Prerna Arora

Background: The SARS-CoV-2 variants KP.3.1.1 and XEC currently dominate the COVID-19 epidemic. However, their cell tropism, proteolytic processing, and susceptibility to neutralization by monoclonal antibodies remain incompletely characterized. Methods: We employed pseudotyped viruses to assess the entry efficiency of KP.3.1.1 and XEC in various cell lines, their dependence on TMPRSS2 for lung cell entry, and their ability to use ACE2 for infection. Additionally, we evaluated their susceptibility to neutralization by monoclonal antibodies BD55-4637 and BD55-5514. Results: KP.3.1.1 and XEC entered cell lines with similar efficiency as the parental JN.1 lineage and utilized TMPRSS2 for Calu-3 lung cell entry. Unlike JN.1, KP.3.1.1 and XEC failed to efficiently use murine ACE2 for cell entry. Both variants were effectively neutralized by the monoclonal antibodies BD55-4637 and BD55-5514, suggesting therapeutic potential. Conclusions: Our findings demonstrate that JN.1, KP.3.1.1, and XEC, like their predecessor BA.2.86, rely on TMPRSS2 for lung cell entry and remain sensitive to certain neutralizing monoclonal antibodies. However, these variants differ in their ability to utilize ACE2 species orthologs for cell entry.

The naturally occurring mutation E484D in the spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can render viral entry ACE2 independent and imdevimab resistant. Here, we investigated whether the cellular proteins ASGR1, DC-SIGN, and TMEM106B, which interact with the viral S protein, can contribute to these processes. Employing S protein-pseudotyped particles, we found that expression of ASGR1 or DC-SIGN jointly with TMEM106B allowed for robust entry of mutant E484D into otherwise non-susceptible cells, while this effect was not observed upon separate expression of the single proteins and upon infection with SARS-CoV-2 wild type (WT). Furthermore, expression of ASGR1 or DC-SIGN conferred ACE2 independence and imdevimab resistance to entry of mutant E484D but not WT, and entry under those conditions was dependent on endogenous TMEM106B. These results suggest that engagement of certain cellular lectins can direct SARS-CoV-2 mutant E484D to an ACE2-independent, TMEM106B-dependent entry pathway that is not inhibited by imdevimab. IMPORTANCE: The interaction of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein with the ACE2 receptor determines the viral cell tropism and is the key target of the neutralizing antibody response. Here, we show that SARS-CoV-2 with a single, naturally occurring mutation in the spike protein, E484D, can use the cellular lectins ASGR1 and DC-SIGN in conjunction with TMEM106B for ACE2-independent entry and evasion of therapeutic antibodies. These results suggest that engagement of cellular lectins might modulate target cell choice of SARS-CoV-2 and might allow evasion of certain neutralizing antibodies.

New SARS-CoV-2 lineages continue to evolve and may exhibit new characteristics regarding host cell entry efficiency and potential for antibody evasion. Here, employing pseudotyped particles, we compared the host cell entry efficiency, ACE2 receptor usage, and sensitivity to antibody-mediated neutralization of four emerging SARS-CoV-2 lineages, KP.2, KP.2.3, KP.3, and LB.1. The XBB.1.5 and JN.1 lineages served as controls. Our findings reveal that KP.2, KP.2.3, KP.3, and LB.1 lineages enter host cells efficiently and in an ACE2-dependent manner, and that KP.3 is more adept at entering Calu-3 lung cells than JN.1. However, the variants differed in their capacity to employ ACE2 orthologues from animal species for entry, suggesting differences in ACE2 interactions. Moreover, we demonstrate that only two out of seven therapeutic monoclonal antibody (mAbs) in preclinical development retain robust neutralizing activity against the emerging JN.1 sublineages tested, while three mAbs displayed strongly reduced neutralizing activity and two mAbs lacked neutralizing activity against any of the lineages tested. Furthermore, our results show that KP.2, KP.2.3, KP.3, and LB.1 lineages evade neutralization by antibodies induced by infection or vaccination with greater efficiency than JN.1, particularly in individuals without hybrid immunity. This study indicates that KP.2, KP.2.3, KP.3, and LB.1 differ in ACE2 interactions and the efficiency of lung cell entry and suggest that evasion of neutralizing antibodies drove the emergence of these variants.

The SARS-CoV-2 KP.3.1.1 lineage is currently the dominating lineage on several continents. In parallel, the XEC lineage, a recombinant of KS.1.1 and KP.3.3, is on track of becoming the next dominant lineage in Europe and North America. Here we performed a rapid virological characterisation of the XEC lineage and studied the impact of JN.1 mRNA booster vaccination on KP.3.1.1 and XEC neutralisation.

Recently, a recombinant SARS-CoV-2 lineage, XD, emerged that harbors a spike gene that is largely derived from the Omicron variant BA.1 in the genetic background of the Delta variant. This finding raised concerns that the recombinant virus might exhibit altered biological properties as compared to the parental viruses and might pose an elevated threat to human health. Here, using pseudotyped particles, we show that ACE2 binding and cell tropism of XD mimics that of BA.1. Further, XD and BA.1 displayed comparable sensitivity to neutralization by antibodies induced upon vaccination with BNT162b2/Comirnaty (BNT) or BNT vaccination followed by breakthrough infection. Our findings reveal important biological commonalities between XD and Omicron BA.1 host cell entry and its inhibition by antibodies.