Targeted Killing of Virally Infected Cells by Radiolabeled Antibodies to Viral Proteins

BACKGROUND: The HIV epidemic is a major threat to health in the developing and western worlds. A modality that targets and kills HIV-1-infected cells could have a major impact on the treatment of acute exposure and the elimination of persistent reservoirs of infected cells. The aim of this proof-of-principle study was to demonstrate the efficacy of a therapeutic strategy of targeting and eliminating HIV-1-infected cells with radiolabeled antibodies specific to viral proteins in vitro and in vivo. METHODS AND FINDINGS: Antibodies to HIV-1 envelope glycoproteins gp120 and gp41 labeled with radioisotopes bismuth 213 ((213)Bi) and rhenium 188 ((188)Re) selectively killed chronically HIV-1-infected human T cells and acutely HIV-1-infected human peripheral blood mononuclear cells (hPBMCs) in vitro. Treatment of severe combined immunodeficiency (SCID) mice harboring HIV-1-infected hPBMCs in their spleens with a (213)Bi- or (188)Re-labeled monoclonal antibody (mAb) to gp41 resulted in a 57% injected dose per gram uptake of radiolabeled mAb in the infected spleens and in a greater than 99% elimination of HIV-1-infected cells in a dose-dependent manner. The number of HIV-1-infected thymocytes decreased 2.5-fold in the human thymic implant grafts of SCID mice treated with the (188)Re-labeled antibody to gp41 compared with those treated with the (188)Re-control mAb. The treatment did not cause acute hematologic toxicity in the treated mice. CONCLUSIONS: The current study demonstrates the effectiveness of HIV-targeted radioimmunotherapy and may provide a novel treatment option in combination with highly active antiretroviral therapy for the eradication of HIV

W-Curve Alignments for HIV-1 Genomic Comparisons

The W-curve was originally developed as a graphical visualization technique for viewing DNA and RNA sequences. Its ability to render features of DNA also makes it suitable for computational studies. Its main advantage in this area is utilizing a single-pass algorithm for comparing the sequences. Avoiding recursion during sequence alignments offers advantages for speed and in-process resources. The graphical technique also allows for multiple models of comparison to be used depending on the nucleotide patterns embedded in similar whole genomic sequences. The W-curve approach allows us to compare large numbers of samples quickly.We are currently tuning the algorithm to accommodate quirks specific to HIV-1 genomic sequences so that it can be used to aid in diagnostic and vaccine efforts. Tracking the molecular evolution of the virus has been greatly hampered by gap associated problems predominantly embedded within the envelope gene of the virus. Gaps and hypermutation of the virus slow conventional string based alignments of the whole genome. This paper describes the W-curve algorithm itself, and how we have adapted it for comparison of similar HIV-1 genomes. A treebuilding method is developed with the W-curve that utilizes a novel Cylindrical Coordinate distance method and gap analysis method. HIV-1 C2-V5 env sequence regions from a Mother/Infant cohort study are used in the comparison.The output distance matrix and neighbor results produced by the W-curve are functionally equivalent to those from Clustal for C2-V5 sequences in the mother/infant pairs infected with CRF01_AE.Significant potential exists for utilizing this method in place of conventional string based alignment of HIV-1 genomes, such as Clustal X. With W-curve heuristic alignment, it may be possible to obtain clinically useful results in a short time-short enough to affect clinical choices for acute treatment. A description of the W-curve generation process, including a comparison technique of aligning extremes of the curves to effectively phase-shift them past the HIV-1 gap problem, is presented. Besides yielding similar neighbor-joining phenogram topologies, most Mother and Infant C2-V5 sequences in the cohort pairs geometrically map closest to each other, indicating that W-curve heuristics overcame any gap problem

Different Pattern of Immunoglobulin Gene Usage by HIV-1 Compared to Non-HIV-1 Antibodies Derived from the Same Infected Subject

