Early Study Supports Nanoparticle Delivery of LIF Protein to Brain
LIF, a protein with anti-inflammatory and neuroprotective properties, can be successfully delivered to immune cells in the brain using a nanoparticle formulation, and partially reverses induced paralysis in a mouse model of multiple sclerosis (MS), a proof-of-concept study has found.
These findings validate LIF-loaded nanoparticles as a therapeutic candidate to treat MS.
The study, “Multiple Sclerosis: LIFNano-CD4 for Trojan Horse Delivery of the Neuro-Protective Biologic ‘LIF’ Into the Brain: Preclinical Proof of Concept,” was published in the journal Frontiers in Medical Technology.
MS consists of an inflammatory response that targets the myelin sheath — a fatty coating surrounding nerve fibers (axons) that improves the transmission of electrical impulses. Damage to the myelin sheath in the brain disrupts signals traveling along the nerve fibers between the brain and the body, causing a variety of disease symptoms.
Chronic inflammation is driven, in part, by the immune signaling protein (cytokine) interleukin-6 (IL-6), which activates immune T-cells to become the pro-inflammatory Th17 cells, which cause damage.
Leukemia inhibitory factor (LIF) is a protein produced by stem cells that plays an opposing role to IL-6 because it triggers T-cells to become regulatory T-cells (Tregs), which have anti-inflammatory properties, stimulate tissue repair, and suppress the autoimmune response.
Recently, to leverage LIF as a potential MS therapeutic, it was packaged into PLGA nanoparticles (LIFNano) — an approved medicinal delivery system with controlled and sustained-release properties, low toxicity, and compatibility with tissue. Over one week, PLGA nanoparticles degrade to carbon dioxide gas and water.
When targeted to bind to immune cells called CD4 T-cells, the LIFNano formulation promoted Treg growth, suppressed inflammatory Th17 cells, and induced myelin repair, leading to improved symptoms in animal models of MS.
In advance of human trials, researchers at LIFNano Therapeutics in the U.K., which is developing the therapy, tested whether LIFNano crossed the blood-brain barrier — a selective membrane that allows only specific molecules in the bloodstream to enter the brain. For LIFNano to be an effective MS therapeutic, it must gain access to the brain.
Additionally, the team conducted a preclinical study in two mouse models of MS to further validate LIFNano’s efficacy and safety.
First, to determine whether the nanoparticles targeting CD4 T-cells accessed the brain, they were packaged with a visible colored dye and injected into the veins (intravenously) of normal mice. After six hours, the team found the dye in all tissues, including the brain, eye, liver, pancreas, and blood. After 24 hours, the dye remained in the brain only.
Next, the nanoparticles were loaded with human LIF, and administered to healthy mice as before. The researchers directly measured LIF released from the nanoparticle and found that after administration of LIFNano, LIF levels peaked in the brain after two hours and decreased to less than half by six hours. Once released, LIF’s levels reduced to half in about 20 minutes (half-life).
Then, two mouse models were used. The Hooke EAE model mimics progressive MS, with total paralysis of tail and hind limbs by day 14 after disease induction. On day 15, one group of these mice were given empty nanoparticles, another received LIFNano nanoparticles, and a control group was without disease and not given treatment.
Compared with untreated animals, LIFNano therapy significantly reversed some of the paralysis as early as four days after the first administration, based on EAE disease scores, a measure of paralysis in this model.
Blood and brain samples were tested for IL-6 and GM-CSF, an immune signaling protein linked to inflammation by enhancing IL-6-dependent Th17 cell development.
Although the levels of IL-6 in blood were similar in treated and untreated MS mice, they were significantly reduced in the brain of treated mice, reaching levels similar to those of healthy mice. Thus, “for IL-6, an effect of LIF derived from [LIFNano] was confirmed, and this effect was selective for the brain,” the team wrote.
LIFNano treatment also significantly reduced GM-CSF. But in contrast to IL-6, these effects occurred outside the brain, as the levels of brain GM-CSF in all tested groups were similar with or without disease.
The second mouse model (Biozzi EAE model) better mimicked relapsing-remitting MS, with a slower progression of myelin damage and disability. Here, LIFNano was injected directly into the brain (intracranial) and given immediately before an induced relapse.
While there were no effects on relapsing paralysis, the researchers observed a significant dose-dependent effect on the rate of recovery from the induced relapse, “which is a hallmark of neuro-protection,” the team wrote.
Finally, three increasing doses of LIFNano (50, 100, and 150 mg per kg body weight) were given to healthy mice to assess safety with tests for toxicity and blood problems.
There were mild transient effects seen at the two higher doses, but not at the lowest dose, which “is on a par to that shown to be neuro-protective in the Biozzi mouse EAE model of relapsing MS via intra-cranial delivery,” the researchers wrote.
The “delivery of the biologic ‘LIF’ into the brain using LIF-loaded PLGA nanoparticles functionalized with anti-CD4 is a validated therapeutic candidate to treat multiple sclerosis,” the team concluded.