18th Conference on Retroviruses and
Opportunistic Infections CROI 2011
February 27th-March 2nd, Boston, USA
Effector CD8+ T cells expressing Granzyme B Surround JC Virus-Infected
Oligodendrocytes in PML-IRIS Lesions
G. Martin-Blondel1,2, J. Bauer3, V. Cuvinciuc1, P. Delobel1,2, E. Uro-Coste1, F. Bonneville1, M.B. Delisle1, H. Lassmann3, L.T. Mars2, and B. Marchou1 Infectious diseases department, Toulouse University Hospital, Toulouse, F-31000 France.
Tél. : 33 5 61 77 23 53 Fax : 33 5 61 77 21 38
1Toulouse University Hospital, Toulouse, France 2INSERM U1043 – CNRS UMR 5282, Toulouse, France 3Medical University of Vienna, Vienna, Austria Corresponding author: G. Martin-Blondel
Background: Antiretroviral therapy (ART)-induced immune recovery is not always beneficial to
progressive multifocal leukoencephalopathy (PML). We analyzed features of PML associated with Immune Clinical features of PML-IRIS patients Histological features of PML-IRIS patients Prognosis of PML-IRIS patients
Reconstitution Inflammatory Syndrome (PML-IRIS) in HIV-infected patients.
• Among 40 HIV-infected patients diagnosed with PML over the ten-year study period, eleven cases fulfilled criteria for • Stereotactic brain biopsies were performed in five patients, mainly because the diagnosis of PML-IRIS was not made after • Six patients received steroid therapy (54.5%). Median dose and duration were 100 mg
Methods: Retrospective cohort study of all HIV-infected patients diagnosed with PML-IRIS in a single
PML-IRIS: nine paradoxical PML-IRIS and two unmasking PML-IRIS. analysis of MRI. All biopsies were performed before any steroid therapy. [Range 60-1000] and 10.5 days [Range 3-60]. There was no significant difference between
center between January 2000 and January 2010.
• Median age was 42 years [IQ25-75 40-47]. • In all cases, histological analysis confirmed PML and demonstrated demyelinating lesions associated with inflammatory patients treated or not by steroids.
Results: Among 40 consecutive patients diagnosed with PML, 11 developed PML-IRIS (9 paradoxical and
• Ten patients were males (91%). infiltrates dominated by CD8+ cells, in association in a lesser extent with CD68+ and CD20+ cells (table 2). • The overall survival of PML-IRIS patients since IRIS onset was 21 months [Range
2 unmasking). Median time between ART initiation and onset of IRIS was 36 days [Range 18-66]. At IRIS
• Ten patients were previously naive of ART (91%). • In three patients, CD3+ T cells express Granzyme B. In the patient n°4 triple labeling confocal microscopy showed 0.4-105.6].
onset, median CD4 T cell count and HIV viral load were 181/mm3 [IQ25-75 92,5-252,5] (+104/mm3 [IQ25-75
• Initiated ART was based on NRTI associated with boosted PI in nine patients (three receiving also enfuvirtide) or NNRTI concentration of CD3+ Granzyme B+ T cells around JC-infected oligodendrocytes, without apposition of these cytotoxic T • Four patients (36.3%) deceased of PML-IRIS with a median time to death of 1 month after
47,5-182]) and 2,8 log10 copies/mL [IQ25-75 2,2-3,42] (-3 log10 copies/mL [IQ25-75 3,2-2,4]). Brain MRI
in two patients. cells neither polarization of granules to oligodendrocytes. the diagnosis of PML-IRIS [Range 0.4-1.3].
showed contrast-enhanced lesions and/or mass effect in 7 patients (64%) but neither in 4 patients (36%).
