Immunoliposomes (Ligand-Targeted Liposomes) Production

CD Bioparticles offers custom services to produce special bioparticles by advanced bionanotechnology. Our experienced scientists have created a comprehensive platform to synthesize different bioparticles and ensure product quality. Among these bioparticles, immunoliposome is one of the enormous potential drug targeting carrier systems.

Introduction to Immunoliposomes

Immunoliposomes are liposomes which have antibody attached to the membrane surface. The functionalization of immunoliposomes has emerged as a promising strategy for targeted delivery to and uptake by cells overexpressing the antigens to these antibodies, with a consequent reduction in side effects. Antibodies or other ligands can be attached to liposomes either before or after their preparation. Binding is achieved either covalently or noncovalently. For covalent attachment, it must be taken into consideration that the antibody molecule contains functional chemical groups. Furthermore, the sulfhydryl group plays a key role as a targeting group and has been extensively reported. Nonetheless, this group occurs infrequently in antibody molecules and must be generated by either the reduction of disulfide groups or through appropriate thiolation agents. Thiolated antibodies containing sulfhydryl groups will then react with antibodies containing a chemically reactive molecule, like a lipid-containing maleimide, forming a thioether linkage. The lipid can be synthesized prior to its incorporation into the liposome’s bilayer, and a reaction is carried out between the ligand and the anchor followed by mixing the resulting ligand with the other constituents of the liposomes. Alternatively, the anchor may be previously incorporated in the liposomal membrane and then a coupling reaction is carried out on the surface of vehicles. In addition, antibodies can be non-covalently linked to liposomes, although this strategy is less common. The reaction between biotin and neutravidin or streptavidin is particularly useful. In the process, the ligand is bound to the surface of liposomes through hydrophobic anchor with functional groups.

Immunoliposomes share the key features are as follows:

• Antibodies can be covalently and non-covalently attached to liposomes.
• The covalence of antibodies and liposomes commonly occurs between the sulfhydryl groups of thiolated antibodies and maleimide-containing liposomes.
• Immunoliposomes can target pathologic site by coupling targeting ligands to the surface of liposomes, such as monoclonal antibodies or their fragments.
• Preclinical studies evidenced selective targeting by immunoliposomes to pathologic site.

Figure.1 The structure and application of immunoliposomes (Josimar O. E., et al. Immunoliposomes: A review on functionalization strategies and targets for drug delivery. Colloids and Surfaces B: Biointerfaces. 2017）

Immunoliposomes Applications

With its specific targeting strategy, immunoliposomes have been focused on cancer therapy, inflammatory therapy and cardiovascular diseases therapy, infectious diseases therapy, and autoimmune therapy and neurodegenerative diseases therapy. Although immunoliposomes have not yet received clinical approval, the design of immunoliposomes incorporating a variety of chemotherapeutics that simultaneously exhibit specific target-cell interactions and stimuli-sensitivity has been shown a great potential to improve controlling release of drug formulations and specific targeting at the same time. And its versatile application could be found below:

• Immunoliposomes can carry contrast agent to accumulate in the target area, and help for electron paramagnetic resonance imaging and magnetic resonance imaging.
• Immunoliposomes can deliver a drug to pass the blood-brain barrier (BBB).
• Immunoliposomes is an effective way to deliver shRNA to cell and has the ability to induce gene silencing in mammalian cells.
• Immunoliposomes have been also found useful in the treatment of leukemia.
• Immunoliposomes have been applied to many other cancer types such as gastric, neuroblastoma, colorectal cancer, lung cancer, prostate cancer, and hepatic cancer.
• Immunoliposomes show an advantage in the treatment of HIV.

Figure 2. Immunoliposome-ligand-targeted liposome structure diagram.

Our Immunoliposomes Featured Services

• Liposome formulation design: we customize liposomes design based on our clients’ demand by varying lipid compositions, vesicle sizes, surface charges, etc.
• Liposome encapsulation: we employ customized protocols to encapsulate drug molecules into liposome with high encapsulation efficiency.
• Liposome-drug complex analysis and characterization: we can comprehensive analysis assays for liposomes before and after encapsulation, which includes visual appearance, size distribution, stability, Zeta potential, lamellarity, entrapment efficiency, and release rate.

