Newly emerging SARS-CoV-2 variants may escape monoclonal antibodies (mAbs) and antiviral drugs. By using live virus assays, we assessed the ex vivo inhibition of the B.1 wild-type (WT), delta and omicron BA.1 and BA.2 lineages by post-infusion sera from 40 individuals treated with bamlanivimab/etesevimab (BAM/ETE), casirivimab/imdevimab (CAS/IMD), and sotrovimab (SOT) as well as the activity of remdesivir, nirmatrelvir and molnupiravir. mAbs and drug activity were defined as the serum dilution (ID₅₀) and drug concentration (IC₅₀), respectively, showing 50% protection of virus-induced cytopathic effect. All pre-infusion sera were negative for SARS-CoV-2 neutralizing activity. BAM/ETE, CAS/IMD, and SOT showed activity against the WT (ID₅₀ 6295 (4355–8075) for BAM/ETE; 18,214 (16,248–21,365) for CAS/IMD; and 456 (265–592) for SOT) and the delta (14,780 (ID₅₀ 10,905–21,020) for BAM/ETE; 63,937 (47,211–79,971) for CAS/IMD; and 1103 (843–1334) for SOT). Notably, only SOT was active against BA.1 (ID₅₀ 200 (37–233)), whereas BA.2 was neutralized by CAS/IMD (ID₅₀ 174 (134–209) ID₅₀) and SOT (ID₅₀ 20 (9–31) ID₅₀), but not by BAM/ETE. No significant inter-variant IC₅₀ differences were observed for molnupiravir (1.5 ± 0.1/1.5 ± 0.7/1.0 ± 0.5/0.8 ± 0.01 μM for WT/delta/BA.1/BA.2, respectively), nirmatrelvir (0.05 ± 0.02/0.06 ± 0.01/0.04 ± 0.02/0.04 ± 0.01 μM) or remdesivir (0.08 ± 0.04/0.11 ± 0.08/0.05 ± 0.04/0.08 ± 0.01 μM). Continued evolution of SARS-CoV-2 requires updating the mAbs arsenal, although antivirals have so far remained unaffected.

Despite the major target of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, being the respiratory system, clinical evidence suggests that the male reproductive system may represent another viral target organ. Revealing the effect of SARS-CoV-2 infection on testis and sperm is a priority for reproductive biology, as well as for reproductive medicine. Here, we confirmed that the SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2) is highly expressed on human testis and ejaculated sperm; moreover, we provide evidence for the expression of the co-receptors transmembrane protease/serine (TMPRSS2), Basigin (BSG), and Catepsin L (CTSL). Human sperm were readily infected, both in vivo and in vitro, by SARS-CoV-2, as demonstrated by confocal and electron microscopy. The demonstration that the seminiferous epithelium and sperm support SARS-CoV-2 viral replication suggests the possibility that the spermatogenetic process may be detrimentally affected by the virus, and at the same time, supports the need to implement safety measures and guidelines to ensure specific care in reproductive medicine.

Combination antiviral therapy may be helpful in the treatment of SARS-CoV-2 infection; however, no clinical trial data are available, and combined use of direct-acting antivirals (DAA) and monoclonal antibodies (mAb) has been reported only anecdotally. To assess the cooperative effects of dual drug combinations in vitro, we used a VERO E6 cell-based in vitro system with the ancestral B.1 or the highly divergent BQ.1.1 virus to test pairwise combinations of the licensed DAA, including nirmatrelvir (NRM), remdesivir (RDV) and the active metabolite of molnupiravir (EIDD-1931) as well the combination of RDV with four licensed mAbs (sotrovimab, bebtelovimab, cilgavimab, tixagevimab; tested only with the susceptible B.1 virus). According to SynergyFinder 3.0 summary and weighted scores, all the combinations had an additive effect. Within DAA/DAA combinations, paired scores with the B.1 and BQ.1.1 variants were comparable. In the post hoc analysis weighting synergy by concentrations, several cases of highly synergistic scores were detected at specific drug concentrations, both for DAA/DAA and for RDV/mAb combinations. This was supported by in vitro confirmation experiments showing a more than a linear shift of a drug-effective concentration (IC₅₀) at increasing concentrations of the companion drug, although the effect was prominent with DAA/DAA combinations and minimal or null with RDV/mAb combinations. These results support the cooperative effects of dual drug combinations in vitro, which should be further investigated in animal models before introduction into the clinic.

