Unleashing the Potential: Designing Antibody-Targeted Lipid Nanoparticles for Industrial Applications with CMC Considerations and Clinical Outlook.

Antibody-targeted lipid nanoparticles (Ab-LNPs) are rapidly gaining traction as multifaceted platforms in precision medicine, adept at delivering a diverse array of therapeutic agents, including nucleic acids and small molecules. This review provides an incisive overview of the latest developments in the field of Ab-LNP technology, with a special emphasis on pivotal design aspects such as antibody engineering, bioconjugation strategies, and advanced formulation techniques. Furthermore, it addresses critical chemistry, manufacturing, and controls (CMC) considerations and thoroughly examines the in vivo dynamics of Ab-LNPs, underscoring their promising potential for clinical application. By seamlessly blending scientific advancements with practical industrial perspectives, this review casts a spotlight on the burgeoning role of Ab-LNPs as an innovative and potent tool in the realm of targeted drug delivery.

Drug Stability and Minimized Acid-/Drug-Catalyzed Phospholipid Degradation in Liposomal Irinotecan.

Therapeutics at or close to the nanoscale, such as liposomal irinotecan, offer significant promise for the treatment of solid tumors. Their potential advantage over the unencapsulated or free form of the drug is due in part to their altered biodistribution. For slow and sustained release, significant optimization of formulation is needed to achieve the required level of stability and allow long-term storage of the drug product. Gradient-based liposomal formulation of camptothecins such as irinotecan poses unique challenges owing to the camptothecin- and acid-catalyzed hydrolysis of phospholipid esters in the inner monolayer of the liposomal membrane. We demonstrated that a narrow set of conditions related to the external pH, temperature, intraliposomal concentration, identity of the drug-trapping agent, physical form of the drug inside the liposomes, and final drug load have a marked impact on the stability of the liposome phospholipid membrane. The physical form of the drug inside the liposome was shown to be an insoluble gel with an irinotecan-to-sulfate ratio approximating 1:1, reducing the potential for irinotecan-catalyzed phospholipid hydrolysis in the internal phospholipid monolayer. As a result of this work, a stable and active liposome formulation has been developed that maintains phospholipid chemical stability following long-term storage at 2-8°C.

Liposomal encapsulation of trans-crocetin enhances oxygenation in patients with COVID-19-related ARDS receiving mechanical ventilation.

Current therapeutic treatments improving the impaired transportation of oxygen in acute respiratory distress syndrome (ARDS) have been found to be relevant and beneficial for the therapeutic treatment of COVID-19 patients suffering from severe respiratory complications. Hence, we report the preclinical and the preliminary results of the Phase I/II clinical trial of LEAF-4L6715, a liposomal nanocarrier encapsulating the kosmotropic agent trans-crocetin (TC), which, once injected, enhance the oxygenation of vascular tissue and therefore has the potential to improve the clinical outcomes of ARDS and COVID-19 in severely impacted patients. We demonstrated that the liposomal formulation enabled to increase from 30 min to 48 h the reoxygenation properties of free TCs in vitro in endothelial cells, but also to improve the half-life of TC by 6-fold in healthy mice. Furthermore, we identified 25 mg/kg as the maximum tolerated dose in mice. This determined concentration led to the validation of the therapeutic efficacy of LEAF-4 L6715 in a sepsis mouse model. Finally, we report the preliminary outcomes of an open-label multicenter Phase I/II clinical trial (EudraCT 2020-001393-30; NCT04378920), which was aimed to define the appropriate schedule and dosage of LEAF-4L6715 and to confirm its tolerability profile and preliminary clinical activity in COVID-19 patients treated in intensive care unit.

Targeting EphA2 in Bladder Cancer Using a Novel Antibody-Directed Nanotherapeutic.

Ephrin receptor A2 (EphA2) is a member of the Ephrin/Eph receptor cell-to-cell signaling family of molecules, and it plays a key role in cell proliferation, differentiation, and migration. EphA2 is overexpressed in a broad range of cancers, and its expression is in many cases associated with poor prognosis. We recently developed a novel EphA2-targeting antibody-directed nanotherapeutic encapsulating a labile prodrug of docetaxel (EphA2-ILs-DTXp) for the treatment of EphA2-expressing malignancies. Here, we characterized the expression of EphA2 in bladder cancer using immunohistochemistry in 177 human bladder cancer samples and determined the preclinical efficacy of EphA2-ILs-DTXp in four EphA2-positive patient-derived xenograft (PDX) models of the disease, either as a monotherapy, or in combination with gemcitabine. EphA2 expression was detected in 80-100% of bladder cancer samples and correlated with shorter patient survival. EphA2 was found to be expressed in tumor cells and/or tumor-associated blood vessels in both primary and metastatic lesions with a concordance rate of approximately 90%. The EphA2-targeted antibody-directed nanotherapeutic EphA2-ILs-DTXp controlled tumor growth, mediated greater regression, and was more active than free docetaxel at equitoxic dosing in all four EphA2-positive bladder cancer PDX models. Combination of EphA2-ILs-DTXp and gemcitabine in one PDX model led to improved tumor growth control compared to monotherapies or the combination of free docetaxel and gemcitabine. These data demonstrating the prevalence of EphA2 in bladder cancers and efficacy of EphA2-ILs-DTXp in PDX models support the clinical exploration of EphA2 targeting in bladder cancer.

