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AAV-expressed eCD4-Ig provides durable protection from multiple SHIV challenges.

Gardner MR, Kattenhorn LM, Kondur HR, von Schaewen M, Dorfman T, Chiang JJ, Haworth KG, Decker JM, Alpert MD, Bailey CC, Neale ES, Fellinger CH, Joshi VR, Fuchs SP, Martinez-Navio JM, Quinlan BD, Yao AY, Mouquet H, Gorman J, Zhang B, Poignard P, Nussenzweig MC, Burton DR, Kwong PD, Piatak M, Lifson JD, Gao G, Desrosiers RC, Evans DT, Hahn BH, Ploss A, Cannon PM, Seaman MS, Farzan M - Nature (2015)

Bottom Line: Rhesus macaques inoculated with an AAV vector stably expressed 17-77 μg ml(-1) of fully functional rhesus eCD4-Ig for more than 40 weeks, and these macaques were protected from several infectious challenges with SHIV-AD8.Rhesus eCD4-Ig was also markedly less immunogenic than rhesus forms of four well-characterized bNAbs.Our data suggest that AAV-delivered eCD4-Ig can function like an effective HIV-1 vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Diseases, The Scripps Research Institute, Jupiter, Florida 33458, USA.

ABSTRACT
Long-term in vivo expression of a broad and potent entry inhibitor could circumvent the need for a conventional vaccine for HIV-1. Adeno-associated virus (AAV) vectors can stably express HIV-1 broadly neutralizing antibodies (bNAbs). However, even the best bNAbs neutralize 10-50% of HIV-1 isolates inefficiently (80% inhibitory concentration (IC80) > 5 μg ml(-1)), suggesting that high concentrations of these antibodies would be necessary to achieve general protection. Here we show that eCD4-Ig, a fusion of CD4-Ig with a small CCR5-mimetic sulfopeptide, binds avidly and cooperatively to the HIV-1 envelope glycoprotein (Env) and is more potent than the best bNAbs (geometric mean half-maximum inhibitory concentration (IC50) < 0.05 μg ml(-1)). Because eCD4-Ig binds only conserved regions of Env, it is also much broader than any bNAb. For example, eCD4-Ig efficiently neutralized 100% of a diverse panel of neutralization-resistant HIV-1, HIV-2 and simian immunodeficiency virus isolates, including a comprehensive set of isolates resistant to the CD4-binding site bNAbs VRC01, NIH45-46 and 3BNC117. Rhesus macaques inoculated with an AAV vector stably expressed 17-77 μg ml(-1) of fully functional rhesus eCD4-Ig for more than 40 weeks, and these macaques were protected from several infectious challenges with SHIV-AD8. Rhesus eCD4-Ig was also markedly less immunogenic than rhesus forms of four well-characterized bNAbs. Our data suggest that AAV-delivered eCD4-Ig can function like an effective HIV-1 vaccine.

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Comparison of eCD4-Ig variants and HIV-1 neutralizing antibodiesa, HIV-1 pseudotyped with the Envs of the indicated isolates were incubated with TZM-bl cells and varying concentrations of the indicated entry inhibitors, and the resulting IC50 values are plotted. IC90 values and standard errors are presented in Extended Data Figs. 6a and 6b. b, Experiments similar to those in (a) except that HIV-1 pseudotyped with the SIVmac239 Env was incubated with varying concentrations of CD4-Ig, eCD4-Ig variants, or CD4bs antibodies. Extended Data Fig. 6c shows a similar study using the HIV-2 ST Env. Errors bars of triplicates denote one s.e.m. of triplicates. c, ADCC activity was assessed using CEM.NKR-CCR5 target cells incubated with infectious HIV-1 NL4-3, SHIVKB9 or SIVmac239 for four days. Cells were then incubated with KHYG-1 NK effector cells26 for 8 hours in the presence of the indicated inhibitors. ADCC activity was measured as loss of luciferase activity from the target cells. All experiments represented in this figure were performed at least twice with each isolate and inhibitor with similar results.
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Figure 2: Comparison of eCD4-Ig variants and HIV-1 neutralizing antibodiesa, HIV-1 pseudotyped with the Envs of the indicated isolates were incubated with TZM-bl cells and varying concentrations of the indicated entry inhibitors, and the resulting IC50 values are plotted. IC90 values and standard errors are presented in Extended Data Figs. 6a and 6b. b, Experiments similar to those in (a) except that HIV-1 pseudotyped with the SIVmac239 Env was incubated with varying concentrations of CD4-Ig, eCD4-Ig variants, or CD4bs antibodies. Extended Data Fig. 6c shows a similar study using the HIV-2 ST Env. Errors bars of triplicates denote one s.e.m. of triplicates. c, ADCC activity was assessed using CEM.NKR-CCR5 target cells incubated with infectious HIV-1 NL4-3, SHIVKB9 or SIVmac239 for four days. Cells were then incubated with KHYG-1 NK effector cells26 for 8 hours in the presence of the indicated inhibitors. ADCC activity was measured as loss of luciferase activity from the target cells. All experiments represented in this figure were performed at least twice with each isolate and inhibitor with similar results.

