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PyCB lipid
Formulation & Performance
Formulation & Performance
Study-specific compositions, preparation conditions and experimental outcomes for PyCB lipid.
Full article and user-supplied SI reviewed. Final three-component ThrCo, screening recipes and four-component controls are distinguished.
1. Formulations and component amounts
Reported composition and calculated mass are separate. Molecular-weight inputs use the current catalog values; DMG-PEG2000 is fixed initially at 2507. Edit inputs only when using a verified alternative material.
Final ThrCo
mol%Main Fig. 2H; SI section 2.1
| Component | Molecular weight | Reported molar entry | Calculated mass |
|---|---|---|---|
| ALC-0315 | 766.27 | 35.6 | Not calculated |
| PyCB lipid | 929.42 | 44.4 | Not calculated |
| DSPC | 790.15 | 20 | Not calculated |
No complete mass recipe is generated for this formulation while composition or identity-dependent inputs remain unresolved.
Four-component study control
mol%Main study-control description
| Component | Molecular weight | Molar parts | Normalized mol% | Amount (µmol) | Weight (mg) |
|---|---|---|---|---|---|
| ALC-0315 | 50.000 | 77.0873 | 59.0697 | ||
| DSPC | 10.000 | 15.4175 | 12.1821 | ||
| Cholesterol | 38.500 | 59.3572 | 22.9505 | ||
| DMG-PEG2000 | 1.500 | 2.3126 | 5.7977 |
Calculated total: 100.0000 mg lipid. RNA, solvent and process loss are excluded.
How component weights are calculated
For total lipid mass M (mg), molar parts rᵢ and molecular weight MWᵢ: component mass = M × rᵢ × MWᵢ / Σ(r × MW). Component amount (µmol) = component mass (mg) × 1000 / MWᵢ. Molar parts are normalized only for the displayed percentage. The calculation does not infer N/P or RNA dose.
2. Preparation and experimental setup
| Parameter | Reported procedure / limitation | Source |
|---|---|---|
| Assembly method | Ethanol dilution is explicitly reported; exact phase ratio, mixing rate and assembly-buffer pH are not specified. | SI section 2.1 |
| Recipe selection | PyCB:ALC-0315 ratio optimized to 5:4, followed by DSPC optimization to 20%; final rounded composition 44.4:35.6:20 in that order. | Main Fig. 1 and optimization text |
| Screen versus final recipe | The initial screening entry uses 45% PyCB; final ThrCo uses 44.4%. | Main Results; not combined into one exact recipe |
| N/P and processing | Exact N/P, phase ratio, mixing flow and assembly-buffer pH are not established in the reviewed article/SI passages. | No generic LNP protocol is substituted |
| Identity-dependent calculations | A final ThrCo mass recipe is withheld pending resolution of the supplied PyCB structural / salt molecular-weight basis. The existing product identity fields are retained. | Identity limitation; comparator mass calculation remains available |
3. Particle characterization
| Measurement / batch | Reported result | Evidence context |
|---|---|---|
| Final ThrCo | Approximately 200 nm | Main Fig. 3 characterization |
| Composition dependence | Initial zero-PyCB screen had encapsulation 25.23%; increasing PyCB to 67.5% reduced translation | Main optimization Results; not final ThrCo encapsulation |
| Particle stability | Particle size followed in PBS at 37 °C for 48 h and in serum-containing conditions | Main characterization; size stability alone does not prove preserved RNA expression |
4. Performance by experiment
| Experiment and dose | Readout and comparator | Result and source |
|---|---|---|
| Lipid-label distribution C57BL/6 mice; IV study dose 5 µg RNA/mouse; DiR lipid tracer | Readout: 6 h Comparator: ALC-0315 four-component study control | Liver tracer signal was 30.9% of comparator; spleen signal was 2.6-fold higher. These tracer results are not the luciferase-expression fold change.Fig. 2J–M; n=3 |
| Functional reporter expression Same study context; luciferase mRNA | Readout: 6 h Comparator: ALC-0315 four-component study control | Spleen luciferase expression was 4.5-fold higher. Cy5 RNA tracking and DiR lipid tracking are separate measurements.Fig. 2J–M; n=3 |
| Alternative spleen formulations Luciferase mRNA 0.25 mg/kg IV | Readout: 6 h Comparator: 18PA and 93-O17O study formulations | Reported spleen expression was 42.1-fold and 9.3-fold higher, respectively. Comparator compositions differ.SI Fig. S6 |
| Cell uptake and expression DC2.4 cells; labeled / reporter RNA 1 µg/mL | Readout: 3 h uptake; later 24 h expression assay Comparator: Study formulations and inhibitor controls | Uptake and translated reporter expression were measured independently; inhibitor responses do not establish an exclusive entry route.Main mechanistic experiments |
| Formulated-RNA storage ThrCo formulations stored at 4 °C for 48 h or −80 °C for 1 week | Readout: Expression after storage Comparator: Fresh particles and four-component study control | Expression remained comparable under the reported cold-storage conditions. A separate 37 °C / 24 h comparison favored ThrCo over the benchmark.SI Fig. S5D–G |
| Repeat administration 5 µg RNA IV once weekly: EGFP cargo for the first two doses, luciferase for the third | Readout: 24 h after the final dose Comparator: Four-component study control | ThrCo retained reporter expression more effectively in the reported repeat-dose comparison. Cargo-switch design and dose schedule are part of this result.Main repeat-dose study |
| Acute liver markers Male and female mice; IV RNA 1 or 5 mg/kg | Readout: 24 h Comparator: Study control LNPs | ALT/AST were assessed; no significant increase at 1 mg/kg in the reported comparison. This does not establish long-term or repeated high-dose safety.Fig. 2N–Q; n=4 |
Mechanistic evidence
The paper combines pH-dependent behavior, membrane-fusion/FRET and cellular trafficking observations. Those measurements support a formulation mechanism; they do not establish identical behavior for all PyCB-containing LNPs.
5. Interpretation and unresolved details
- SI section 2.1 labels ThrCo(22.5%) as ALC-0315:PyCB:DSPC = 22.5:67.5:10 and ThrCo(67.5%) as 67.5:22.5:10. These labels conflict with the written PyCB percentages. The final 35.6:44.4:20 recipe is listed separately and is not reconstructed from those ambiguous screen labels.
- The study’s ALC-0315 four-component control is a research formulation; it is not the finished commercial vaccine composition.
- PyCB-assisted ThrCo is a complete three-component system; the performance cannot be assigned to neat PyCB alone.
- An apparent LNP pKa has not been established by the cited pH-response experiment.
- Final ThrCo mass calculation remains unavailable until the PyCB identity/MW basis is resolved; no placeholder molecular weight is used.
Results apply to the stated LNP composition, cargo, process, dose, route and model. LNP experimental measurements are not batch-release specifications for the isolated lipid.
6. References and data access
Zhao Y et al. Replacing cholesterol and PEGylated lipids with zwitterionic ionizable lipids in LNPs for spleen-specific mRNA translation. Science Advances. 2025;11:eady6460.
Download experiment table (CSV) · Open printable Formulation & Performance guide
Revision FP-20261022 · Source review: 22 October 2026 · DC Chemicals