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BIOMARKERS IN ACUTE KIDNEY INJURY

Dr MÜGE AYDOĞDUGAZİ UNIVERSITY MEDICAL FACULTY

PULMONARY DISEASES DEPARTMENT AND CRITICAL CARE UNIT13.10.2014

Acute Kidney Injury (AKİ): Epidemiology • AKI occurs in 5% of hospitalised patients and in

30-50% of critical care patients * ;

• Incidence is increasing with extensive ICU procedures!!!!

• Mortality among the ICU patients with AKI requiring RRT is; > %50*.

• Among the ICU patients receiving RRT; CKD or ESRD develops with in 3-5 years*

*Cole vd., 2000; Gursel ve Demir, 2006; Hoste vd., 2006; KDIGO, 2012; Uschino vd., 2005

AKI-Why Early Diagnosis? • AKI can be reversible when diagnosed earlier. • With eary diagnosis;

• It will be possible to start treatment earlier• Nephrotoxic procedures or medications can be avoided

AKI-Why Early Diagnosis?

Hoste et al. Crit Care 2006; 10(3): R73

AKI- Problems of Diagnosis in ICU

• Etiology of AKI is multifactorial (nephrotoxic drug use, sepsis, ischemia etc)

RIFLE ve AKIN diagnostic criteria depend on serum creatinin and urine output levels

These will lead to misdiagnosis or delayed diagnosis in ICU

AKI: Problems of Early DiagnosisSerum Creatinin Caused Delayed Diagnosis

AKI: Problems of Early Diagnosis

Factors decreasing creatinin levels: Muscle mass and activity Hepatic failure Sepsis Body weight Age Sex Gender Protein intake Excess volume Pregnancy IADH Blood transfusion

Factors increasing creatinin levels : Trauma, rhabdomyolisis

Excessive protein intake

Fever

Immobilisation

Drugs (TMP, Cimetidine, Triamterene, Probenecide, Spironolacton, Amiloride)


• BUN• Low molecular weight,water soluble by-product of protein

metablolism

• Serum marker of uremic solute retansion and elimination.

• Not a sensitive marker of renal injury

• Increased BUN levels with extrarenal factors :• High protein diet

• Critical illness (sepsis, burn, trauma)

• GI bleeding

• Glucocorticoid treatment and tetracycline use

• Decreased BUN levels with extrarenal factors• Chronic hepatic disease

• Low protein intake

Oliguria

• At least 6 hours are necessary for detection of oliguria

• In most of the patients excretory function of kidneys are not disturbed during AKI

• Misdiagnosis can be made by just looking oliguria in acute tubular injury

• Trend should be followed in ICU

• Has low sensitivity and specificity


AKI-Clinical Continuity

Normal

İnjury ????

GFR↓

SCr ↑

Renal Failure

AKI Biomarkers :Why Necessary? • To make earlier diagnosis of structural abnormalities during

renal injury before funtional abnormalities take place

• To make differential diagnosis of reversible AKI and CKD

• Risk classification and triage

• To start treatment and to take preventive measure as soon as possible

• To define indications and criteria for future therapies.

• To identify prognosis (dialysis, death, ICU and hospital stay)

• Similar to Troponin and BNP

• Monitorization of treatment response

How Biomarkers Are Developed?• Phase 1-Discovery Phase

• To find the candidate biomarkes and to evaluate the biological competence by using scientific technologies• Gene analysis (cDNA microarray ve NextGen

sequencing)• Protein analysis (Proteomics)

• Phase 2-Translational Phase• To define and develop the assay methods, tests for

candidate biomarkers. • Phase 3-Validation Phase

• To evalute the prognostic value and clinical applicability of the biomarker with large clinical trials.

Devarajan, Biomarkers Med 201; 4: 265-80

How can be the ideal biomarker?1. Noninvasive2. Easy, rapid result, cheap, can be studied in an already

present sample like urine3. High sensitivity and specificity4. Rapid and reliable rise secondary to renal injury 5. Correlated with the level of renal injury6. Provide prognostic information and risk classification 7. Specific to kidneys8. Can be applied to differen populations. 9. Can show possible causes of renal injury (Prerenal,

intrarenal, postrenal)10. Stabile during time, at different temperatures and pHs11. Not affected from other drugs

AKI –Biomarker StudiesIn recent years many studies have being performed for

AKI early diagnosis, both from urine and plasma samples.

[ serum Cystatin C, urine IL-18 and neutrophil gelatinase associated lipocalin (NGAL), urine KIM-1(kidney injury marker-1) and urine L-FABP (liver fatty acid binding protein) ]

AKI Biomarkers

Neutrophil gelatinase associated lipocalin-2 (NGAL)

• 25 kDa protein, covalenty bound to neutrophil gelatinase

• Expressed at low levels in lungs, stomach, colon and renal proximal tupular epithelial cellls

• It was found to be increasing so rapidly with in 2 hours after an ischemic insult in (Mishra et al, 2003)

• In 2005, Mori et al identified that in AKI developing in ICU patients, plasma NGAL levels increase 10 times, and urine NGALlevels increase 100 times when compared with non AKI controls.