Recurrence of coronavirus disease 19 (COVID-19) symptoms and SARS-CoV-2 viral load relapse have been reported in people treated with nirmatrelvir/ritonavir (NM/r). However, little is understood about the etiology of this phenomenon. Our aim was to investigate the relation between the host’s immune response and viral rebound. We described three cases of COVID-19 rebound that occurred after treatment with nirmatrelvir/ritonavir (group A). In addition, we compared spike-specific antibody response and plasma cytokine/chemokine patterns of the rebound cases with those of (i) control patients treated with nirmatrelvir/ritonavir who did not show rebound (group B), and (ii) subjects not treated with any anti-SARS-CoV-2 drug (group C). The anti-spike antibodies and plasma cytokines/chemokines were similar in groups A and B. However, we observed a higher anti-BA.2 spike IgG response in patients without antiviral treatment (group C) [geometric mean titer 210,807, 5.1- and 8.2-fold higher compared to group A (p = 0.039) and group B (p = 0.032)]. Moreover, the patients receiving antiviral treatment (groups A-B) showed higher circulating levels of platelet-derived growth factor subunit B (PDGF-BB) and vascular endothelial growth Factors (VEGF) and lower levels of interleukin-9 (IL-9), interleukine-1 receptor antagonist (IL-1 RA), and regulated upon activation normal T cell expressed and presumably secreted chemokine (RANTES) when compared to group C. In conclusion, we observed lower anti-spike IgG levels and different cytokine patterns in nirmatrelvir/ritonavir-treated patients compared to those not treated with anti-SARS-CoV-2 drugs. This suggests that early antiviral treatment, by reducing viral load and antigen presentation, could mitigate the immune response against SARS-CoV-2. The clinical relevance of such observation should be further investigated in larger populations.

Newly emerging SARS-CoV-2 variants may escape monoclonal antibodies (mAbs) and antiviral drugs. By using live virus assays, we assessed the ex vivo inhibition of the B.1 wild type (WT), delta and omicron BA.1 and BA.2 lineages by post-infusion sera from 40 individuals treated with bamlanivimab/etesevimab (BAM/ETE), casirivimab/imdevimab (CAS/IMD) and sotrovimab (SOT) as well as the activity of remdesivir, nirmatrelvir and molnupiravir. mAbs and drug activity were defined as the serum dilution (ID50) and drug concentration (IC50), respectively, showing 50% protection of virus-induced cytopathic effect. All pre-infusion sera were negative for SARS-CoV-2 neutralizing activity. BAM/ETE, CAS/IMD and SOT showed activity against the WT (ID50 6,295 [4,355-8,075] for BAM/ETE; 18,214 [16,248-21,365] for CAS/IMD and 456 [265-592] for SOT) and the delta (14,780 [ID50 10,905-21,020] for BAM/ETE, 63,937 [47,211-79,971] for CAS/IMD and 1,103 [843-1,334] for SOT). Notably, only SOT was active against BA.1 (ID50 200 [37-233]) while BA.2 was neutralized by CAS/IMD (ID50 174 [134-209] ID50) and SOT (ID50 20 [9-31] ID50) but not by BAM/ETE. No significant inter-variant IC50 differences were observed for molnupiravir (1.5±0.1/1.5±0.7/1.0±0.5/0.8±0.01 μM for WT/delta/BA.1/BA.2, respectively); nirmatrelvir (0.05±0.02/0.06±0.01/0.04±0.02/0.04±0.01 μM) and remdesivir (0.08±0.04/0.11±0.08/0.05±0.04/0.08±0.01 μM). Continued evolution of SARS-CoV-2 requires updating the mAbs arsenal, however antivirals have so far remained unaffected.

Live virus neutralization is the gold standard to investigate immunity. This prospective observational study aimed to determine the magnitude of response against the original B.1 lineage and against the BA.5 lineage six months after the third BNT162b2 mRNA vaccine dose in patients with HIV infection on successful antiretroviral treatment and no previous SARS-CoV-2 infection. A total of 100 subjects (M/F 83/17, median age 54 years) were included in the analysis: 95 had plasma HIV RNA <40 copies/mL, the median CD4+ T cell count at the administration of the third dose was 580 cells/mm³, and the median nadir CD4+ T cell count was 258 cells/mm³. Neutralizing antibodies (NtAb) against B.1 were detectable in all the subjects, but those to BA.5 were only detected in 88 (p < 0.001). The median NtAb titer to B.1 was significantly higher than that to BA.5 (393 vs. 60, p < 0.0001), and there was a strong positive correlation between the paired measurements (p < 0.0001). Linear regression on a subset of 87 patients excluding outlier NtAb titers showed that 48% of the changes in NtAb titers to BA.5 are related to the changes in value titers to B.1. SARS-CoV-2 variants evolve rapidly, challenging the efficacy of vaccines, and data on comparative NtAb responses may help in tailoring intervals between vaccine doses and in predicting vaccine efficacy.