**10. The future of biomarkers**

Development of new technologies involved in molecular biology, analysis of m-RNA expres‐ sion, proteomics and metabolomics create a possibility of discovery of new markers for early diagnosis of AKI and IF/TA. Relatively new method of microarrays (microarrays of cDNA and oligonuclotides- DNA chips) are sets of molecular probes attached to solid background in strictly determined order constituting two dimensional system of microscopic areas with defined sequences of nucleic acid. Microarray technology allow for detection of thousands of molecules of nucleic acids due to possibility of performing simultaneously many hybridization experiments (Dean et al., 2012). DNA microarrays technology permit for simultaneous monitoring expression of many genes (Scian et al., 2011). Identification of these genes constitute further step in earlier diagnosis and better prognosis of TA/IF(tubular atrophy/interstitial fibrosis).

**Proteomic techniques** Recently broadly applied proteomic techniques facilitate discovery of new biomarkers useful in evaluation of transplanted kidney function. Proteomics apply protein analysis using techniques such as MS e.g. (MALDI-TOF-Matrix Assisted Laser Desorption Ionisation - Time of Flight; SELDI-TOF-Surface Enhanced Laser Desorption Ionisation - Time of Flight; ES multielementary I - LTQ – FTICR-Electrospray Ionisation - Linear Trap Quadrupole - Fourier Transform Ion Cyclotron Resonance). Proteomics combine series of techniques for simultaneous analysis of hundreds or thousands of cells proteins. Proteomics objective is not only creation of the list of important proteins, but first of all exploration of differences in protein profiles of healthy and diseased people. Proteomic identification of urinary protein profiles is an noninvasive method for detection of renal proximal tubules dysfunction of transplanted kidney (Srivastava et al., 2011; Gwinner, 2007). Proteomic techniques are alternative for diagnostics based on single markers, because it allows for simultaneous analysis of large numbers of protein and peptide markers creating specific "finger print" of disease. Proteomics determines pattern of expression or secretion taking into account qualitative and quantitative relations between peptides and proteins produced in defined pathophysiological conditions.

**Metabolomics** based on analysis sets of metabolites connected with proteins, lipids, carbohy‐ drates, hormones, etc. evaluate qualitative and quantitative relations between particular metabolites. Due to metabolomics it is possible to determine definite metabolites characteristic for specific groups of diseases and changes occurring under influence of genetic and patho‐ physiological stimuli (Wishart, 2006).

New technologies and bioinformatics tools offer tremendous research possibilities which should make possible now and in the future precise monitoring of kidney graft, allow early detection and treatment of renal graft rejection and allow both for preventing and treatment of renal transplant complications as well as to improve number of long term patients survival.


(AUC 0.984) or albuminuria (AUC 0.984). μ-GST is an early biomarker for Henle's loop and

Development of new technologies involved in molecular biology, analysis of m-RNA expres‐ sion, proteomics and metabolomics create a possibility of discovery of new markers for early diagnosis of AKI and IF/TA. Relatively new method of microarrays (microarrays of cDNA and oligonuclotides- DNA chips) are sets of molecular probes attached to solid background in strictly determined order constituting two dimensional system of microscopic areas with defined sequences of nucleic acid. Microarray technology allow for detection of thousands of molecules of nucleic acids due to possibility of performing simultaneously many hybridization experiments (Dean et al., 2012). DNA microarrays technology permit for simultaneous monitoring expression of many genes (Scian et al., 2011). Identification of these genes constitute further step in earlier diagnosis and better prognosis of TA/IF(tubular atrophy/interstitial

**Proteomic techniques** Recently broadly applied proteomic techniques facilitate discovery of new biomarkers useful in evaluation of transplanted kidney function. Proteomics apply protein analysis using techniques such as MS e.g. (MALDI-TOF-Matrix Assisted Laser Desorption Ionisation - Time of Flight; SELDI-TOF-Surface Enhanced Laser Desorption Ionisation - Time of Flight; ES multielementary I - LTQ – FTICR-Electrospray Ionisation - Linear Trap Quadrupole - Fourier Transform Ion Cyclotron Resonance). Proteomics combine series of techniques for simultaneous analysis of hundreds or thousands of cells proteins. Proteomics objective is not only creation of the list of important proteins, but first of all exploration of differences in protein profiles of healthy and diseased people. Proteomic identification of urinary protein profiles is an noninvasive method for detection of renal proximal tubules dysfunction of transplanted kidney (Srivastava et al., 2011; Gwinner, 2007). Proteomic techniques are alternative for diagnostics based on single markers, because it allows for simultaneous analysis of large numbers of protein and peptide markers creating specific "finger print" of disease. Proteomics determines pattern of expression or secretion taking into account qualitative and quantitative relations between peptides and proteins produced in

**Metabolomics** based on analysis sets of metabolites connected with proteins, lipids, carbohy‐ drates, hormones, etc. evaluate qualitative and quantitative relations between particular metabolites. Due to metabolomics it is possible to determine definite metabolites characteristic for specific groups of diseases and changes occurring under influence of genetic and patho‐

New technologies and bioinformatics tools offer tremendous research possibilities which should make possible now and in the future precise monitoring of kidney graft, allow early detection and treatment of renal graft rejection and allow both for preventing and treatment of renal transplant complications as well as to improve number of long term patients survival.

distal tubules damage (Tonomura et al., 2010).

74 Current Issues and Future Direction in Kidney Transplantation

**10. The future of biomarkers**

defined pathophysiological conditions.

physiological stimuli (Wishart, 2006).

fibrosis).