A biased usage of immunoglobulin (Ig) genes is observed in human anti-HIV-1 monoclonal antibodies (mAbs) resulting probably from compensation to reduced usage of the VH3 family genes, while the other alternative suggests that this bias usage is due to antigen requirements. If the antigen structure is responsible for the preferential usage of particular Ig genes, it may have certain implications for HIV vaccine development by the targeting of particular Ig gene-encoded B cell receptors to induce neutralizing anti-HIV-1 antibodies. To address this issue, we have produced HIV-1 specific and non-HIV-1 mAbs from an infected individual and analyzed the Ig gene usage. Green-fluorescence labeled virus-like particles (VLP) expressing HIV-1 envelope (Env) proteins of JRFL and BaL and control VLPs (without Env) were used to select single B cells for the production of 68 recombinant mAbs. Ten of these mAbs were HIV-1 Env specific with neutralizing activity against V3 and the CD4 binding site, as well as non-neutralizing mAbs to gp41. The remaining 58 mAbs were non-HIV-1 Env mAbs with undefined specificities. Analysis revealed that biased usage of Ig genes was restricted only to anti-HIV-1 but not to non-HIV-1 mAbs. The VH1 family genes were dominantly used, followed by VH3, VH4, and VH5 among anti-HIV-1 mAbs, while non-HIV-1 specific mAbs preferentially used VH3 family genes, followed by VH4, VH1 and VH5 families in a pattern identical to Abs derived from healthy individuals. This observation suggests that the biased usage of Ig genes by anti-HIV-1 mAbs is driven by structural requirements of the virus antigens rather than by compensation to any depletion of VH3 B cells due to autoreactive mechanisms, according to the gp120 superantigen hypothesis

Nucleoside Reverse Transcriptase Inhibitor Resistance Mutations Associated with First-Line Stavudine-Containing Antiretroviral Therapy: Programmatic Implications for Countries Phasing Out Stavudine

Background The World Health Organization Antiretroviral Treatment Guidelines recommend phasing-out stavudine because of its risk of long-term toxicity. There are two mutational pathways of stavudine resistance with different implications for zidovudine and tenofovir cross-resistance, the primary candidates for replacing stavudine. However, because resistance testing is rarely available in resource-limited settings, it is critical to identify the cross-resistance patterns associated with first-line stavudine failure. Methods We analyzed HIV-1 resistance mutations following first-line stavudine failure from 35 publications comprising 1,825 individuals. We also assessed the influence of concomitant nevirapine vs. efavirenz, therapy duration, and HIV-1 subtype on the proportions of mutations associated with zidovudine vs. tenofovir cross-resistance. Results Mutations with preferential zidovudine activity, K65R or K70E, occurred in 5.3% of individuals. Mutations with preferential tenofovir activity, ≥two thymidine analog mutations (TAMs) or Q151M, occurred in 22% of individuals. Nevirapine increased the risk of TAMs, K65R, and Q151M. Longer therapy increased the risk of TAMs and Q151M but not K65R. Subtype C and CRF01_AE increased the risk of K65R, but only CRF01_AE increased the risk of K65R without Q151M. Conclusions Regardless of concomitant nevirapine vs. efavirenz, therapy duration, or subtype, tenofovir was more likely than zidovudine to retain antiviral activity following first-line d4T therap

Human Non-neutralizing HIV-1 Envelope Monoclonal Antibodies Limit the Number of Founder Viruses during SHIV Mucosal Infection in Rhesus Macaques

HIV-1 mucosal transmission begins with virus or virus-infected cells moving through mucus across mucosal epithelium to infect CD4+ T cells. Although broadly neutralizing antibodies (bnAbs) are the type of HIV-1 antibodies that are most likely protective, they are not induced with current vaccine candidates. In contrast, antibodies that do not neutralize primary HIV-1 strains in the TZM-bl infection assay are readily induced by current vaccine candidates and have also been implicated as secondary correlates of decreased HIV-1 risk in the RV144 vaccine efficacy trial. Here, we have studied the capacity of anti-Env monoclonal antibodies (mAbs) against either the immunodominant region of gp41 (7B2 IgG1), the first constant region of gp120 (A32 IgG1), or the third variable loop (V3) of gp120 (CH22 IgG1) to modulate in vivo rectal mucosal transmission of a high-dose simian-human immunodeficiency virus (SHIV-BaL) in rhesus macaques. 7B2 IgG1 or A32 IgG1, each containing mutations to enhance Fc function, was administered passively to rhesus macaques but afforded no protection against productive clinical infection while the positive control antibody CH22 IgG1 prevented infection in 4 of 6 animals. Enumeration of transmitted/founder (T/F) viruses revealed that passive infusion of each of the three antibodies significantly reduced the number of T/F genomes. Thus, some antibodies that bind HIV-1 Env but fail to neutralize virus in traditional neutralization assays may limit the number of T/F viruses involved in transmission without leading to enhancement of viral infection. For one of these mAbs, gp41 mAb 7B2, we provide the first co-crystal structure in complex with a common cyclical loop motif demonstrated to be critical for infection by other retroviruses