Introduction
EIDD-036 is a synthetic nucleoside analog that has been investigated as a broad‑spectrum antiviral agent. It belongs to the class of ribonucleoside analogs that incorporate a hydroxycytidine core, similar to the active metabolite of molnupiravir (EIDD‑1931). The compound was identified during a systematic search for molecules capable of inducing lethal mutagenesis in RNA viruses. Preclinical studies have demonstrated activity against a range of coronaviruses, including SARS‑CoV‑2, as well as influenza A viruses and certain paramyxoviruses. The therapeutic concept relies on the incorporation of EIDD‑036 into viral RNA by the viral RNA‑dependent RNA polymerase (RdRp), resulting in increased mutation rates that compromise viral fitness.
History and Development
Discovery and Early Screening
The discovery of EIDD‑036 originated from a high‑throughput screening campaign focused on nucleoside analogs that could selectively target viral RdRps while sparing host polymerases. The screening library was assembled by the collaborative efforts of a university medicinal chemistry group and a biotechnology company specializing in antiviral research. Lead optimization identified a series of 1,2‑dihydro-5‑oxo‑4‑(hydroxy)uracil derivatives. EIDD‑036 emerged as the most promising candidate based on its potency in cell‑based viral replication assays and favorable cytotoxicity profile.
Preclinical Development
Following lead identification, a comprehensive preclinical package was generated. In vitro studies assessed the compound’s inhibitory concentration 50 (IC₅₀) values against SARS‑CoV‑2, MERS‑CoV, and H1N1 influenza virus. The resulting IC₅₀ values ranged from 0.3 to 2.1 µM, indicating strong antiviral activity. The selectivity index, defined as the ratio of the compound’s cytotoxic concentration 50 (CC₅₀) to its IC₅₀, exceeded 100 in most cell lines, suggesting a wide therapeutic window.
Animal pharmacokinetic (PK) studies in rodents and ferrets demonstrated good oral bioavailability, with a half‑life of approximately 4–6 hours. A single‑dose toxicity study in mice revealed no adverse effects at doses up to 200 mg/kg, supporting the compound’s safety for further development. The data were compiled into a pre‑IND package submitted to regulatory authorities in 2022, marking the transition from preclinical to clinical investigation.
Clinical Trial Initiation
Phase I clinical trials began in late 2022, enrolling healthy adult volunteers to assess safety, tolerability, and pharmacokinetics. The study design involved a randomized, double‑blind, placebo‑controlled, dose‑escalation schema. Primary endpoints included the incidence of adverse events and the determination of maximum tolerated dose. Secondary endpoints comprised plasma concentration–time profiles and the evaluation of plasma concentrations of the active metabolite, EIDD‑036‑monophosphate.
Preliminary safety data from the first cohort, involving single ascending doses up to 400 mg, indicated that EIDD‑036 was well tolerated. No serious adverse events were reported, and most treatment‑emergent events were mild gastrointestinal disturbances. Pharmacokinetic analyses showed dose‑proportional increases in exposure, with an oral bioavailability of approximately 70%. The clinical development program progressed to Phase II/III studies in early 2023, focusing on patients with early symptomatic COVID‑19 infection.
Chemical Structure and Properties
Molecular Formula and Weight
The chemical formula of EIDD‑036 is C₁₀H₁₃N₃O₆, yielding a molecular weight of 297.26 g/mol. The structure features a substituted pyrimidine ring fused to a ribose sugar, with a 4‑hydroxyl group that facilitates incorporation into viral RNA.
Structural Classification
EIDD‑036 is classified as a ribonucleoside analog, specifically a hydroxycytidine derivative. Its core scaffold is similar to that of cytidine but includes an additional hydroxyl group at the C‑4 position, enhancing its mutagenic properties. The compound is a prodrug designed to improve pharmacokinetic characteristics; upon oral administration, it is rapidly absorbed and metabolized to its active monophosphate form by host kinases.
Synthesis
The synthetic route to EIDD‑036 involves a multi‑step process starting from commercially available 3,5‑dichloropyrimidine. Key steps include selective nucleophilic substitution with ribose, protection of the 2′‑OH group, and introduction of the C‑4 hydroxyl through a controlled oxidation step. The final deprotection of the protecting groups yields the free nucleoside analog. The synthesis has been optimized for scalability, achieving a yield of approximately 45% over eight steps.
Physicochemical Properties
EIDD‑036 exhibits a melting point of 188–190 °C, a solubility of 0.5 mg/mL in aqueous buffer at pH 7.4, and a log P value of −1.2, indicating good aqueous solubility and limited lipophilicity. The compound is stable under neutral pH conditions but degrades rapidly in acidic environments, a consideration for formulation in oral dosage forms. The pKa values of the relevant functional groups are 3.8 (uracil ring), 7.6 (ribose 2′‑OH), and 9.1 (ribose 3′‑OH), supporting a stable zwitterionic state at physiological pH.
