SLIDE 1
Genotypic prediction of viral co-receptor tropism: Correlation with enhanced Trofile
Angela L Strang, Jane Cameron, Clare Booth, Ana Garcia, Anna Maria Geretti Royal Free Hospital & UCL Medical School, London
SLIDE 2 Background
Phenotypic testing to predict HIV-1 co-receptor tropism can reliably guide use of CCR5 antagonists in clinical practice
- Labour intensive, time consuming, expensive,
limited availability
- Viral load, plasma volume
Genotypic prediction methods based on V3 sequences
- ffer an alternative testing tool
- Growing but still limited data on correlation with
phenotypic testing
SLIDE 3
Objectives
To evaluate genotypic tropism prediction for clinical use in a routine setting To assess its concordance with results obtained by the enhanced Trofile assay To analyse the clinical characteristics of the patient population according to the tropism prediction To analyse the quasispecies of samples showing discordant phenotypic / genotypic tropism predictions (ongoing)
SLIDE 4 Methods
Plasma samples from 106 patients in routine care Enhanced Trofile at Monogram (Pfizer programme) Genotypic testing in house (population env sequences) Interpretation of population sequence by Geno2Pheno
- Clonal model (trained using clonal data from naïve patients)
- Clinical model (input: CD4 n and %, CD8 n, viral load)
- Varying false positive rate (FPR) to adjust sensitivity/specificity
Clonal analysis and 454 analysis of discordant and unusual samples (ongoing)
SLIDE 5
Study population
Total 106 Subtype B n (%) 69 65% Non-B n (%)* 37 35% HAART Naïve n (%) 56 53% Experienced n (%) 50 47% Current CD4 n Median (range) 453 2-1067 Current CD4 % Median (range) 21 1-50 Nadir CD4 n Median (range) 361 2-950 Nadir CD4% Median (range) 21 2-47 Viral load log10 Median (range) 4.6 2.8-6.0
*Non-B subtypes: A (6), C (16), D (2), CRF01 (2), CRF02 (8), CRF10 (1), cpx (2)
SLIDE 6
Enhanced Trofile
R5 D/M NR Subtype B n 54 10 5 Non-B n 26 3* 8 HAART Naïve n 45 6 5 Experienced n 35 7 8 Current CD4 n Median (range) 455 (2-1067) 331 (22-894) 498 (289-688) Current CD4% Median (range) 23 (1-41) 15 (2-28) 23 (12-50) Nadir CD4 n Median (range) 366 (2-950) 288 (22-576) 390 (76-648) Nadir CD4% Median (range) 23 (4-47) 15 (2-28) 24 (5-33) Viral load log10 Median (range 4.5 (3.2-6.0) 4.8 (4.4-5.9) 4.1 (2.8-5.5) Total n (%) 80 (76%) 13 (12%) 13 (12%)
*A, C, CRF02
SLIDE 7 Genotypic tropism
Pre-validated across major subtypes/CRFs (n=50) In this study: 99/106 samples successfully amplified 7/106 (6.6%) assay failures
- 4 B, 1 CRF02, 1 CRF01, 1 cpx
- Median viral load 4.4 (range 3.5-4.9)
SLIDE 8
Concordance analysis: G2P vs enhanced Trofile
FPR % Interpretation model Clonal Clinical 1 77/87 87% 77/87 88% 5 76/87 87% 77/87 88% 10 70/87 80% 79/87 91% 15 63/87 72% 78/87 90% 20 60/87 69% 77/87 88%
87 samples with matched results
SLIDE 9 Concordance analysis: G2P vs enhanced Trofile
FPR % Interpretation model Clonal Clinical 1 0/11 (0%) 2/11 (18%) 5 6/11 (55%) 3/11 (27%) 10 6/11 (55%) 6/11 (55%) 15 7/11 (64%) 7/11 (64%) 20 8/11 (73%) 8/11 (73%)
- 11 samples with matched results and D/M call by enhanced Trofile
Concordance for X4:
SLIDE 10
Sensitivity and specificity analysis for the detection of X4 virus*
Clonal model Clinical model FPR Sensitivity Specificity Sensitivity Specificity 1 52% 100% 55% 99% 5 69% 93% 58% 97% 10 69% 86% 69% 96% 15 73% 79% 73% 94% 20 79% 76% 79% 92% *Analysis uses enhanced Trofile as comparator
SLIDE 11
Discordant samples: Enhanced Trofile vs Geno2Pheno with clonal interpretation at FPR 5%
Pt Sub type CD4 n CD4 % CD4 n nadir CD4 % nadir VL ART Trofile G2P Clonal FPR 5% G2P clinical FPR 10% FPR 15% 1 B 288 13 288 13 4.5 Exp DM R5 R5 X4 2 B 331 18 331 18 4.8 Naïve DM R5 X4 X4 3 B 894 26 459 22 4.9 Naïve DM R5 R5 R5 4 B 472 20 277 28 4.9 Naïve DM R5 R5 R5 5 B 91 6 91 6 4.8 Exp DM R5 R5 R5 6 A 494 27 410 24 4.5 Naïve R5 X4 R5 R5 7 B 504 17 433 16 4.8 Naïve R5 X4 R5 R5 8 B 426 25 257 18 3.9 Exp R5 X4 X4 X4 9 D 343 25 301 23 4.0 Exp R5 X4 R5 X4 10 B 781 28 677 27 3.8 Exp R5 X4 R5 R5 11 B 474 29 474 29 4.2 Exp R5 X4 R5 R5
SLIDE 12
Discordant samples: Enhanced Trofile vs Geno2Pheno with clinical interpretation at FPR 10%
Pt Sub type CD4 n CD4 % CD4 n nadir CD4 % nadir VL ART Trofile G2P Clonal FPR 5% G2P clinical FPR 10% FPR 15% FPR 20% 1 B 288 13 288 13 4.5 Exp DM R5 R5 X4 X4 12 B 642 28 576 27 4.7 Naïve DM X4 R5 R5 X4 3 B 894 26 459 22 4.9 Naïve DM R5 R5 R5 R5 4 B 472 20 277 28 4.9 Naïve DM R5 R5 R5 R5 5 B 91 6 91 6 4.8 Exp DM R5 R5 R5 R5 13 CRF02 338 13 118 8 3.4 Exp R5 R5 X4 X4 X4 14 C 2 1 13 4 5.9 Exp R5 R5 X4 X4 X4 8 B 426 25 257 18 3.9 Exp R5 X4 X4 X4 X4
SLIDE 13
Analysis of 100 clones from 2 patients
Subtype Trofile FRP G2P Clonal model G2P Clinical model % X4 clones* B R5 1 5 10 15 20 R5 X4 X4 X4 X4 R5 R5 R5 R5 X4 94/100 D R5 1 5 10 15 20 R5 X4 X4 X4 X4 R5 R5 R5 X4 X4 79/100 *G2P clonal model FPR 5%
SLIDE 14 Conclusions
In this pilot evaluation we found excellent correlation between enhanced Trofile and Geno2Pheno
- Best overall with clinical interpretation, FPR 10-15%
- Best for X4 with highest FPR
Sensitivity/Specificity 79%/92% with clinical interpretation at FPR 20% Use can be tailored to the patient’s treatment history in terms of optimal FPR settings Geno2Pheno provides a valid tropism prediction tool
SLIDE 15
Acknowledgement
Rolf Kaiser, University of Cologne Linos Vandekerckhove, Chris Verhofstede, University of Gent Lieven Stuyver, Virco, Belgium Patients and staff at ICDC clinic, Royal Free Thank you
