2.1 Patients, setting, and human ethical approvals

This retrospective observational study, conducted at the Shanghai Geriatric Medical Center (Meilong Branch of Zhongshan Hospital, Fudan University), examined individuals with COVID-19 who were hospitalized from April 26 to June 30, 2022. The Shanghai Geriatric Medical Center is a specialized medical facility that admits patients with COVID-19 in Shanghai for treatment. The omicron sublineage BA.2.2 was the primary cause of COVID-19 cases reported in Shanghai during this time frame.¹⁹

The patient's infection severity was categorized into 5 types: no symptom, non-severe, general, and severe types, based on the Diagnosis and Treatment Guideline for COVID-19 (Trial Version 9) issued by the China National Health Commission.¹⁸ The high-risk group for severe COVID-19 includes individuals aged 60 years or older who have a medical history of hypertension, cardiovascular diseases, cerebral infarction, chronic liver or kidney diseases, cancers, smoking, or obesity.^20-23 This analysis included individuals who were diagnosed with no symptom, non-severe, general and severe types of COVID-19 and were deemed to be at a high risk of developing severe illnesses. Patients were excluded if they had one reverse-transcription polymerase chain reaction (RT-PCR) test during their hospitalization and the result is negative, a severe COVID-19 diagnosis upon hospitalization, or insufficient information in the electronic medical record, contraindications to nirmatrelvir–ritonavir, severe renal impairment (estimated glomerular filtration rate [eGFR] <30 mL/min per 1.73 m², undergoing dialysis, or had undergone renal transplantation), or severe liver impairment (cirrhosis, hepatocellular carcinoma, or had undergone liver transplantation).

2.2 Baseline characteristics of the patients

Upon hospital admission, patients' baseline characteristics were gathered, including sex, age, clinical classification (i.e., severity of symptoms), underlying health conditions, risk factors of developing severe illness, and immunization record. The number of days from a positive RT-PCR test to initiation of treatment were calculated. Within 2 days of being admitted to the hospital, respiratory samples were obtained from the upper respiratory tract of patients using a nasopharyngeal swab. The samples underwent testing using a real-time fluorescence quantitative RT-PCR method to detect the presence of the SARS-CoV-2 virus. The targets for the RT-PCR assay were open reading frame 1ab (ORF), nucleocapsid protein gene (N gene) and envelope protein gene (E gene). All the targets were recorded with the cycle threshold (Ct) value, which represents the number of cycles required to amplify the viral nucleic acid to a detectable level. A Ct value below 35 for ORF, N gene or E gene was used to define a positive sample of SARS-CoV-2.²⁴

2.3 Intervention and control

All of the patients were provided with standard care, which involved resting in bed, monitoring vital signs, measuring oxygen saturation, conducting routine blood chemistry and urine analysis, examining biochemical indicators such as liver and myocardial enzymes and renal function, assessing coagulation parameters, analyzing arterial blood gas, conducting chest imaging and cytokine detection, and administering oxygen therapy if necessary. Individuals diagnosed with mild-to-moderate COVID-19 and had one of the high risks were administered nirmatrelvir at a dosage of 300 mg alongside ritonavir at a dosage of 100 mg, twice a day, for a duration of 5 days. Alternatively, if patients' eGFR ranged from 30 to 59 mL/min per 1.73 m², the dosage of nirmatrelvir was reduced to 150 mg while ritonavir remained at 100 mg. Patients in the control group only obtained standard care and were not administered nirmatrelvir–ritonavir. Patients in both groups did not receive any other medications specifically for COVID-19, such as remdesivir or monoclonal antibodies.

2.4 Primary outcome

The primary outcome was the duration until conversion, which is defined as the time between the initial positive RT-PCR test and the second negative RT-PCR test outcome. From the second day of hospitalization onwards, all patients underwent daily RT-PCR testing using nasopharyngeal swabs until two consecutive negative results were obtained. According to China's COVID-19 diagnosis and treatment guideline Trial Version 9, criteria for ending isolation and discharge involve obtaining two consecutive tests result with Ct values of ≥35 for ORF, N, and E genes with an interval of more than 24 h.^{20, 25}

2.5 Secondary outcomes and safety assessment

Secondary outcomes included the proportion of individuals with a negative RT-PCR test for respiratory SARS-CoV-2 on the 15th day after the initial positive RT-PCR test, in-hospital morality, incidence of turning critically ill, needs for mechanical ventilation and oxygen therapy. In addition, a safety evaluation of the nirmatrelvir–ritonavir was conducted by examining the medical records of patients to identify potential adverse effects (AEs) such as diarrhea and stomach discomfort. Monitoring for AEs began at the start of treatment and continued until the patient was released from the hospital.

2.6 Statistical analyses

Data were expressed as frequencies (percentages) for categorical variables, mean (SD) for continuous normally distributed variables, and median (interquartile range, IQR) for continuous non-normally distributed variables. Propensity score matching was employed based on age, COVID-19 type upon admission, comorbidities (including diabetes mellitus, malignancy, stroke, hypertension, cardiovascular diseases, pulmonary diseases), vaccination status, and medications administered during hospitalization such as corticosteroids (prednisone, dexamethasone, hydrocortisone, methylprednisolone), antibiotics (β-lactams, moxifloxacin, levofloxacin, metronidazole, vancomycin, azithromycin, linezolid, tigecycline, caspofungin, voriconazole), antiviral drugs (entecavir, valacyclovir), Chinese traditional medications (Lianhua Qingwen Granules, Qingkailing Soft Capsules, Jinhua Qinggan Granules, Shufeng Jiedu Capsules), immunomodulator (recombinant human interferon α2b injection, human interleukin-11 for injection, human Granulocyte colony-stimulating factor injection, intravenous human immunoglobulin, thymalfasin), and Chinese decoctions in a logistic regression model. The propensity-score matching utilized a caliper width of 0.2 without replacement. The standardized mean differences (SMDs) were calculated for each covariate between the groups before and following the propensity-score matching. These differences were considered balanced if the SMD was less than the threshold of 0.1.

