Biology subjects

Nittayacharn, P.

Publications and source records attributed to Nittayacharn, P..

6 recordsLinked to original sources

bioRxiv2026

Nonviral, ultrasound-triggered gene delivery platform via gas-core cationic nanobubbles

Despite their promise, lipid nanoparticle gene delivery systems have repeatedly failed clinical trials and struggle to achieve efficient, localized transfection in target tissues. The majority of endocytosed nanoparticles are degraded before nucleic acid release, and an inability to track particle distribution in vivo prevents validation of successful delivery. Alternatively, nanobubbles (NBs) are lipid-shelled, gas-core preclinical ultrasound contrast agents and stimuli-responsive drug delivery vehicles. Under varying acoustic pressures, NBs expand, contract, and burst, releasing cargo in an externally controlled, site-specific manner while scattering unique echoes for simultaneous ultrasound visualization. Here, we introduce a cationic nanobubble (CNB) formulation with a +42.3 mV zeta potential, 265 nm diameter, and 2.43x1011 NBs/mL concentration. CNBs produce stable ultrasound contrast, electrostatically load plasmid DNA onto their surface, and internalize into >99% of human prostate cancer cells within 15 minutes in vitro. CNBs remain brightly echogenic intracellularly and induce sonication-dependent expression of green fluorescent protein (GFP). In vivo, CNBs generate contrast in mouse livers for 50 minutes after intravenous administration. Therapeutic ultrasound stimulation over the liver causes a sharp reduction in ultrasound contrast, visualizing localized cavitation in the target organ and inducing a 2.5-fold increase in anti-GFP mean fluorescence intensity relative to the untransfected control. Importantly, no GFP expression is observed without ultrasound stimulation, supporting a mechanism for selective and site-specific gene delivery. This study presents a highly stable CNB capable of efficient DNA loading and ultrasound-dependent gene expression. These results provide a foundation for the future development of CNB platforms to advance image-guided, ultrasound-triggered gene therapy.

bioengineering↗

bioRxiv2026

Ultrasound Activated Nanobubbles Induce Durable Systemic Antitumor Immunity

Clinical outcomes in aggressive breast cancer vary widely, in part because the tumor microenvironment is structured to exclude immune infiltration. Low antigen load, dysfunctional antigen-presenting cells, T cell exclusion and exhaustion, and a stiff extracellular matrix that physically restricts immune cell trafficking work together to form a suppressive barrier that current immunotherapies struggle to overcome. We addressed this barrier using ultrasound (US)-activated nanobubbles (NBs), a drug-free intervention based on perfluoropropane-filled nanoparticles. The size and deformable phospholipid shell enable NBs to achieve deep tumor penetration and a uniform distribution throughout the entire tumor. Upon ultrasound activation, NBs generate localized mechanical forces that restore extracellular matrix elasticity, disrupt tumor transport barriers, and drive HMGB1 release, re-engaging endogenous antitumor immunity without pharmacological agents. In a syngeneic triple-negative breast cancer model, US-NB treatment depleted immunosuppressive myeloid cells 3-fold within 3 hours, followed by a greater than 5-fold increase in the ratio of antigen-experienced to suppressive T cells at 48 hours. US-NB drives rapid infiltration of CD4+ and CD8+ T cells within 48 hours. US-NB treatment achieved an 85% cure rate in the D2A1 model; cured animals maintained durable systemic immune memory, rejecting both local and systemic tumor rechallenge. Consistent therapeutic benefit was observed in a luminal B-like mammary tumor model (E0771), supporting activity across breast cancer subtypes. These results establish US-NB mechanical immunomodulation as a drug-free therapeutic strategy capable of generating robust and durable antitumor immunity, acting through biophysical tissue properties rather than tumor-specific molecular targets. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=90 SRC="FIGDIR/small/714247v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@8f274eorg.highwire.dtl.DTLVardef@5da575org.highwire.dtl.DTLVardef@19defborg.highwire.dtl.DTLVardef@7e630e_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

Nanobubbles for Precision Oncology: Preclinical Evaluation of Molecular Targeted Ultrasound Contrast Agents in a Rabbit Model

