STATENS SERUM INSTITUT Antiretroviral therapy and development of resistant HIV Louise Bruun Jørgensen Department of Virology Statens Serum Institut

STATENS SERUM INSTITUT

Antiretroviral therapy and Antiretroviral therapy and development of resistant HIVdevelopment of resistant HIV

Louise Bruun JørgensenLouise Bruun JørgensenDepartment of VirologyDepartment of VirologyStatens Serum InstitutStatens Serum Institut



Diagnostic Monitorering

* HIV antibodies * HIV RNA (Viral load) (ELISA – WB) (PCR)

* HIV antigen * CD4 counts (ELISA) (FACS)

* HIV DNA * Resistance (PCR) (PCR-sequencing)

Culture

Virological diagnosis and monitoring of HIV-1 infektion


MACS study: 209 pt inrolled in 1984 and 1985


MACS study: 209 pt inrolled in 1984 and 1985




Drugs approved for treatment of HIV

Pommier, Nature Reviews, 2005


NRTI


NNRTI


Protease inhibitors


Protease inhibitors


Treatment of HIV patients


Recommendations for HIV treatment in Denmark

HAART3-4 antiviral drugs

1. EFV + AZT + 3TC2. NEV + AZT + 3TC3. LOP/Rit + AZT + 3TC

http://www.dsinfm.dk

•CD4 cell count < 200-300 cells/ul•Acute infection•Clinical Symptoms•Pregnancy


Combination therapy:

Drugs Clinical effect Virological effect

HAART Highly Active Antiretroviral Therapy(3-4 drugsNRTI+NNRTI or PI)




0

20000

40000

60000

80000

100000

120000

140000

160000

31-01-1993 15-06-1994 28-10-1995 11-03-1997 24-07-1998 06-12-1999

Vira

l load

Måneder

HAARTHAART





0

20000

40000

60000

80000

100000

120000

140000

160000

31-01-1993 15-06-1994 28-10-1995 11-03-1997 24-07-1998 06-12-1999

Vira

l load

Måneder

HAARTHAART





0

20000

40000

60000

80000

100000

120000

140000

160000

31-01-1993 15-06-1994 28-10-1995 11-03-1997 24-07-1998 06-12-1999

Vira

l load

Months

HAARTHAART

ComplianceSite-effectsDevelopment of resistance



Compliance and Site-effects


Mutations in the Pro and RT gene associated with resistance

Primary: D30N, M46I/L, G48V, I50V, V82F/A/T/S, I84V, L90M

Secondary: L10F/I/R/V, K20R/M, L24I/V, V32I, L33F, M36I, I47V, F53L, I54V/M/L, L63P, A71V/T, G73S/A, V77I, N88D/S

NRTI: M41L, E44D, A62V, K65R, D67N, T69D/(S+xx), K70R, L74V, V75T/M/A/I, F77L, Y115F, F116Y, V118I, Q151M, M184V/I, L210W, T215Y/F, K219Q/E

NNRTI: A98G, L100I, K101E, K103N, V106A/I, V108I, V179D, Y181C/I, Y188L/C/H, G190A/S, P225H, M230L, P236L

Development of resistance


Time

Viral load

Fitness of the resistant virus

Speed of development of resistance

Effectivity of the drug against wild type

Development of resistant HIV against antiretroviral drugs


Methods for genotyping

Commercial services:VircoVirologicetc.

Commercial kitsAbbott ViroSeqVisible Genetics

In House:Home brewed asays


EDTA-blood sample

Extraction of HIV-RNA from plasma

cDNA syntesis by MuLV reverse transcriptase

Single PCR-amplification of 1,8 kb of the pol-gene

Direct sequencing of the protease- and RT-gene

Electroforesis (ABI 3100)

Nucleotide sequence

Translation to amino acidsequence

Identification of mutations associated to resistance90

10

54

84

82

71

63

20

32

46

24

4850

33

36

73

47

77

30

88

Genotypic resistance by direct sequencing


Genotypic resistance testing on routine basis Genotypic resistance testing on routine basis in a quality insured (ISO 17025) laboratory: in a quality insured (ISO 17025) laboratory:

