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Sponholtz, M. R.

Publications and source records attributed to Sponholtz, M. R..

3 recordsLinked to original sources

A human cytomegalovirus prefusion-like glycoprotein B subunit vaccine elicits similar humoral immunity to that of postfusion gB in mice

Human cytomegalovirus (HCMV) is the leading infectious cause of birth defects. Despite the global disease burden, there is no FDA-approved HCMV vaccine. The most efficacious HCMV vaccine candidates to date have used glycoprotein B (gB), a class III viral fusion protein, in its postfusion form. While some viral fusion proteins have been shown to elicit stronger neutralizing responses in their prefusion conformation, HCMV prefusion-like and postfusion gB were recently shown to elicit antibodies with similar fibroblast neutralization titers in mice. We aimed to define and compare the specificity and functionality of plasma IgG elicited by distinct prefusion-like and postfusion gB constructs. Prefusion-like and postfusion gB elicited comparable IgG responses that predominantly mapped to AD-5 antigenic domain known to elicit neutralizing antibodies. Interestingly, postfusion gB elicited significantly higher plasma IgG binding to cell-associated gB and antibody dependent cellular phagocytosis than that of prefusion-like gB. The vaccines elicited comparable neutralization titers of heterologous HCMV strain AD169r in fibroblasts, yet neither elicited neutralizing titers against the vaccine-matched strain Towne in fibroblasts. Our data indicate that gB in this prefusion-like conformation elicits similar specificity and functional humoral immunity to that of postfusion gB, unlike certain class I viral fusion proteins which have been used as vaccine antigens. These findings deepen our understanding of the immune response elicited by class III fusion proteins and may inform further design and testing of conformationally-dependent herpesvirus glycoprotein vaccine candidates.

immunology↗

Structure-based design of a soluble human cytomegalovirus glycoprotein B antigen stabilized in a prefusion-like conformation

Human cytomegalovirus (HCMV) glycoprotein B (gB) is a class III membrane fusion protein required for viral entry. HCMV vaccine candidates containing gB have demonstrated moderate clinical efficacy, but no HCMV vaccine has been FDA-approved. Here, we used structure-based design to identify and characterize amino acid substitutions that stabilize gB in its metastable prefusion conformation. One variant containing two engineered interprotomer disulfide bonds and two cavity-filling substitutions (gB-C7), displayed increased expression and thermostability. A 2.8 [A] resolution cryo-electron microscopy structure shows that gB-C7 adopts a prefusion-like conformation, revealing additional structural elements at the membrane-distal apex. Unlike previous observations for several class I viral fusion proteins, mice immunized with postfusion or prefusion-stabilized forms of soluble gB protein displayed similar neutralizing antibody titers, here specifically against an HCMV laboratory strain on fibroblasts. Collectively, these results identify initial strategies to stabilize class III viral fusion proteins and provide tools to probe gB-directed antibody responses. TeaserA structure-based design campaign leads to stabilization of the class III viral fusion protein from HCMV in a prefusion-like conformation.

molecular biology↗

A single, improbable B cell receptor mutation confers potent neutralization against cytomegalovirus

Cytomegalovirus (CMV) is a leading cause of infant hearing loss and neurodevelopmental delay, but vaccine candidates have faced challenges eliciting neutralizing antibodies. One of the most well-studied targets for CMV vaccines is the viral fusogen glycoprotein B (gB), which is required for viral entry into host cells. Within gB, antigenic domain 2 site 1 (AD-2S1) is a target of potently neutralizing antibodies, but gB-based candidate vaccines have yet to elicit robust responses against this region. We mapped the genealogy of B cells encoding potently neutralizing anti-gB AD-2S1 antibodies from their inferred unmutated common ancestor (UCA) and characterized the binding and function of early lineage ancestors. Remarkably, we found that the single amino acid heavy chain mutation A33N, an improbable mutation rarely generated by somatic hypermutation machinery, conferred broad CMV neutralization to the UCA antibody. Structural studies revealed that this mutation mediated key contacts with the gB AD-2S1 epitope. Collectively, these results provide insight into potently neutralizing gB-directed antibody evolution and a foundation for designing next-generation CMV vaccines. One-sentence summaryThis manuscript identifies an early B cell lineage mutation that confers neutralizing function to antibodies targeting CMV fusogen gB.

microbiology↗