Whole-body immunoPET reveals active SIV dynamics in viremic and antiretroviral therapy-treated macaques

Abstract

The detection of viral dynamics and localization in the context of controlled HIV infection remains a challenge and is limited to blood and biopsies. We developed a method to capture total-body simian immunodeficiency virus (SIV) replication using immunoPET (antibody-targeted positron emission tomography). The administration of a poly(ethylene glycol)-modified, (64)Cu-labeled SIV Gp120-specific antibody led to readily detectable signals in the gastrointestinal and respiratory tract, lymphoid tissues and reproductive organs of viremic monkeys. Viral signals were reduced in aviremic antiretroviral-treated monkeys but detectable in colon, select lymph nodes, small bowel, nasal turbinates, the genital tract and lung. In elite controllers, virus was detected primarily in foci in the small bowel, select lymphoid areas and the male reproductive tract, as confirmed by quantitative reverse-transcription PCR (qRT-PCR) and immunohistochemistry. This real-time, in vivo viral imaging method has broad applications to the study of immunodeficiency virus pathogenesis, drug and vaccine development, and the potential for clinical translation.

Figures

Figure 1

PET/CT results from uninfected control and four chronically SIV infected, viremic rhesus macaques. (a,b) Frontal, sagittal and axial views and magnifications of a viremic monkey, Viremic 1, and of a representative uninfected monkey, Control 1, imaged with the modified 7D3 probe. Sites of axial sections are marked by a yellow line within the sagittal view. Scale bars (white): axial and frontal view of torso and sagittal view of head, 100 mm; sagittal view of whole body, 50 mm; lymph nodes (LN) and genital tract, 20 mm. (c,d) IHC against SIV Gag in tissues from Viremic 1 (c) and in spleen from an SIV-infected monkey with high viremia and in uninfected colon control tissues (d). Arrows indicate infected mononuclear cells. Scale bars: colon uninfected (d), 50 μm; all other images (c,d), 20 μm. (e) Quantification of the PET data from various tissues from 4 SIV-infected and viremic monkeys (magenta) as well as 2 uninfected monkeys administered the 7D3 probe and 2 monkeys administered an isotype IgG probe (blue). The maximum standard uptake value (SUVmax) within each organ was compared directly with the qRT-PCR results. Mean ± s.d. shown for PET and SIV RNA copies for each organ across all animals tested. (f) Rectal biopsy results from 2 chronically infected and 2 uninfected animals. Activity is reported in counts per min (CPM) adjusted for tissue weight and the activity of the injected probe. (g) Comparison of SUVmax results in the same viremic and uninfected monkeys as in e; a repeated measures ANOVA confirmed that the animal conditions were statistically different (P = 7.39 × 10−6). SUVmax values obtained with nonspecific IgG control in uninfected animals are also included. (h) Measurement of the SUV ratio at two time points following probe injection (PI, post-injection).

Figure 2

PET/CT results from a chronically infected macaque, before and at 5 weeks of ART. (a) Standard uptake value (SUV) maps of GI tract, lymph nodes (LN), genital tract, spleen and small bowel. NALT, nasal-associated lymphoid tissue. Arrows indicate areas for which specific PET signals decreased during ART. Scale bars: frontal view of torso, 100 mm; sagittal view of head, 50 mm; LN and genital tract, 20 mm; small bowel, 15 mm. (b) SUVmax values before and after 5 weeks of ART compared with background uptake in 2 uninfected animals. (c) qRT-PCR verification of residual virus compared with SUVmax PET data at 5 weeks of ART in one representative treated macaque.

Figure 3

PET/CT results from SIV-infected elite controllers (ECs). (a) Frontal, sagittal and axial views and magnifications from one SIV-infected EC macaque 36 h after injection of the labeled antibody. The axial section is denoted by a yellow line within the sagittal view. Macaque EC 1 was infected for over 6 years and displayed plasma viral loads of fewer than 60 copies of viral RNA per ml for the last 5 years. Scale bars (white): axial and frontal view of torso and sagittal view of head, 100 mm; sagittal view of whole body, 50 mm; lymph nodes (LN) and genital tract, 20 mm. (b) SUVmax quantification results from the PET/CT imaging, comparing 4 viremic and 4 EC monkeys with 2 uninfected controls (mean ± s.d.). See Supplementary Note for a description of the statistical analysis of this data. (c) IHC results against the SIVMAC239 Gag for macaque EC 1. Arrows indicate infected mononuclear cells in select tissue biopsies. Scale bar, 20 μm.

Figure 4

Comparison of viremic with elite controller (EC) macaques. (a) Comparison of SUVmean and SUV voxel fraction (fraction of total volume of GI tract) within the GI tract of 4 chronic SIV-positive and 4 EC macaques. The voxel fractions that contained SUVs above 1 and below 3.3 (excludes interfering signals) were included. (b) Haralick texture function measurements of angular second moment and contrast for regions of interest (ROIs; depicted by red boxes in c,d). The angular second moment is a measure of homogeneity, whereas the contrast metric represents the local variations within an image or ROI. Data reported as mean ± s.d. (c,d) Representative axial cross sections of a chronically infected macaque (Viremic 1) and one controller macaque (EC 2). Red arrows indicate uptake within the liver; white arrows in c point toward the GI tract; white arrows in d point to mesenteric lymph nodes and other foci. Scale bars, 50 mm.