In the relentless battle against COVID-19, scientists are devising clever strategies to lock the cellular door against the ever-evolving virus.
The COVID-19 pandemic has been a stark reminder of our vulnerability to emerging viruses. As SARS-CoV-2 continues to evolve, producing variants like Delta and Omicron that are more contagious and adept at evading immunity, the quest for effective treatments has intensified. While vaccines train our immune systems to recognize the pathogen, antiviral drugs directly interfere with the virus's ability to infect our cells.
Among the most promising strategies is targeting the very first step of infection: viral entry. Preventing the virus from entering our cells is like stopping an intruder at the gate—it halts the infection before it can even begin. This article explores the scientific frontier of identifying drugs that block SARS-CoV-2 entry, offering a potential powerful tool against current and future variants.
Blocking viral entry prevents infection at the earliest stage, stopping the virus before it can replicate inside cells.
Entry inhibitors work alongside vaccines, providing protection especially for immunocompromised individuals.
To understand how entry inhibitors work, we must first understand the virus's entry mechanism. SARS-CoV-2's key into our cells is its spike (S) protein, which studs the viral surface and gives coronaviruses their crown-like appearance 2 6 .
The entry process is a complex, multi-step dance:
Each of these steps represents a vulnerable point that can be targeted by drugs. Entry inhibitors could work by blocking the ACE2 receptor, disabling the priming proteases, or preventing the membrane fusion process itself.
Viruses like SARS-CoV-2 are not static; they mutate. The Delta and Omicron variants carry a worrying number of mutations in their spike proteins. These mutations can enhance the virus's ability to bind to ACE2, make it more efficient at entering cells, or help it hide from antibodies generated by previous infection or vaccination 1 6 .
Notable for increased transmissibility and potential reduction in neutralization by some antibodies.
Characterized by extensive mutations in spike protein, leading to significant immune evasion.
For instance, research on chronic infections has shown that mutations like P812S in the spike's fusion peptide domain can help the virus evade neutralizing antibodies, a hallmark of Omicron-like adaptation 1 . This constant evolution makes the development of broad-spectrum antivirals—drugs that remain effective across multiple variants—a critical priority for pandemic preparedness.
So, how do scientists actually find these microscopic doorstops? One of the most powerful methods involves high-throughput screening (HTS), a technique that allows researchers to rapidly test thousands of compounds for antiviral activity.
Working with live SARS-CoV-2 requires stringent Biosafety Level 3 (BSL-3) containment, which is not accessible to most research labs. To overcome this, scientists have engineered ingenious surrogate viruses called pseudotyped particles (PPs) 5 .
These PPs are safe to handle under BSL-2 conditions. They are crafted to carry the SARS-CoV-2 spike protein on their surface but lack the actual viral genome. Instead, they package a harmless reporter gene, such as luciferase (the enzyme that makes fireflies glow). When these PPs successfully enter a cell, the cell lights up, providing a clear and measurable signal of infection 5 .
A typical HTS campaign, as described in recent literature, follows a rigorous multi-step process to identify genuine hits 5 :
Thousands of compounds are tested against PPs bearing the SARS-CoV-2 spike protein. Any compound that reduces the luminescence signal is a potential entry inhibitor.
The "hits" from the first round are then tested against PPs with a different viral protein (e.g., VSV-G). This crucial step eliminates compounds that generally disrupt all viral entry, pinpointing only those specific to the SARS-CoV-2 spike.
The most promising compounds are tested against PPs displaying the spike proteins from variants of concern, such as Delta and Omicron, to assess broad-spectrum potential.
Finally, top candidates are moved into a BSL-3 lab to confirm their efficacy against authentic, replicating SARS-CoV-2.
| Screening Stage | Description | Number of Compounds Identified |
|---|---|---|
| Primary Screen | Tested for inhibition of SARS-CoV-2 S pseudovirus | Nearly 200,000 |
| Initial Hits | Inhibited SARS-CoV-2 S entry | Not Specified |
| Counter-Screen | Specific to SARS-CoV-2 S (not VSV-G) | 65 |
| Broad-Screen | Also inhibited MERS-CoV S pseudovirus | 5 (including known inhibitor Nafamostat) |
One rigorous screening study, published in iScience in 2024, exemplifies this pipeline. Researchers screened nearly 200,000 small molecules to find those that block entry after the virus binds to the receptor, targeting the fusion step itself .