Synergy between EphA2-ILs-DTXp, a Novel EphA2-Targeted Nanoliposomal Taxane, and PD-1 Inhibitors in Preclinical Tumor Models.

Combinations of chemotherapy with immunotherapy have seen recent clinical success, including two approvals of anti-PD-1/L1 agents in combination with taxane-based chemotherapy in non-small cell lung cancer and triple-negative breast cancer. Here, we present a study on the combination activity and mechanistic rationale of a novel EphA2-targeted liposomal taxane (EphA2-ILs-DTXp) and anti-PD-1. This combination was highly active in mouse syngeneic tumor models, with complete responses observed in 3 of 5 models. In the EMT-6 tumor model, combination of EphA2-ILs-DTXp with anti-PD-1 resulted in a 60% complete response rate, with durable responses that were resistant to rechallenge. These responses were not observed in the absence of CD8+ T cells. Characterization of the immune infiltrates in EMT-6 tumors reveals increased CD8+ T cells, increased CD8+ IFNγ+ CTLs, and an increased CD8/regulatory T-cell (Treg) ratio. These immunomodulatory effects were not observed in mice treated with a combination of docetaxel and anti-PD-1. Pharmacokinetic analysis revealed that the AUC of docetaxel was increased 15 times, from 52.1 to 785 ng/mL/hour, when delivered by EphA2-ILs-DTXp. A dose reduction study of EphA2-ILs-DTXp showed a dose-response relationship for both tumor growth inhibition and the CD8/Treg ratio. Our data indicate that synergism between docetaxel and anti-PD-1 is achievable with nanoliposomal delivery.

Formulation optimization of an ephrin A2 targeted immunoliposome encapsulating reversibly modified taxane prodrugs.

Ephrin A2 targeted immunoliposomes incorporating pH-sensitive taxane prodrugs were developed for sustained delivery of active drug to solid tumors. Here we describe the systematic formulation development and characterization of these immunoliposomes. We synthesized both paclitaxel and docetaxel prodrugs to formulate as ephrin A2-targeted liposomes stabilized in the aqueous core with sucroseoctasulfate (SOS). The optimized lipid formulation was comprised of egg-sphingomyelin, cholesterol, and polyethylene glycol distearoyl glycerol (PEG-DSG). The formulations examined had a high efficiency of prodrug encapsulation (as high as 114 mol% taxane per mole phospholipid) and subsequent stability (>3 years at 2-8 °C). The taxane prodrug was stabilized with extraliposomal citric acid and subsequently loaded into liposomes containing a gradient of SOS, resulting in highly stable SOS-drug complexes being formed inside the liposome. The internal prodrug and SOS concentrations were optimized for their impact on in vivo drug release and drug degradation. Cryo-electron microscope images revealed dense prodrug-SOS complex in the aqueous core of the immunoliposomes. Ephrin A2-targeted taxane liposomes exhibited sub-nanomolar (0.69 nM) apparent equilibrium dissociation constant toward the extracellular domain of the ephrin A2 receptor, long circulation half-life (8-12 h) in mouse plasma, a release rate dependent on intraliposomal drug concentration and stable long-term storage. At an equitoxic dose of 50 mg taxane/kg, ephrin A2-targeted liposomal prodrug showed greater antitumor activity than 10 mg/kg of docetaxel in A549 non-small cell lung, as well as MDA-MB-436 and SUM149 triple negative breast cancer xenograft models. The lead molecule entered a Phase I clinical trial in patients with solid tumors (NCT03076372).

Antitumour activity and tolerability of an EphA2-targeted nanotherapeutic in multiple mouse models.

Antibody-mediated tumour targeting and nanoparticle-mediated encapsulation can reduce the toxicity of antitumour drugs and improve their efficacy. Here, we describe the performance of a nanotherapeutic encapsulating a hydrolytically sensitive docetaxel prodrug and conjugated to an antibody specific for EphA2-a receptor overexpressed in many tumours. Administration of the nanotherapeutic in mice led to slow and sustained release of the prodrug, reduced exposure of active docetaxel in the circulation (compared with administration of the free drug) and maintenance of optimal exposure of the drug in tumour tissue. We also show that administration of the nanotherapeutic in rats and dogs resulted in minimal haematological toxicity, as well as the absence of neutropenia and improved overall tolerability in multiple rodent models. Targeting of the nanotherapeutic to EphA2 improved tumour penetration and resulted in markedly enhanced antitumour activity (compared with administration of free docetaxel and non-targeted nanotherapeutic controls) in multiple tumour-xenografted mice. This nanomedicine could become a potent and safe therapeutic alternative for cancer patients undergoing chemotherapy.