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NDT Advance Access published online on March 23, 2009
Nephrology Dialysis Transplantation, doi:10.1093/ndt/gfp010
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© The Author [2009]. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org


Kidney injury molecule-1 (KIM-1): a urinary biomarker and much more

Joseph V. Bonventre1

1 Renal Division, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA

Correspondence and offprint requests to: Joseph V. Bonventre, Brigham and Women's Hospital, Harvard Institutes of Medicine Rm 576, 4 Blackfan Circle, Boston, MA 02115, USA. Tel: +1-617-525-5960; Fax: +1-617-525-5965; E-mail: joseph_bonventre{at}hms.harvard.edu

Keywords: acute kidney injury; TIM-1; acute renal failure; phagocytosis; apoptosis



   KIM-1 kidney expression and function
Top
 KIM-1 kidney expression and...
KIM-1 as a biomarker
 References
 
The kidney injury molecule-1 (designated as Kim-1 in rodents, KIM-1 in humans) mRNA was identified using techniques of representational difference analysis, a PCR-based technique [1], which we carried out to find genes whose expression was markedly upregulated 24–48 h after ischaemia in the rat [2]. Kim-1 was the gene found to be most highly upregulated in this screen. A large pharmaceutical company consortium, using an unbiased genomic approach to evaluate genes upregulated with the nephrotoxin cisplatin, determined that Kim-1 was upregulated more than any other of the 30 000 genes tested [3]. There are a large number of studies in animals showing robust Kim-1 protein production in the affected segments of the proximal tubule whenever a toxin or pathophysiological state results in dedifferentiation of the epithelium (e.g. [4–6]). Dedifferentiation is a very early manifestation of the epithelial cell response to injury [7]. KIM-1 is also expressed, at much lower levels, in lymphocytes and has also been referred to as T-cell immunoglobulin mucin (TIM)-1 and HAVCR-1, hepatitis A virus cellular receptor-1. The protein has also been reported to be expressed in the cochlea in response to cisplatin-induced injury [8]. The KIM/TIM family consists of eight members in mice, six in rats and three in humans [9,10].

Using standard northern or western blot analyses and immunocytochemistry, KIM-1 gene or protein expression is undetectable in the normal kidney. With injury KIM-1 mRNA is rapidly made and protein is generated and localized at very high levels on the apical membrane of proximal tubule in that region where the tubule is most affected. In the case of experimental ischaemia in rodents, Kim-1 expression is predominantly in the S3 segment of the proximal tubule. In human ischaemic and toxic acute kidney injury (AKI) it is found in the three segments of the proximal tubule.

KIM-1 is a type I cell membrane glycoprotein which contains, in its extracellular portion, a novel six-cysteine immunoglobulin-like domain, two N-glycosylation sites and a T/SP rich domain characteristic of mucin-like O-glycosylated proteins. The structure of the protein led us to initially believe it had adhesion molecule properties [11] (Figure 1). KIM-1 is also expressed at high levels in patients with clear cell-type renal cell carcinoma (RCC) [12]. RCC, like renal tubular injury, is associated with proximal tubule cell dedifferentiation.


Figure 1
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  		Fig. 1 Structure of KIM-1. KIM-1 is a type 1 membrane glycoprotein that contains, in its extracellular portion, a novel six-cysteine immunoglobulin-like domain, two N-glycosylation sites and a T/SP rich domain characteristic of mucin-like O-glycosylated proteins. There is one transmembrane domain and a short intracellular domain with a tyrosine phosphorylation signalling motif present in the renal form (KIM-1b).

 
We are beginning to appreciate the functional role of KIM-1 in the kidney. KIM-1 confers on epithelial cells the ability to recognize and phagocytose dead cells that are present in the post-ischaemic kidney and contribute to the obstruction of the tubule lumen that characterizes AKI. KIM-1 is a phosphatidylserine receptor that recognizes apoptotic cells directing them to lysosomes. It also serves as a receptor for oxidized lipoproteins and hence is adept at recognizing apoptotic cell ‘eat me’ signals. Given these properties KIM-1 is unique in being the first non-myeloid phosphatidylserine receptor that transforms epithelial cells into semi-professional phagocytes [13,14]. In addition to the facilitation of clearance of the apoptotic debris from the tubular lumen, KIM-1 may play an important role in limiting the autoimmune response to injury since it is known in many systems that phagocytosis of apoptotic bodies is one mechanism for limiting the proinflammatory response. Acute protective responses, however, may not necessarily translate to chronic effects of KIM-1 expression, a clinically relevant issue, since we [15] and others [16] have found that many individuals with chronic renal failure express the KIM-1 protein in their proximal tubules.

