bims-novged Biomed News
on Non-viral vectors for gene delivery
Issue of 2023–08–06
six papers selected by
the Merkel lab, Ludwig-Maximilians University
  1. Inhalable dry powder product (DPP) of mRNA lipid nanoparticles (LNPs) for pulmonary delivery.
  2. Biophysical characterization of siRNA-loaded Lipid Nanoparticles with different PEG content in an aqueous system.
  3. Lipid nanoparticles for siRNA delivery in cancer treatment.
  4. The interplay of quaternary ammonium lipid structure and protein corona on lung-specific mRNA delivery by selective organ targeting (SORT) nanoparticles.
  5. Lipid nanoparticle-targeted mRNA formulation as a treatment for ornithine-transcarbamylase deficiency model mice.
  6. Get out or die trying: peptide- and protein-based endosomal escape of RNA therapeutics.
  1. Drug Deliv Transl Res. 2023 Aug 01.
    Inhalable dry powder product (DPP) of mRNA lipid nanoparticles (LNPs) for pulmonary delivery.
    Ashish Sarode, Priyal Patel, Natalia Vargas-Montoya, Ayed Allawzi, Alisa Zhilin-Roth, Saswata Karmakar, Lianne Boeglin, Hongfeng Deng, Shrirang Karve, Frank DeRosa.
      Pulmonary delivery of mRNA via inhalation is a very attractive approach for RNA-based therapy for treatment of lung diseases. In this work, we have demonstrated successful development of an mRNA-lipid nanoparticle (LNP) dry powder product (DPP), wherein the LNPs were spray dried using hydroalcoholic solvent along with mannitol and leucine as excipients. The desired critical attributes for the DPP were accomplished by varying the excipients, lipid composition, concentration of LNPs, and weight percentage of mRNA. Leucine alone or in combination with mannitol improved the formulation by increasing the mRNA yield as well as decreasing the particle size. Intratracheal administration of the DPP in mice resulted in luciferase expression in the trachea and lungs indicating successful delivery of functional mRNA. Our results show formulation optimization of mRNA LNPs administered in the form of DPP results in an efficacious functional delivery with great promise for future development of mRNA therapeutics for lung diseases.
    Keywords:  Dry powder; Leucine; Lipid nanoparticle (LNP); Mannitol; Pulmonary; Spray drying; mRNA
    DOI:  https://doi.org/10.1007/s13346-023-01402-y
  2. Eur J Pharm Biopharm. 2023 Jul 27. pii: S0939-6411(23)00195-9. [Epub ahead of print]
    Biophysical characterization of siRNA-loaded Lipid Nanoparticles with different PEG content in an aqueous system.
    Ki Hyun Kim, Ripesh Bhujel, Ravi Maharjan, Jae Chul Lee, Hun Soon Jung, Hye Jeong Kim, Nam Ah Kim, Seong Hoon Jeong.
      Although lipid nanoparticles (LNP) are potential carriers of various pharmaceutical ingredients, further investigation for maintaining their stability under various environmental stressors must be performed. This study evaluated the influence of PEGylation and stress conditions on the stability of siRNA-loaded LNPs with different concentrations of PEG (0.5 mol%; 0.5% PEG-LNP and 1.0 mol%; 1.0% PEG-LNP) anchored to their surface. We applied end-over-end agitation, elevated temperature, and repeated freeze and thaw (F/T) cycles as physicochemical stressors of pH and ionic strength. Dynamic light scattering (DLS), flow imaging microscopy (FIM), and ionic-exchange chromatography (IEX) were to determine the degree of aggregation and change in siRNA content. The results indicate that 0.5% PEG-LNP resisted aggregation only at low pH levels or with salt, whereas 1.0% PEG-LNP had increased colloidal stability except at pH 4. 0.5% PEG-LNP withstood aggregation until 71°C and three cycles of F/T. In contrast, 1.0% PEG-LNP maintained colloidal stability at 90°C and seven F/T cycles. Moreover, 1.0% PEG-LNP had higher siRNA stability under all stress conditions. Therefore, to ensure the stability of LNP and encapsulated siRNA, the PEG concentration must be carefully controlled while considering LNPs' colloidal instability mechanisms under various stress conditions.