Department of Virology SSIDepartment of Virology SSI

EDTA-blood sample


* Abbott ViroSeq 2 system

Nucleotide sequence

* Quality control by phylogenetic analysis

* Identification of mutations associated to resistance

* Interpretation of resistance

* Report





EDTA-blood sample



Nucleotide sequence

* Quality control and subtyping by phylogenetic analysis



* Report


Quality control by phylogenetic analysis

Nucleotide Substitutions (x100)0

12.2

24681012

pt1906PT1857G

pt1900pt1861

pt1908pt1911

pt1914gpt1889

pt1854pt1893

pt1895pt1901

pt1771pt1801

pt1262PT1915

PT1868pt1904B-hxb2

PT1864pt1912

PT1867pt1896

pt1903pt1852

pt1894gB-rf

pt1910D-eli

D-ndkpt1856

pt1858pt1880gg

pt1891C-BRO25

CH2220pt1909

pt1883gpt1897

pt1899F1-BR

F2MP255pt1902

pt1881AUG037pt1862

AETH253HCF056

GNG083pt1882g

JSE7022

Subtyping

eg. subtype D




EDTA-blood sample



Nucleotide sequence




* Report


Identification of mutationer associated to resistance




EDTA-blood sample



Nucleotide sequence




* Report


Interpretation of genotypic resistance

Tables www.iasusa.org

Rulebased algoritms www.hivdb.stanford.edu/hiv/

Virtual phenotypes www.virco.be

Neural networks


Interpretation of genotypic resistanceDept. of Virology, SSI




EDTA-blood sample



Nucleotide sequence




* Report


ReportDept. of Virology, SSI


New phenotypic assay:Recombinant Virus Assay (RVA)

Recombinant virus

MTT-assay(IC-50)


Cross-resistance

Susceptible Genotypic WT (K103)

Phenotypic Fold = 1

J.Virology 2001, 75(11)


Cross-resistance

Susceptible K103N Genotypic WT (K103)

Resistant

Phenotypic Fold = 1 Fold = 57-437X

J.Virology 2001, 75(11)


0

100

200

300

400

500

600

31-01-93 15-06-94 28-10-95 11-03-97 24-07-98 06-12-99

CD

4 c

ou

nt

0

50000

100000

150000

200000

250000

300000

350000

Vir

al

loa

d

AZT3TC SAQ ddC

D4TCRX NOR

ddINEVNEL EFA

ABCHYD

M184VM41LD67NK70R

L210WT215YK219EG190AY181C

L10IM46IL63PA71VV77II84V

L90M

M184VM41LD67NK70R

L210WT215YK219EL74I

Y181C

L10IM46IL63PL90M

0.1

1.0

10.0

100.0

1000.0NEV

ABC

AZT

3TC

D4T

CRX

NOR

SAQ

0.1

1.0

10.0

100.0

1000.0NEV

ABC

D4T

AZT

3TCNEL

CRX

NOR

SAQ


Viral load

0

50

100

150

200

250

300

350ja

n-9

6

jul-

96

jan

-97

jul-

97

jan

-98

jul-

98

jan

-99

jul-

99

jan

-00

jul-

00

jan

-01

jul-

01

jan

-02

Viral load

AZT3TCd4TABCddINevIndNelRit

RNA/DNA

TI-2 start

(x 1000 kopier/ml)

RNA/DNA

W184,W215

W184,M215W184,R215

M184,R215

M184,M215

W184,M215

M=mutantW=wildtypeR=revertant

W184,M215

M184,M215

TI-1 start

M184,M215(jul 2002)