Mentions: We compared eCD4-Ig, eCD4-Igmim2 and eCD4-IgQ40A,mim2 with a panel of 12 antibodies and inhibitors using three additional HIV-1 isolates (Fig. 2a; Extended Data Fig. 6a and b). eCD4-Ig and its variants neutralized the SG3 and YU2 isolates more efficiently than any of these inhibitors. Five bNAbs neutralized JR-CSF more efficiently than any eCD4-Ig variant, but four of these could not neutralize SG3. All eCD4-Ig variants neutralized these isolates with IC50s less than 0.3 μg/ml, more efficiently than CD4-Ig, the tetrameric CD4-Ig variant PRO-54212,14, or the antibodies 2G12, 4E10, and VRC01. eCD4-Ig and its variants, but not three CD4-binding site bNAbs, neutralized the neutralization-resistant SIVmac239 as well as HIV-2 ST (Fig. 2b; Extended Data Fig. 6c). As observed with SIVmac251, the Q40A variant was less efficient at neutralizing SIVmac239 and HIV-2. The potency of these eCD4-Ig variants was also reflected in their abilities to mediate antibody-dependent cell-mediated cytotoxicity (ADCC). eCD4-Ig, eCD4-Igmim2, and eCD4-IgQ40A,mim2 each facilitated 30–40 times more killing of infected cells by CD16+ natural killer cells26 than did CD4-Ig or the antibody IgGb12 (Fig. 2c). Thus the carboxy-terminal modification of eCD4-Ig did not interfere with the ADCC effector function of its Fc domain.


AAV-expressed eCD4-Ig provides durable protection from multiple SHIV challenges.

Gardner MR, Kattenhorn LM, Kondur HR, von Schaewen M, Dorfman T, Chiang JJ, Haworth KG, Decker JM, Alpert MD, Bailey CC, Neale ES, Fellinger CH, Joshi VR, Fuchs SP, Martinez-Navio JM, Quinlan BD, Yao AY, Mouquet H, Gorman J, Zhang B, Poignard P, Nussenzweig MC, Burton DR, Kwong PD, Piatak M, Lifson JD, Gao G, Desrosiers RC, Evans DT, Hahn BH, Ploss A, Cannon PM, Seaman MS, Farzan M - Nature (2015)

Comparison of eCD4-Ig variants and HIV-1 neutralizing antibodiesa, HIV-1 pseudotyped with the Envs of the indicated isolates were incubated with TZM-bl cells and varying concentrations of the indicated entry inhibitors, and the resulting IC50 values are plotted. IC90 values and standard errors are presented in Extended Data Figs. 6a and 6b. b, Experiments similar to those in (a) except that HIV-1 pseudotyped with the SIVmac239 Env was incubated with varying concentrations of CD4-Ig, eCD4-Ig variants, or CD4bs antibodies. Extended Data Fig. 6c shows a similar study using the HIV-2 ST Env. Errors bars of triplicates denote one s.e.m. of triplicates. c, ADCC activity was assessed using CEM.NKR-CCR5 target cells incubated with infectious HIV-1 NL4-3, SHIVKB9 or SIVmac239 for four days. Cells were then incubated with KHYG-1 NK effector cells26 for 8 hours in the presence of the indicated inhibitors. ADCC activity was measured as loss of luciferase activity from the target cells. All experiments represented in this figure were performed at least twice with each isolate and inhibitor with similar results.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4352131&req=5