The ectodomain of KIM-1 is shed from cells in vitro [11] and in vivo into the urine in rodents [4,5] and humans [17] after proximal tubular kidney injury or in patients with RCC [12]. This shedding is regulated, at least in part, by MAP kinase signalling pathways that are activated with stress [18]. Metalloproteinase activity results in the release of soluble KIM-1. Mutagenesis studies demonstrate that the juxtamembrane protein secondary structure affects susceptibility to the metalloproteinase-mediated KIM-1 cleavage [18]. The role of shed KIM-1 within the tubule remains unknown and the implications of the long-term expression of KIM-1 in patients with chronic kidney disease, albeit, at levels lower than that found in AKI, remain to be defined.



   KIM-1 as a biomarker
Top
 KIM-1 kidney expression and...
 KIM-1 as a biomarker
References
 
There were a number of characteristics of KIM-1 that led us to believe that the protein might make it an ideal biomarker of kidney injury: the absence of KIM-1 expression in the normal kidney; its marked upregulation and insertion into the apical membrane of the proximal tubule; its persistence in the epithelial cell until the cell has completely recovered; the rapid and robust cleavage of the ectodomain and the ex vivo room temperature stability of the ectodomain. There is an urgent need for better biomarkers for acute kidney injury (AKI) for its timely diagnosis, for the prediction of severity and outcome and for the monitoring of proximal tubule injury in AKI as well as chronic kidney disease. The classical methods of assessing renal function involve the measurement of serum blood urea nitrogen and creatinine, biomarkers that are insensitive and nonspecific, especially in the setting of AKI. It is also important to recognize that changes in serum creatinine and blood urea nitrogen concentrations primarily reflect functional changes in filtration capacity and are not true ‘injury markers’. We have recently reviewed KIM-1, and other biomarkers of kidney injury that are currently being evaluated, in the context of the definitions and diagnosis of AKI [19,20]. The restrictions of length in this commentary dictate only a brief discussion of animal data and the increasing volume of clinical data on this marker. There are a growing number of studies in animals that demonstrate the utility of Kim-1 in the urine as a noninvasive marker for kidney injury. A partial list of the settings in which the utility of Kim-1 has been demonstrated includes: ischaemia and angiotensin-mediated injury in the Ren rat, various toxins including cisplatin, S-(1,1,2,2-tetrafluoroethyl)-l-cysteine (TFEC), folic acid [4], gentamicin, mercury, chromium [6], cadmium [21], iodinated contrast agents [22], vancomycin, ochratoxin A, cyclosporine [23], d-serine, protein overload nephropathy [24] and ageing-induced nephropathy. Kim-1 othologues are present in many species besides rodents and man, including zebrafish, monkeys and dogs.

The first human subjects studies with KIM-1 were published in 2002 [17] where we demonstrated that there was a markedly increased expression of KIM-1 in kidney biopsy specimens from patients with a pathology diagnosis of acute tubular necrosis, and there were very high levels of KIM-1 ectodomain in the urine of patients with clinically significant AKI. Urinary levels of KIM-1 increased prior to the appearance of casts in the urine. There was less KIM-1 in the urine in less severe cases of AKI related to contrast exposure, but even in a limited number of these cases it was apparent that the level of urinary KIM-1 tracked the severity of disease as estimated by peak serum creatinine (analysis not included in paper). We subsequently showed that KIM-1 is also a sensitive marker for kidney injury in children undergoing cardiac surgery [25]. Liangos et al. evaluated urinary KIM-1 and another biomarker, N-acetylglucosaminidase (NAG) in 201 patients with clinically established AKI and reported that elevated levels of urinary KIM-1 and NAG were significantly associated with the clinical composite endpoint of death or dialysis requirement, even after adjustment for disease severity or comorbidity [26]. We evaluated the diagnostic performance of nine urinary biomarkers of AKI—KIM-1, neutrophil gelatinase-associated lipocalin (NGAL), interleukin-18 (IL-18), hepatocyte growth factor (HGF), cystatin C (Cys), N-acetyl-β-D-glucosaminidase (NAG), vascular endothelial growth factor (VEGF), chemokine interferon-inducible protein 10 (IP-10; CXCL10) and total protein—in a cross-sectional comparison of 204 patients with or without clinically documented AKI. In the case of each biomarker, the median urinary concentrations were significantly higher in patients with AKI than in those without AKI. The area under the receiver operating characteristics curve (AUC-ROC) for KIM-1 was 0.95 when the AKI patients were compared to healthy control patients. A combination of biomarkers using a logic regression model [risk score of 2.93 x (NGAL > 5.72 and HGF > 0.17) + 2.93 x (PROTEIN > 0.22) –2 x (KIM < 0.58)] was significantly greater (0.94) than individual biomarker AUC-ROCs when a number of hospitalized control groups were included (some of which likely had clinically silent AKI). Age-adjusted levels of urinary KIM-1, NAG, HGF, VEGF and total protein were significantly higher in patients who died or required renal replacement therapy (RRT) when compared to those who survived and did not require RRT [15].

