targeting inflammation, fibrosis and signaling pathways. Among agents currently undergoing trials that may modify the clinical practice on top of RAS blockade in a 5-year horizon, anti-inflammatory agents currently hold the most promise while anti-fibrotic agents have so far disappointed. Pentoxifylline, an anti-inflammatory agent already in clinical use, was recently reported to delay estimated glomerular filtration rate (eGFR) loss in chronic kidney disease (CKD) stage 3–4 diabetic kidney disease when associated with RAS blockade and promising phase 2 data are available for the pentoxifylline derivative CTP-499. Among agents targeting chemokines or chemokine receptors, the oral small molecule C-C chemokine receptor type 2 (CCR2) inhibitor CCX140 decreased albuminuria and eGFR loss in phase 2 trials. A dose-finding trial of the anti-IL-1β antibody gevokizumab in diabetic kidney disease will start in 2015. However, clinical development is most advanced for the endothelin receptor A blocker atrasentan, which is undergoing a phase 3 trial with a primary outcome of preserving eGFR. The potential for success of these approaches and other pipeline agents is discussed in detail.

CXCL16 in kidney and cardiovascular injury

CXC chemokine ligand 16 (CXCL16) is a CXC soluble chemokine, an adhesion molecule and a cell surface scavenger receptor. CXCL16 regulates inflammation, tissue injury and fibrosis. Parenchymal renal cells, vascular wall cells, leukocytes and platelets express and/or release CXCL16 under the regulation of inflammatory mediators. CXCL16 expression is increased in experimental and human nephropathies. Targeting CXCL16 protected from experimental glomerular injury or interstitial fibrosis. Conflicting results were reported for experimental cardiovascular injury. High circulating CXCL16 levels are associated to human kidney and cardiovascular disease and urinary CXCL16 may increase in kidney injury. In conclusion, mounting evidence suggests a role of CXCL16 in kidney and cardiovascular disease. However, a better understanding is still required before exploring CXCL16 targeting in the clinic.