Analysis of HIV1 Pol Sequences in CSF and Plasma Shows Increasing Compartmentalization of Infection

1
Analysis of HIV-1 Pol Sequences in CSF and
Plasma Shows Increasing Compartmentalization of
Infection with Disease Stage and ADC

• S.Sala, S. Spudich, M. Gisslen, L.Hagberg,
  S.Presi, A.Bestetti, S.Bossolasco, S.Sala, R.W.
  Price, P. Cinque
• San Raffaele Scientific Institute, Milan, Italy,
• University of California San Francisco, USA,
• University of Goteborg, Sweden

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Background and Objectives

• Viral load studies of paired CSF and plasma
  specimens indicate that HIV replication can be
  variably segregated in these two compartments
• Objective of this study was to evaluate the
  extent of variation between CSF and plasma HIV-1
  pol sequences and its association with
  HIV-induced CNS disease, stage of infection and
  patient variables

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Patients and Samples
Paired CSF and plasma from 107 HIV-infected
patients admitted at 3 clinical sites from 1994
to 2004 (Milan, San Francisco, Goteborg)

• According to anti-HIV treatment
• - RT inhibitors (RTI) 51
• - No RTI 56 (31 RTI naïve)
• - Protease inhibitors (PI) 23
• - No PI 84 (69 PI naïve)

• According to HIV-1 infection stage and CNS-D
• - Primary HIV infection 3
• - HIV and CNS asymptomatic 16
• - AIDS, CNS asymptomatic 11
• - ADC 0.5 14
• - ADC 1 14
• - CNS-OIs 49

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Methods

• 1. Amplification of RT and protease sequences by
  RT-PCR of RNA extracted from CSF and plasma
  samples (Milan, San Francisco)
• 2. Direct sequencing of RT-PCR products (Milan,
  San Francisco)
• 3. Alignment of CSF/plasma sequences by ClustalX
• ? RT 546 nt ( 102/107 patients)
• ? protease 261 nt (88/107 patients)
• 4. Separate data analysis for RT and protease
  calculation of diversity

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Calculation of percent nucleotide diversity
G ? A SCORE 1
CSF
G
A
AGCTCTATTAGATACAGGAGCAGATGATACAGTATTAGAAGAG
G
TGGCTTTGCCAGGA
PL
GAA
G
CTCTRTTAGATACAGGAGCAG
A
TGATACAGTATTAGAAGAGATGGCTTTGCCAGGA



R A / G SCORE 0.5
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RT CSF / plasma sequence diversity
CSF/ plasma diversity
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Protease CSF / plasma sequence diversity
CSF/ plasma diversity
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RT CSF / plasma sequence diversity in untreated
and treated patients
NO RTIs (n 54)
RTI-treated (n48)
CSF/ plasma diversity
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Protease CSF / plasma sequence diversity in
untreated and treated patients
NO PIs (n 67)
PI-treated (n 21)
CSF/ plasma diversity
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Correlation between CSF/plasma sequence diversity
and patient variables
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Summary

• CSF/plasma diversity increased with HIV-1
  infection stage
• CSF/plasma diversity was higher in patients with
  ADCgt1 than in CNS asymptomatic or ADC O.5
  subjects
• These differences were more evident in untreated
  patients
• RT and protease CSF/plasma diversity correlated
  with CSF HIV-RNA and CD4 cell count

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Conclusions

• CSF/plasma diversity increase with disease stage
  and correlation with CD4 cells support the
  presence of compartmentalized viral evolution
  through the natural course of HIV-1 infection
• Higher CSF/plasma diversity in ADC supports the
  hypothesis of an autonomous HIV replication in
  the CNS in this condition
• Evaluation of CSF/plasma nucleotide diversity
  might represent a marker to differentiate the
  source of CSF virus

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THANKS TO
Paola Cinque Simona Bossolasco
Bestetti Arabella Stefania Sala Silvia
Presi Richard W Price Serena Spudich
Magnus Gisslen Lars Hagberg
Department of Infectious Diseases San Raffaele
Hospital, Milan Department of
Neurovirology UCSF, USA Department of
Infectious Diseases University of Goteborg,
Sweden