ABSTRACT
The treatment of patients with advanced-stage solid tumours typically involves a multimodality approach (including surgery, chemotherapy, radiotherapy, targeted therapy and/or immunotherapy), which is often ultimately ineffective. Nucleic acid-based drugs, either as monotherapies or in combination with standard-of-care therapies, are rapidly emerging as novel treatments capable of generating responses in otherwise refractory tumours. These therapies include those using viral vectors (also referred to as gene therapies), several of which have now been approved by regulatory agencies, and nanoparticles containing mRNAs and a range of other nucleotides. In this Review, we describe the development and clinical activity of viral and non-viral nucleic acid-based treatments, including their mechanisms of action, tolerability and available efficacy data from patients with solid tumours. We also describe the effects of the tumour microenvironment on drug delivery for both systemically administered and locally administered agents. Finally, we discuss important trends resulting from ongoing clinical trials and preclinical testing, and manufacturing and/or stability considerations that are expected to underpin the next generation of nucleic acid agents for patients with solid tumours.
Subject(s)
Neoplasms , Nucleic Acids , Humans , Neoplasms/drug therapy , Neoplasms/genetics , Neoplasms/therapy , Nucleic Acids/therapeutic use , Tumor Microenvironment/drug effects , Genetic Therapy/methods , Nanoparticles/therapeutic use , Antineoplastic Agents/therapeutic use , Drug Delivery Systems/methodsABSTRACT
Adding a cationic helper lipid to a lipid nanoparticle (LNP) can increase lung delivery and decrease liver delivery. However, it remains unclear whether charge-dependent tropism is universal or, alternatively, whether it depends on the component that is charged. Here, we report evidence that cationic cholesterol-dependent tropism can differ from cationic helper lipid-dependent tropism. By testing how 196 LNPs delivered mRNA to 22 cell types, we found that charged cholesterols led to a different lung:liver delivery ratio than charged helper lipids. We also found that combining cationic cholesterol with a cationic helper lipid led to mRNA delivery in the heart as well as several lung cell types, including stem cell-like populations. These data highlight the utility of exploring charge-dependent LNP tropism.
Subject(s)
Liver , Stem Cells , Heart , Cations , Cholesterol , RNA, MessengerABSTRACT
Poly(ethylene glycol) (PEG)-lipids are used in Food-and-Drug-Administration-approved lipid nanoparticle (LNP)-RNA drugs, which are safe and effective. However, it is reported that PEG-lipids may also contribute to accelerated blood clearance and rare cases of hypersensitivity; this highlights the utility of exploring PEG-lipid alternatives. Here, it is shown that LNPs containing poly(2-ethyl-2-oxazoline) (PEOZ)-lipids can deliver messenger RNA (mRNA) to multiple cell types in mice inside and outside the liver. In addition, it is reported that LNPs formulated with PEOZ-lipids show reduced clearance from the bloodstream and lower levels of antistealth lipid immunoglobulin Ms than LNPs formulated with PEG-lipids. These data justify further exploration of PEOZ-lipids as alternatives to PEG-lipids in LNP-RNA formulations.
Subject(s)
Lipids , Nanoparticles , Polyamines , Polyethylene Glycols , Polyethylene Glycols/chemistry , Animals , Polyamines/chemistry , Nanoparticles/chemistry , Mice , Lipids/chemistry , Humans , RNA, Messenger/genetics , RNA, Messenger/metabolismABSTRACT
The underlying neural mechanisms underpinning the association between age-related hearing loss (ARHL) and dementia remain unclear. A limitation has been the lack of functional neuroimaging studies in ARHL cohorts to help clarify this relationship. In the present study, we investigated the neural correlates of feature binding in visual working memory with ARHL (controls = 14, mild HL = 21, and moderate or greater HL = 23). Participants completed a visual change detection task assessing feature binding while their neural activity was synchronously recorded via high-density electroencephalography. There was no difference in accuracy scores for ARHL groups compared to controls. There was increased electrophysiological activity in those with ARHL, particularly in components indexing the earlier stages of visual cognitive processing. This activity was more pronounced with more severe ARHL and was associated with maintained feature binding. Source space (sLORETA) analyses indicated greater activity in networks modulated by frontoparietal and temporal regions. Our results demonstrate there may be increased involvement of neurocognitive control networks to maintain lower-order neurocognitive processing disrupted by ARHL.
