FormalPara Key Summary Points

Why carry out this study?

Studies have suggested that ainuovirine/lamivudine/tenofovir might be an effective and safe primary treatment regimen for human immunodeficiency virus-1 (HIV-1)-infected patients. However, the pharmacokinetics of ainuovirine/lamivudine/tenofovir in elderly and young HIV-1-infected individuals remains unclear.

This study aimed to compare the pharmacokinetics of ainuovirine/lamivudine/tenofovir in HIV-1-infected patients aged ≥ 65 (elderly patients) and ≤ 40 years (young patients).

What was learned from the study?

This study demonstrates that the ainuovirine/lamivudine/tenofovir regimen is an efficacious and well-tolerated treatment regimen for HIV-1-infected patients aged ≥ 65 years and ≤ 40 years.

This study was a single-center, prospective, open-label, parallel-controlled clinical study with a small sample size and short follow-up period. Larger, longer, and more rigorously designed clinical trials and real-world studies are needed to validate the results of this study and to further explore the applicability and benefits of this regimen in different populations.

Introduction

Human immunodeficiency virus-1 (HIV-1) infection is a global health challenge that affects millions of people worldwide. Since the first case of AIDS was reported, more than 79.3 million people worldwide have been infected with HIV-1 and 36.3 million have died from HIV-related illnesses [1, 2]. By 2022, a total of 39.0 million people were living with HIV-1 [3, 4]. Current guidelines recommend that people with HIV-1 infection receive antiretroviral therapy (ART) immediately after they are diagnosed [5, 6]. ART has improved the survival and quality of life of HIV-1-infected patients, but also poses challenges such as drug resistance, toxicity, and poor adherence [7, 8]. Therefore, it is important to develop new and improved ART regimens to overcome these challenges and meet the needs of different populations.

The main drug classes used for ART treatment include nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), protease inhibitors (PIs), integrase inhibitors (INSTIs) and fusion/invasion inhibitors. Typically, ART regimens consist of a combination of two or three classes of drugs to enhance efficacy and reduce the incidence of drug resistance [9]. Ainuovirine/lamivudine/tenofovir is a novel ART regimen comprising two NRTIs (lamivudine and tenofovir) and an NNRTI (ainuovirine) approved by the Chinese health authorities for the treatment of HIV [10]. A preclinical study demonstrated that ainuovirine exhibited potent antiviral activity against various HIV strains in vitro, as well as synergistic effects with lamivudine and tenofovir [11]. In addition, a phase III clinical trial found that the ainuovirine/tenofovir/lamivudine group was not inferior to the efavirenz/tenofovir/lamivudine group in terms of viral suppression at both 48 and 96 weeks and had a better tolerability and safety profile, particularly in terms of liver function, lipids, neuropsychiatric symptoms, and skin rashes [12]. This trial suggests that ainuovirine/lamivudine/tenofovir is an effective and safe primary treatment regimen for HIV-infected patients.

With an increase in the aging population in China, the incidence of HIV-1 infection in the elderly is increasing year by year [13]. Elderly HIV-1-infected patients may have decreased renal function, increased body fat, decreased lean body mass, altered drug metabolism and clearance, and are at a high risk of cardiovascular disease, osteoporosis, diabetes mellitus, and cognitive impairment [14,15,16], thus affecting the absorption, distribution, metabolism, and excretion processes of drugs in the body. In addition, studies have found that people living with HIV aged > 50 years have more concomitant medications and are at a higher risk of potential drug–drug interactions compared to people living with HIV aged ≤ 50 years [17, 18]. Tyrberg et al. also found higher steady-state levels of plasma Darunavir and Atazanavir in people aged ≥ 65 years living with HIV treated with darunavir compared to controls aged ≤ 49 years [19]. It is not clear whether there are differences in the pharmacokinetics of ainuovirine/lamivudine/tenofovir treatment between elderly and young HIV-1-infected people. Therefore, we compared the pharmacokinetic parameters and safety profile of ainuovirine/lamivudine/tenofovir in HIV-1-infected patients aged 65 and 40 years.