    Keywords:  Aggregation; Colloidal stability; Flow imaging microscopy; PEGylated LNPs; siRNA
    DOI:  https://doi.org/10.1016/j.ejpb.2023.07.013
  3. J Control Release. 2023 Jul 31. pii: S0168-3659(23)00478-9. [Epub ahead of print]
    Lipid nanoparticles for siRNA delivery in cancer treatment.
    Souhaila H El Moukhtari, Elisa Garbayo, Ane Amundarain, Simón Pascual-Gil, Arantxa Carrasco-León, Felipe Prosper, Xabier Agirre, María J Blanco-Prieto.
      RNA-based therapies, and siRNAs in particular, have attractive therapeutic potential for cancer treatment due to their ability to silence genes that are imperative for tumor progression. To be effective and solve issues related to their poor half-life and poor pharmacokinetic properties, siRNA require adequate drug delivery systems that protect them from degradation and allow intracellular delivery. Among the various delivery vehicles available, lipid nanoparticles have emerged as the leading choice. These nanoparticles consist of cholesterol, phospholipids, PEG-lipids and most importantly ionizable cationic lipids. These ionizable lipids enable the binding of negatively charged siRNA, resulting in the formation of stable and neutral lipid nanoparticles with exceptionally high encapsulation efficiency. Lipid nanoparticles have demonstrated their effectiveness and versatility in delivering not only siRNAs but also multiple RNA molecules, contributing to their remarkable success. Furthermore, the advancement of efficient manufacturing techniques such as microfluidics, enables the rapid mixing of two miscible solvents without the need for shear forces. This facilitates the reproducible production of lipid nanoparticles and holds enormous potential for scalability. This is shown by the increasing number of preclinical and clinical trials evaluating the potential use of siRNA-LNPs for the treatment of solid and hematological tumors as well as in cancer immunotherapy. In this review, we provide an overview of the progress made on siRNA-LNP development for cancer treatment and outline the current preclinical and clinical landscape in this area. Finally, the translational challenges required to bring siRNA-LNPs further into the clinic are also discussed.
    Keywords:  Endosomal escape; Gene inhibition; Lipid nanoparticles; Microfluidics; Small interfering RNAs (siRNAs); cancer
    DOI:  https://doi.org/10.1016/j.jconrel.2023.07.054
  4. J Control Release. 2023 Aug 01. pii: S0168-3659(23)00482-0. [Epub ahead of print]
    The interplay of quaternary ammonium lipid structure and protein corona on lung-specific mRNA delivery by selective organ targeting (SORT) nanoparticles.
    Sean A Dilliard, Yehui S Sun, Madeline O Brown, Yun-Chieh Sung, Sumanta Chatterjee, Lukas Farbiak, Amogh Vaidya, Xizhen Lian, Xu Wang, Andrew Lemoff, Daniel J Siegwart.
      Messenger RNA (mRNA) can treat genetic disease using protein replacement or genome editing approaches but requires a suitable carrier to circumnavigate biological barriers and access the desired cell type within the target organ. Lipid nanoparticles (LNPs) are widely used in the clinic for mRNA delivery yet are limited in their applications due to significant hepatic accumulation due to the formation of protein corona enriched in apolipoprotein E (ApoE). Our lab developed selective organ targeting (SORT) LNPs that incorporate a supplementary component, termed a SORT molecule, for tissue-specific mRNA delivery to the liver, spleen, and lungs of mice. Mechanistic work revealed that the biophysical class of SORT molecule added to the LNP forms a distinct protein corona that helps determine where in the body mRNA is delivered. To better understand which plasma proteins could drive tissue-specific mRNA delivery, we characterized a panel of quaternary ammonium lipids as SORT molecules to assess how chemical structure affects the organ-targeting outcomes and protein corona of lung-targeting SORT LNPs. We discovered that variations in the chemical structure of both the lipid alkyl tail and headgroup impact the potency and specificity of mRNA delivery to the lungs. Furthermore, changes to the chemical structure alter the quantities and identities of protein corona constituents in a manner that correlates with organ-targeting outcomes, with certain proteins appearing to promote lung targeting whereas others reduce delivery to off-target organs. These findings unveil a nuanced relationship between LNP chemistry and endogenous targeting, where the ensemble of proteins associated with an LNP can play various roles in determining the tissue-specificity of mRNA delivery, providing further design criteria for optimization of clinically-relevant nanoparticles for extrahepatic delivery of genetic payloads.