Subject(s)
Memory, Short-Term , Presbycusis , Humans , Visual Perception , ElectroencephalographyABSTRACT
Recent epidemiological research has linked decline in multiple sensory functions with an increased risk of dementia. However, the pathways linking such factors with neurodegenerative disorders remain unclear. Studies that assess this are crucial for guiding the distribution of public health resources and the development of clinical trials aiming to delay or prevent dementia. This commentary examines the contribution of a study by Dintica and colleagues to the field of sensory-cognitive research.
Subject(s)
Alzheimer Disease , Cognitive Dysfunction , Dementia , Humans , Dementia/epidemiology , Dementia/psychology , Sensation , Health Policy , Cognitive Dysfunction/psychologyABSTRACT
Lipid nanoparticles (LNPs) are a clinically relevant way to deliver therapeutic mRNA to hepatocytes in patients. However, LNP-mRNA delivery to end-stage solid tumors such as head and neck squamous cell carcinoma (HNSCC) remains more challenging. While scientists have used in vitro assays to evaluate potential nanoparticles for HNSCC delivery, high-throughput delivery assays performed directly in vivo have not been reported. Here we use a high-throughput LNP assay to evaluate how 94 chemically distinct nanoparticles delivered nucleic acids to HNSCC solid tumors in vivo. DNA barcodes were used to identify LNPHNSCC, a novel LNP for systemic delivery to HNSCC solid tumors. Importantly, LNPHNSCC retains tropism to HNSCC solid tumors while minimizing off-target delivery to the liver.
Subject(s)
Head and Neck Neoplasms , Nanoparticles , Humans , Squamous Cell Carcinoma of Head and Neck , RNA, Messenger/genetics , Lipids , Head and Neck Neoplasms/genetics , RNA, Small Interfering/geneticsABSTRACT
Stereochemistry can alter small-molecule pharmacokinetics, safety and efficacy. However, it is unclear whether the stereochemistry of a single compound within a multicomponent colloid such as a lipid nanoparticle (LNP) can influence its activity in vivo. Here we report that LNPs containing stereopure 20α-hydroxycholesterol (20α) delivered mRNA to liver cells up to 3-fold more potently than LNPs containing a mixture of both 20α- and 20ß-hydroxycholesterols (20mix). This effect was not driven by LNP physiochemical traits. Instead, in vivo single-cell RNA sequencing and imaging revealed that 20mix LNPs were sorted into phagocytic pathways more than 20α LNPs, resulting in key differences between LNP biodistribution and subsequent LNP functional delivery. These data are consistent with the fact that nanoparticle biodistribution is necessary, but not sufficient, for mRNA delivery, and that stereochemistry-dependent interactions between LNPs and target cells can improve mRNA delivery.
Subject(s)
Lipids , Nanoparticles , Lipids/chemistry , RNA, Messenger/genetics , Tissue Distribution , Nanoparticles/chemistryABSTRACT
Lipid nanoparticles (LNPs) have delivered RNA to hepatocytes in patients, underscoring the potential impact of nonliver delivery. Scientists can shift LNP tropism to the lung by adding cationic helper lipids; however, the biological response to these LNPs remains understudied. To evaluate the hypothesis that charged LNPs lead to differential cellular responses, we quantified how 137 LNPs delivered mRNA to 19 cell types in vivo. Consistent with previous studies, we observed helper lipid-dependent tropism. After identifying and individually characterizing three LNPs that targeted different tissues, we studied the in vivo transcriptomic response to these using single-cell RNA sequencing. Out of 835 potential pathways, 27 were upregulated in the lung, and of these 27, 19 were related to either RNA or protein metabolism. These data suggest that endogenous cellular RNA and protein machinery affects mRNA delivery to the lung in vivo.
Subject(s)
Lipids , Nanoparticles , Humans , Liposomes/metabolism , Hepatocytes/metabolism , RNA, Messenger/genetics , RNA, Small InterferingABSTRACT
Lipid nanoparticles (LNPs) have delivered siRNA and mRNA drugs in humans, underscoring the potential impact of improving the therapeutic window of next-generation LNPs. To increase the LNP therapeutic window, we applied lessons from small-molecule chemistry to ionizable lipid design. Specifically, given that stereochemistry often influences small-molecule safety and pharmacokinetics, we hypothesized that the stereochemistry of lipids within an LNP would influence mRNA delivery. We tested this hypothesis in vivo using 128 novel LNPs that included stereopure derivatives of C12-200, an ionizable lipid that when formulated into LNPs delivers RNA in mice and non-human primates but is not used clinically due to its poor tolerability. We found that a novel C12-200-S LNP delivered up to 2.8-fold and 6.1-fold more mRNA in vivo than its racemic and C12-200-R controls, respectively. To identify the potential causes leading to increased delivery, we quantified LNP biophysical traits and concluded that these did not change with stereochemistry. Instead, we found that stereopure LNPs were better tolerated than racemic LNPs in vivo. These data suggest that LNP-mediated mRNA delivery can be improved by designing LNPs to include stereopure ionizable lipids.