Immunoliposomes Production Process

The production of immunoliposomes involves a series of optimized formulation and modification steps to create ligand-targeted liposomal systems with enhanced specificity, stability, and drug delivery performance. The process integrates liposome preparation, surface functionalization, purification, and quality evaluation to ensure consistent and reliable targeted delivery.

1. Lipid & Reagent Preparation

High-quality lipids, targeting molecules, therapeutic cargos, and functional excipients are selected based on the desired application. The lipid composition and component ratios are optimized to achieve suitable membrane properties, encapsulation efficiency, and formulation stability.

2. Thin Film Hydration

Lipids are dissolved in organic solvents and converted into a thin lipid film through solvent evaporation. Hydration with an aqueous phase enables lipid self-assembly and forms preliminary liposomal structures for further optimization.

3. Size Reduction & Homogenization

Newly formed liposomes are processed through extrusion, microfluidics or homogenization techniques to control particle size and improve uniformity. Optimized particle characteristics contribute to enhanced stability, circulation behavior, and reproducible delivery performance.

4. Ligand Conjugation (Targeting Ligand)

Targeting ligands, such as antibodies, peptides, or other recognition molecules, are attached to the liposome surface through suitable conjugation strategies. Controlled ligand density and orientation improve selective interaction with target cells or tissues.

5. Purification

Unreacted ligands, free drugs, residual reagents, and impurities are removed using purification methods such as dialysis, ultrafiltration, or tangential flow filtration (TFF). This step improves product purity while maintaining liposome integrity.

6. Sterile Filtration

The purified immunoliposome formulation is handled under controlled aseptic conditions throughout the preparation process, including the use of sterile reagents, equipment, and appropriate aseptic techniques to minimize contamination risks. Additional measures, such as a low-volume rinse with sterile water or other suitable terminal treatments, may be applied when appropriate to the formulation. If sterile filtration is incorporated, the filtration parameters are carefully controlled to minimize potential effects on particle size, drug retention, and biological activity.

7. Quality Control

Comprehensive characterization is performed to evaluate key quality attributes, including particle size, PDI, zeta potential, ligand density, binding affinity, encapsulation efficiency, and stability. These analyses ensure formulation consistency and targeting performance.

8. Aseptic Filling & Packaging

Qualified immunoliposome formulations are filled under aseptic conditions and packaged using suitable containers to protect against degradation, aggregation, and environmental stress.

9. Storage & Delivery

Final products are stored and transported under optimized conditions to maintain liposome structure, ligand functionality, and drug encapsulation performance throughout their intended shelf life.

Quotations and Ordering

References:
1. S. Zalipsky, et al. Polyethylene Glycol-Grafted Immunoliposomes. Journal of Controlled Release. 1996, 39: 153–61.
2. A. S. Manjappa, et al. Antibody Derivatization and Conjugation Strategies: Application in Preparation of Stealth Immunoliposome to Target Chemotherapeutics to Tumor. Journal of Controlled Release. 2011, 150: 2–22.
3. Zhang, Y., et al. In vivo knockdown of gene expression in brain cancer with intravenous RNAi in adult rats. J. Gene Med. 2003, 5: 1039-1045.
4. Mendelsohn, J., Baselga, J., Status of epidermal growth factor receptor antagonists in the biology and treatment of cancer. J. Clin. Oncol. 2003, 21: 2787-2799.
5. Guin, S., Yao, H.P., Wang, M.H., RON Receptor Tyrosine Kinase as a Target for delivery of Chemodrugs by Antibody Directed Pathway for Cancer Cell Cytotoxicity. Mol. Pharm. 2009, 7: 386-397.
6. Torchilin, V. Antibody-modified liposomes for cancer chemotherapy. Expert Opin. Drug Deliv. 2008, 5: 1003-1025.
7. Clayton, R., et al. Sustained and specific in vitro inhibition of HIV-1 replication by a protease inhibitor encapsulated in gp-120 targeted liposomes. Antiviral Res. 2009, 84: 142-149.
8. Josimar O. E., et al. Immunoliposomes: A review on functionalization strategies and targets for drug delivery. Colloids and Surfaces B: Biointerfaces. 2017, http://dx.doi.org/10.1016/j.colsurfb.2017.07.085.