    Keywords:  Endogenous targeting; Lipid nanoparticle; Lungs; Protein corona; mRNA delivery
    DOI:  https://doi.org/10.1016/j.jconrel.2023.07.058
  5. Mol Ther Nucleic Acids. 2023 Sep 12. 33 210-226
    Lipid nanoparticle-targeted mRNA formulation as a treatment for ornithine-transcarbamylase deficiency model mice.
    Kazuto Yamazaki, Kenji Kubara, Satoko Ishii, Keita Kondo, Yuta Suzuki, Takayuki Miyazaki, Kaoru Mitsuhashi, Masashi Ito, Kappei Tsukahara.
      Ornithine transcarbamylase (OTC) plays a significant role in the urea cycle, a metabolic pathway functioning in the liver to detoxify ammonia. OTC deficiency (OTCD) is the most prevalent urea cycle disorder. Here, we show that intravenously delivered human OTC (hOTC) mRNA by lipid nanoparticles (LNP) was an effective treatment for OTCD by restoring the urea cycle. We observed a homotrimer conformation of hOTC proteins produced by the mRNA-LNP in cells by cryo-electron microscopy. The immunohistochemistry revealed the mitochondria localization of produced hOTC proteins in hepatocytes in mice. In livers of mice intravenously injected with hOTC-mRNA/LNP at 1.0 mg/kg, the delivered hOTC mRNA levels steeply decreased with a half-life (t1/2) of 7.1 h, whereas the produced hOTC protein levels retained for 5 days and then declined with a t1/2 of 2.2 days. In OTCD model mice (high-protein diet-fed Otcspf-ash hemizygous males), a single dose of hOTC-mRNA/LNP at 3.0 mg/kg ameliorated hyperammonemia and weight loss with prolonged survival rate (22 days) compared with that of untreated mice (11 days). Weekly repeated doses at 0.3 and 1.0 mg/kg were well tolerated in wild-type mice and showed a dose-dependent amelioration of survival rate in OTCD mice, thus, showing the therapeutic potential of LNP-formulated hOTC mRNA for OTCD.
    Keywords:  MT: delivery strategies: ornithine transcarbamylase; Otcspf-ash mice; cryo-electron microscopy; homotrimer conformation; lipid nanoparticles; messenger RNA therapy; mitochondria localization
    DOI:  https://doi.org/10.1016/j.omtn.2023.06.023
  6. Adv Drug Deliv Rev. 2023 Aug 01. pii: S0169-409X(23)00362-9. [Epub ahead of print] 115047
    Get out or die trying: peptide- and protein-based endosomal escape of RNA therapeutics.
    Alexander Klipp, Michael Burger, Jean-Christophe Leroux.
      RNA therapeutics offer great potential to transform the biomedical landscape, encompassing the treatment of hereditary conditions and the development of better vaccines. However, the delivery of RNAs into the cell is hampered, among others, by poor endosomal escape. This major hurdle is often tackled using special lipids, polymers, or protein-based delivery vectors. In this review, we will focus on the most prominent peptide- and protein-based endosomal escape strategies with focus on RNA drugs. We discuss cell penetrating peptides, which are still incorporated into novel transfection systems today to promote endosomal escape. However, direct evidence for enhanced endosomal escape by the action of such peptides is missing and their transfection efficiency, even in permissive cell culture conditions, is rather low. Endosomal escape by the help of pore forming proteins or phospholipases, on the other hand, allowed to generate more efficient transfection systems. These are, however, often hampered by considerable toxicity and immunogenicity. We conclude that the perfect enhancer of endosomal escape has yet to be devised. To increase the chances of success, any new transfection system should be tested under relevant conditions and guided by assays that allow direct quantification of endosomal escape.
    Keywords:  Cell penetrating peptides (CPPs); Endosomal escape; Endosomal rupture; Gene delivery; Phospholipase; Pore formation; Protein-based endosomal escape; RNA therapeutics
    DOI:  https://doi.org/10.1016/j.addr.2023.115047
This page is being maintained by Thomas Krichel. It was last updated on 2025‒01‒13 at 11:36.