Tests and monitoring in HIV infection

Tests and monitoring in HIV infection

UK standard of care and some other useful tests

Matthew WilliamsUK CAB




CD4 countViral load

ResistanceTherapeutic drug monitoring


CD4 count

●Blood test●Used to judge how far HIV disease has advanced●Helps predict the risk of opportunistic infections●Most useful when it is compared with the count obtained from an earlier test.


CD4 count

CD4 dips on HIV infection from a normal count of 500-1,500 cells in a cubic millimeter (mm3) of blood (a drop, more or less), recovers somewhat, then falls over time down to as low as 0.


CD4 count

CD4 <200 = greater risk of opportunistic infections (OIs) = “AIDS” = threshold for prophylaxis eg for PCP = bottom of UK threshold for starting combination therapy

CD4 <50 = very great risk of OIs


CD4 count

CD4 over 350: treatment not recommended

CD4 any count: treatment recommended if “symptomatic”

BHIVA, Treatment of HIV-infected adults with antiretroviral therapy (2006)


CD4 count

CD4 count lower in pregnancy – temporary drop of 50 cells/mm3


CD4 count

CD4% - CD4 cells as proportion of all lymphocytes (white blood cells), normally about 40% in adults

CD4% is used to monitor babies and children who have higher CD4 counts


Viral load

●Blood test (can be other serum eg CSF)●Used to judge whether treatment is working (early infection?)●Helps predict the risk of disease progression?●Most useful when it is compared with the count obtained from an earlier test.


Viral load

After infection, viral load surges to a very high for the first weeks or months

Often 1,000,000+ copies in a millilitre (mL) of blood, when you are very infectious

Viral load falls as the body controls HIV infection then rises over time as immunity is damaged


Viral load

When you are on HIV treatment, your viral load should be reduced to “undetectable” = <50 copies/mL = 1.7 log10


Viral load

Blip = 1 viral load test detectable (over 50 copies) – 2 of these may be a trend and indicate “virological failure”


Viral load

Viral load is usually a PCR (polymerase chain reaction) quantitative (counting) test for HIV RNA - research tests can measure viral load below 50 copies

Other tests: bDNA (branched DNA), NASBA (nucleic acid sequence based amplification)


Resistance test

●Blood test (can be other serum eg CSF)●Used to judge whether treatment will work●Involves interpretation●Two types: genotype and phenotype●Viral load needs to be over 500 copies/mL for the test to work


Resistance test

Resistance test


Resistance test

Genotype looks at genetic make up of HIV viruses in infection and compares this to what is known about which mutations lead to resistance

Phenotype looks at whether the drugs work in a test tube


Resistance test

Minority species of resistant virus may be missed by conventional resistance testing (ie if less than 10% of your virus is resistant the test may not pick this up).

In patients without evidence of transmitted resistance, a suboptimal virological response to first-line therapy (<1 log10 copies/mL drop in viral load by 4–8 weeks) should prompt resistance testing at that time.BHIVA, Treatment of HIV-infected adults with antiretroviral therapy (2006)


Resistance test


Therapeutic drug monitoring (TDM)

Measures drug levels in the blood - requires blood samples at recorded time intervals after a drug dose

Practical uses for NNRTIs and PIs



Freely available at low (£45/drug) or no cost (if covered by drug company) from University of Liverpool Department of Pharmacology.

http://www.hiv-druginteractions.org/

http://www.delphicdiagnostics.com/



Drug levels vary – much evidence of this – recommended dose is based on averages

University of Liverpool TDM audit – 20-25% of children on NNRTIs or PIs using drugs below therapeutic level.

http://www.i-base.info/htb/v7/htb7-6/University.html


UK standard of care (BHIVA)

All patients should have:i) a resistance test at diagnosis, ii) before starting HAART, iii) if viral load does not drop by <1log10 after 4-

8 weeks after starting HAART, (genotype) iv) after virological failure


UK standard of care

BHIVA – viral load before and 4-8 weeks after starting treatment (as necessary

thereafter)


UK standard of care

BHIVA – CD4 count before treatment (as necessary thereafter)


UK standard of care

BHIVA – TDM for management of drug interactions, pregnancy and paediatrics,

highly treatment-experienced patients when TDM and resistance test results can be integrated, kidney or liver impairment,

transplant patients, drug toxicity, alternative dosing where safety and efficacy has not

been established



Tests and monitoring in HIV infection● Liver enzymes● Kidney function● Urine● Albumin● Bone density● Glucose● Platelets● Red blood count● White blood count● Lipids

Tests and monitoring in HIV infection● C-reactive protein● DEXA scan● Chest x-ray● Sputum serology● Lactic acid and blood pH● Haemoglobin and iron● Serum urea● Creatine phosphokinase and lactate

dehydrogenase● Electrolytes


Tests = blood, blood and more blood...

