Antigenic target, antibody subclass, function and glycans and SARS-CoV-2 coinfection impact the placental transfer of herpes simplex virus antibodies

Primary versus recurrent herpes simplex virus 1 or 2 (HSV-1 or HSV- 2) infection during pregnancy carries a higher risk of neonatal herpes suggesting that placental transfer of antibodies protects against transmission and infection. Murine and clinical studies demonstrate that antibody-dependent cellular cytotoxicity (ADCC) provides greater protection than neutralizing antibodies (nAbs) against disseminated neonatal disease. To quantify the relative transfer of HSV-specific Abs with different functions and targets and whether SARS-CoV-2 coinfection modified transfer, we conducted a prospective cohort study of mother-infant dyads prior to and during COVID-19. Total and HSV lysate, glycoprotein D (gD) and glycoprotein B (gB)- specific IgG, IgG1 and IgG3, nAbs, and ADCC were quantified in paired 3rd trimester maternal and cord blood. Transfer ratios (TR) were defined as cord: maternal Ab levels. IgG1 and IgG3 subclass and gD or gB-specific Abs were isolated by column purification and glycan profiles were assessed using mass spectrometry. The pre-COVID study population included 21 term and 15 preterm dyads who were HSV-1 (± HSV-2) seropositive (+) enrolled between 2018-2019 and the peri- COVID cohort included 25 HSV-1 (±HSV-2)+term dyadswhosemothers were also SARS-CoV-2 PCR and COVID Ab+ at delivery;14 were asymptomatic and 11 had mild-moderate COVID disease. None of the mothers had active genital HSV lesions during delivery. HSV-specific IgG, IgG1, and IgG3 TR were higher in term compared to preterm pre-COVID dyads (all p< 0.05). Similarly, the neutralizing Ab TR was 2.4[1.5, 4.0] in term vs 0.8[0.6, 1] in preterm (median [95%CI], p< 0.0001) but the ADCC TR was < 1.0 for both groups. To determine if the low ADCC TR reflected antigenic target, subclass, and/or glycans, we enriched for anti-gD and anti-gB specific and IgG1 and IgG3 Abs. These envelope glycoproteins are primary targets of neutralizing and ADCC responses, respectively. The anti-gD Abs were exclusively IgG1 and had only neutralizing activity. In contrast, anti-gB Abs were both IgG1 and IgG3;the IgG1 gB Abs had both neutralizing and ADCC activity whereas the IgG3 were only neutralizing. The anti-gD Abs were enriched for glycans associated with an affinity for FcRn, whereas anti-gB Abs expressed glycans associated with both FcRn and FcγRIIIa (receptor-associated with ADCC activity) binding. There was no significant difference in HSV-specific IgG TR in pre-COVID vs COVID dyads (0.42) but the nAb TR was lower (p = 0.018) and ADCC TR higher (p<0.001) in COVID compared to pre-COVID patients. Studies are in progress to assess whether this reflects increased placental colocalization of FcRn and FcgRIIIA, which would favor the transfer of ADCCAbs or modified Fc glycans. ADCC Abs transfer relatively inefficiently compared to nAbs, particularly in preterm infants and this may contribute to an increased risk of HSV disease. ADCC Ab transfer increased with SARS-CoV-2 coinfection, which may reflect differences in glycans and/or alterations in the placental architecture. Defining the determinants of ADCC transfer has implications for future vaccine and monoclonal Ab strategies to prevent/treat neonatal herpes. We speculate that increasing the transfer of ADCC may be a key element in providing immune protection.

Antibody function, antigenic target and glycans determine the transfer of herpes simplex virus (HSV) antibodies (Abs) from mothers to newborns and transfer is altered by SARS-CoV-2