• Median time between ART initiation and onset of IRIS was 36 days [Range 18-66]. At onset of IRIS, compared to the patients who survived, the deceased patients
Brain biopsies were obtained from 5 patients and allowed a better understanding of mechanisms of PML-
• Clinical manifestations of PML-IRIS were occurrence or worsening of motor impairment (11/11), cranial nerve palsies demonstrated a trend to:
IRIS. Histological analysis and immunohistochemistry confirmed PML and showed demyelination (varying Brain infiltrating cells
(5/11), ataxia (2/11), seizures (2/11) or vigilance disorders (2/11). A higher amplification of CD8+ T cell (+ 417/mm3 [IQ25-75 53-478] versus – 46/mm3
of 15 to 100%) and CD8+ T cell infiltration (varying of 29 to 1264 cells/mm2). In 3 patients Granzyme B
PML-IRIS Demyelination CD8+ cells/mm2 Granzyme B+
was detected in up to 22.5% of CD8+ T cells, testifying their cytotoxic differentiation. Confocal microscopy [IQ25-75 -161-+590])
CD3+ cells/mm2 (% of CD3+ cells/mm2 (% of CD20+ cells CD68+ cells
studies in one patient demonstrated that concentration of CD8+ Granzyme B+ T cells surround JC-infected
oligodendrocytes. Six patients received steroid therapy (54.5%). Four patients deceased of PML-IRIS
Biological and neuro-imaging features of PML-IRIS patients cells) CD3+ cells)
A larger number of cerebral areas affected (5 [IQ25-75 4-5.5] versus 3 [IQ25-75 2-4])
A more frequent mass effect on brain MRI (4/4 versus 1/7)
(36.3%). At onset of IRIS, compared to the patients who survived, the deceased patients demonstrated a 1 Unmasking 100% 256 236 (92) 8 (3.1) + ++ Less prescription of steroids therapy (1/4 versus 5/7).
trend to a higher amplification of CD8+ T cell (Median increase + 417/mm3 [IQ25-75 53-478] versus – 46/ 2 Unmasking 15% 672 756 (112) 8 (1) + ++ • With a median follow-up of 45.2 months [Range 4.9-105.6], all survivors present a mild
mm3 [IQ25-75 -161-+590]), a larger number of cerebral areas affected (5 [IQ25-75 4-5.5] versus 3 [IQ25-75 2-4]) PML-IRIS Initiation of ART Onset of IRIS Delta neurological disability (Mean modified Rankin scale: 2).
3 Paradoxical 100% 29 33 (113) 0 + ++
and a more frequent mass effect on brain MRI (4/4 versus 1/7); they also received less often steroids therapy
(1/4 versus 5/7). With a median follow-up of 45.2 months [Range 4.9-105.6], all survivors present a mild CD4 T cells/mm3 51 [42-94] 181 [92-252] + 104 [47-182] 4 Paradoxical >50% 852 980 (105) 192 (22.5) + ++
neurological disability (Mean modified Rankin scale: 2). 5 Paradoxical 100% 1264 1420 (112) 0 + ++ CONCLUSION & PERSPECTIVES
Conclusions: IRIS is a frequent complication of PML in HIV-infected patients initiating ART. Here we CD8 T cells/mm3 542 [428-852] 796 [556-1047] + 231 [-25-+478]
Table 2: Main histological characteristics of the five PML-IRIS patients in whom a stereotactic biopsy was performed • PML-IRIS is a frequent and life-threatening complication in PML patients initiating
report that Granzyme B+ CD8+ T cells dominate in lesions of PML-IRIS where they surround JC virus-
CD4/CD8 ratio 0.11 [0.08-0.13] 0.23 [0.16-0.30] + 0.11 [0.08-0.16] ART.
infected oligodendrocytes. These data support an important role of JC-specific CD8+ T cell responses in the
• As there is no biomarker, PML-IRIS is presently diagnosed on the basis of a
pathogenesis of PML-IRIS.
HIV viral load (log10 copies/mL) 5.7 [5.3-6.1] 2.8 [2.2-3.4] - 3 [3.2-2.4]
Figure 2: Histological analysis of a
stereotactic brain biopsy at PML-IRIS
A B C multiparametric assessment, including brain MRI.
However, performances of brain MRI are insufficient since 36% of patients in our
study did not demonstrate at the onset of PML-IRIS gadolinium-enhanced lesions or
Contrast-enhanced lesions (%)* 0/9 5/11 (45) - A: Luxol fast blue staining showing
extensive demyelination (x2.5), mass effect.
