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Incorporation of a GPI-anchored engineered cytokine as a molecular adjuvant enhances the immunogenicity of HIV VLPs.

Feng H, Zhang H, Deng J, Wang L, He Y, Wang S, Seyedtabaei R, Wang Q, Liu L, Galipeau J, Compans RW, Wang BZ - Sci Rep (2015)

Bottom Line: Co-delivery of mucosal adjuvants has been shown to be essential to induce effective mucosal immunity by non-replicating vaccines.A novel cytokine, GIFT4, engineered by fusing GM-CSF and interleukin-4, was previously found to simulate B cell proliferation and effector function.Incorporation of GPI-anchored GIFT4 into VLPs as a molecular adjuvant represents a novel approach to increase their immunogenicity.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Emory Vaccine Center, Emory University, Atlanta, GA 30322, USA.

ABSTRACT
HIV vaccines should elicit immune responses at both the mucosal portals of entry to block transmission and systemic compartments to clear disseminated viruses. Co-delivery of mucosal adjuvants has been shown to be essential to induce effective mucosal immunity by non-replicating vaccines. A novel cytokine, GIFT4, engineered by fusing GM-CSF and interleukin-4, was previously found to simulate B cell proliferation and effector function. Herein a membrane-anchored form of GIFT4 was constructed by fusing a glycolipid (GPI)-anchoring sequence and incorporated into Env-enriched HIV virus-like particles (VLPs) as a molecular adjuvant. Guinea pigs were immunized with the resulting HIV VLPs through an intramuscular priming-intranasal boosting immunization route. The GIFT4-containing VLPs induced higher levels of systemic antibody responses with significantly increased binding avidity and improved neutralizing breadth and potency to a panel of selected strains, as well as higher levels of IgG and IgA at several mucosal sites. Thus, the novel GPI-GIFT4-containging VLPs have the potential to be developed into a prophylactic HIV vaccine. Incorporation of GPI-anchored GIFT4 into VLPs as a molecular adjuvant represents a novel approach to increase their immunogenicity.

No MeSH data available.


Related in: MedlinePlus

Diagram of GPI-anchored form of GIFT4 and insect cell expression.a, a schematic presentation of the GPI-GIFT4 encoding gene. Coding sequences for the melittin SP and murine CD59-GPI anchor were fused at the 5′ and 3′-ends of GIFT4 encoding DNA, respectively, to form the full-length gene. b, GPI-GIFT4 cellular expression. Western blots were developed with anti-GM-CSF antibody. Lane 1, whole cell lysate of GPI-GIFT4-expressing rBV infected insect cell sample; Lane 2, purified GM-CSF protein; Lane 3, whole cell lysate of HIV Env-expressing rBV infected cells as a control. c, FACS analysis of GPI-GIFT4 cell surface expression in rBV-infected insect cell samples. Red, isotype antibody control; Blue, Control Cells + anti-GMCSF antibodies; Green, GPI-GIFT4–expressing cells + anti-GMCSF antibodies.
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f1: Diagram of GPI-anchored form of GIFT4 and insect cell expression.a, a schematic presentation of the GPI-GIFT4 encoding gene. Coding sequences for the melittin SP and murine CD59-GPI anchor were fused at the 5′ and 3′-ends of GIFT4 encoding DNA, respectively, to form the full-length gene. b, GPI-GIFT4 cellular expression. Western blots were developed with anti-GM-CSF antibody. Lane 1, whole cell lysate of GPI-GIFT4-expressing rBV infected insect cell sample; Lane 2, purified GM-CSF protein; Lane 3, whole cell lysate of HIV Env-expressing rBV infected cells as a control. c, FACS analysis of GPI-GIFT4 cell surface expression in rBV-infected insect cell samples. Red, isotype antibody control; Blue, Control Cells + anti-GMCSF antibodies; Green, GPI-GIFT4–expressing cells + anti-GMCSF antibodies.

Mentions: A diagram of the membrane-bound form of GIFT4 gene is shown in Fig. 1a. The melittin signal peptide (SP) and CD59 GPI anchoring signal-coding sequences were fused to 5′ and 3′-ends of the GIFT4 encoding sequence in frame to facilitate the membrane insertion of GPI-GIFT421. Western blot (Fig. 1b) using anti-GM-CSF antibody detected a band migrating at 37 kDa in the lysate of sf9 cells infected by recombinant baculoviruses (rBVs) expressing the GPI-GIFT4 gene (lane 1 in Fig. 1b), corresponding to the expected size of GPI-anchored GIFT4. The membrane anchoring of the expressed GPI-GIFT4 was further demonstrated by the enhanced fluorescent intensity measured by FACS analysis of the rBV-infected cells after cell surface staining with anti-GM-CSF antibodies, followed by PE-conjugated secondary antibodies (Fig.1c).


