Biology subjects

Malim, M. H.

Publications and source records attributed to Malim, M. H..

5 recordsLinked to original sources

bioRxiv2022

Modular capsid decoration boosts adenovirus vaccine-induced humoral and cellular immunity against SARS-CoV-2

Adenovirus vector vaccines have been widely and successfully deployed in response to COVID-19. However, despite inducing potent T cell immunity, improvement of vaccine-specific antibody responses upon homologous boosting is modest compared to other technologies. Here, we describe a system to enable modular decoration of adenovirus capsid surfaces with protein antigens and demonstrate induction of potent humoral immunity against these displayed antigens. Ligand attachment via a covalent isopeptide bond was achieved in a rapid and spontaneous reaction, requiring simple co-incubation of ligand and vector components. We used a recently described protein superglue, DogTag/DogCatcher, which is similar to the widely used SpyTag/SpyCatcher ligation system but performs better in loop structures. DogTag was inserted into surface-exposed loops in the adenovirus hexon protein to allow attachment of DogCatcher-fused ligands on virus particles. Efficient coverage of the capsid surface was achieved using a variety of ligands and vector infectivity was retained in each case. Capsid decoration shielded particles from anti-adenovirus neutralizing antibodies. In prime-boost regimens, proof-of-concept COVID-19 adenovirus vaccines decorated with the receptor-binding domain (RBD) of SARS-CoV-2 spike induced >10-fold higher SARS-CoV-2 neutralization titers compared to an undecorated adenovirus vector encoding spike. Importantly, decorated vectors retained robust T cell immunogenicity to encoded antigens, a key hallmark of adenovirus vector vaccines. We propose capsid decoration via protein superglue-mediated covalent ligation as a novel strategy to improve the efficacy and boostability of adenovirus-based vaccines and therapeutics. One Sentence SummaryDecorating the capsid surface of adenovirus vaccine vectors using a spontaneous protein superglue induces potent pathogen-specific immunity

bioengineering↗

bioRxiv2022

MX2 viral substrate breadth and inhibitory activity are regulated by protein phosphorylation

Human immunodeficiency virus type-1 (HIV-1) infection is potently inhibited by human myxovirus resistance 2 (MX2/MxB), which binds to the viral capsid and blocks the nuclear import of viral DNA. We have recently shown that phosphorylation is a key regulator of MX2 antiviral activity, with phosphorylation of serine residues at positions 14, 17 and 18 repressing MX2 function. Here, we extend the study of MX2 post-translational modifications, and identify serine and threonine phosphorylation in all domains of MX2. By substituting these residues with aspartic acid or alanine, hence mimicking the presence or absence of a phosphate group, respectively, we identified key positions that control MX2 antiviral activity. Aspartic acid substitutions of residues Ser306 or Thr334 and alanine substitutions of Thr343 yielded proteins with substantially reduced antiviral activity, whereas the presence of aspartic acid at positions Ser28, Thr151 or Thr343 resulted in enhanced activity: referred to as hypermorphic mutants. In some cases, these hypermorphic mutations, particularly when paired with other MX2 mutations (e.g., S28D/T151D or T151D/T343A) acquired the capacity to inhibit HIV-1 Capsid mutants known to be insensitive to wild type MX2, such as P90A or T210K, as well as MX2-resistant retroviruses such as equine infectious anaemia virus (EIAV) and murine leukaemia virus (MLV). This work highlights the complexity and importance of MX2 phosphorylation in the regulation of antiviral activity, and in the selection of susceptible viral substrates. Author summaryProductive infection by human immunodeficiency virus type-1 (HIV-1) requires the import of viral replication complexes into the nuclei of infected cells. Myxovirus resistance 2 (MX2/MxB) blocks this step, halting nuclear accumulation of viral DNA and virus replication. We recently demonstrated how phosphorylation of a stretch of three serines in the amino-terminal domain of MX2 inhibits the antiviral activity. Here, we identify additional positions in MX2 whose phosphorylation status reduces or enhances antiviral function (hypomorphic and hypermorphic variants, respectively). Importantly, hypermorphic mutant proteins not only increased inhibitory activity against wild type HIV-1, but can also exhibit antiviral capabilities against HIV-1 Capsid mutant viruses that are resistant to wild type MX2. Furthermore, some of these proteins were also able to inhibit retroviruses that are insensitive to MX2. Therefore, we propose that phosphorylation comprises a major element of MX2 regulation and substrate determination.

