Sex-specific associations of ACE2 variants with COVID-19 hospitalization, viral load, and mortality: an Iranian hospital-based case-control study

Angiotensin-converting enzyme 2 ( ACE2 ) is the cellular receptor for SARS-CoV-2, and its genetic polymorphisms may influence COVID-19 susceptibility. This hospital-based case-control study investigated the association between three ACE2 SNPs (rs2285666, rs4646127, rs971249) and normalized viral load, as well as risk of hospitalization for SARS-CoV-2 infection (comparing hospitalized COVID-19 patients versus hospitalized SARS-CoV-2-negative patients with other severe acute conditions), in 441 participants (226 cases, 215 hospital controls). Genotyping was performed by PCR-RFLP, and viral load was quantified by qRT-PCR and normalized to the RNase P gene. Associations were assessed using sex-stratified logistic regression with Benjamini-Hochberg correction for multiple testing. In females, the rs971249 T allele showed a trend toward higher frequency in controls (48.9% vs. 39.5%; raw p = 0.007), but this association did not remain significant after Benjamini-Hochberg correction (q = 0.12). In males, the T alleles of rs971249 and rs4646127 were associated with reduced infection risk and remained significant after correction for multiple testing (raw p = 0.003, q = 0.048 and raw p = 0.002, q = 0.036, respectively). No significant associations were observed for rs2285666. While no ACE2 polymorphism was significantly associated with normalized viral load after correction for multiple testing (all q > 0.05), rs971249 showed a trend toward higher viral load in heterozygous females that did not reach statistical significance (raw p = 0.052; q = 0.18). For mortality analysis, the reported associations for rs971249 and rs4646127 in females (adjusted OR = 2.54 and 2.73, respectively) are based on covariate-adjusted logistic regression (age, comorbidities, sampling time) but did not undergo Benjamini-Hochberg correction due to the exploratory nature of this subgroup analysis; these findings should be considered hypothesis-generating and are clearly labeled as ‘nominally significant’ in the Results section (raw p < 0.05). However, these findings should be interpreted cautiously due to the limited sample size and the exploratory nature of the subgroup analyses and because the rs971249 variant deviated from Hardy-Weinberg equilibrium in female controls ( p = 0.010). These findings suggest that the rs971249 and rs4646127 T alleles may confer sex-specific protection against hospitalized SARS-CoV-2 infection in this cohort of hospitalized patients. However, because controls were hospitalized patients with severe non-COVID-19 conditions, these associations reflect differences between two hospitalized groups rather than population-level susceptibility. Therefore, large-scale studies with population-based controls and diverse ethnic groups are warranted to validate these sex-specific associations and to clarify the role of ACE2 variants in population-level COVID-19 susceptibility.