Biology subjects

Papp, T. E.

Publications and source records attributed to Papp, T. E..

5 recordsLinked to original sources

bioRxiv2026

Rapid receptor internalization potentiates CD7-targeted lipid nanoparticles for efficient mRNA delivery to T cells and in vivo CAR T-cell engineering

Targeted lipid nanoparticles (tLNPs) enable efficient mRNA delivery to T cells, allowing for in situ generation of chimeric antigen receptor (CAR) T cells without ex vivo manipulation. This strategy has shown promising therapeutical efficacy in preclinical studies of cardiac fibrosis, cancer, and autoimmune diseases. While multiple T-cell surface receptors have been targeted across studies for tLNP-mediated in vivo CAR T-cell generation and exhibit diverse efficiencies, their comparative performance and the mechanisms underlying these differences remain unclear. Here, we systematically compared tLNPs with antibody-based moieties targeting T-cell receptors including CD2, CD4, CD5, CD7, CD8, or a CD4/8 dual-targeting combination under identical conditions, assessing their mRNA delivery efficiency in human T cells and PBMCs in vitro, and subsequently validating the best performer in vivo in humanized mice. Among all moieties tested, CD7-targeting tLNPs achieved the highest mRNA delivery to T cells and efficiently generated functional CAR T cells in vivo. Mechanistic analysis revealed that receptor internalization, rather than the receptor abundance, is the primary determinant of delivery efficiency, a property intrinsic to each receptor and largely independent of antibody clone. These findings provide a rational framework for selecting optimal targeting moiety to enable highly efficient in vivo CAR T-cell engineering. HighlightsO_LITargeting CD7 outperforms other receptors for tLNP-mRNA delivery to T cells C_LIO_LIReceptor abundance does not predict tLNP-mRNA delivery efficiency C_LIO_LIReceptor internalization kinetics governs tLNP-mRNA delivery efficiency C_LIO_LICD7-targeting LNP-mRNA enables efficient in vivo CAR T-cell engineering C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=90 SRC="FIGDIR/small/701374v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@9c2c4eorg.highwire.dtl.DTLVardef@120ced7org.highwire.dtl.DTLVardef@eb9b5forg.highwire.dtl.DTLVardef@25b6f7_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

SRSF3 knockdown-induced cellular senescence as a possible therapeutic strategy for non-small cell lung cancer

Tyrosine kinase (TK) inhibitors improve clinical outcomes in non-small cell lung cancer (NSCLC) with targetable mutations. However, such NSCLC cases only consist of about 50% in the western populations. This study, for the first time in NSCLC cells including those without a targetable TK mutation, explores a tumor-suppressive activity of siRNA knockdown of a splicing factor SRSF3, which was reportedly effective in other cancer cell types. The knockdown of SRSF3 increased cellular senescence, indicated by senescence-associated {beta}-galactosidase activity and reduced cell proliferation, in all NSCLC cell lines examined, including A549 (no TK mutation; TP53 wild-type), NCI-H1975 (EGFR L858R/T790M; TP53 R273H mutant), NCI-H322 (no TK mutation; TP53 R248L mutant) and NCI-H596 (no TK mutation; TP53 G245C mutant). An increase in apoptotic cleavage of caspase-3 and poly(ADP-ribose) polymerase was also observed in A549 cells. p53{beta}, a tumor-suppressive p53 isoform generated via alternative mRNA splicing, was upregulated by SRSF3 knockdown, as previously reported in normal fibroblasts. However, neither cellular senescence nor apoptosis was increased by overexpression of p53{beta}, suggesting no or minimum contribution of this p53 isoform to the tumor-suppressive activity of SRSF3 knockdown in NSCLC cells. Our gene expression assay indicated that the SRSF3 knockdown-induced senescence in NSCLC cells may be mediated by downregulation of TOP2A, UBE2C or ASPM, which are known to be oncogenic and are associated with poor patient prognosis. We also generated SRSF3 siRNA-encapsulating lipid nanoparticles as a future therapeutic tool. This study suggests a therapeutic strategy for NSCLC irrespective of the mutation status of TP53 and TK-encoding genes. SummaryKnockdown of a splicing factor SRSF3 increases cellular senescence in NSCLC cells including those with no targetable mutation of tyrosine kinases and thus may represent a novel therapeutic strategy for a hard-to-treat group of NSCLC.

cancer biology↗

bioRxiv2025

In vivo anti-FAP CAR T therapy reduces fibrosis and restores liver homeostasis in metabolic dysfunction-associated steatohepatitis