Methods

Study Design

This was a single-center, prospective, open-label, parallel-controlled clinical study to evaluate the pharmacokinetics and safety profile of ainuovirine 150 mg/lamivudine 300 mg/tenofovir 300 mg in HIV-1-infected patients. The participants were divided into two groups based on age (elderly group, ≥ 65 years; young group, 18–40 years). The study was conducted at the Guiyang Public Health Treatment Center (China) from October 2022 to February 2023, in accordance with the Declaration of Helsinki of 1964 and its later amendments. The study protocol, amendments, and written consent forms were reviewed and approved by the Ethics Committee of the Guiyang Public Health Treatment Center prior to study initiation (ID 202152). All the participants provided written informed consent for participation and publication.

Study Population

The inclusion criteria for this study were patients (1) aged 18–40 years or ≥ 65 years, regardless of gender; (2) diagnosed with HIV-1 infection, and received initial treatment with ainuovirine/lamivudine/tenofovir for at least 7 days; (3) able to understand and comply with the study protocol requirements, and able to voluntarily sign a written informed consent form.

Patients were excluded if they met any of the following conditions: (1) frailty or other conditions that may jeopardize the safety of the subjects; (2) patients who received drugs and/or foods that may potentially have affected CYP2C19 activity (mango, grapefruit, pomelo, etc.) in the past 7 days, or missed at least one dose of the study drug in the past 7 days; (3) history of drug abuse, recent alcohol or drug dependence, or participated in other drug or therapeutic device clinical trials within the 30 days prior to enrollment in this study; (4) used systemic immunosuppressive therapy or immunomodulators within the 30 days prior to treatment in this study, or could not avoid using them during the clinical trial; (5) cirrhosis and severe liver dysfunction, or creatinine ≥ upper limit of normal range and a glomerular filtration rate calculated by Chronic Kidney Disease-Epidemiology Collaboration creatinine formula ≤ 60 (ml/min/1.73 m2); (6) any grade 3 or 4 manifestations according to the Division of AIDS grading table; (7) subjects who had active tuberculosis and were undergoing treatment at screening, or who had taken or were taking antifungal, corticosteroid, or anti-tuberculosis drugs within the 14 days prior to screening; (8) history of allergy or hypersensitivity to any component or excipient of the study drug; (9) pregnant or lactating women, women of childbearing age who did not take effective contraceptive measures, men with active heterosexual behavior who did not undergo vasectomy and did not take birth control measures, or men and women who were unwilling to continue taking contraceptive measures during the trial period and up to at least 30 days after its end; (10) undergoing any surgery or operation that would change the pharmacokinetics of the experimental drug.

Pharmacokinetic Sampling

Intravenous blood samples (1 mL) were obtained 30 min before the administration of the study drug and at various time points (0.5, 1, 1.5, 2, 3, 4, 8, 12, 16, and 24 h) after dosing to measure the pharmacokinetics (PK) parameters of ainuovirine, lamivudine, and tenofovir. Patients fasted for 2 h after administration. Lunch was taken after 4 h and dinner was taken after about 10 h.

Safety Assessments

Safety was assessed by monitoring all adverse events (AEs), physical examinations, and clinical laboratory tests (routine blood, blood biochemistry, and urine tests) during the study period and 30 days after the PK test. In addition, a medical evaluation was performed for any AEs that occurred.

Sample Size

The study sample size was calculated on the basis of the expected result that the maximum concentration (Cmax,ss) in elderly subjects was 30% higher than that in young subjects (526 ± 143 ng/ml) under steady-state conditions. The study set a two-sided α = 0.05 and β = 0.2 for sample size calculation. Therefore, this study requires a sample size of 14–15 for both elderly and young subject groups. Each group comprised 15 participants.

Bioanalytical Methods

The concentrations of ainuovirine, lamivudine, and tenofovir in human (sodium heparin) plasma samples were determined using high-performance liquid chromatography–tandem mass spectrometry by Yu Jing Technology (Shanghai) Co., Ltd. (Shanghai, China). Lamivudine, ainuovirine, and tenofovir had quantification ranges of 20.0–5000 ng/mL, 4.0–1000 ng/mL, and 4.0–1000 ng/mL, respectively.