Scientiscs are still looking for the "kidney troponin". Actually, more than ten promising biomarkers for kidney damage have been identified. The most relevant and the best studied substances are neutrophil gelatinase-associated lipocalin (NGAL), cystatin C, kidney injury molecule-1 (KIM-1), beta-2 microglobulin (β2M), and interleukin-18 (IL- 18). In kidney allograft recipients, urinary KIM-1 expression provides prognostic information in relation to the rate of renal function decline, irrespective of the kidney pathology (Ting et al., 2012; Han

Utility of Urinary Biomarkers in Kidney Transplant Function Assessment

http://dx.doi.org/10.5772/54746

77

Validation of those kidney markers in various pathologic conditions is actually ongoing. However, the majority of publications reviewed are small cross-sectional studies, and there are only a handful of longitudinal studies. Another important point is that biomarkers only have clinical value if the results are reproducible. However none of the biomarkers reviewed here have been studied in more than 2 longitudinal trials so their clinical applicability needs

Among enzymes which retain high diagnostic value in diagnostics of renal diseases are: hexosaminidase and its isoenzyme B as a marker of the proximal tubular damage as well as AAP or GST as a marker of the tubular brush border membrane. Cytosolic FBP-1,6 is of great diagnostic value for assessment of graft function. It is commonly believed that appropriate panel of urinary proteins and enzymes may by a practical marker for evaluation of the nephron function of transplant kidney and prognosis of the renal allograft fate. In the future, discovery of new biomarkers and research techniques may change practical approach to treating patients with renal grafts. In summary we feel it is necessary for an international body to develop a renal marker utility grading system, to evaluate the usefulness of particular markers of nephron function and to make recommendations for the use of renal transplant markers,

similar to those instilled for tumor markers (Hayes et al., 1996; Locker et al., 2006).

, Sylwia Chojnowska3

, Krzysztof Zwierz7

3 Medical Institute, College of Computer Science and Business Administration, Łomża, Po‐

4 Department of Radiology, Children Hospital, Medical University of Białystok, Poland

6 Department of Pediatric Nephrology, Medical University of Białystok, Poland

5 Department of Emergency Medicine and Disasters, Medical University, Białystok, Poland

, Beata Zalewska-Szajda4

and Sławomir Dariusz Szajda<sup>5</sup>

,

to be confirmed in good quality, long-term, large longitudinal trials.

et al., 2002; Szeto et al., 2010).

**Author details**

Jerzy Robert Ładny<sup>5</sup>

, Napoleon Waszkiewicz2

, Anna Wasilewska6

1 The Children's Memorial Health Institute, Warsaw, Poland

2 Department of Psychiatry, Medical University, Białystok, Poland

7 Medical College the Universal Education Society, Łomża, Poland

Alina Kępka<sup>1</sup>

land

**Table 1.** Urinary biomarkers for the early detection of acute and chronic allograft dysfunction.

#### **11. Conclusion**

In this chapter we presented traditional and new biomarkers for diagnostics and monitoring condition of transplant kidneys. Urine is practical, easy to obtain, noninvasive material for diagnosis of kidney diseases. Numerous reports from molecular biology, genetics, proteomics and metabolomics disclosed an array of new markers specifically connected with damage of specific nephron segments in the course of successive steps of disease. Particular expectations are connected with proteins represented particular nephron section, or produced locally in the place of nephron damage. Presence of cytokines and chemokines in urine is an early sign of renal inflammatory state, due to influx of granulocytes to the damaged nephron area. Majority of traditional biomarkers, particularly enzymuria retains diagnostic value in an evaluation of the renal tubules function. Multitude of presented biomarkers suggest their limited diagnostic value. Discovering universal marker seems to be very difficult. However, it is potentially more fruitful to identity the putative biomarker proteins useful in diagnostics of kidney disease. Scientiscs are still looking for the "kidney troponin". Actually, more than ten promising biomarkers for kidney damage have been identified. The most relevant and the best studied substances are neutrophil gelatinase-associated lipocalin (NGAL), cystatin C, kidney injury molecule-1 (KIM-1), beta-2 microglobulin (β2M), and interleukin-18 (IL- 18). In kidney allograft recipients, urinary KIM-1 expression provides prognostic information in relation to the rate of renal function decline, irrespective of the kidney pathology (Ting et al., 2012; Han et al., 2002; Szeto et al., 2010).

Validation of those kidney markers in various pathologic conditions is actually ongoing. However, the majority of publications reviewed are small cross-sectional studies, and there are only a handful of longitudinal studies. Another important point is that biomarkers only have clinical value if the results are reproducible. However none of the biomarkers reviewed here have been studied in more than 2 longitudinal trials so their clinical applicability needs to be confirmed in good quality, long-term, large longitudinal trials.

Among enzymes which retain high diagnostic value in diagnostics of renal diseases are: hexosaminidase and its isoenzyme B as a marker of the proximal tubular damage as well as AAP or GST as a marker of the tubular brush border membrane. Cytosolic FBP-1,6 is of great diagnostic value for assessment of graft function. It is commonly believed that appropriate panel of urinary proteins and enzymes may by a practical marker for evaluation of the nephron function of transplant kidney and prognosis of the renal allograft fate. In the future, discovery of new biomarkers and research techniques may change practical approach to treating patients with renal grafts. In summary we feel it is necessary for an international body to develop a renal marker utility grading system, to evaluate the usefulness of particular markers of nephron function and to make recommendations for the use of renal transplant markers, similar to those instilled for tumor markers (Hayes et al., 1996; Locker et al., 2006).