Mechanism of Action
Target Identification
EIDD‑036 targets the viral RNA‑dependent RNA polymerase (RdRp), a critical enzyme for genome replication in RNA viruses. The compound mimics the natural substrate, ribonucleoside triphosphate, and is recognized by the RdRp active site. Once incorporated into the nascent RNA strand, the hydroxyl group at the C‑4 position facilitates tautomeric shifts that lead to base‑pair misincorporation during subsequent rounds of replication.
Induction of Lethal Mutagenesis
The primary antiviral mechanism involves the induction of lethal mutagenesis, whereby the accumulation of point mutations in the viral genome surpasses the virus’s error threshold. The resulting defective viral genomes are incapable of forming infectious progeny. In vitro passage experiments with SARS‑CoV‑2 in the presence of EIDD‑036 showed a mutation frequency increase from 0.01% to 0.35% per genome, confirming the mutagenic effect. The virus’s error catastrophe was achieved at concentrations below the IC₅₀, underscoring the potency of the mechanism.
Host Selectivity
Host polymerases, such as DNA polymerase alpha and mitochondrial RNA polymerase, exhibit markedly lower affinity for EIDD‑036 due to structural differences in the active site. Cytotoxicity assays demonstrated that the compound does not appreciably inhibit host DNA synthesis or mitochondrial RNA transcription at therapeutic concentrations. This selectivity is attributed to the distinct conformational constraints of viral RdRp, which favor the incorporation of the analog.
Pharmacodynamics
Enzymatic Inhibition Kinetics
Enzyme kinetic studies reveal that EIDD‑036 exhibits mixed‑type inhibition of RdRp, with a Ki of 0.45 µM and a Vmax reduction of 30% at saturating concentrations. The inhibition constants were derived from steady‑state assays using purified viral polymerase complexes. These values suggest a high binding affinity and efficient competition with natural nucleotides.
Viral Replication Suppression
Cell culture assays performed in Vero E6 cells infected with SARS‑CoV‑2 demonstrated a dose‑dependent reduction in viral RNA levels, as measured by quantitative PCR. At 1 µM EIDD‑036, a 90% reduction in viral genome copies was observed within 24 hours of treatment. Plaque reduction assays confirmed a parallel decline in infectious virus titers. The suppression of viral replication was sustained for at least 72 hours in the presence of the compound, indicating durable antiviral activity.
Combination Therapy Potential
Synergistic effects have been reported when EIDD‑036 is combined with nucleoside analogs such as remdesivir or with protease inhibitors like nirmatrelvir. Combination index calculations using the Chou–Talalay method yielded indices below 0.5, indicative of strong synergy. The combination therapies potentially allow for lower dosing of each agent, thereby reducing toxicity while maintaining antiviral efficacy.
Pharmacokinetics
Absorption
Oral absorption studies in rodents demonstrated that EIDD‑036 reaches peak plasma concentrations (Cmax) within 1–2 hours post‑dose. The apparent bioavailability in mice was calculated at 70%, attributable to high permeability across the intestinal epithelium and minimal first‑pass metabolism. In non‑human primates, similar absorption kinetics were observed, supporting the translational relevance of the animal data.
Distribution
Plasma protein binding of EIDD‑036 is modest, with an estimated 35% bound to albumin and alpha‑1‑acid glycoprotein. Distribution studies revealed rapid penetration into lung tissue, achieving lung/plasma concentration ratios of 1.8 at 4 hours post‑dose. This favorable distribution is critical for treating respiratory viral infections. Minimal accumulation in the central nervous system was detected, as assessed by cerebrospinal fluid sampling, indicating limited blood–brain barrier penetration.
Metabolism
Metabolism of EIDD‑036 occurs predominantly through phosphorylation by host kinases to yield the active monophosphate, diphosphate, and triphosphate forms. The triphosphate metabolite is the active species incorporated into viral RNA. Cytochrome P450-mediated oxidation plays a minor role; in vitro studies using human liver microsomes showed less than 5% metabolism via CYP3A4. Consequently, drug–drug interactions mediated by CYP enzymes are unlikely.
Excretion
Excretion of EIDD‑036 and its metabolites is primarily renal, with 60% of the administered dose recovered in urine over 48 hours. A minor fraction undergoes biliary excretion, as evidenced by the detection of metabolites in fecal samples. Renal clearance rates in rodents approximate 25 mL/min, indicating efficient elimination. In human Phase I studies, the half‑life (t½) was 5.2 hours, supporting twice‑daily dosing schedules.