In this study, we aimed to determine the efficacy of in vivo chimeric antigen receptor (CAR) T cell therapy, generated by targeted lipid nanoparticles (t-LNPs), as an anti-fibrotic in metabolic dysfunction-associated steatotic liver disease. Hepatic fibrosis is a key predictor of mortality in liver disease, driven by fibrogenic hepatic stellate cells (HSCs). In heart, chimeric antigen receptor (CAR) T cells targeting fibroblast activation protein alpha (FAP) reduce murine cardiac fibrosis. However, the value of this approach in liver is unknown. We explored the anti-fibrotic potential of in vivo-generated anti-FAP CAR T cells in metabolic dysfunction-associated steatohepatitis (MASH), a highly prevalent disease with no approved anti-fibrotic therapies. We first established that FAP expression in both human and murine MASH is specific to HSCs. We then used flow cytometry, Sirius Red morphometry, digital pathology analysis, and single nuclear RNA-sequencing to assess the impact of anti-FAP CAR T cell therapy on murine MASH. Anti-CD5 targeted-LNPs carrying anti-FAPCAR mRNA generate activated, transient anti-FAP CAR T cells, which significantly reduced fibrosis by depleting pro-fibrogenic HSCs, and by modulating immune cells, endothelial cells and hepatocytes in a non-cell autonomous manner to mitigate inflammation and restore hepatic homeostasis. These findings reinforce the potential of in vivo CAR T therapy to attenuate a highly morbid and pervasive liver disease through integrated, multicellular salutary effects. One Sentence SummaryRNA-based treatment transiently reprograms immune cells to target scar-forming cells in fatty liver disease, thus improving liver health overall.

cell biology↗

bioRxiv2024

Lipid Nanoparticle-Associated Inflammation is Triggered by Sensing of Endosomal Damage: Engineering Endosomal Escape without Side Effects

Lipid nanoparticles (LNPs) have emerged as the dominant platform for RNA delivery, based on their success in the COVID-19 vaccines and late-stage clinical studies in other indications. However, we and others have shown that LNPs induce severe inflammation, and massively aggravate pre-existing inflammation. Here, using structure-function screening of lipids and analyses of signaling pathways, we elucidate the mechanisms of LNP-associated inflammation and demonstrate solutions. We show that LNPs hallmark feature, endosomal escape, which is necessary for RNA expression, also directly triggers inflammation by causing endosomal membrane damage. Large, irreparable, endosomal holes are recognized by cytosolic proteins called galectins, which bind to sugars on the inner endosomal membrane and then regulate downstream inflammation. We find that inhibition of galectins abrogates LNP-associated inflammation, both in vitro and in vivo. We show that rapidly biodegradable ionizable lipids can preferentially create endosomal holes that are smaller in size and reparable by the endosomal sorting complex required for transport (ESCRT) pathway. Ionizable lipids producing such ESCRT-recruiting endosomal holes can produce high expression from cargo mRNA with minimal inflammation. Finally, we show that both routes to non-inflammatory LNPs, either galectin inhibition or ESCRT-recruiting ionizable lipids, are compatible with therapeutic mRNAs that ameliorate inflammation in disease models. LNPs without galectin inhibition or biodegradable ionizable lipids lead to severe exacerbation of inflammation in these models. In summary, endosomal escape induces endosomal membrane damage that can lead to inflammation. However, the inflammation can be controlled by inhibiting galectins (large hole detectors) or by using biodegradable lipids, which create smaller holes that are reparable by the ESCRT pathway. These strategies should lead to generally safer LNPs that can be used to treat inflammatory diseases.

bioengineering↗

bioRxiv2023

Targeting lipid nanoparticles to the blood brain barrier to ameliorate acute ischemic stroke

After more than 100 failed drug trials for acute ischemic stroke (AIS), one of the most commonly cited reasons for the failure has been that drugs achieve very low concentrations in the at-risk penumbra. To address this problem, here we employ nanotechnology to significantly enhance drug concentration in the penumbras blood-brain barrier (BBB), whose increased permeability in AIS has long been hypothesized to kill neurons by exposing them to toxic plasma proteins. To devise drug-loaded nanocarriers targeted to the BBB, we conjugated them with antibodies that bind to various cell adhesion molecules on the BBB endothelium. In the transient middle cerebral artery occlusion (tMCAO) mouse model, nanocarriers targeted with VCAM antibodies achieved the highest level of brain delivery, nearly 2 orders of magnitude higher than untargeted ones. VCAM-targeted lipid nanoparticles loaded with either a small molecule drug (dexamethasone) or mRNA (encoding IL-10) reduced cerebral infarct volume by 35% or 73%, respectively, and both significantly lowered mortality rates. In contrast, the drugs delivered without the nanocarriers had no effect on AIS outcomes. Thus, VCAM-targeted lipid nanoparticles represent a new platform for strongly concentrating drugs within the compromised BBB of penumbra, thereby ameliorating AIS. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/544645v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@86762dorg.highwire.dtl.DTLVardef@1b89a2aorg.highwire.dtl.DTLVardef@916d85org.highwire.dtl.DTLVardef@1ad4670_HPS_FORMAT_FIGEXP M_FIG C_FIG Acute ischemic stroke induces upregulation of VCAM. We specifically targeted upregulated VCAM in the injured region of the brain with drug- or mRNA-loaded targeted nanocarriers. Nanocarriers targeted with VCAM antibodies achieved the highest brain delivery, nearly orders of magnitude higher than untargeted ones. VCAM-targeted nanocarriers loaded with dexamethasone and mRNA encoding IL-10 reduced infarct volume by 35% and 73%, respectively, and improved survival rates.

pharmacology and toxicology↗