Selective organ targeting (SORT) is a novel approach in the field of

targeted drug delivery Targeted drug delivery, sometimes called smart drug delivery, is a method of delivering Pharmaceutical drug, medication to a patient in a manner that increases the Dose (biochemistry), concentration of the medication in some parts of the body relat ...

that systematically engineers multiple classes of lipid nanoparticles (LNPs) to enable targeted delivery of therapeutics to specific

organ Organ and organs may refer to: Biology * Organ (biology), a group of tissues organized to serve a common function * Organ system, a collection of organs that function together to carry out specific functions within the body. Musical instruments ...

s in the body. The SORT molecule alters tissue tropism by adjusting the composition and physical characteristics of the nanoparticle. Adding a permanently cationic lipid, a permanently anionic lipid, or ionizable amino lipid increases delivery to the

lung The lungs are the primary Organ (biology), organs of the respiratory system in many animals, including humans. In mammals and most other tetrapods, two lungs are located near the Vertebral column, backbone on either side of the heart. Their ...

spleen The spleen (, from Ancient Greek '' σπλήν'', splḗn) is an organ (biology), organ found in almost all vertebrates. Similar in structure to a large lymph node, it acts primarily as a blood filter. The spleen plays important roles in reg ...

, and

liver The liver is a major metabolic organ (anatomy), organ exclusively found in vertebrates, which performs many essential biological Function (biology), functions such as detoxification of the organism, and the Protein biosynthesis, synthesis of var ...

, respectively. SORT LNPs utilize SORT molecules to accurately tune and mediate gene delivery and editing, resulting in predictable and manageable protein synthesis from

mRNA In molecular biology, messenger ribonucleic acid (mRNA) is a single-stranded molecule of RNA that corresponds to the genetic sequence of a gene, and is read by a ribosome in the process of Protein biosynthesis, synthesizing a protein. mRNA is ...

in particular organ(s), which can potentially improve the efficacy of drugs while reducing side effects.

Overview

LNPs are non-viral synthetic nanoparticles that can carry and deliver different functional molecules to specific tissues. Traditionally, LNPs are composed of four indispensable lipid components: an ionizable amino lipid that aids in both escaping the endosomes and binding nucleic acids to the particle, an amphipathic

phospholipid Phospholipids are a class of lipids whose molecule has a hydrophilic "head" containing a phosphate group and two hydrophobic "tails" derived from fatty acids, joined by an alcohol residue (usually a glycerol molecule). Marine phospholipids typ ...

that promotes fusion with the target cell and endosomes,

cholesterol Cholesterol is the principal sterol of all higher animals, distributed in body Tissue (biology), tissues, especially the brain and spinal cord, and in Animal fat, animal fats and oils. Cholesterol is biosynthesis, biosynthesized by all anima ...

to enhance nanoparticle stability, and a

polyethylene glycol Polyethylene glycol (PEG; ) is a polyether compound derived from petroleum with many applications, from industrial manufacturing to medicine. PEG is also known as polyethylene oxide (PEO) or polyoxyethylene (POE), depending on its molecular wei ...

lipid that improves colloidal stability and reduces clearance of the particle by the reticuloendothelial system. LNPs have demonstrated safety and effectiveness but are limited to intramuscular and intravenous administration targeting the liver. This limitation largely stems from LNPs' resemblance to

very-low-density lipoprotein Very-low-density lipoprotein (VLDL), density relative to extracellular water, is a type of lipoprotein made by the liver. VLDL is one of the five major groups of lipoproteins (chylomicrons, VLDL, intermediate-density lipoprotein, low-density lip ...

, leading to a propensity for adsorbing

apolipoprotein E Apolipoprotein E (Apo-E) is a protein involved in the metabolism of fats in the body of mammals. A subtype is implicated in Alzheimer's disease and cardiovascular diseases. It is encoded in humans by the gene ''APOE''. Apo-E belongs to a family ...

(ApoE) present in

blood plasma Blood plasma is a light Amber (color), amber-colored liquid component of blood in which blood cells are absent, but which contains Blood protein, proteins and other constituents of whole blood in Suspension (chemistry), suspension. It makes up ...

. Consequently, LNPs accumulate in the

by binding to the low-density lipoprotein receptor found in

hepatocyte A hepatocyte is a cell of the main parenchymal tissue of the liver. Hepatocytes make up 80% of the liver's mass. These cells are involved in: * Protein synthesis * Protein storage * Transformation of carbohydrates * Synthesis of cholesterol, bi ...

s. SORT LNPs overcome this limitation by augmenting the LNP with an additional component (termed a SORT molecule), allowing delivery to targeted tissues beyond the liver.