Subject(s)
Lipids , Nanoparticles , Mice , Animals , Lipids/chemistry , RNA, Messenger/genetics , RNA, Small Interfering/genetics , RNA, Small Interfering/chemistry , Nanoparticles/chemistry , RNA, Double-StrandedABSTRACT
Lipid nanoparticles (LNPs) have delivered therapeutic RNA to hepatocytes in humans. Adsorption of apolipoprotein E (ApoE) onto these clinical LNP-mRNA drugs has been shown to facilitate hepatocyte entry via the low-density lipoprotein receptor (LDLR). Since ApoE-LDLR trafficking is conserved in mice, non-human primates, and humans, characterizing this mechanism eased clinical transition. Recently, LNPs have delivered mRNA to non-hepatocytes in mice and non-human primates, suggesting they can target new cell types via ApoE- and LDLR-independent pathways. To test this hypothesis, we quantified how 60 LNPs delivered mRNA with cell type resolution in wild-type mice and three knockout mouse strains related to lipid trafficking: ApoE-/-, LDLR-/-, and PCSK9-/-. These data suggest that the hydrophobic tail length of diketopiperazine-based lipids can be changed to drive ApoE- and LDLR-independent delivery in vivo. More broadly, the results support the hypothesis that endogenous LNP trafficking can be tuned by modifying lipid chemistry.
Subject(s)
Apolipoproteins E , Lipoproteins, LDL , Nanoparticles , Animals , Mice , Apolipoproteins E/genetics , Lipoproteins, LDL/genetics , Mice, Knockout , Nanoparticles/chemistry , RNA, Messenger/chemistryABSTRACT
In humans, lipid nanoparticles (LNPs) have safely delivered therapeutic RNA to hepatocytes after systemic administration and to antigen-presenting cells after intramuscular injection. However, systemic RNA delivery to non-hepatocytes remains challenging, especially without targeting ligands such as antibodies, peptides, or aptamers. Here we report that piperazine-containing ionizable lipids (Pi-Lipids) preferentially deliver mRNA to immune cells in vivo without targeting ligands. After synthesizing and characterizing Pi-Lipids, we use high-throughput DNA barcoding to quantify how 65 chemically distinct LNPs functionally delivered mRNA (i.e., mRNA translated into functional, gene-editing protein) in 14 cell types directly in vivo. By analyzing the relationships between lipid structure and cellular targeting, we identify lipid traits that increase delivery in vivo. In addition, we characterize Pi-A10, an LNP that preferentially delivers mRNA to the liver and splenic immune cells at the clinically relevant dose of 0.3 mg/kg. These data demonstrate that high-throughput in vivo studies can identify nanoparticles with natural non-hepatocyte tropism and support the hypothesis that lipids with bioactive small-molecule motifs can deliver mRNA in vivo.
Subject(s)
Lipids , Nanoparticles , Humans , Lipids/chemistry , Liposomes , Nanoparticles/chemistry , Piperazine , RNA, Messenger/metabolism , RNA, Small Interfering/metabolismABSTRACT
Cells that were previously described as homogeneous are composed of subsets with distinct transcriptional states. However, it remains unclear whether this cell heterogeneity influences the efficiency with which lipid nanoparticles (LNPs) deliver messenger RNA therapies in vivo. To test the hypothesis that cell heterogeneity influences LNP-mediated mRNA delivery, we report here a new multiomic nanoparticle delivery system called single-cell nanoparticle targeting-sequencing (SENT-seq). SENT-seq quantifies how dozens of LNPs deliver DNA barcodes and mRNA into cells, the subsequent protein production and the transcriptome, with single-cell resolution. Using SENT-seq, we have identified cell subtypes that exhibit particularly high or low LNP uptake as well as genes associated with those subtypes. The data suggest that cell subsets have distinct responses to LNPs that may affect mRNA therapies.