Van Timmeren et al. stained for the presence of KIM-1 protein in tissue specimens from 102 patients who underwent a kidney biopsy for a variety of kidney diseases and showed that positive KIM-1 staining in dedifferentiated proximal tubular cells correlated with tubulo-interstitial fibrosis and inflammation. In a subset of patients who underwent urine collection near the time of biopsy, urinary KIM-1 levels correlated with the tissue expression of KIM-1 [16]. In an analysis of a small number of patients with non-diabetic renal disease, urinary KIM-1 levels were increased in proteinuric patients and were reduced if the proteinuric patients were treated with renin–angiotensin–aldosterone inhibition, sodium restriction and or diuretic therapies that reduced the proteinuria, thus linking the degree of proteinuria to proximal tubule injury as quantitated by KIM-1 in the urine [27].

In studies of transplantation we quantified human KIM-1 protein expression in renal transplant biopsies by immunohistochemistry and correlated these findings with renal functional indices [28]. Although protocol biopsies showed no detectable tubular injury on histological examination, we found focal positive KIM-1 expression in 28% of the cases. This does not reflect a false positive test but rather the result of superior sensitivity of KIM-1 expression in detecting proximal tubule injury when compared to morphology alone. In this study of renal allografts KIM-1 expression was detected in 100% of biopsies from patients with deterioration in kidney function and histological changes indicative of tubular damage. KIM-1 expression was significantly correlated with levels of serum creatinine and BUN concentrations and inversely correlated with estimated glomerular filtration rate on the biopsy day. KIM-1 was expressed focally in affected tubules in 92% of kidney biopsies from patients with acute cellular rejection reflecting the epithelial cell injury that results from a severe cellular rejection.

Van Timmeren et al. also evaluated the utility of urinary KIM-1 in renal transplant recipients. They looked at a cohort with a median of 6 years post-transplant, and we measured baseline KIM-1 excretion in stored urine samples. Graft loss was monitored over time [29]. The occurrence of graft loss increased with increasing tertiles of KIM-1 excretion. High KIM-1 levels were associated with low creatinine clearance, proteinuria and high donor age. KIM-1 levels predicted graft loss independent of creatinine clearance, proteinuria and donor age.

In summary, urinary KIM-1 is a scavenger receptor on renal epithelial cells which converts the normal proximal tubule cell into a phagocyte. KIM-1 expression is not measurable in normal proximal tubule cells and is markedly upregulated with injury/dedifferentiation. It is highly expressed on the apical domain of the cell and its large ectodomain is cleaved and is stable as it is excreted in the urine. Its presence in the urine is highly specific for kidney injury. No other organs have been shown to express KIM-1 to a degree that would influence kidney excretion. It has been shown to be much more sensitive than BUN and creatinine as a marker for injury in a large number of preclinical studies with a large number of kidney insults, including various toxins. It is a true translational biomarker in that its behaviour in man mirrors its behaviour in animals and hence is likely to be very useful in drug development and kidney safety monitoring. In fact the FDA and EMEA have included KIM-1 in the small list of kidney injury biomarkers that they will now consider in the evaluation of kidney damage as part of their respective drug review processes of new drugs [30]. More prospective studies are in progress in humans to delineate the many kidney disease processes that will be better detected and followed by using a simple urinary test that may serve as a surrogate for the kidney biopsy. While the role of KIM-1 as a biomarker has a robust future, it will be particularly interesting to explore further its role in acute and chronic kidney disease as this knowledge may lead to a role not only as a biomarker but also as a therapeutic target.



   Acknowledgments
 
The author thanks Drs Takaharu Ichimura, Vishal Vaidya and Suskrat Waikar as well as many collaborators in my laboratory and around the world who have contributed to the work presented. Work described that has been performed in Dr Bonventre's laboratory has been supported by US National Institutes of Health grants DK39773, DK72831 and DK74099.

Conflict of interest statements. Dr Bonventre is a coinventor on KIM-1 patents that have been licensed by Partners Health Care to Johnson and Johnson Inc. and Biogen Idec. He serves as a consultant to Johnson and Johnson, Inc.



   References
Top
 KIM-1 kidney expression and...
 KIM-1 as a biomarker
 References
 

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Received for publication: 30.12.08
Accepted in revised form: 5. 1.09


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