Mechanism

Traditionally, LNPs utilize an optimal balance of ionizable amines and nanoparticle-stabilizing hydrophobicity to deliver functional molecules to cells effectively but are limited to liver hepatocytes. In the SORT strategy, these nanoparticles are systematically engineered without altering the molar ratio of the core four components in LNPs, ensuring that the ability to encapsulate

RNA Ribonucleic acid (RNA) is a polymeric molecule that is essential for most biological functions, either by performing the function itself (non-coding RNA) or by forming a template for the production of proteins (messenger RNA). RNA and deoxyrib ...

and escape from endosomes remains intact. The addition of a SORT molecule alters the biodistribution and redirects the molecules facilitating the uptake in specific organs via endogenous targeting mechanisms of action or by influencing the binding affinity to specific serum proteins. Tissue tropism is determined by the distinct chemical functional groups present on the surface of the nanoparticle, which alter the physicochemical properties of the LNP. These properties encompass factors such as

molarity Molar concentration (also called molarity, amount concentration or substance concentration) is the number of moles of solute per liter of solution. Specifically, It is a measure of the concentration of a chemical species, in particular, of a solu ...

, percentage added, and various other characteristics. The critical factor that governs tissue tropism is the modulation of the surface's

acid dissociation constant In chemistry, an acid dissociation constant (also known as acidity constant, or acid-ionization constant; denoted ) is a quantitative property, quantitative measure of the acid strength, strength of an acid in Solution (chemistry), solution. I ...

(pKa), which corresponds to the pH at which the proportion of charged and uncharged ionizable lipids at the particle's surface is equal and depends on the type of ionizable lipids and charged helper lipids used in the nanoparticle formulation. The shift from liver tissues is attributed to the alteration in the surface pKa induced by the addition of an anionic head group, which subsequently reduced the strength of interactions with ApoE. Change in surface pKa promotes the adsorption of plasma proteins such as β2- glycoprotein I (β2-GPI) instead of ApoE, resulting in altered

protein corona A protein corona is a dynamic coating of biomolecules, usually proteins, around the surface of a nanoparticle that forms spontaneously in colloidal nanomaterials upon exposure to biological mediums. Protein coronas can form in many different patte ...

that mediates tissue-specific delivery towards the spleen and lung. Adding a cationic quaternary amino lipid, such as 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), in an increasing molar percentage, was able to shift the distribution progressively from the liver to spleen and then the lung, with a threshold that allowed for exclusive lung delivery. Negatively charged SORT lipids allow for direct delivery to the spleen.

Synthesis of SORT LNPs

To prepare self-assembled SORT LNPs, the lipids are mixed in

ethanol Ethanol (also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound with the chemical formula . It is an Alcohol (chemistry), alcohol, with its formula also written as , or EtOH, where Et is the ps ...

to create a dissolved lipid mixture solution, ensuring that the initial relative molar ratios of the four fundamental components remain unaltered. mRNA is dissolved in citrate buffer separately. To encourage that uniform LNPs are formed, it is necessary to rapidly mix both solutions: the lipid solution containing all lipids and the buffer solution containing mRNA. By employing high-speed mixing, the environmental polarity is enhanced, facilitating the formation of homogenous LNPs. Mixing methods include pipetting, vortex or microfluidic mixing. After mixing, characterize SORT LNPs to measure the particle size and encapsulation efficiency and proceed to delivery into the organism. Delivery can be

intrathecal Intrathecal administration is a route of administration for drugs via an injection into the spinal canal, or into the subarachnoid space (sin. ''intrathecal space'') so that it reaches the cerebrospinal fluid (CSF). It is useful in several applic ...

intravenous Intravenous therapy (abbreviated as IV therapy) is a medical technique that administers fluids, medications and nutrients directly into a person's vein. The intravenous route of administration is commonly used for rehydration or to provide nutr ...

intramuscular Intramuscular injection, often abbreviated IM, is the injection of a substance into a muscle. In medicine, it is one of several methods for parenteral administration of medications. Intramuscular injection may be preferred because muscles hav ...

or through nebulization.