Isocratic chromatographic separation of ainuovirine, lamivudine, and tenofovir was achieved on a Waters ACQUITY UPLC HSS T3 (2.1 × 100 mm, 1.8 μm) analytical column. The mobile phase consisted of 0.1% formic acid in water and 2% methanol delivered at a flow rate of 0.4 mL/min. The gradient elution program was as follows: 0–0.6 min, 2–20% B; 0.6–2 min, 20–45% B; 2–2.01 min, 45–95% B; 2.01–4 min, 95–95% B; and 4–5.5 min, 95–2% B. The volume of the sample was 10 μL and the autosampler tray was kept at 40 °C .

The following setup was used for analysis: capillary voltage, 1.0 kV; source temperature, 150 °C; the desolvation gas flow rate and temperature were 1000 L/h and 550 °C, respectively. The analytes were detected and quantified in multiple reaction-monitoring (MRM) modes. The m/z values for the precursor-to-product ion conversions of ainuovirine, lamivudine, and tenofovir are shown in Table 1.

Table 1 Ion transitions and other optimized parameters for ainuovirine, lamivudine, and tenofovir
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In addition, the between-run precisions (coefficient of variation [%CV]) of ainuovirine, lamivudine, and tenofovir were 17.31%, 15.28%, and 11.76%, respectively. The between-run precisions (%CV) for ainuovirine, lamivudine, and tenofovir were 11.09%, 11.84%, and 10.42%, respectively.

Pharmacokinetic and Statistical Analysis

Non-compartmental pharmacokinetic analysis was performed using Phoenix WinNonLin (Certara) to calculate the PK parameters for ainuovirine, lamivudine, and tenofovir. The Cmax, time to Cmax (tmax), area under the plasma concentration–time curve (AUC0–24, AUC0–inf), elimination rate constant (Ke), and elimination half-life (T1/2Ke) were obtained directly from the observed concentration–time data.

Data were analyzed using the SPSS 21 software. Categorical variables were described as percentages (%) and compared using the chi-square test. Normally distributed data were described with mean ± standard deviation and compared with the Student’s t test. Non-normally distributed data were described as median (25th percentile, 75th percentile) and compared using nonparametric tests. Differences were considered statistically significant at P < 0.05.

Results

Demographic and Baseline Characteristics

Thirty patients with HIV were enrolled in this study. Of these, 15 were young patients (aged 30 ± 6.93 years) and 15 were elderly patients (aged 68.53 ± 3.80 years). The mean weights and heights of the young patients were 61.47 ± 14.63 kg and 164.87 ± 8.40 cm, while the mean weights and heights of the elderly patients were 58.83 ± 9.32 kg and 154.4 ± 27.14 cm, with no statistical difference between the two groups (P > 0.05). The blood platelet count, estimated glomerular filtration rate, and serum albumin were significantly lower in the elderly patients than those in the young patients (P < 0.05). In the elderly group, two patients were prescribed hypolipidemic drugs and hypoglycemic drugs, and four patients were prescribed hypotensive drugs during the same period. No concomitant drugs were taken in the young patients. All patients received a single oral dose of ainuovirine (150 mg/lamivudine 300 mg/tenofovir 300 mg) and completed all study assessments. Demographic and baseline characteristics are shown in Table 2.

Table 2 Baseline characteristics
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Pharmacokinetics

The mean plasma concentration–time profiles of ainuovirine/lamivudine/tenofovir are shown in Fig. 1. The plasma levels of each component of ainuovirine/lamivudine/tenofovir, a combination medicine used to treat HIV, reached their highest point within 1–4 h after drug administration and then gradually declined during the rest of the observation period (Fig. 1).

Fig. 1
figure 1

Mean (± standard deviation) concentration–time profiles of ainuovirine/lamivudine/tenofovir. a Ainuovirine, b lamivudine, c tenofovir

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Ainuovirine

The Cmax (698.18 ± 270.16 vs. 586 ± 235.04 ng/ml) and Tmax (3.0 (1.0, 4.0) vs. 2.0 (1.5, 4) h) of the elderly group were similar to those of the young group (P > 0.05, Table 3). The AUC0–24 and AUC0–inf of ainuovirine were significantly increased in elderly patients (9098.71 ± 3572.91 and 22,933.07 ± 19,026.61 ng h/ml, respectively) compared to young patients (6235.32 ± 2065.53 and 8989.11 ± 3439.60 ng h/ml, respectively) (P < 0.05). The Ke value of the elderly group was smaller than that of the young group (0.031 ± 0.017 vs. 0.053 ± 0.018, respectively, P = 0.002), while the T1/2Ke of the elderly group was twice as high as that of the young group (29.98 ± 17.82 vs. 14.64 ± 5.16, P < 0.05).