Resumed treatment

W184,W215

STATENS SERUM INSTITUT120194

281191

300502D

150502D

150502

150296

300502

250597*

201097*

170698*

070499*

121200

030701*

180702*

170102*

201099

201099D

ASE7253

AUG037

Treatment interuption 1

Treatment interuption 2

T215YT215C/S*M184V

97

56

61

29

Fylogeni af Pol-genet


Detection limit of Genotypic resistance assay


0

1000

2000

3000

4000

5000

6000

17

.03

.97

16

.04

.97

28

.07

.97

24

.09

.97

29

.12

.97

01

.04

.98

02

.07

.98

08

.09

.98

07

.12

.98

01

.03

.99

07

.06

.99

07

.09

.99

01

.11

.99

07

.02

.00

Copies/ml

Protease

RT

L63PV77I

L74VM184V

New ultra sensitive assay for genotypic resistance


0

1000

2000

3000

4000

5000

6000

17

.03

.97

16

.04

.97

28

.07

.97

24

.09

.97

29

.12

.97

01

.04

.98

02

.07

.98

08

.09

.98

07

.12

.98

01

.03

.99

07

.06

.99

07

.09

.99

01

.11

.99

07

.02

.00

Copies/ml

Protease

RT

L63PV77I

L74VM184V

Improved extractionNested / double PCR=> 20 copies pr ml



0

1000

2000

3000

4000

5000

6000

17

.03

.97

16

.04

.97

28

.07

.97

24

.09

.97

29

.12

.97

01

.04

.98

02

.07

.98

08

.09

.98

07

.12

.98

01

.03

.99

07

.06

.99

07

.09

.99

01

.11

.99

07

.02

.00

Copies/ml

Protease

RT

L63PV77I

L74VM184V

L74VM184V

L63PV77I

M36IL63PV77IN88DL90M*

L33FM36IL63PV77II84VN88DL90M*

L33FM36IL63PV77II84VN88D

L33FM36IL63PV77II84VN88D


M36IL63PV77II84VL90M*


K20TL24ID30N*M36IL63PV77IN88DL90M*

Improved extractionNested / double PCR=> 20 copies pr ml



Therapy

Development ofDrug resistance


Therapy

Development of Drug resistance

Transmission of resistance


Transmission of resistant HIV-1 strains

RTI:PI:

Spain 3% 1% Italy 11% 2% Belgium 13% 4% Luxembourg 12% 0% Germany 16% 3% France 17% 2%

Europe

Total MDR: Primary HIV:

1995-1998 8% 4%

1999-2000 23% 10%

RTI: PI: Seroconv. 6% 1% IDVU 0% 0% New diagn. 13% 3% Military 10% 10%

USA


Reversion of genotypic resistance

AZT 215

ACC(T)

WT

A. DeRonde J.Virol.2001

0 1 2 År



AZT 215

TAC(Y)ACC(T)

WT


0 1 2 År

STOP of therapy



AZT 215

TCC(S)

GAC(D)

TAC(Y)ACC(T) AAC(N)

WT


0 1 2 År

STOP of therapy



AZT 215

TCC(S)

GAC(D) AGC(S)

TAC(Y)ACC(T) AAC(N) GAC(D)

WT


0 1 2 År

STOP of therapy


2000: 2,0% (2/104) 41L, 210S, 215S 215S

2001: 4,8% (7/147) 90M, 63P, 71V, 77I 215E

215L (x2) 215S 215S 82F, 10I, 63P

98G, 215S

2002: 5,0% (7/140) 103N, 215Y 103N, 188C

41L, 215D 90M, 10I, 63P, 71T, 77I215D (x2)215S67N

2003: 5,5% (7/127) 67N, 219Q 67N, 219E 84V, 10V,33F,36I,63P

69D 215N215S103N181C

2004: 0,7% (1/137) 190A

Total prv. RT PI

Prevalence of drug resistance mutations in newly diagnosed HIV-1 patients in Denmark 2000-2004


HIV-1 subtypes

Based on the variation ofthe genome HIV can be divided in to different subtypes

D

CF

E

O

B

G

A

H


HIV-1 subtypes

9 identified subtypes (A-K) and 16 CRF (circulating recombinantforms)

In the industrialised world subtype B is mostprevalent. However, globally subtype C is the most prevalent subtype.