After the multi-stage filtering process, they identified a previously unreported small molecule and its derivative as potent inhibitors. These compounds were effective not only against the original SARS-CoV-2 strain but also against pseudoviruses bearing the spikes of the Alpha, Delta, and Omicron variants. This broad-spectrum activity suggests they target a part of the spike protein that is conserved and less prone to mutation, making them exciting candidates for further development .
A novel small molecule identified through HTS shows potent inhibition against multiple SARS-CoV-2 variants by targeting the conserved fusion mechanism of the spike protein.
While most research focuses on the spike protein, scientists are also exploring other, more conserved viral targets to outflank the virus's evolution.
Another promising approach targets the spike protein's transmembrane domain (TMD), the anchor that holds the spike in the viral membrane. A 2025 study identified a compound called 261 that binds to an aromatic-rich region next to the TMD. Using nuclear magnetic resonance (NMR) and molecular dynamics simulations, the researchers mapped this binding site and found it to be highly conserved across many human coronaviruses 3 4 . This makes compound 261 a potential pan-coronavirus inhibitor, which could be effective against future emergent coronaviruses.
In a novel twist, a 2025 paper in Nature described a small molecule (JNJ-9676) that targets the viral membrane (M) protein 7 . The M protein is the most abundant protein in the virus's envelope and acts as a master organizer for viral assembly. JNJ-9676 stabilizes the M protein in an altered shape, preventing the release of infectious viral particles. This molecule showed excellent efficacy in animal models and is active against SARS-CoV, SARS-CoV-2 variants, and even bat and pangolin coronaviruses 7 .
| Treatment Scenario | Dosage | Key Outcome |
|---|---|---|
| Pre-exposure Prophylaxis | 25 mg/kg, twice daily | Reduced viral load in lung by 3.5 log10 (∼99.97%) |
| Post-exposure Treatment | 75 mg/kg, twice daily (started 48h after infection) | Significant reduction in viral load, even after peak infection |
The search for entry inhibitors relies on a specialized set of tools and reagents. The table below details some of the key components used in the experiments discussed.
| Reagent / Tool | Function in Research | Example from Search Results |
|---|---|---|
| Pseudotyped Particles (PPs) | Safe, BSL-2 compatible surrogates for live virus that measure entry efficiency. | MLV-based PPs with spike protein and luciferase reporter 5 . |
| Cell Lines Expressing ACE2 | Model human cells that are susceptible to SARS-CoV-2 infection. | HEK293-ACE2 cells, VeroE6 cells 3 5 . |
| Molecular Dynamics (MD) Simulation | Computational method to visualize protein dynamics and drug binding at an atomic level. | Used to map the binding pocket of compound 261 in the TMD 3 . |
| Cryo-Electron Microscopy (Cryo-EM) | High-resolution imaging technique to determine the 3D structure of proteins and protein-drug complexes. | Used to solve the structure of the M protein bound to JNJ-9676 7 . |
The discovery of entry inhibitors against SARS-CoV-2 represents a vital arm in our arsenal against COVID-19. As the virus continues to evolve, the need for broad-spectrum therapeutics that can withstand these changes becomes ever more pressing. The promising research on fusion inhibitors, TMD-targeting molecules, and innovative targets like the M protein, illuminates a path forward.
Future efforts will focus on combining drugs with different mechanisms of action to create potent therapies less vulnerable to resistance.
Targeting conserved viral regions could yield drugs effective against future coronaviruses beyond SARS-CoV-2.
AI-powered screening and structure-based drug design will accelerate discovery of next-generation inhibitors.
Future efforts will likely focus on combining drugs with different mechanisms of action—for example, an entry inhibitor with a replication inhibitor—to create potent combination therapies that are less vulnerable to drug resistance. The tools and knowledge gained in this fight not only help us against SARS-CoV-2 but also prepare us for the next coronavirus, or the next pandemic pathogen, that may emerge from nature's reservoir.