Subject(s)
Lipids , Nanoparticles , Liposomes , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA, Small Interfering/geneticsABSTRACT
To predict whether preclinical lipid nanoparticle (LNP) delivery will translate in humans, it is necessary to understand whether the mechanism used by LNPs to enter cells is conserved across species. In mice, non-human primates, and humans, LNPs deliver RNA to hepatocytes by adsorbing apolipoprotein E (ApoE), which binds low-density lipoprotein receptor (LDLR). A growing number of LNPs can deliver RNA to nonhepatocytes, suggesting that ApoE- and LDLR-independent interactions could affect LNP tropism. To evaluate this hypothesis, we developed a universal DNA barcoding system that quantifies how chemically distinct LNPs deliver small interfering RNA in any mouse model, including genetic knockouts. We quantified how 98 different LNPs targeted 11 cell types in wildtype, LDLR-/-, very low-density lipoprotein receptor, and ApoE-/- mice, studying how these genes, which traffic endogenous lipids, affected LNP delivery. These data identified a novel, stereopure LNP that targets Kupffer cells, endothelial cells, and hepatocytes in an ApoE-independent manner. These results suggest that non-ApoE interactions can affect the tropism of LNP-RNA drugs.
Subject(s)
Lipids , Nanoparticles , Animals , Apolipoproteins E/genetics , Apolipoproteins E/metabolism , Endothelial Cells/metabolism , Lipoproteins, LDL , Liposomes , Mice , Nanoparticles/metabolism , RNA, Small Interfering/geneticsABSTRACT
Music interventions may represent an effective approach to improving symptoms and delaying progression of MCI to dementia. This review identified nine studies (8 RCT's, 1 observational study) that explored the benefits of music interventions to those with MCI. Studies included five music-playing interventions (sample size (n) ranged from 35 to 201, age ranged from 62 to 94), one music listening intervention (n = 100, mean age = 77 (music intervention) mean age = 76 (dance intervention), one music with movement intervention (n = 16, age range 65-84 years) and two music reminiscence interventions (n = 68; 72, age range = 60-85 years). Only individuals with a clinical diagnosis of MCI were included, no individuals with a diagnosis of dementia were included. Studies were limited due to their sample size, failure to consider confounding variables (i.e. socialization), inconsistency with therapist led sessions, failure to match conditions across interventions, limited follow-up period post-intervention and the tendency to focus on depression exclusively as a measure of behavioural symptoms. Different types of music interventions have differential results on cognitive and behavioural symptoms. The different pattern of brain activation and cognitive abilities which support each type of music activity (e.g. listening vs playing music) may offer some explanation towards these differences. A standardised protocol is needed for each type of music intervention to address how music interventions are studied, taking these limitations into consideration.
Subject(s)
Cognitive Dysfunction , Dementia , Music Therapy , Music , Aged , Aged, 80 and over , Cognition , Cognitive Dysfunction/psychology , Cognitive Dysfunction/therapy , Dementia/psychology , Dementia/therapy , Depression , Humans , Music Therapy/methods , Observational Studies as TopicABSTRACT
Systemically delivered lipid nanoparticles are preferentially taken up by hepatocytes. This hinders the development of effective, non-viral means of editing genes in tissues other than the liver. Here we show that lipid-nanoparticle-mediated gene editing in the lung and spleen of adult mice can be enhanced by reducing Cas9-mediated insertions and deletions in hepatocytes via oligonucleotides disrupting the secondary structure of single-guide RNAs (sgRNAs) and also via their combination with short interfering RNA (siRNA) targeting Cas9 messenger RNA (mRNA). In SpCas9 mice with acute lung inflammation, the systemic delivery of an oligonucleotide inhibiting an sgRNA targeting the intercellular adhesion molecule 2 (ICAM-2), followed by the delivery of the sgRNA, reduced the fraction of ICAM-2 indels in hepatocytes and increased that in lung endothelial cells. In wild-type mice, the lipid-nanoparticle-mediated delivery of an inhibitory oligonucleotide, followed by the delivery of Cas9-degrading siRNA and then by Cas9 mRNA and sgRNA, reduced the fraction of ICAM-2 indels in hepatocytes but not in splenic endothelial cells. Inhibitory oligonucleotides and siRNAs could be used to modulate the cell-type specificity of Cas9 therapies.
Subject(s)
Gene Editing , Nanoparticles , Animals , Antigens, CD , CRISPR-Cas Systems , Cell Adhesion Molecules/genetics , Endothelial Cells , Lipids/chemistry , Liposomes , Liver , Lung , Mice , Nanoparticles/chemistry , SpleenABSTRACT
Nanoparticles are tested in mice and non-human primates before being selected for clinical trials. Yet the extent to which mRNA delivery, as well as the cellular response to mRNA drug delivery vehicles, is conserved across species in vivo is unknown. Using a species-independent DNA barcoding system, we have compared how 89 lipid nanoparticles deliver mRNA in mice with humanized livers, primatized livers and four controls: mice with 'murinized' livers as well as wild-type BL/6, Balb/C and NZB/BlNJ mice. We assessed whether functional delivery results in murine, non-human primate and human hepatocytes can be used to predict delivery in the other species in vivo. By analysing in vivo hepatocytes by RNA sequencing, we identified species-dependent responses to lipid nanoparticles, including mRNA translation and endocytosis. These data support an evidence-based approach to making small-animal preclinical nanoparticle studies more predictive, thereby accelerating the development of RNA therapies.