Prevalence of subtype non-B in newly diagnosed HIV-1 patients in Denmark

Subtype 2000

A8%

AE4% AG

5%

C18%

D3%

G2%

B59%

Subtype 2001

A9% AE

7% AG3%

B63%

C12%

D2%

G1%

cpx3%

30%

12%47%

5% 6%

Global subtype distribution

C

DAndre

A

B

Ref.: J.Clin. Virol. 29 (2004)


EpidemilogyTransmission route of infection 2001

0%

10%

20%

30%

40%

50%

2001-SUR 33% 47% 9% 7%

2001 36% 46% 10% 8%

MSM HTX IVDU Unknown

Subtype 2001

A9% AE

7% AG3%

B63%

C12%

D2%

G1%

cpx3%

Country of infection 2001

Denmark51%

Europe2%

Afrika22%

Asia8%

NorthAmerika1%

Unknown16%


2000: 2,0% (2/104) 41L, 210S, 215S B 41 % 0/2 215S B

2001: 4,8% (7/147) 90M, 63P, 71V, 77I B 39 % 0/7 215E B

215L (x2) B+B 215S B 215S 82F, 10I, 63P B

98G, 215S B

2002: 5,0% (7/140) 103N, 215Y B 37 % 4/7 103N, 188C AG

41L, 215D 90M, 10I, 63P, 71T, 77I B215D (x2) B+D215S AE67N A

2003: 5,5% (7/127) 67N, 219Q B 34 % 4/7 67N, 219E 84V, 10V,33F,36I,63P AE

69D AE215N B215S B103N A181C AE

2004: 0,7% (1/137) 190A AD (34 %) 0/1

Total prv. RT PI Subtype % non-B Prv. Non-B

Prevalence of drug resistance mutations in newly diagnosed HIV-1 patients in Denmark 2000-2004


Transmission of HIV


Case

African/danish family African mother - HIV pos in 2004 African daugter – HIV pos in 2005

– Probably infected at birth – Arrived as teenager in DK– Has no HIV symptoms

Danish stepfather – HIV positiv 2003– Infected by african wife


Questions:

Can we confirm the mother has infected the daughter?

Is it possible to identify af transmission chain after 17 years?