For the primary outcome, A Kaplan–Meier curve was drawn after propensity score matching. Hazard ratios (HRs) with 95% (confidence intervals, CIs) between nirmatrelvir–ritonavir users and non-users were estimated using Cox regression models, adjusted for age, sex, vaccination status, COVID-19 type upon admission, value of first positive PCR, comorbidities, medication use, before and after propensity score matching. Results were stratified by the patients' age (≤60 years vs. >60 years), clinical classification, and vaccination history (fully vaccinated vs. partially vaccinated) at baseline. Additionally, the subgroup analysis was stratified based on the time interval between the administration of nirmatrelvir–ritonavir and the first positive RT-PCR test.

All the analyses were performed using software SPSS 26.0 and R 4.2.1. Statistical tests were 2-side and the statistical significance level set at p < .05.

3.1 Demographic characteristics of the patients

During the designated study period, Shanghai Geriatric Medical Centre admitted a total of 5524 patients who were diagnosed with SARS-CoV-2 infections. After removing subjects that did not meet inclusion criteria, the study included 3025 patients. Nirmatrelvir–ritonavir was given to 324 patients (5.8%). The control group consisted of 324 patients from the retrospective cohort, with baseline characteristics used as matching criteria (Figure 1). Following the matching process, patient characteristics were comparable between the nirmatrelvir–ritonavir and control cohorts at baseline, with all SMDs below 0.1, except for RT-PCR Ct value upon admission (SMD = −0.59), administration of corticosteroids (SMD = 0.11) and administration of antiviral medication (SMD = 0.10). The baseline characteristics of patients treated with or without nirmatrelvir–ritonavir are displayed in Table 1. Overall, less than one-third of the group were completely immunized, with 194 individuals (29.9%) having been administered at least one dose of the SARS-CoV-2 vaccine. Most of the patients were over the age of 65, and the median age was 77 years old (IQR, 67-88 years). 314 (48.4%) patients were men. 547 (84.4%) were classified as mild cases. The prevalent underlying conditions consisted of individuals aged ≥65 years (530 [81.8%]), hypertension (311 [48.0%]), and diabetes (144 [22.2%]); more than one risk factor was present in 432 (66.7%) patients. There was no significant difference found between the nirmatrelvir–ritonavir and matched control groups.

3.2 Application of nirmatrelvir–ritonavir for primary and secondary outcomes

The mean time from the first positive RT-PCR result to negative RT-PCR results was similar in both the treatment and control groups (16.2 ± 5.0 vs. 16.1 ± 6.3 days, p = .83, Table 2). Figure 2 displays the Kaplan–Meier curve for the primary outcome, and suggests that nirmatrelvir–ritonavir use was not significantly associated with the shorter time to conversion of RT-PCR results. Cox regression models showed no association of nirmatrelvir–ritonavir and negative conversion of RT-PCR value (Table 3).

Following the initial RT-PCR confirmation, 92% of was administered within 14 days, while 82% and 64% were taken within 10 and 7 days respectively. Hence, the subgroup analysis was further stratified based on the time interval between the administration of nirmatrelvir–ritonavir and the first positive RT-PCR test. Patients who received nirmatrelvir–ritonavir within 10 days of experiencing symptoms demonstrated quicker conversion of RT-PCR results compared to the control group that was matched (Figure 3). Cox regression analysis also demonstrated that patients treated with nirmatrelvir–ritonavir ≤10 days was associated with a shorter negative conversion time of RT-PCR value (Table 3).

Stratified analyses are shown in Figure 4. No significant correlation was observed between the time to conversion of RT-PCR results and patients' age, clinical classification, concurrent medication usage, or prior vaccination in the analysis of individuals treated with nirmatrelvir–ritonavir, except in the case of the usage of antibiotics (Figure 4).

No significant difference was found between the two groups in the percentage of patients who tested negative for respiratory SARS-CoV-2 using RT-PCR 15 days after the onset of symptoms (47.2% vs. 50.0%, p = .53) (Figure 5). Additionally, the two groups did not differ significantly with respect to the progression to more severe conditions, including in-hospital mortality (3.1% vs. 3.1%, p = 1.00), turning to critically ill (1.9% vs. 1.9%, p = 1.00), need for mechanical ventilation (0.3% vs. 1.5%, p = .21), and the need for oxygen therapy (2.5% vs. 3.4%, p = .64).

3.3 Evaluation of the safety of nirmatrelvir–ritonavir in COVID patients at high risk

Among patients treated with nirmatrelvir–ritonavir, 64 patients reported the AEs. Out of these patients, 19 (29.7%) encountered at least one form of AEs, with gastrointestinal disorders (21.8%) being the prevailing side effects. The detailed information is found in Table 4.