The immune reconstitution inflammatory syndrome affecting the central nervous system (neuro-IRIS) is an
Mass effect (%)* 0/9 5/11 (45) - Serial proton-magnetic-resonance-spectroscopy (PMRS) by demonstrating an
emerging complication of antiretroviral therapy (ART)-induced immune recovery in HIV-infected patients1. B: JC-positive oligodendrocytes (SV40
inflammatory profile during the clinical evolution from PML to paradoxical PML-IRIS
While a neuro-IRIS affects 16.7% (2.3–50.7) of HIV-infected patients with progressive multifocal immunohistochemistry, x20),
Contrast-enhanced lesions or mass effect (%)* 0/9 7/11 (64) under ART may be used to discriminate between PML and PML-IRIS5. Moreover PML
leukoencephalopathy (PML) initiating ART (PML-IRIS)2, there is no homogeneous series describing C: Macrophage/microglia infiltration (CD68
patients with detectable JC virus-specific CD8+ T cells have an increased likelihood of
clinical, radiological and histological features of PML-IRIS3. We report herein eleven consecutive cases of immunohistochemistry, x40),
Neither contrast-enhanced lesions or mass effect (%)* 9/9 4/11 (36) - D: CD8 T cells infiltration (CD8 having PML-IRIS (odds ratio 7.8, 95% CI 1.16-52.35)6. Assessing the inflammatory
PML-IRIS and focus on pathophysiology, diagnostic procedures and prognosis of PML-IRIS.
D E F profile by PMRS and the reactivity of circulating CD8+ T cells against JC virus at the
Table 1: Main biological and neuro-imaging features of PML-IRIS patients time of PML worsening may represent an accessible method to improve diagnosis of
AIM All continuous values are expressed as median and interquartile 25-75. E: Granzyme B positive cells (Granzyme B
* Concerning neuro-imaging data, the denominator is 9 at the initiation of ART (Patients with PML who develop a • Here we show that CD8+ T cells dominating inflammatory lesions are cytotoxic and
To describe immunopathological features of PML-IRIS in HIV-infected patients. F: B cells in perivascular cuffs (CD20
paradoxical PML-IRIS) and 11 at the onset of IRIS (patients who developed paradoxical and unmasking PML-IRIS). surround JC-infected oligodendrocytes. As JC-specific CD8+ T cells are detected in the
METHODS Figure 1: Brain MRI of a PML-IRIS patient
blood of patients with PML-IRIS, concomitant with contrast enhancement on brain MRI
and/or brain inflammation6,7, our data highlight that JC-specific CD8+ T cells are the main
• Retrospective cohort study of all HIV-infected patients diagnosed with PML in Toulouse University A and B: At PML diagnosis in an HIV-infected patient: effectors of tissular damages in PML-IRIS patients. These data support that neuro-IRIS is a
Hospital, France, between January 2000 and January 2010. A: Axial FLAIR showed multiple patchy areas of subcortical dysregulated host immune response provoked by pathogen-derived antigens that cause
• The diagnosis of PML relies on typical clinical and MRI findings (presumptive diagnosis), and on evidence hyperintensities, without mass effect disproportionate tissue damage.
of JC virus DNA in cerebrospinal fluid (CSF) or on consistent neuropathology in brain tissues with JC virus B: Axial T1 with gadolinium showed marked right frontal Figure 3: Triple labeling confocal
DNA or protein detected by in situ techniques (definite diagnosis)4. subcortical hypointensity, without enhancement. microscopy performed in patient n°4:
• C and D: At PML-IRIS onset (day 18 after ART initiation): Concentration of CD3+ Granzyme B+ T FUNDING
Diagnostic criteria for PML-IRIS cells around JC-infected oligodendrocytes
C: Axial FLAIR showed progression of the hyperintensities and
(SV40 (Purple), CD3 (Blue) and Granzyme This work was supported by the French national institute for medical research (INSERM),
i) the absence of neurological complaints in a patient starting ART and presenting afterward with right frontal sulcal effacement, suggesting a mass effect.
B (Green) immunohistochemistry, x100). the Agence Nationale pour la Recherche sur le Sida et les hépatites virales (ANRS) and the
neurological abnormalities due to PML (unmasking PML-IRIS), or pre-existing PML worsening after ART D: Axial T1 with gadolinium revealed peripheral contrast
Collège des Universitaires de Maladies Infectieuses et Tropicales (CMIT).
initiation (paradoxical PML-IRIS) enhancement. Note also the enhancement of a subcortical left
ii) a close temporal relationship between ART initiation and disease onset parietal lesion (arrow). REFERENCES
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