20.0

24681012141618

sero386A1.KE.00.MSA4076

A1.KE.00.KER2009SERO630

A1.KE.00.KNH1211A1.KE.99.KSM4021

A1.KE.00.MSA4069A1.KE.00.KNH1209sero736

A1.KE.00.KNH1199sero531g

sero913zse1520

se762se1712A1.KE.00.KNH1144

A1.KE.00.KER2008A1.KE.94.Q23_17

A1.KE.99.KNH1135A1.KE.97.ML605_3_1997

se1071A1.KE.00.MSA4079A1.KE.97.ML752se1033

se983vsero553

A1.KE.86.ML013_10_1986A1.KE.97.ML013_2_1997

sero350ggA1.KE.00.KSM4024

sero434sero838

sero369AUG037A1.KE.00.NKU3005

sero394A1.KE.00.KER2012

A1.KE.99.KNH1088A1.KE.00.MSA4070

A1.KE.86.ML170_1986sero530gsero540

SERO705se1084

sero801sero904

sero379sero519

sero655sero411p

sero560A1.KE.00.KNH1207

SERO670UGA1.KE.00.MSA4072

A1.KE.00.KSM4030sero558

sero1035Sero360

se1037SERO744UG

SERO1122SERO878

AETH253sero454

sero943PatientrefB-hxb2

B-rfD-eli

D-ndkF1-BR

F2MP255GNG083

JSE7022HCF056C-BRO25CH2220

SE1501(se1498)Omvp5180


20.0

24681012141618

sero386A1.KE.00.MSA4076

A1.KE.00.KER2009SERO630

A1.KE.00.KNH1211A1.KE.99.KSM4021

A1.KE.00.MSA4069A1.KE.00.KNH1209sero736

A1.KE.00.KNH1199sero531g

sero913zse1520

se762se1712A1.KE.00.KNH1144

A1.KE.00.KER2008A1.KE.94.Q23_17

A1.KE.99.KNH1135A1.KE.97.ML605_3_1997

se1071A1.KE.00.MSA4079A1.KE.97.ML752se1033

se983vsero553

A1.KE.86.ML013_10_1986A1.KE.97.ML013_2_1997

sero350ggA1.KE.00.KSM4024

sero434sero838

sero369AUG037A1.KE.00.NKU3005

sero394A1.KE.00.KER2012

A1.KE.99.KNH1088A1.KE.00.MSA4070

A1.KE.86.ML170_1986sero530gsero540

SERO705se1084

sero801sero904

sero379sero519

sero655sero411p

sero560A1.KE.00.KNH1207

SERO670UGA1.KE.00.MSA4072

A1.KE.00.KSM4030sero558

sero1035Sero360

se1037SERO744UG

SERO1122SERO878

AETH253sero454

sero943PatientrefB-hxb2

B-rfD-eli

D-ndkF1-BR

F2MP255GNG083

JSE7022HCF056C-BRO25CH2220

SE1501(se1498)Omvp5180

94100

78 (Daughter)

(Mother)

(Stepfather)

POL

Metode: Neighbour joining tree, 100 bootstrap replikations 1200 bp All HIV-1 subtype A POL sequences from Los Alamos included Subtype A sequences from SERO project

Bootstrapvalues over 50% is shown



18.1

24681012141618

AY772968AY772969

AY772985AY322184

AY322185L11774AF159877

L11772AF095833

AY322193L11775

L11757AF159878

AY772954L11786

L11794AF361873U56356

se762se1520

se1712AY772970

L11773AY322190AF095832AY772958

AY772955L11770

AF539405L11768

AB098332AB098333

U56359AY772953

L11788L11791AF004885

U56360AF159876

AF481779AB098330AB098331

AY772971U51190

AF481778AY772986

AY187680AY456310

AY676592AY676591

AY676593AY676594

AY456312AY456313AY456314

AY456316AY456315

L11798M62320

L11777AY772973

AY772975AY772983

AY772974AY772976

AY772978K03455

AB081213AY456330

AF286238L11783


18.1

24681012141618

AY772968AY772969

AY772985AY322184

AY322185L11774AF159877

L11772AF095833

AY322193L11775

L11757AF159878

AY772954L11786

L11794AF361873U56356

se762se1520

se1712AY772970

L11773AY322190AF095832AY772958

AY772955L11770

AF539405L11768

AB098332AB098333

U56359AY772953

L11788L11791AF004885

U56360AF159876

AF481779AB098330AB098331

AY772971U51190

AF481778AY772986

AY187680AY456310

AY676592AY676591

AY676593AY676594

AY456312AY456313AY456314

AY456316AY456315

L11798M62320

L11777AY772973

AY772975AY772983

AY772974AY772976

AY772978K03455

AB081213AY456330

AF286238L11783

10068

StepfatherMother

Daughter

Gag

Metode: Neighbour joining tree, 100 bootstrap replikations 370 bp All HIV-1 subtype A gag sequences from Los Alamos included Bootstrapvalues over 50% is shown


GAG

Daughter!


Dublication

GAG


Epidemiological Study of HIV-1 in Greenland using Phylogenetic Analyses

TV Madsen, N Obel, N Lohse, J Gerstoft, AB Petersen, C Nielsen & LB Jørgensen

•90 samples from Greenland HIV-1 positive patients with high viral load, and as early in the infection as possible, was chosen for the study. HIV-1 sequences from Danish patients are also included in order to compare the genetic composition of the two populations.

•Sequences were obtained from selected regions (gag: p17-region; pol: protease and 250 aa of RT; env: V3-region) using in-house PCR and sequencing methods, and ViroSeqTM HIV Genotyping System (Abbott).

•In order to study the genetic variation over time and to identify local outbreaks and chains of infection in the Greenland HIV-1 population the phylogeny was correlated with epidemiological data from the Danish HIV Cohort.