Incorporation of a GPI-anchored engineered cytokine as a molecular adjuvant enhances the immunogenicity of HIV VLPs.

Feng H, Zhang H, Deng J, Wang L, He Y, Wang S, Seyedtabaei R, Wang Q, Liu L, Galipeau J, Compans RW, Wang BZ - Sci Rep (2015)

Diagram of GPI-anchored form of GIFT4 and insect cell expression.a, a schematic presentation of the GPI-GIFT4 encoding gene. Coding sequences for the melittin SP and murine CD59-GPI anchor were fused at the 5′ and 3′-ends of GIFT4 encoding DNA, respectively, to form the full-length gene. b, GPI-GIFT4 cellular expression. Western blots were developed with anti-GM-CSF antibody. Lane 1, whole cell lysate of GPI-GIFT4-expressing rBV infected insect cell sample; Lane 2, purified GM-CSF protein; Lane 3, whole cell lysate of HIV Env-expressing rBV infected cells as a control. c, FACS analysis of GPI-GIFT4 cell surface expression in rBV-infected insect cell samples. Red, isotype antibody control; Blue, Control Cells + anti-GMCSF antibodies; Green, GPI-GIFT4–expressing cells + anti-GMCSF antibodies.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493578&req=5

f1: Diagram of GPI-anchored form of GIFT4 and insect cell expression.a, a schematic presentation of the GPI-GIFT4 encoding gene. Coding sequences for the melittin SP and murine CD59-GPI anchor were fused at the 5′ and 3′-ends of GIFT4 encoding DNA, respectively, to form the full-length gene. b, GPI-GIFT4 cellular expression. Western blots were developed with anti-GM-CSF antibody. Lane 1, whole cell lysate of GPI-GIFT4-expressing rBV infected insect cell sample; Lane 2, purified GM-CSF protein; Lane 3, whole cell lysate of HIV Env-expressing rBV infected cells as a control. c, FACS analysis of GPI-GIFT4 cell surface expression in rBV-infected insect cell samples. Red, isotype antibody control; Blue, Control Cells + anti-GMCSF antibodies; Green, GPI-GIFT4–expressing cells + anti-GMCSF antibodies.
Mentions: A diagram of the membrane-bound form of GIFT4 gene is shown in Fig. 1a. The melittin signal peptide (SP) and CD59 GPI anchoring signal-coding sequences were fused to 5′ and 3′-ends of the GIFT4 encoding sequence in frame to facilitate the membrane insertion of GPI-GIFT421. Western blot (Fig. 1b) using anti-GM-CSF antibody detected a band migrating at 37 kDa in the lysate of sf9 cells infected by recombinant baculoviruses (rBVs) expressing the GPI-GIFT4 gene (lane 1 in Fig. 1b), corresponding to the expected size of GPI-anchored GIFT4. The membrane anchoring of the expressed GPI-GIFT4 was further demonstrated by the enhanced fluorescent intensity measured by FACS analysis of the rBV-infected cells after cell surface staining with anti-GM-CSF antibodies, followed by PE-conjugated secondary antibodies (Fig.1c).

Bottom Line: Co-delivery of mucosal adjuvants has been shown to be essential to induce effective mucosal immunity by non-replicating vaccines.A novel cytokine, GIFT4, engineered by fusing GM-CSF and interleukin-4, was previously found to simulate B cell proliferation and effector function.Incorporation of GPI-anchored GIFT4 into VLPs as a molecular adjuvant represents a novel approach to increase their immunogenicity.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Emory Vaccine Center, Emory University, Atlanta, GA 30322, USA.

ABSTRACT
HIV vaccines should elicit immune responses at both the mucosal portals of entry to block transmission and systemic compartments to clear disseminated viruses. Co-delivery of mucosal adjuvants has been shown to be essential to induce effective mucosal immunity by non-replicating vaccines. A novel cytokine, GIFT4, engineered by fusing GM-CSF and interleukin-4, was previously found to simulate B cell proliferation and effector function. Herein a membrane-anchored form of GIFT4 was constructed by fusing a glycolipid (GPI)-anchoring sequence and incorporated into Env-enriched HIV virus-like particles (VLPs) as a molecular adjuvant. Guinea pigs were immunized with the resulting HIV VLPs through an intramuscular priming-intranasal boosting immunization route. The GIFT4-containing VLPs induced higher levels of systemic antibody responses with significantly increased binding avidity and improved neutralizing breadth and potency to a panel of selected strains, as well as higher levels of IgG and IgA at several mucosal sites. Thus, the novel GPI-GIFT4-containging VLPs have the potential to be developed into a prophylactic HIV vaccine. Incorporation of GPI-anchored GIFT4 into VLPs as a molecular adjuvant represents a novel approach to increase their immunogenicity.

No MeSH data available.


Related in: MedlinePlus