Woo. K

. Han C

urrr ent Biom

a rker Find ings 201 2

Woo. K. Han Currrent Biomarker Findings 2012

Extrarenal Factors for NGAL Rise: Systemic stress CKD Acute bacterial infection, sepsis Gastric malignity Pancreas carcinoma Endometrial hyperplasia Down Syndrome Pelvic inflamatory disease, inflam. Bowel disease Glioma Esential thrombocytemia, policytemia vera CML with molecular remission ST segment elevation MI, as a prognostic indicator Acute vs stabile coronary arterial disease

NGAL Valuable for detecting subclinical AKI

Does sublinical AKI affects long term prognosis????

Not an ideal biomarker due to low specificity

Serum Cystatin C• Cystatin C; low molecular weight endogenous systein

proteinase inhibitor.

• Cystatin C; can be synthesized and released to plasma at a stabile rate from all nucleated cells in the body

• Not affected from the age, gender, muscle mass and diet protperties of the patients .

• Superior to serum creatinine in predicting GFR and in follow up of renal functions. *

*Dhamidharka VR et al. Am J Kidney Disease 2002; 40: 221-6*Coll E et al. Am J Kidney Disease 2000; 36: 29-34

Serum Cystatin C• Has better performance in predicting GFR in some

special patient groups;• Elderly• Children• Renal transplant patients• Cirhosis• Malnutrition

• Serum Cystatin C, when compared with creatinin more sensitive to mild and earlier renal dysfunction

Serum Cystatin C

• In a small cohort study of critically ill patiens; rise in Cystatin C levels can be identified 1-2 days before the rise in serum creatinine (> 50% of the v-baseline) in AKI patients.

• Due to developing standart immunonephelometric assays serum Cystatin C is being used more commonly in our daily practice and gives rapid results.

*Herget-Rosenthal S. et al. Kidney Int 2004; 66: 115-1122

Serum Cystatin C• Among a large cross sectional study with 8058 patients some

other factors were found to be associated with increased serum Cystatin C levels*;• Old age, • Male gender, • Being tall and overweight, • Active smoking, • Increased CRP levels

• Cystatin C affected at the same time from**;• Thyroid functional abnormalities• Immunosuppressive treatment • Systemic inflammation

*Knight EL et al. Kidney Int 2004; 66: 1115-22**Manetti L et al. Clim Chim Acta 2005; 356:227-228

Manetti L et al. J Endocr Invest 2005; 28:346-349

Urine Cystatin C• Cystatin C; also identified in urine of AKI patients. This

can be used for evaluating the severity of tubular injury..

• In a small prospective study among the critically ill patients, increased urine Cystatin C levels were identified to be more successfull in predicting the need for RRT than other urinary biomarkers*

* Herget Rosenthal S et al. Clin Chem 2004; 50:552-558

Woo. K. Han Currrent Biomarker Findings 2012

AKI Biomarkers• TIMP-2;

• Matrix metalloproteinase inhibitor• Inhibits endothelial cell proliferation directly .

• IGFBP7: • Stimulates cell adhesion, plays a role in cell cycle arrest

AKI Biomarkers

TIMP2 ve IGFBP7

•IGFBP7 (Insulin Like Growth Factor Binding Protein-7) & TIMP-2 (Tissue Inhibitor of Metalloproteinase-2);G1 cell cycle arrest markers showing early cellular injury

•After injury renal tubular cells enter a short period of G1 cell cycle arrest

•G1 cell cycle arrest is a kind of protective mechanismi preventing the division and multiplication of injured DNA

•IGFBP7 & TIMP-2 at the same time plays a role of “alarm” released by otocrine and paracrine ways from the site of injury Kashani et al. Critical Care 2013, 17:R25

In the earlier period of sepsis , p53 ve p21

proteins will stop cell cycle, and together with

released IGFBP-7 ve TIMP-2 cause the G1 cell cycle

arrest.

If no new injury occurs and if sepsis treated properly cyclin/CDK4

complex will allow cell cycle to continue into

G1/S, and will lead to renal cellular regeneration

AKI Biomarkers

Urine IGFBP-7 ve TIMP-2 first studied in; Early diagnosis of urinary system malignancies Animal studies Sapphire Study of Kashani K et al. in identifying AKI in early

period of AKI in septic ICU patients

*Are g g e r F, Ue hling e r DE, Wito ws ki J, Brunis ho lz RA, Hunz ike r P, Fre y FJ, Jö rre s A. 2 0 1 3 . “Id e ntific a tio n o f IG FBP-7 by urina ry p ro te o m ic s a s a no ve l p ro g no s tic m a rke r in e a rly a c ute

kidne y injury ” . Kidne y Inte rna tio na l; 8 5 : 9 0 9 -9 1 9

**Biho ra c A, Cha wla L, Sha w AD, AL-Kha fa ji A, Da vis o n DL, De Muth G E, e t a l. 2 0 1 4. “Va lida tio n o f c e ll c y c le a rre s t bio m a rke rs fo r a c ute kidne y injury us ing c linic a l a d jud ic a tio n” . Am J o f Re s p ira to ry

a nd Critic a l Ca re Me d ic ine ; 1 8 9 (8 ): 9 3 2 -9 3 9 .