Applications

SORT LNPs can mediate therapeutically relevant protein production levels and safely deliver proteins to specific tissues and even particular cell populations. The tissue specificity occurs quickly and is not dependent on time. Further benefits of SORT LNPs include formulation stability and conservation of physiochemical properties over time, including a maintained in vivo efficacy after storage at 4 degrees Celsius. LNPs, in general, are well tolerated in mice and humans, and no alterations in kidney and liver function or alteration of serum proteins have been found in studies with murine models evaluating ''in vivo'' toxicity. SORT has the potential to revolutionize drug delivery by improving the efficacy and

pharmacokinetics Pharmacokinetics (from Ancient Greek ''pharmakon'' "drug" and ''kinetikos'' "moving, putting in motion"; see chemical kinetics), sometimes abbreviated as PK, is a branch of pharmacology dedicated to describing how the body affects a specific su ...

of drugs while reducing side effects. SORT molecules can reach deep tissues that were previously inaccessible for treatment, enhancing tissue penetration. This holds significant promise in benefiting a wide range of

genetic disorder A genetic disorder is a health problem caused by one or more abnormalities in the genome. It can be caused by a mutation in a single gene (monogenic) or multiple genes (polygenic) or by a chromosome abnormality. Although polygenic disorders ...

s, enabling advancements in protein replacement therapy and gene editing, as this strategy allows for gene editing without local administration. The benefits of targeted delivery of protein products or gene editing machinery to the liver are shown in genetic diseases affecting the liver or in which the altered gene product is produced in the liver, such as

tyrosinemia Tyrosinemia or tyrosinaemia is an error of metabolism, usually inborn, in which the body cannot effectively break down the amino acid tyrosine. Symptoms of untreated tyrosinemia include liver and kidney disturbances. Without treatment, tyrosinemi ...

, and transthyretin amyloidosis, respectively, and the addition of a SORT molecule has been shown further improve liver-targeting LNP systems further. However, the SORT strategy could potentially extrapolate these benefits to other organs. One promising target for gene editing is

cystic fibrosis Cystic fibrosis (CF) is a genetic disorder inherited in an autosomal recessive manner that impairs the normal clearance of Sputum, mucus from the lungs, which facilitates the colonization and infection of the lungs by bacteria, notably ''Staphy ...

, as a tailored therapy with an effective delivery system could significantly rescue '' CFTR'' expression. Other possible applications include restoration of

gene expression Gene expression is the process (including its Regulation of gene expression, regulation) by which information from a gene is used in the synthesis of a functional gene product that enables it to produce end products, proteins or non-coding RNA, ...

in other organs, such as restoring

dystrophin Dystrophin is a rod-shaped cytoplasmic protein, and a vital part of a protein complex that connects the cytoskeleton of a muscle fiber to the surrounding extracellular matrix through the cell membrane. This complex is variously known as the costa ...

expression in muscle for

Duchenne muscular dystrophy Duchenne muscular dystrophy (DMD) is a severe type of muscular dystrophy predominantly affecting boys. The onset of muscle weakness typically begins around age four, with rapid progression. Initially, muscle loss occurs in the thighs and pe ...

. Targeted approaches for bone marrow and brain tropism are currently in development One of the most promising applications of SORT is

cancer Cancer is a group of diseases involving Cell growth#Disorders, abnormal cell growth with the potential to Invasion (cancer), invade or Metastasis, spread to other parts of the body. These contrast with benign tumors, which do not spread. Po ...

treatment. By targeting the cancerous cells in a specific organ, SORT may be able to deliver drugs or gene therapies directly to the cancerous cells while sparing the healthy cells in other organs. Selectivity for the spleen could also be applicable in treating cancer via chimeric antigen receptor (CAR)-T cell therapy and opens a new path for developing ''in vivo''

T-cell T cells (also known as T lymphocytes) are an important part of the immune system and play a central role in the adaptive immune response. T cells can be distinguished from other lymphocytes by the presence of a T-cell receptor (TCR) on their ce ...

targeted mRNA delivery systems able to induce robust and transient CAR expression. There are promising applications in the combination of SORT and different delivery methods besides intravenous administration, such as nebulization, intrathecal or intramuscular administration, as these will deliver deliver the SORT molecules directed to targeted organs and further reduce systemic exposure. Additionally, SORT technology is applicable to several classes of established four-component LNPs, and various non-lipid nanoparticle components. This broadens the spectrum of its applications and enables the delivery of diverse therapeutics, encompassing not only nucleic acids but also single or multiple proteins, and even entire genome editors.

Limitations

At present, the SORT strategy is capable of achieving targeted delivery exclusively to specific organs such as the liver, lungs, and spleen. Establishing the SORT LNP formulation is a fine-tuning process, as some concentrations of SORT molecules may aid in delivery to other organs, whereas different concentrations completely select delivery to another organ. However, this fine-tuning mechanism is limited as it can also alter the molecule's activity and render it ineffective. Moreover, it is difficult to accurately predict the biodistribution of LNPs based on their physicochemical parameters, and biodistribution alone cannot predict mRNA-induced protein expression in a specific tissue. There is no indication that a massive accumulation of LNPs in a given tissue will necessarily lead to a high degree of protein expression in the targeted cells

References