OBJECTIVES/GOALS: Murine and clinical data suggest that antibody-dependent cellular cytotoxicity (ADCC) is associated with greater protection against disseminated neonatal HSV disease. To quantify the relative transfer of Abs with different functions and targets, we conducted a prospective study of mother-infant term and preterm dyads pre and during COVID-19 METHODS/STUDY POPULATION: Total and HSV lysate, glycoprotein D (gD) and glycoprotein B (gB)-specific IgG, IgG1 and IgG3 as well as HSV neutralizing Abs (nAbs) and ADCC were quantified in paired 3rd-trimester pregnant women and their newborns (cord) blood. Transfer ratios (TR) were defined as cord:maternal Ab levels. IgG1 and IgG3 subclass and gD or gB-specific Abs were isolated by column purification and glycan profiles were assessed by mass spectrometry. The study population included 21 term and 15 preterm dyads who were HSV-1 (+/- HSV-2) seropositive enrolled between 2018-2019 (pre-COVID) and 25 additional HSV-1 (+/- HSV-2) seropositive term dyads whose mothers were SARS-CoV-2 PCR and COVID Ab+ at delivery;14 were asymptomatic and 11 had mild-moderate COVID disease. None of the mothers had active genital HSV lesions during delivery RESULTS/ANTICIPATED RESULTS: Anti-HSV IgG, IgG1 and IgG3 TR were higher in term vs. preterm dyads (p<0.05). The nAb TR was 2.4 in term vs. 0.8 in preterm (p<0.001) but the ADCC TR was < 1.0 for both. To determine if the latter reflected antigenic target, subclass or glycans, we enriched for gD and gB specific and IgG1 and IgG3 Abs. The gD Abs were IgG1 and had only neutralizing activity. In contrast, gB Abs were polyfunctional and included IgG1 and IgG3 but only the IgG1 Abs had ADCC activity. The gD Abs were enriched for glycans associated with an affinity for the neonatal Fc receptor (FcRn);gB Abs expressed glycans associated with both FcRn and FcγRIIIa binding. There was no significant difference in total HSV-specific IgG TR in pre-COVID vs post-COVID dyads but the nAb TR was lower (p=0.018) and ADCC TR higher (p<0.001) in the COVID compared to pre-COVID cohort DISCUSSION/SIGNIFICANCE: HSV ADCC Abs, which may provide greater protection than nAbs against neonatal disease, transfer poorly particularly to preterm newborns. However, in the setting of SARS-CoV-2, the TR of HSV ADCC is significantly higher. This may reflect alterations in the placental architecture and/or glycan composition which is currently being investigated.

Antibodies to Crucial Epitopes on HSV-2 Glycoprotein D as a Guide to Dosing an mRNA Genital Herpes Vaccine.

The toxicity of mRNA-lipid nanoparticle (LNP) vaccines depends on the total mRNA-LNP dose. We established that the maximum tolerated dose of our trivalent mRNA-LNP genital herpes vaccine was 10 µg/immunization in mice. We then evaluated one of the mRNAs, gD2 mRNA-LNP, to determine how much of the 10 µg total dose to assign to this immunogen. We immunized mice with 0.3, 1.0, 3.0, or 10 µg of gD2 mRNA-LNP and measured serum IgG ELISA, neutralizing antibodies, and antibodies to six crucial gD2 epitopes involved in virus entry and spread. Antibodies to crucial gD2 epitopes peaked at 1 µg, while ELISA and neutralizing titers continued to increase at higher doses. The epitope results suggested no immunologic benefit above 1 µg of gD2 mRNA-LNP, while ELISA and neutralizing titers indicated higher doses may be useful. We challenged the gD2 mRNA-immunized mice intravaginally with HSV-2. The 1-µg dose provided total protection, confirming the epitope studies, and supported assigning less than one-third of the trivalent vaccine maximum dose of 10 µg to gD2 mRNA-LNP. Epitope mapping as performed in mice can also be accomplished in phase 1 human trials to help select the optimum dose of each immunogen in a multivalent vaccine.

Natto extract, a Japanese fermented soybean food, directly inhibits viral infections including SARS-CoV-2 in vitro.

Natto, a traditional Japanese fermented soybean food, is well known to be nutritious and beneficial for health. In this study, we examined whether natto impairs infection by viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as well as bovine herpesvirus 1 (BHV-1). Interestingly, our results show that both SARS-CoV-2 and BHV-1 treated with a natto extract were fully inhibited infection to the cells. We also found that the glycoprotein D of BHV-1 was shown to be degraded by Western blot analysis and that a recombinant SARS-CoV-2 receptor-binding domain (RBD) was proteolytically degraded when incubated with the natto extract. In addition, RBD protein carrying a point mutation (UK variant N501Y) was also degraded by the natto extract. When the natto extract was heated at 100 °C for 10 min, the ability of both SARS-CoV-2 and BHV-1 to infect to the cells was restored. Consistent with the results of the heat inactivation, a serine protease inhibitor inhibited anti-BHV-1 activity caused by the natto extract. Thus, our findings provide the first evidence that the natto extract contains a protease(s) that inhibits viral infection through the proteolysis of the viral proteins.