In a recent study published in the journal Nature Communications, researchers in the United States investigated the benefits of liquid foam as an alternative to conventional liquid-based gene delivery agents. They accessed the safety, practicality, and accessibility (including cost) of these novel vectors. Their findings highlight that a liquid foam comprised of methylcellulose and xanthan gum (both approved by the US Food and Drug Administration [FDA]) as being safe for human use) depicted transfection efficiency improvements between 2.9- and 384-fold over liquid-based approaches in nonviral gene delivery to murine model systems.
Together, they estimate that foam-based vectors can outperform liquid-based ones by preventing the leaking of the vector's DNA cargo to non-target cells (practicality), can reduce the cost of treatment by tenfold or more (cost and accessibility), and can shield the vector from the host immune system (safety), thereby preventing immune-system-mediated toxicity or oncogenesis (common concerns in conventional liquid vector-based approaches).
Schematic illustration depicting how gene therapy foam is freshly prepared and applied therapeutically to supply new genetic material or change existing DNA in cells. A Nonviral or viral vector (Therapeutic) is added to foam precursor in a syringe connected to a second syringe filled with air. The air and foam precursor are mixed by vigorously drawing the syringe plungers back and forth at least 30 times, creating a uniform microfoam consisting of gas bubbles separated by a network of interconnected liquid film structures called lamellae. Gene therapy vectors are concentrated in this liquid phase as the foam matures. B Once applied to tissue, the foam gradually deploys its therapeutic cargo and either supplies new genetic material or changes the endogenous DNA in the target cell. Study: Liquid foam improves potency and safety of gene therapy vectors
What are foams, and how can they benefit medicine?
Foams are materials formed by the colloidal dispersion of packets of air trapped within liquid or solid layers (called 'lamellae'), with bath sponges and the head (froth) of beer being commonly observed examples. Medical researchers are taking a keen interest in foams (especially liquid-based foams) as drug-delivery systems due to their unique physicochemical properties – their large gas volumes interspersed by low-volume liquid lamellae ensure that their therapeutic payloads are concentrated in the lamellae. This, in turn, provides these materials with the advantages of high stability, sustained drug delivery at the target site, and low- to no leakage of the therapeutic agent to non-target tissue.
Schematic explaining the key advantages of foam as a gene delivery system in comparison to conventional liquid formulations. A Foam is mostly gas, so the embedded vector particles become heavily concentrated in its liquid component, which ensures high-density exposure of target tissue to the gene therapy vector. B Foam remains at the application site longer, thereby enhancing the delivery of the gene therapy drug to the intended cells and minimizing unwanted off-target effects. C Higher vector density combined with longer contact time results in higher transfection rates and deeper tissue penetration.
Given these advantages, a growing number of foam-based therapeutics (e.g., Varithena®, Uceris®, or Luxiq®) have entered the clinical market. Notably, these products have been validated as safe for human use (by the United States [US] Food and Drug Administration [FDA]). Encouragingly, clinical trials investigating efficacy comparisons between these foams and conventional liquid-based drug-delivery agents have revealed that the former outperforms the latter to such an extent that in most medical areas where foams are used, these novel drug-delivery agents have effectively replaced their liquid counterparts.
Despite these substantial advantages and their rising popularity across pharmaceutical and cosmetic industries, foam-based vector delivery agents have surprisingly not been investigated for clinical gene therapy applications. As the first wave of gene-correcting drugs begins to reach patients, the limitations of their liquid vectors become more apparent – liquid-based agents are prone to tissue leakage, oftentimes spilling over to non-target tissue.
This introduces many problems, including 1. Manufacturers deliberately increase drug concentrations (to account for leakage). This, in turn, 2. significantly increases the prices of these cutting-edge and extremely costly drugs, reducing their accessibility to the general public. 3. Delivery of these highly cell-type-specific gene therapies to non-target tissue has been known to cause immune-system mediated toxicity or oncogenesis, affecting the drug's safety.