Many tests require a blood sample


Blood glucoseBlood cholesterols and triglycerides

Kidney functionC-reactive protein

Liver enzymes

AnaemiaLactic acidosis


Blood glucose

Normal range 4-8 millimoles per litre (µmol/L)

High glucose ?= kidney disease, neuropathy, insulin resistance, cardiovascular disease

Drugs: PIs, tenofovir, AZT?


Blood cholesterols and triglycerides

Low-density lipoproteins (LDLs) or "bad" cholesterol, and high-density lipoproteins (HDLs) or "good" cholesterol – ratio is key measure

Insulin resistance, metabolic syndrome, cardiovascular disease

Drugs: PIs, AZT, efavirenz



Total cholesterol

Target level under 5.2 µmol/L (4-6.4 µmol/L)



LDL cholesterol

Target level under 3.4 µmol/L



HDL cholesterol

Target level over 0.9 µmol/L



Cholesterol – CV risk cannot be judged on cholesterol levels alone, HDL:LDL ratio, lifestyle, BMI, age and other factors are important

Smoking!



Triglycerides

Fasted levels <2.2 µmol/L normal, 2.2–4.4 µmol/l borderline, >11 µmol/L very high

Each +1.1 µmol/L increased the risk of a heart attack by about 25% in men and 60% in women


Urine

Dipstick test

●Protein – kidney (tenofovir) << mostly albumin●Glucose – insulin resistance (PIs), kidney (tenofovir, indinavir?, atazanavir?)●Bilirubin – liver ( X atazanavir)

Sensitive but not specific


Kidney function

Urine dipstick test or blood test = first test and is indicative but inconclusive


Kidney function

●Serum creatinine (blood, not very specific or sensitive)●Creatinine clearance (blood and/or urine samples over 24 hours)●Calculated creatinine clearance (1 blood sample) – also called estimated glomular filtration rate (eGFR)

Creatinine clearance can increase in pregnancy


Kidney function

Different formulas to calculate GFRhttp://en.wikipedia.org/wiki/Glomerular_filtration_rate


C-reactive protein

Risk of diabetes, hypertension and CV disease

Low risk: <1mg/LHigh risk: >3mg/L

Drugs: PIs?, AZT?

C-reactive protein is a plasma protein produced by the liver (so, another blood test...)


C-reactive protein

C-reactive protein is also marker of inflammation from infection but seems to be a reliable marker for CV disease in HIV regardless of HAART

http://gateway.nlm.nih.gov/MeetingAbstracts/102261383.html


Liver enzymes

AST, ALT, ALP, GGT and bilirubin

Many conditions, some specific indications, complicated by hepatitis B and C coinfection, pharmacological agents (all kinds) and food

Drugs: ritonavir, nevirapine, efavirenz, tipranavir, atazanavir, indinavir, d4T - most ARVS and many other drugs may affect the liver


Liver enzymes

ALT (alanine aminotransferase)

Normal range 7-30 units/L W, 10-55 units/L M

May be more reliable sign of liver damage


Liver enzymes

AST (aspartate aminotransferase)


Unreliable sign of liver damage

Pregnancy may decrease AST


Liver enzymes

ALP (alkaline phosphatase)


Non-specific sign of liver damage

Atazanavir and indinavir can raise ALP


Liver enzymes

GGT (gamma glutamyl transferase)

Normal range >50 units/L W, >65 units/L M

Can be specific sign of liver damage


Liver enzymes

Interpretation requires experience and the whole picture

●ALP+ GGT normal = bone disease?●ALP+ GGT+ = bile ducts? liver damage?●10 x ALT/AST = viral hepatitis? ARVs?


Liver enzymes

Bilirubin

Direct (unconjugated) 0-7 µmol/L Total 0-17 µmol/L

Bilirubin levels slightly higher in males than females, black Africans.

Drugs: atazanavir, indinavir


Liver enzymes

Bilirubin

Jaundice clinically detectable at levels above 40 µmol/l.

Exception: with atazanavir (or ritonavir) if bilirubin levels around 60-70 µmol/l


Liver enzymes

Bilirubin ++ jaundice

Other enzymes ++ may show no outward sign


Liver enzymes

Q: what is the most liver-damaging over-the-counter (OTC) medicine?


