the 6 th national scientific conference on hiv/aids evaluation of two techniques for viral load...

THE 6TH NATIONAL SCIENTIFIC CONFERENCE ON HIV/AIDS

Evaluation of two techniques for viral load monitoring on DBS ANRS 12338 project

Phase I - Laboratory evaluation phase (The MOVIDA Vietnam project:

Improving access to viral load monitoring in HIV-infected patients on ART in decentralised area using Dried Blood spot )

TAIEB F1-2, TRAN HONG T6., HO THI H4, PHAM VA4, NGUYEN L5, THONG LA3, TUAILLON E7, NGUYEN TA6, BUI DUC D3, DO THI N.3, MADEC Y.1

1.Emerging Diseases Epidemiology Unit-Institut Pasteur, Paris (France)

2.Direction de la Recherche Clinique et du Développement-Assistance Publique des Hôpitaux de Paris- Saint-Louis Hospital, Paris (France)

3.Vietnam Authority of HIV/AIDS Control (VAAC), Hanoi (Vietnam)

4.Hanoi School of Public Health- HIV/AIDS Prevention and Control department-Hanoi (Vietnam)

5.DONG DA OPC, Hanoi (Vietnam)

6.National Institute of Hygiene and Epidemiology- Virology laboratory-HIV/AIDS Department, Hanoi (Vietnam)

7.INSERM U1058 unit- Pathogenesis and Control of Chronic Infections-Virology department, Montpellier University Hospital, Montpellier (France)

The 6th National Scientific Conference on HIV/AIDS

Outline

1. Background

2. Objectives

3. Methods

4. Results & Discussions

5. Conclusions

6. Recommendations


Follow-up of patients on ART• Essentially clinical and immunological• Virological monitoring:

– still scarce

– especially in decentralized areas

BACKGROUND


Benefit of VL monitoring:

• Early and specific detection of therapeutic failure

• Objective measure (and improvement) of adherence to ART

• Adapted change of ART

• Prevents HIV drug resistance acquisition


Challenges for VL monitoring in decentralized area

• Several type of constraints:

– Technical

– Human

– Financial

Only reference laboratories able to provide VL measurements.


Challenges for VL monitoring in decentralized area

• Plasma (gold standard) transfer is possible but:

– Rapid RNA destruction at ambient temperature

– Costly (cold-chain)

– Infectious sample

Patients followed in decentralized area (>50% of patients) have low or no access to VL monitoring


Advantage for VL monitoring Dried Blood Spots (DBS)

• At the HIV care site, DBS are:– Easy to perform

– Easy to maintain and safe (sample not contagious)

– Easy to transfer:

No cold chain

By the existing postal network


Advantage for VL monitoring Dried Blood Spots (DBS)

• At the virology laboratory (centralised):– No extra equipment needed

– Capacity development of the personnel • 2 techniques of VL measurement on DBS have been

validated:– Nuclisens by Biomerieux®

– Abbott® m2000rt

• Development of new techniques– Roche® FVE protocol

– First results are very promising (Xiaoning Wu X et al. Poster at 2014 IAS conference)


Evaluation of VL measurement on DBS in routine practice

• To evaluate Sensitivity and Specificity of:

– DBS vs. plasma (gold standard)

– at the threshold of 1000 copies/mL .

OBJECTIVES


Study site and population:

• HIV-1 positive adults,

• Followed at Dong Da OPC in Hanoï

• Who agreed to participate

METHODS


Laboratory procedures

• Transfer of Whole blood (10 mL EDTA tube) from Dong Da OPC to NIHE

• DBS cards performed at NIHE laboratory (min of 15 days before VL measurement)

• Centrifugation and conservation (-80°C) of plasma on cryotubes (2 mL)

• Testing VL plasma and DBS samples by:

– Roche: Cobas®AmpliPrep/Cobas® TaqMan HIV-1 Test v2.0

– Abbott: Abbott m2000rt


Laboratory proceduresTraining VL DBS testing for laboratory staff

• 2 trainings for each technique

• performed by the manufacturer teams

• before the beginning of the study


RESULTs Population description

• July 1st to October 9th, 2015: 198 patients enrolled

• 145 (73,3%) males

• Median [IQR] age: 38 years [34-41]

• Median [IQR] time on ART: 38 months [3-74]

Patients category N (%)

Pre ART 19 (9.6)

1-6 months 34 (17.2)

> 6 months 51 (25.8)

Suspected treatment failure 94 (47.4)


Roche technique

Plasma≥1000 cp/mL

<1000 cp/mL Total

DBS (FEV protocol)

≥1000 cp/mL 23 0 23

<1000 cp/mL 30 145 175

Total 53 145 198

Sensitivity 43.4% [29.8-57.7]

Specificity 100% [97.4-100]

Concordance 85.6%


Roche technique

• Pearson correlation coefficient: r=0.81 (p<0.001)

Plasma Viral Load (log10 cp/mL)


Abbott technique

Plasma≥1000 cp/mL

<1000 cp/mL Total

DBS

≥1000 cp/mL 42 8 148

<1000 cp/mL 3 145 50

Total 45 153 198

Sensitivity 93.3% [81.7-98.6]

Specificity 94.8% [90.0-97.7]

Concordance 94.4%


Abbott technique

• Pearson correlation coefficient: r=0.91 (p<0.001)

Plasma Viral Load (log10 cp/mL)


DISCUSSION

• Good correlation between plasma and DBS with both techniques

• Sensitivity is much better with Abbott technique than with Roche technique

• Specificity is slightly better with Roche technique than with Abbott technique


Sensitivity: Ability to detect virological failure

• False virological success (i.e. VL <1000 cp/ml on DBS) explanation:– Destruction of RNA

• Condition of conservation?: room temperature

• same conditions for both techniques


Sensitivity: Ability to detect virological failure

• False virological success (i.e. VL <1000 cp/ml on DBS) explanation:– Destruction of RNA

• Delay of conservation?

– Median [IQR] delay (days):

» Roche: 23 [15-71]

» Abbott: 27 [22-70]


Sensitivity: capacity to detect virological failure

• False virological success (i.e. VL <1000 cp/ml on DBS) explanation:– Quantity of whole blood used?

• Roche: 1 spot of 70µL

• Abbott: 2 spots of 50µL


Specificity: Ability to detect virological success

• False virological failure (i.e. VL >1000 cp/ml on DBS) explanation:– Dried Blood Spot (whole blood)

– Over estimation of VL: Amplification of proviral DNA (Monleau M, et al. J. Clin. Microbiol. 2009 and J. Antimicrob. Chemother. 2010)


CONCLUSIONS

• Study which aims at replicate routine condition of VL monitoring (delay between collection sample and VL measurement; condition of DBS storage, etc.)


Dried Blood Spot• can overcome challenges to VL monitoring

• Is available immediately

• Allow to perform a large panel of analyze (Drug resistance genotyping, HCV-HBV serology; HCV VL (current studies).

• Better Sensitivity with Abbott technique than with Roche technique

• Sensitivity is the most important criterion in our opinion

• Indication to switch the treatment: 2 VL > 1000 cp/mL


RECOMMENDATIONS

• Further studies in “real life” to validate similar results

• Studies of other elution protocol with DBS• Use of DBS for VL monitoring in decentralized

area (eg: mountain area)


Thank you for your attention

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