Table 3 Comparison of pharmacokinetic parameters after oral administration of ainuovirine/lamivudine/tenofovir in the elderly and young groups
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Lamivudine

Elderly patients showed a significant increase in the Cmax, AUC0–24, and AUC0–inf of lamivudine (3011.88 ± 1100.60 ng/ml, 21,238.89 ± 7299.89 ng h/ml, and 22,630.62 ± 7312.37 ng h/ml, respectively) in comparison to young patients (2009.86 ± 693.90 ng/ml, 11,652.82 ± 2745.43 ng h/ml, and 12,222.69 ± 2876.82 ng h/ml, respectively) (all P < 0.05). However, the elderly group were similar to the young group in terms of Tmax (2.30 ± 1.21 vs. 2.60 ± 1.04 h), Ke (0.118 ± 0.027 vs. 0.118 ± 0.021), and T1/2Ke (6.38 ± 2.47 vs. 6.05 ± 1.22), with no statistically significant differences (all P > 0.05).

Tenofovir

Tenofovir was rapidly absorbed (median Tmax = 1.0 h for the elderly group and 1.5 h for the young group; P > 0.05). Elderly patients had a higher Cmax (337.87 ± 81.61 vs. 262.61 ± 93.16 ng/ml, P = 0.008) and AUC0–inf (4713.69 ± 1659.63 vs. 3347.68 ± 924.13 ng h/ml, P = 0.006) than young patients, but no differences were observed in AUC0–24 (2952.67 ± 1113.01 vs. 2465.11 ± 703.87 ng h/ml, P = 0.163). In addition, there were no statistically significant differences in Ke (0.044 ± 0.015 vs. 0.056 ± 0.012, P = 0.15) or T1/2Ke (12.03 (5.65, 17.05) vs. 12.54 (10.9, 13.32), P = 0.775) between the elderly group and the young group.

Safety

No abnormalities were observed in the physical examinations and laboratory tests for either the young or elderly groups 30 days after oral administration of ainuovirine/lamivudine/tenofovir (Table 4).

Table 4 Comparison of physical examinations after 30 days of oral administration of ainuovirine/lamivudine/tenofovir combination tablets in young and elderly patients with human immunodeficiency virus-1 infection
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Ainuovirine/lamivudine/tenofovir was well tolerated and no unexpected safety signals were observed. After treatment, the overall incidence of AEs in the young group was 20%, while that in the elderly group was 33.33%; the difference between the two groups was not statistically significant (P > 0.05, Table 5). There were two drug-related AEs in the elderly group (sleep disorders and cough) and two in the young group (sleep disorders and chest stuffiness). There were no statistically significant differences in drug-related AEs between the two groups (P > 0.05). Sleep disorders were more severe in the elderly group (grade 3–4) than in the young group (grade 1–2). All drug-related AEs were improved after treatment in elderly group and improved without treatment in young group.

Table 5 Adverse event summary
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Discussion