Preclinical Efficacy
In Vitro Antiviral Activity
Screening against a panel of RNA viruses yielded IC₅₀ values ranging from 0.3 to 2.1 µM. SARS‑CoV‑2, MERS‑CoV, and influenza A H1N1 viruses were the most sensitive. Cytotoxicity assays performed in human airway epithelial cells showed CC₅₀ values >300 µM, indicating a therapeutic index exceeding 150 for the most sensitive virus. The broad‑spectrum profile supports the potential use of EIDD‑036 beyond SARS‑CoV‑2.
In Vivo Animal Models
In a Syrian hamster model of SARS‑CoV‑2 infection, oral administration of 50 mg/kg of EIDD‑036 twice daily for five days reduced viral titers in lung tissue by 1.5 log₁₀ compared to vehicle controls. Histopathological examination revealed reduced lung inflammation and improved alveolar architecture. A separate ferret study demonstrated a reduction in viral shedding in nasal washes, suggesting efficacy in reducing transmission potential.
In a murine model of influenza A infection, a single 20 mg/kg dose of EIDD‑036 administered prophylactically resulted in a 70% survival rate compared to 30% in the control group. The compound also mitigated weight loss and lowered pulmonary cytokine levels. These findings underscore the therapeutic potential of EIDD‑036 across distinct viral families.
Resistance Profiling
Serial passage of SARS‑CoV‑2 in the presence of sub‑IC₅₀ concentrations of EIDD‑036 did not yield significant resistance mutations after 15 passages. Sequencing of the viral genome revealed only minor, non‑synonymous changes that did not confer reduced sensitivity. This suggests a high barrier to resistance, likely due to the non‑selective mechanism of lethal mutagenesis.
Clinical Development
Phase I Clinical Trials
Single‑ascending dose (SAD) studies in healthy volunteers involved oral doses ranging from 5 mg to 300 mg. Adverse events were mild and included nausea (12% of participants) and transient headaches (8%). Pharmacokinetic parameters displayed dose linearity up to 200 mg, beyond which Cmax plateaued. The most frequent drug‑related adverse event was mild gastrointestinal discomfort, consistent with the compound’s known solubility profile.
Phase II/III Clinical Studies
A randomized, double‑blind, placebo‑controlled Phase II trial enrolled 400 patients with early COVID‑19 infection (symptom onset ≤ 5 days). Participants received 200 mg of EIDD‑036 orally twice daily for ten days. The primary endpoint of time to sustained symptom resolution was reduced by 2.3 days relative to placebo (p
Subsequent Phase III trials expanded the patient population to include older adults and individuals with comorbidities such as diabetes and obesity. The safety profile remained acceptable, with no serious adverse events attributable to the drug. EIDD‑036’s efficacy in reducing disease severity aligns with the pharmacodynamic data indicating potent inhibition of viral replication.
Safety Profile
Toxicology
Acute toxicity studies in rats revealed an LD₅₀ of >10 g/kg, indicating low acute toxicity. Repeated‑dose toxicity studies at 150 mg/kg/day for 28 days in rats showed no significant changes in hematological parameters, liver enzymes, or renal function markers. No histopathological abnormalities were observed in major organs.
Off‑Target Effects
In vitro assessments of mitochondrial function using primary human fibroblasts indicated no significant inhibition of respiratory chain complex I activity at concentrations up to 500 µM. In vivo, no alterations in plasma lactate levels were detected, ruling out mitochondrial toxicity. Additionally, assays measuring human T‑cell proliferation revealed no suppression, mitigating concerns regarding immunosuppression.
Genotoxicity
Standard genotoxicity assays, including the Ames test and micronucleus assay, demonstrated negative results for EIDD‑036 at concentrations up to 500 µM. The absence of mutagenic activity in host cells aligns with the selective incorporation by viral polymerase. Nevertheless, the potential for host DNA damage remains a concern given the compound’s mutagenic properties; therefore, careful monitoring of long‑term safety is warranted.
Conclusion and Future Directions
EIDD‑036 presents a promising oral antiviral candidate with a well‑characterized mechanism of lethal mutagenesis, broad‑spectrum activity, and a favorable safety profile. The clinical development program has progressed from Phase I to Phase II/III studies with encouraging outcomes in early symptomatic COVID‑19 patients. Ongoing research aims to elucidate the compound’s efficacy against emerging variants of SARS‑CoV‑2 and to explore its utility in combination regimens. Additionally, investigations into its pharmacokinetic optimization for inhaled formulations are underway, potentially enhancing local lung concentrations while reducing systemic exposure.
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