Subject(s)
Lipids , Nanoparticles , Animals , Liposomes , Mice , RNA, Messenger/geneticsABSTRACT
RNA-based gene therapy requires therapeutic RNA to function inside target cells without eliciting unwanted immune responses. RNA can be ferried into cells using non-viral drug delivery systems, which circumvent the limitations of viral delivery vectors. Here, we review the growing number of RNA therapeutic classes, their molecular mechanisms of action, and the design considerations for their respective delivery platforms. We describe polymer-based, lipid-based, and conjugate-based drug delivery systems, differentiating between those that passively and those that actively target specific cell types. Finally, we describe the path from preclinical drug delivery research to clinical approval, highlighting opportunities to improve the efficiency with which new drug delivery systems are discovered.
Subject(s)
Drug Delivery Systems , Genetic Therapy , Genetic Vectors , RNA, Small Interfering/geneticsABSTRACT
mRNA drugs can preempt infectious disease and treat Mendelian disorders, such as sickle cell anemia, muscular dystrophy, and cystic fibrosis, as well as autoimmunity and cancer. The three major therapeutic areas for which mRNA delivery is currently being explored are antigen production, including the COVID-19 vaccine, protein replacement therapy, and genome engineering. It was demonstrated 30 years ago that introducing in vitro transcribed mRNA intramuscularly results in detectable protein expression for specific antigens protecting against the likes of influenza and cancer. Utilizing mRNA as a therapeutic modality, however, is challenging. mRNA is large and anionic and, as a result, cannot passively diffuse across the negatively charged plasma membrane. In addition, RNases present in the bloodstream and tissues rapidly degrade mRNA, and its administration induces the innate immune response. In consequence, lipid-, polymer-, dendrimer-, and natural membrane-based mRNA drug delivery systems have been developed to deliver mRNA to target cells. Significant efforts and investments have been made to translate some of these systems into the clinic. Specifically, systemically administered lipid nanoparticles (LNPs) have delivered mRNA to the liver, and intramuscularly administered LNPs have delivered mRNA to immune cells to protect against coronavirus disease of 2019. However, clinically relevant delivery in non-liver tissues such as the spleen, lungs, heart, eye, central nervous system, and lymphatics requires improved drug delivery systems.In this Account, we provide an overview of key advances that have led us to Food and Drug Administration approval for the Pfizer/BioNTech mRNA-based vaccine against SARS-CoV-2 and Emergency Use Authorization for the Moderna mRNA-based vaccine against the same disease, and we explain how these developments will contribute to the clinical translation of mRNA therapeutics targeted outside of the liver. We first focus on the chemical modifications and sequence optimization that can improve the potency of mRNA, resulting in greatly improved pharmacokinetics. After detailing what makes an ideal mRNA payload, we review drug delivery systems used to deliver the payload into target cells. We describe efforts to reduce clearance by the liver, a key obstacle to the development of non-liver therapies. We then consider recent examples of nanoparticles that have delivered mRNA to non-liver tissues. Finally, we discuss current clinical mRNA programs, focusing on the COVID vaccines and highlighting lessons that may be applied to future mRNA drugs.
Subject(s)
COVID-19 Vaccines , COVID-19 , Humans , Liposomes , Liver , Nanoparticles , RNA, Messenger/genetics , SARS-CoV-2ABSTRACT
Objectives: To investigate whether the relationship between subjective age-related hearing loss (SARHL) and episodic memory functioning is mediated by measures of social functioning.Methods: Using data from 8,163 adults over 50 that participated in the Irish Longitudinal Study of Ageing (three waves, each two years apart), we used a multiple mediation model within a Structural Equation Modelling framework to explore potential social mediators of the relationship between SARHL and episodic memory functioning, controlling for demographic and health covariates.Results: Neither the direct effect of self-reported hearing difficulties on memory functioning (ß = -.03), nor the total effect (ß = .01), were significant. A small inconsistent indirect effect of self-reported hearing difficulties on episodic memory via weekly social activity engagement (ß = -.002) was found.Conclusions: Self-reported hearing difficulties may exert an indirect effect on episodic memory via weekly social activity engagement. The findings may have implications for identification of individuals at risk of memory decline in later life.