Figure 2: Comparison of eCD4-Ig variants and HIV-1 neutralizing antibodiesa, HIV-1 pseudotyped with the Envs of the indicated isolates were incubated with TZM-bl cells and varying concentrations of the indicated entry inhibitors, and the resulting IC50 values are plotted. IC90 values and standard errors are presented in Extended Data Figs. 6a and 6b. b, Experiments similar to those in (a) except that HIV-1 pseudotyped with the SIVmac239 Env was incubated with varying concentrations of CD4-Ig, eCD4-Ig variants, or CD4bs antibodies. Extended Data Fig. 6c shows a similar study using the HIV-2 ST Env. Errors bars of triplicates denote one s.e.m. of triplicates. c, ADCC activity was assessed using CEM.NKR-CCR5 target cells incubated with infectious HIV-1 NL4-3, SHIVKB9 or SIVmac239 for four days. Cells were then incubated with KHYG-1 NK effector cells26 for 8 hours in the presence of the indicated inhibitors. ADCC activity was measured as loss of luciferase activity from the target cells. All experiments represented in this figure were performed at least twice with each isolate and inhibitor with similar results.
Mentions: We compared eCD4-Ig, eCD4-Igmim2 and eCD4-IgQ40A,mim2 with a panel of 12 antibodies and inhibitors using three additional HIV-1 isolates (Fig. 2a; Extended Data Fig. 6a and b). eCD4-Ig and its variants neutralized the SG3 and YU2 isolates more efficiently than any of these inhibitors. Five bNAbs neutralized JR-CSF more efficiently than any eCD4-Ig variant, but four of these could not neutralize SG3. All eCD4-Ig variants neutralized these isolates with IC50s less than 0.3 μg/ml, more efficiently than CD4-Ig, the tetrameric CD4-Ig variant PRO-54212,14, or the antibodies 2G12, 4E10, and VRC01. eCD4-Ig and its variants, but not three CD4-binding site bNAbs, neutralized the neutralization-resistant SIVmac239 as well as HIV-2 ST (Fig. 2b; Extended Data Fig. 6c). As observed with SIVmac251, the Q40A variant was less efficient at neutralizing SIVmac239 and HIV-2. The potency of these eCD4-Ig variants was also reflected in their abilities to mediate antibody-dependent cell-mediated cytotoxicity (ADCC). eCD4-Ig, eCD4-Igmim2, and eCD4-IgQ40A,mim2 each facilitated 30–40 times more killing of infected cells by CD16+ natural killer cells26 than did CD4-Ig or the antibody IgGb12 (Fig. 2c). Thus the carboxy-terminal modification of eCD4-Ig did not interfere with the ADCC effector function of its Fc domain.

Bottom Line: Rhesus macaques inoculated with an AAV vector stably expressed 17-77 μg ml(-1) of fully functional rhesus eCD4-Ig for more than 40 weeks, and these macaques were protected from several infectious challenges with SHIV-AD8.Rhesus eCD4-Ig was also markedly less immunogenic than rhesus forms of four well-characterized bNAbs.Our data suggest that AAV-delivered eCD4-Ig can function like an effective HIV-1 vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Diseases, The Scripps Research Institute, Jupiter, Florida 33458, USA.

ABSTRACT
Long-term in vivo expression of a broad and potent entry inhibitor could circumvent the need for a conventional vaccine for HIV-1. Adeno-associated virus (AAV) vectors can stably express HIV-1 broadly neutralizing antibodies (bNAbs). However, even the best bNAbs neutralize 10-50% of HIV-1 isolates inefficiently (80% inhibitory concentration (IC80) > 5 μg ml(-1)), suggesting that high concentrations of these antibodies would be necessary to achieve general protection. Here we show that eCD4-Ig, a fusion of CD4-Ig with a small CCR5-mimetic sulfopeptide, binds avidly and cooperatively to the HIV-1 envelope glycoprotein (Env) and is more potent than the best bNAbs (geometric mean half-maximum inhibitory concentration (IC50) < 0.05 μg ml(-1)). Because eCD4-Ig binds only conserved regions of Env, it is also much broader than any bNAb. For example, eCD4-Ig efficiently neutralized 100% of a diverse panel of neutralization-resistant HIV-1, HIV-2 and simian immunodeficiency virus isolates, including a comprehensive set of isolates resistant to the CD4-binding site bNAbs VRC01, NIH45-46 and 3BNC117. Rhesus macaques inoculated with an AAV vector stably expressed 17-77 μg ml(-1) of fully functional rhesus eCD4-Ig for more than 40 weeks, and these macaques were protected from several infectious challenges with SHIV-AD8. Rhesus eCD4-Ig was also markedly less immunogenic than rhesus forms of four well-characterized bNAbs. Our data suggest that AAV-delivered eCD4-Ig can function like an effective HIV-1 vaccine.

Show MeSH
Related in: MedlinePlus