***Ka s ha ni K, Kha fa ji AA, Ard ile s T, Artig a s A, Ba g s ha w SM, Be ll M, e t a l. 2 0 1 3 . “Dis c o ve ry a nd va lida tio n o f c e ll c y c le a rre s t b io m a rke rs in hum a n a c ute kidne y injury ” . Crit Ca re ; 1 7 : R2 5

AKI Biomarkers

• Kashani K et al., evaluated the predictive value of TIMP2.IGFBP-7 in AKI development with in 12 hours of admission among the urine samples obtained in first 24 hours of admission to ICU.

• An additive affect was identified when urine TIMP2 and IGFBP7 were evaluated together; AUC was 0.80

(This was 0.76 ve 0.79 when these parameters were evaluated by themselves)).

Kashani et al. Critical Care 2013, 17:R25

AKI Biomarkers

Urine NGAL (ng/mL) [TIMP-2]•[IGFBP7] ((ng/mL)2 / 1000)


AKI Biomarkers

Urine KIM-1 (ng/mL) [TIMP-2]•[IGFBP7] ((ng/mL)2 / 1000)


AKI Biomarkers

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

Risk for AKI (KDIGO Stage 2-3)

A

0

0.1

0.2

0.3

0.4

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0.7

0.8

0.9

1

0.01 0.1 1 10 100

Risk for MAKE30

[TIMP-2]•[IGFBP7] ((ng/mL)²/1000)

B

KDIGO 2-3 in 12h

MAKE30MAKE30 (30 days)DeathRRTPersistently elevated sCr (2x over baseline)


• With the combination of two tests TIMP2-IGFBP7 (NEPHROCHECK® Test) early dignostic tool was developed and a threshold level was searched for the early diagnosis of AKİ;

• <0.3 (ng/ml)2/1000 low risk for (KDIGO 2-3) AKİ

• 0.3-2 (ng/ml)2/1000 high risk,

• > 2 (ng/ml)2/1000 highest risk

• < 0.3 (ng/ml)2/1000 cutt of level has a sensitivity of 0.88 and a negative predictive value of 0.94 for the exclusion of KDIGO 2-3 AKI development with in first 12 hours.

AKI Biomarkers


AKI Biomarkers IGFBP-7 in normal, sepsis ve

septic AKİ groupsTIMP-2 in normal, sepsis and ve

septic AKI groups

Normal Sepsis Sepsis+AKI

IGFBP-7 2,4 5,8 23,1

0

5

10

15

20

25

IGFBP-7 ng/ml

Normal Sepsis Sepsis+AKI

TIMP-2 593,00 478,00 527,00

0,00

100,00

200,00

300,00

400,00

500,00

600,00

700,00

TIMP-2 pg/ml

Aydogdu M ve ark. Septik Akut Böbrek Yetmezliği Erken Tanısında İdrar Hücre Siklusu Duraklama Belirteçleri:TIMP-2 ve IGFBP-7'nin Yeri. 11.DCYB Kongresi Sözlü Sunum(SS001)

Figure 3- ROC curve for the prediction of AKI with urine IGFBP-7 (AUC: 0.83)

IGFBP-7; for 3.5ng/ml cut off level; sensitivity %80, specificty %76

Area Under the Curve

Test Result Variable(s): ýgfbp7

,830 ,046 ,000 ,741 ,920Area Std. Error

aAsymptotic

Sig.b

Lower Bound Upper Bound

Asymptotic 95% ConfidenceInterval

The test result variable(s): ýgfbp7 has at least one tie between thepositive actual state group and the negative actual state group. Statisticsmay be biased.

Under the nonparametric assumptiona.

Null hypothesis: true area = 0.5b.

Aydogdu M ve ark. Septik Akut Böbrek Yetmezliği Erken Tanısında İdrar Hücre Siklusu Duraklama Belirteçleri:TIMP-2 ve IGFBP-7'nin Yeri. 11.DCYB Kongresi Sözlü Sunum(SS001)

TAKE HOME MESSAGES…• Early diagnosis of AKI is so important inorder to prevent

morbidity and mortality. • There are many biomarkers being studied, but no ideal

biomarker has yet been defined. • New cell cycle arrest markers are promising but the

studies are not enough• Whatever biomarker is preferred, it should be evaluated

together with the clinic of the patient…

Thank You…

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