About the study
In the present study, researchers investigate if foam-based drug delivery vectors' unique and often exceptional benefits extend to gene therapies, especially from the lenses of safety, efficacy, and cost/accessibility. They evaluate numerous FDA-approved foam candidates for their use as vectors, notably methylcellulose, sodium caseinate, and human serum albumin. To augment the stability and performance of these candidates, Xanthan gum was added to each.
They tested these potential drug delivery agents in the delivery of a nonviral vector (a modification of Moderna's COVID-19 mRNA vaccine, wherein the mRNA antigen was replaced with firefly Luciferase mRNA) in clinically meaningful in vivo murine model systems (four- to six-week-old female albino B6 mice).
"Cells were incubated with LNP suspensions or LNP foam for two hours, during which the culture dish was placed in a horizontal or tilted position. The horizontal setup is designed to compare gene transfer efficiencies in the absence of any liquid drainage, whereas the angled transfection mimics the more realistic scenario of patient tissue that is shifting in position and lacks defined borders that would prevent drainage of the applied therapeutics."
Transfection efficacy estimates (measured using the intensity of Luciferase-induced bioluminescence) revealed that while all three foams outperformed their liquid counterparts across both horizontal and angled transfection mimics, the performance of Xanthan-augmented methylcellulose was exceptional. In the horizontal scenario, efficacy was estimated at 2.9 times that of liquid vectors. In the more realistic angled scenario, efficacy was an astounding 384-fold that of conventional liquid delivery agents.
"Furthermore, because methylcellulose foam stays in place, the transfections were spatially well-defined. This is vividly illustrated by our ability to inscribe text onto cells grown in a tissue culture plate while holding the dish vertically, which then appeared as an identical pattern of gene expression 24 h later."
Following the selection of methylcellulose as the candidate foam of choice, researchers characterized its foam structure (bubble size, distribution, and rate of foam decay) using an automated Dynamic Foam Analyzer (Krűss Scientific DFA100FSM). They further estimated the dispersion of lipid nanoparticles (LNPs – the 'payload') visually using high-resolution confocal microscopy.
Subsequently, they used the aforementioned murine models to test foam's intraperitoneal benefits, its bioavailability and biocompatibility across multiple tissue types, and its potential for use as a carrier of viral gene therapeutic payloads (herein, Lentivirus [LV]).
Study findings and conclusions
In the first study to evaluate the potential benefits of foam-based drug delivery agents for gene therapy, researchers reveal that foam (herein, Xanthan-augmented methylcellulose) outperforms its liquid counterparts many times over. In in vitro horizontal models, methylcellulose depicted a close to 3-fold efficacy improvement over currently available liquid vectors. This observed foam advantage rose to 384-fold in more realistic, angled models. If even a 10-fold efficacy improvement could be realized, this would translate into substantial cost and time savings (gene therapy drugs are extremely expensive and complex to manufacture), significantly improving the general accessibility of these hitherto 'exclusive' clinical interventions.
"While gene therapy foam is clearly not suited for systemic infusion, the potential clinical applications of this foam platform are numerous and include improving the safety and potency of oncolytic virus therapy, enhancing vaccines, developing in situ gene therapy for gastrointestinal diseases (oral cancer, esophageal cancer, stomach cancer, colorectal cancer, autoimmune diseases that affect the digestive system), gynecological cancer, skin disease (in particular wound healing), mesotheliomas, cancers spreading to the peritoneal cavity, or any kind of in situ gene modification that requires topical application."
Foams were also shown to persist longer at the target tissue site without any leakage commonly observed in liquid vectors, thereby further reducing the amount of therapeutic agent required for meeting dosage requirements and minimizing oncogenesis and autoimmune toxicity due to off-target events.
"…findings establish that liquid foam is a highly versatile delivery platform to enhance localized gene therapy. Incorporated into the clinical workflow, this platform could shift the paradigm on how topical gene therapy is applied for the treatment of a wide range of diseases."
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