key: cord-0985671-m7gmyzcs authors: Su, Danmei; Li, Xinglin; Huang, Xueqin; He, Cui; Zeng, Cheng; Wang, Qiang; Qin, Wenchang; Mu, Zhongquan; Ambrosino, Donna; Siber, George; Clemens, Ralf; Liang, Joshua G.; Liang, Peng; Jackson, Nick; Xu, Rong title: Protection from Omicron and other VOCs by Bivalent S-Trimer COVID-19 Vaccine date: 2022-05-03 journal: bioRxiv DOI: 10.1101/2022.05.03.490428 sha: 4b4963783f6df3a4bca84f99c1864b483577a952 doc_id: 985671 cord_uid: m7gmyzcs The Omicron variant of SARS-COV-2 (GISAID GRA clade [B.1.1.529, BA.1 and BA.2]) is now the single dominant Variant of Concern (VOC). The high number of mutations in the Omicron Spike (S) protein promotes humoral immunological escape. Although a third homologous boost with S, derived from the ancestral strain, was able to increase neutralizing antibody titers and breadth including to Omicron, the magnitude of virus neutralization could benefit from further optimization. Moreover, combining SARS-COV-2 strains as additional valences may address the current antigenicity range occupied by VOCs. Using Trimer-Tag™ platform we have previously demonstrated phase 3 efficacy and safety of a prototypic vaccine SCB-2019 in the SPECTRA trial and have submitted applications for licensure. Here, we successfully generated a bivalent vaccine candidate including both Ancestor and Omicron variant S-proteins. Preclinical studies demonstrate this SARS-CoV-2 bivalent S-Trimer subunit vaccine elicits high titers of neutralizing antibodies against all VOCs, with markedly enhanced Omicron specific neutralizing antibody responses. To address Omicron and to drive even broader protection given the potential threat for other 52 VOC to emerge, using the same Trimer-tag platform technology for SCB-2019, we are 53 developing vaccine candidates based on trimerized S-proteins to screen their potential in pre-54 clinical studies against panels of variants. Based on extensive assessments of immunology and 55 antigenicity, for which the antigenic distance of VOC can be mapped by comparing 56 neutralization values for serum / virus pairs; one can hypothesize that breadth can be achieved by 57 selecting a strain in the centroid range of antigenicity (i.e. the Ancestral strain) and a more distal 58 variant (e.g. Omicron) (11, 12) . Here we demonstrate that our bivalent vaccine candidate with Pseudovirus construction and production 124 The variants of concern of SARS-CoV-2 spike protein genes were optimized using mammalian Table 1 ). The lentiviral 129 packaging plasmid psPAX2 and pLVX-AcGFP-N1-Fluc lentiviral reporter plasmid that 130 expresses GFP and luciferase were obtained from HonorGene (HonorGene, China). Pseudovirions were produced by co-transfection HEK 293T cells with psPAX2, pLVX-AcGFP-132 N1-Fluc, and plasmids encoding various S genes by using Lipofectamine 3000 (Invitrogen, 133 L3000-015). The supernatants were harvested at 24 ± 2 h post transfection and centrifuged at 134 1500rpm for 5 min to remove cell debris and then stored at -80°C. Institutional Review Board (IRB), and all patients had provided written informed consent before 158 serum sample were collected. These patients were recently discharged from hospital and the serum was collected at 1-5 weeks after they have been diagnosed as COVID19. Details of 160 sample sourcing and collection are listed in table S1 and certain data previously reported (14). Statistical analysis 163 Data arrangement was performed by Excel and statistical analyses were performed using the 164 Prism 9.2.0 (GraphPad Software). Two-tailed Mann-Whitney tests were used to compare two 165 experiment groups. P values < 0.05 were considered significant. *P < 0.05, **P < 0.01, ***P < 166 0.001. The serum neutralizing responses were monitored post-3 rd dose boost over three months to assess 233 the durability of protection (Fig.4C) . Serum from the control group (no boost) showed robust 234 neutralizing responses maintained against all VOC except Omicron with a low GMT (95%CI) of significantly improved neutralizing responses against all VOC with responses trending higher 242 than the SCB-2022B monovalent boost; with GMT (95% CI) of 799 against Omicron 243 comparable to those elicited by SCB-2022B. These high Omicron specific titers were maintained 244 over the extended observation period (Fig. 4C and Table 2 ). In this study, we corroborated other reports (15-18) VOCs such as the Omicron lineage (4,5). Therefore, we designed a bivalent vaccine, to address advanced models that attempt to define the Group 3: SCB-2022B CpG 1018/Alum No 3rd dose boost Control Group 3: SCB-2022B CpG 1018/Alum Group 4: Bivalent CpG 1018/Alum No 3rd dose boost Control SCB-2019 CpG 1018/Alum SCB-2022B CpG 1018/Alum Group 4: Bivalent CpG 1018/Alum Group 3: SCB-2022B CpG 1018/Alum No 3rd dose boost Control SCB-2019 CpG 1018/Alum SCB-2022B CpG 1018/Alum Group 4: Bivalent CpG 1018/Alum No 3rd dose boost Control SCB-2019 CpG 1018/Alum SCB-2022B CpG 1018/Alum Group 4: Bivalent CpG 1018/Alum