microbiology↗

bioRxiv2021

ChAdOx1 nCoV-19 (AZD1222) vaccine elicits monoclonal antibodies with potent cross-neutralizing activity against SARS-CoV-2 viral variants

Although the antibody response to COVID-19 vaccination has been studied extensively at the polyclonal level using immune sera, little has been reported on the antibody response at the monoclonal level. Here we isolate a panel of 44 anti-SARS-CoV-2 monoclonal antibodies (mAbs) from an individual who received two doses of the ChAdOx1 nCoV-19 (AZD1222) vaccine at a 12-week interval. We show that despite a relatively low serum neutralization titre, mAbs with potent neutralizing activity against the current SARS-CoV-2 variants of concern (B.1.1.7, P.1, B.1.351 and B.1.617.2) were obtained. The vaccine elicited neutralizing mAbs form 8 distinct competition groups and bind epitopes overlapping with neutralizing mAbs elicited following SARS-CoV-2 infection. AZD1222 elicited mAbs are more mutated than mAbs isolated from convalescent donors 1-2 months post infection. Spike reactive IgG+ B cells were still detectable 9-months post boost. These findings give molecular insights into AZD1222 elicited antibody response.

immunology↗

bioRxiv2021

Drug repurposing based on a Quantum-Inspired method versus classical fingerprinting uncovers potential antivirals against SARS-CoV-2 including vitamin B12

The COVID-19 pandemic has accelerated the need to identify new therapeutics at pace, including through drug repurposing. We employed a Quadratic Unbounded Binary Optimization (QUBO) model, to search for compounds similar to Remdesivir (RDV), the only antiviral against SARS-CoV-2 currently approved for human use, using a quantum-inspired device. We modelled RDV and compounds present in the DrugBank database as graphs, established the optimal parameters in our algorithm and resolved the Maximum Weighted Independent Set problem within the conflict graph generated. We also employed a traditional Tanimoto fingerprint model. The two methods yielded different lists of compounds, with some overlap. While GS-6620 was the top compound predicted by both models, the QUBO model predicted BMS-986094 as second best. The Tanimoto model predicted different forms of cobalamin, also known as vitamin B12. We then determined the half maximal inhibitory concentration (IC50) values in cell culture models of SARS-CoV-2 infection and assessed cytotoxicity. Lastly, we demonstrated efficacy against several variants including SARS-CoV-2 Strain England 2 (England 02/2020/407073), B.1.1.7 (Alpha), B.1.351 (Beta) and B.1.617.2 (Delta). Our data reveal that BMS-986094 and different forms of vitamin B12 are effective at inhibiting replication of all these variants of SARS-CoV-2. While BMS-986094 can cause secondary effects in humans as established by phase II trials, these findings suggest that vitamin B12 deserves consideration as a SARS-CoV-2 antiviral, particularly given its extended use and lack of toxicity in humans, and its availability and affordability. Our screening method can be employed in future searches for novel pharmacologic inhibitors, thus providing an approach for accelerating drug deployment.

microbiology↗

bioRxiv2021

Impact of the B.1.1.7 variant on neutralizing monoclonal antibodies recognizing diverse epitopes on SARS-CoV-2 Spike

The interaction of the SARS-CoV-2 Spike receptor binding domain (RBD) with the ACE2 receptor on host cells is essential for viral entry. RBD is the dominant target for neutralizing antibodies and several neutralizing epitopes on RBD have been molecularly characterized. Analysis of circulating SARS-CoV-2 variants has revealed mutations arising in the RBD, the N-terminal domain (NTD) and S2 subunits of Spike. To fully understand how these mutations affect the antigenicity of Spike, we have isolated and characterized neutralizing antibodies targeting epitopes beyond the already identified RBD epitopes. Using recombinant Spike as a sorting bait, we isolated >100 Spike-reactive monoclonal antibodies from SARS-CoV-2 infected individuals. ~45% showed neutralizing activity of which ~20% were NTD-specific. None of the S2-specific antibodies showed neutralizing activity. Competition ELISA revealed that NTD-specific mAbs formed two distinct groups: the first group was highly potent against infectious virus, whereas the second was less potent and displayed glycan-dependant neutralization activity. Importantly, mutations present in B.1.1.7 Spike frequently conferred resistance to neutralization by the NTD-specific neutralizing antibodies. This work demonstrates that neutralizing antibodies targeting subdominant epitopes need to be considered when investigating antigenic drift in emerging variants.

immunology↗