Liver enzymes

Q: what is the most liver-damaging over-the-counter (OTC) medicine?

A: Paracetamol


Liver enzymes

WHO's top 10 liver-damaging medicines

Paracetamol, troglitazone, valproic acid, d4T, halothene, 3TC, ddI, amiodarone, nevirapine, cotrimoxazole

The ABCs of liver disease, Edwin J Bernard, NAMhttp://www.aidsmap.com/files/file1000630.pdf

Tests and monitoring in HIV infection Liver

PT time – Prothrombin Time

Also called INR - International Normalized Ratio

Evaluate the ability of blood to clot properly

Not an enzyme test


Liver


Monitor anti-coagulants?, bleeding disorders, before surgery

Normal range 11-13.5 seconds1.5-2 times normal = too slow but no consensus on calibration of test as marker of over-fast clotting


Liver


INR = (Pt test / PT normal) ISI

ISI = International Sensitivity Index for tissue factor (1-1.4)


Anaemia

Iron, B12, B6, folic acid, red blood count, heamoglobin (HGB), mean corpuscular haemoglobin (MCH), heamocrit (HCT), mean corpuscular volume (MCV)

Drugs: AZT (pregnant?)


Anaemia

Haemocrit

Normal ranges 40-52% M 35-35% W

Low haematocrit = anaemia?


Anaemia

Haemoglobin

Normal ranges 11.5- to 16.5g W 13-18g M per 100mL blood

Low haemoglobin = anaemia?


Anaemia

Mean corpuscular volume

Larger = anaemic, B6, B12, folic acid deficiency?

Smaller = anaemic, iron deficiency?

AZT (and smoking) can increase MCV without causing anaemia


Anaemia

Red blood count (total erythrocytes)

Normal range 3.8-5 W 4.5-6.5 M billion per litre or million per cubic millimitre of blood (1012/L).

Low count = anaemia? but not sensitive or specific = probably a first test


The rest...

Blood chemistryhttp://www.aidsmeds.com/articles/CSTest_4730.shtml

http://www.aidsmeds.com/articles/CBCTest_4729.shtml

A-Z testshttp://www.aidsmap.com/cms1031936.asp

http://www.labtestsonline.org.uk/

Hepatitis C coinfection – liver and diagnosishttp://www.i-base.info/guides/hepc/livertests.html

http://www.i-base.info/guides/hepc/hcvtesting.html


Tests to avoid during pregnancy

● Amniocentesis● Chorionic villus sampling● Fetal scalp sampling● Cordocentis● Percutaneous umbilical cord sampling● Internal fetal labour monitoring (external ultrasound and fetal monitoring OK)


Plebotomy (having blood taken)

A cruel and unusual punishment - or nice chat with the nurse?


Blood count reference ranges

●Red blood count (RBC) 3.8 to 5 W 4.5-6.5 M million per mm3

●White blood count (WBC) 4-11 per mm3

●Haemoglobin (HGB, Hg) 11.5-16.5 W 13-18 M g per 100mL ●Neutropils 2-7.5 per mm3

●Lymphocytes 1.3-4 per mm3

●Platelets 150-440 per per mm3

●Mean corpuscular volume (MCV) 80-97


Blood count reference ranges

CD4 400-1,600 per mm3

CD4% 32-68%CD8 140-1000 per mm3

CD4:CD8 ratio 0.9-6


Sampling

Biggest causes of odd results are ‘sampling error’, ‘processing error’ or ‘sample

contamination’

wrong tube, wrong person's sample, cross-contamination, sample too hot/cold/old,

wrong reagent, wrongly set up equipment, not reading instructions, misreading output...


Results

One result is rarely conclusive

ANY unusual or unexpected results should ALWAYS be retested before making a

treatment decision

“Normal” is a difficult word – tests refer to reference ranges, can mean doctor/nurse is

happy with results even though high/low


Sensitive v specific

Sensitivity = reacts positively

Specificity = reacts positively in right circumstances



Sensitivity = rule in Sensitivity refers to the proportion of people with disease who have a positive test result

Specificity = rule outSpecificity refers to the proportion of people without disease who have a negative test result.



SnNout: when a sign, test or symptom has a high Sensitivity, a Negative result rules out the diagnosis.

SpPin: when a sign, test or symptom has a high Specificity, a Positive result

rules in the diagnosis.



Many “rapid” tests are highly sensitive but not specific enough to be definitive – prone to sample contamination

●Urine dipstick tests●Fingerprick tests

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Tests and monitoring in HIV infection