Figure 1. Phylogeny on pol-region (NJ, 100 bootstrap replicates). Isolates from Greenland have been marked in green.

0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH

SE212 MASE93 KHSE174 RH

SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE

J.SE.94.SE13 CPX.CM.

11 CPX.FR.11 CPX.CM.11 CPX.GR.

SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH

GR58 DIHGR38 DIHGR67 DIH

GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH

GR57 DIHGR41 SSHGR75 DIH

GR14 SSHGR55 SSH

GR62 SSHGR70 SSH

GR9 SSHGR11 SSH

GR44 SSHGR17 SSH

GR40 ASSGR12 SSH

GR82 SSHGR19 SSHGR92 SSH

GR78 SSHGR34 SSH


B.FR.83.HXSE112 RH

ddI-23 RHSE263 RH

GR22 ISB.US.86.JR

SE181 RHSE136 KH

SE148 RHGR2 DIH

GR54 DIHGR56 DIH

SE131 KHSE249 AL

SE199 RHSE134 RH

SE100 RHSE175 RH

BIO17 KHSE203 RH

93

10080

100

71

100

83

81

83

100100

100

99100

100

0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


GR14 SSHGR55 SSH

GR62 SSHGR70 SSH

GR9 SSHGR11 SSH

GR44 SSHGR17 SSH

GR40 ASSGR12 SSH


GR78 SSHGR34 SSH


B.FR.83.HXSE112 RH

ddI-23 RHSE263 RH

GR22 ISB.US.86.JR

SE181 RHSE136 KH

SE148 RHGR2 DIH

GR54 DIHGR56 DIH

SE131 KHSE249 AL

SE199 RHSE134 RH

SE100 RHSE175 RH

BIO17 KHSE203 RH

0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


GR14 SSHGR55 SSH

GR62 SSHGR70 SSH

GR9 SSHGR11 SSH

GR44 SSHGR17 SSH

GR40 ASSGR12 SSH


GR78 SSHGR34 SSH


B.FR.83.HXSE112 RH

ddI-23 RHSE263 RH

GR22 ISB.US.86.JR

SE181 RHSE136 KH

SE148 RHGR2 DIH

GR54 DIHGR56 DIH

SE131 KHSE249 AL

SE199 RHSE134 RH

SE100 RHSE175 RH

BIO17 KHSE203 RH

93

10080

100

71

100

83

81

83

100100

100

99100

100

9774

8872

98

91


Figure 1. Phylogeny on pol-region (NJ, 100 bootstrap replicates). Isolates from Greenland have been marked in green.