Human immunodeficiency virus (HIV) is a major global health concern. The goal of ART is to reduce the viral load, improve morbidity and mortality, and prevent transmission. Ainuovirine/lamivudine/tenofovir is a novel ART regimen containing ainuovirine (150 mg), lamivudine (300 mg), and tenofovir (300 mg) administered orally once daily as a simple, effective, and safe treatment option for HIV-1-infected patients. Ainuovirine is a novel NNRTI that blocks viral replication by binding to HIV reverse transcriptase and inhibiting its catalytic reaction [20]. Lamivudine and tenofovir are NRTIs that bind competitively to natural nucleosides and inhibit HIV reverse transcriptase activity, thereby impeding provirus synthesis [21, 22]. Elderly HIV-1-infected patients are a special population that may have different pharmacokinetic and pharmacodynamic responses to antiretroviral drugs owing to changes in physiology and metabolism. Therefore, the assessment of the pharmacokinetic profile of ainuovirine/lamivudine/tenofovir in elderly patients is necessary to determine whether dose adjustment or monitoring is required. There is a paucity of data regarding the pharmacokinetics of ainuovirine/lamivudine/tenofovir in elderly patients. This study is the first to compare the pharmacokinetics and safety of the ainuovirine/lamivudine/tenofovir regimen in HIV-1-infected patients aged 65 and 40 years. The results showed no significant differences in the Tmax between the two groups for any of these drugs, suggesting that these three drugs were absorbed at a similar rate and were not influenced by age. This is consistent with the results of a previous study [23] comparing the pharmacokinetics of the tenofovir acetate–emtricitabine combination regimen in women, which found that the rate of absorption of tenofovir in the plasma was independent of age. Meanwhile, elderly patients had a smaller Ke and higher T1/2Ke, AUC0–24, and AUC0–inf for ainuovirine than young patients. This indicates that ainuovirine has a reduced clearance rate in elderly patients, leading to higher drug exposure. Ainuovirine is mainly excreted by the bile/feces (70.91%), and the renal pathway accounts for 23.32% [15]. It has been speculated that elderly people may have a reduced digestive function or renal function, leading to a lower clearance rate of ainuovirine. In this study, we found that although there was no significant difference, the clearance rate tenofovir was lower in the elderly group compared with that in the young group, which presumably as a result of the decline in eGFR. Interestingly, the clearance rate of lamivudine did not decrease significantly in older patients, possibly because lamivudine was primarily cleared by the renal tubule organic cationic transport system, which was not measured in older patients in our study. Generally, exposure to tenofovir and lamivudine was higher in the elderly group. This finding is consistent with those of previous studies that reported higher exposure to these drugs in elderly patients [10, 24, 25]. Therefore, dose adjustment or careful monitoring of drug concentrations may be necessary in elderly patients to ensure the safe and effective use of these drugs.

In terms of safety, ainuovirine/lamivudine/tenofovir was well tolerated in both groups, with no unexpected safety signals. The incidence of AEs was 20% and 33.33% in the young and elderly groups, respectively; however, there was no statistical difference between the two groups. Similarly, a previous phase III clinical trial conducted in China, which compared the efficacy and safety of the ainuovirine/lamivudine/tenofovir regimen with the efavirenz/tenofovir/lamivudine regimen in treatment-naïve HIV-1-positive adults, found that the former was not inferior to the latter in inhibiting viral replication and had better tolerability and safety, particularly in terms of liver function, lipids, neuropsychiatric symptoms, and skin rashes [12]. Moreover, we found a similar incidence of drug-related AEs between the two groups; however, sleep disorders were more severe in the elderly patients than in the young patients. It should be noted that this elderly patient also had concomitant hypertension, diabetes, and coronary heart disease and had been diagnosed with insomnia before the start of the study. These factors may lead to the aggravation of sleep disorders in patients. These conditions also emphasize the issue of comorbidities and combined medications in elderly patients. Therefore, potential interactions and effects of ainuovirine/lamivudine/tenofovir with other drugs should be assessed when prescribing this regimen to elderly patients.

There are some limitations in this study. First, this study was a single-center clinical study with a small sample size and short follow-up period. Second, in order to better investigate the pharmacokinetics, we performed some pre-screening to exclude patients with disease that might affect drug metabolism. Therefore, further real-world data with larger sample size and longer follow-up time is needed to validate the results of this study.

Conclusion

This study suggests that the ainuovirine/lamivudine/tenofovir regimen might be an effective and safe HIV-1 treatment regimen for HIV-1-infected patients aged ≥ 65 years and ≤ 40 years. This study also observed that the exposure level to each drug in the treatment plan was higher in middle-aged and elderly patients; however, the incidence of AEs did not increase, indicating the safety of the treatment plan containing ainuovirine in elderly patients. Moreover, during the clinical application process, attention should be paid to whether elderly patients have increased clinical complications due to comorbidities and combined medications. In addition, this study was a single-center, prospective, open-label, parallel-controlled clinical study with a small sample size and short follow-up period. Larger, longer, and more rigorously designed clinical trials and real-world studies are needed to validate the results of this study and to further explore the applicability and benefits of this regimen in different populations.