0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


GR14 SSHGR55 SSH

GR62 SSHGR70 SSH

GR9 SSHGR11 SSH

GR44 SSHGR17 SSH

GR40 ASSGR12 SSH


GR78 SSHGR34 SSH


B.FR.83.HXSE112 RH

ddI-23 RHSE263 RH

GR22 ISB.US.86.JR

SE181 RHSE136 KH

SE148 RHGR2 DIH

GR54 DIHGR56 DIH

SE131 KHSE249 AL

SE199 RHSE134 RH

SE100 RHSE175 RH

BIO17 KHSE203 RH

93

10080

100

71

100

83

81

83

100100

100

99100

100

0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


GR14 SSHGR55 SSH

GR62 SSHGR70 SSH

GR9 SSHGR11 SSH

GR44 SSHGR17 SSH

GR40 ASSGR12 SSH


GR78 SSHGR34 SSH


B.FR.83.HXSE112 RH

ddI-23 RHSE263 RH

GR22 ISB.US.86.JR

SE181 RHSE136 KH

SE148 RHGR2 DIH

GR54 DIHGR56 DIH

SE131 KHSE249 AL

SE199 RHSE134 RH

SE100 RHSE175 RH

BIO17 KHSE203 RH

0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


0.1

SE297 ALB.US.83.RF

SE484 HHB.US.90.WE

SE123 KHSE476 RH

SE180 RHGR46 DIH

GR83 QASSE161 HH

BIO28 KHSE118 KH


SE237 RHGR72 DIH

SE191 RHSE520 RH

SE348 MAJ.SE.93.SE



SE402 RHSE214 RH

SE178 AL01 AE.CF.9

GR1 UPS01 AE.TH.9

SE138 RHSE140 KH

SE176 RHSE417 RH

GR25 DIHGR73 DIH

GR3 DIHGR4 DIH


GR71 DIHGR63 DIH


GR42 DIHGR43 DIH

GR26 DIHGR27 DIH

GR7 DIHGR47 MANS

GR23 DIHGR65 DIH


GR14 SSHGR55 SSH

GR62 SSHGR70 SSH

GR9 SSHGR11 SSH

GR44 SSHGR17 SSH

GR40 ASSGR12 SSH


GR78 SSHGR34 SSH


B.FR.83.HXSE112 RH

ddI-23 RHSE263 RH

GR22 ISB.US.86.JR

SE181 RHSE136 KH

SE148 RHGR2 DIH

GR54 DIHGR56 DIH

SE131 KHSE249 AL

SE199 RHSE134 RH

SE100 RHSE175 RH

BIO17 KHSE203 RH

93

10080

100

71

100

83

81

83

100100

100

99100

100

9774

8872

98

91

?


Transmission

1991

1994

2001


Three patients:

•GR41: 31 year old female HTX infected in Greenland first positive test is from 27/9-94. Analysed sample from 14/7-99.•GR57: 51 year old male HTX infected in Greenland first

positive test 1/1-91. † 6/3-01. Analysed sample from 9/2-00.•GR75: 52 year old female HTX infected in Greenland first positive test is from 1/8-01. Analysed sample from 14/-02.


Transmission

1991

1994

2001

9/2-2000

14/7-1999

14/1-2002

Analysed


New mutation against NRTI?


History

•Heavily treated patient with non-compliant behaviour

•Four samples sent for HIV-1 genotypic resistance


Jun 2000Jan 2002Jan 2003Mar 2005

Mutation D67G_Y


STATENS SERUM INSTITUTRegalInst algoritme


juli 2000 januar 2002 januar 2003 marts 2005

D67N N N G G

D67GY GY GY

T69N N

K70R R R R

L74V V

V118I I

M184V V

K219Q Q Q Q

K103N KN

V108I I

Y181C C C C

M46I MI

V82A A A A

L90M M M M

L10I I I I

K20R R

M36I I I I

F53L L

I54V V V

L63P P P P

A71V V

V77I I

NRTI NNRTI PI

1998 ddi+abc nvp idv

Apr. 2003 (abc+3tc+azt)+tf rtv+amp

Jun. 2003 (abc+3tc+azt)+tf rtv+saq

Mar. 2005 (abc+3tc+azt)+tf rtv+saq

HIV drug mutations 2000-2005


Treatment history

NRTI NNRTI PI VL Genotypic Resistance

1990-1998 AZT+ddI

1995-dec.1996 AZT+3TC

Dec.1996-1998 d4T+3TC IDV

1998 DDI+ABC NVP NFV

1998 DDI+ABC NVP IDV

juli 2000 DDI+ABC NVP IDV 54100 No insert: D67GY

Jan. 2002 DDI+ABC NVP IDV 56300 No insert: D67GY

Jan. 2003 DDI+ABC NVP IDV 40000 Plus insert: D67GY

Apr. 2003 (Abc+3TC+AZT)+TF RTV+Amp

Jun. 2003 (Abc+3TC+AZT)+TF RTV+SAQ

marts 2005 (Abc+3TC+AZT)+TF RTV+SAQ 16300 Plus insert: D67GY


Virus BL-3 laboratory, Department of Virology, SSI

Deptartment of infectious diseasesRigshospitaletHvidovre HospitalSkejby SygehusOdense Universitets HospitalÅlborg SygehusSygehus Øresund

SupportABI and Abbott

Documents

STATENS SERUM INSTITUT Antiretroviral therapy and development of resistant HIV Louise Bruun Jørgensen Department of Virology Statens Serum Institut