The localization of prostate cancer by ultrasound remains limited by the lack of B-mode conspicuity and the confinement of clinically approved microbubbles (MBs) to the vasculature. This precludes the differentiation of viable tumor tissue, necrotic tissue, and margin-associated disease. We investigate the use of prostate-specific membrane antigen (PSMA)-targeted lipid-shelled perfluorocarbon nanobubbles (PSMA-NBs) in an orthotopic rabbit model using a clinical CEUS system. We implanted PSMA-positive PC3pip-GFP tumors into the prostates of immunosuppressed New Zealand White rabbits and performed transabdominal imaging with PSMA-NBs, MBs, and untargeted nanobubbles (Plain-NB) using the same imaging and analysis protocols. To address tumor heterogeneity and ultrasound boundary ambiguity, ROIs were defined from baseline B-mode images and segmented into the tumor core, rim, and a surrounding peritumoral area. Pixel-wise parametric and decorrelation time (DT) maps were generated and correlated with whole-slide histology (H&E) and PSMA IHC. Compared to MBs, PSMA-NBs exhibited higher peak intensities in the tumor core and rim (1.60-fold and 1.50-fold, respectively) and improved retention (mean transit time: 4.20 to 4.50-fold higher) for up to 10 minutes in the tumor and peritumoral areas. PSMA-NB kinetic and DT parameters also correlated with histology-defined tumor viability and PSMA expression. Compared to Plain-NBs, PSMA-NBs also exhibited improved retention (MTT +21%; AUCwo) in the rim and peritumoral areas. This study demonstrates the capability of PSMA-NBs to characterize prostate cancer by molecular CEUS beyond what is possible with conventional MBs.

bioengineering↗

bioRxiv2025

Enhanced delivery of lipid nanoparticle-based immunotherapy by modulating the tumor tissue stiffness using ultrasound-activated nanobubbles

Tumors often exhibit an extracellular matrix with elevated stiffness due to excessive accumulation and crosslinking of proteins, particularly collagen. This elevated stiffness acts as a physical barrier, impeding the infiltration of immune cells and the effective delivery of various immunotherapeutic agents, such as lipid nanoparticle-based RNA therapeutics. Here, we investigate the ability of ultrasound-activated nanobubbles (US-NBs) to increase the permeability and immunogenicity of tumors. Our results show that US-NBs physically remodel the tumor tissue by decreasing its stiffness by 60% five days after a single treatment. US-NB-treated tumors display randomly oriented collagen with a 5.47-fold lower deposition compared to untreated tumors. This leads to the effective delivery and widespread distribution of lipid nanoparticles (LNPs) in the tumor. When LNPs are assisted by US-NB, they have higher gene-transfection across pan-immune cells relative to LNPs alone. Notably, US-NB enables LNPs to genetically modify T cells directly in vivo. By effectively engaging both arms of the immune system, US-NB-assisted LNPs enhance the tumor immunogenicity and infiltration of cytotoxic cells by 4-fold when compared to LNPs alone. These results indicate that gentle mechanical stimulation of the tumor using US-NB offers a promising strategy to augment the delivery and efficacy of existing immunotherapies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/648512v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@da7fborg.highwire.dtl.DTLVardef@8f5f27org.highwire.dtl.DTLVardef@b002f6org.highwire.dtl.DTLVardef@19265da_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2023

Ultrasound-Mediated Drug-free Theranostics for Treatment of Prostate Cancer

RationaleLipid-shelled nanobubbles (NBs) can be visualized and activated using noninvasive ultrasound (US) stimulation, leading to significant bioeffects. We have previously shown that active targeting of NBs to prostate-specific membrane antigen (PSMA) overexpressed in prostate cancer (PCa) enhances the cellular internalization and prolongs retention of NBs with persistent acoustic activity ([~]hrs.). In this work, we hypothesized that tumor-accumulated PSMA-NBs combined with low frequency therapeutic US (TUS) will lead to selective damage and induce a therapeutic effect in PSMA-expressing tumors compared to PSMA-negative tumors. MethodsPSMA-targeted NBs were formulated by following our previously established protocol. Cellular internalization of fluorescent PSMA-NBs was evaluated by confocal imaging using late endosome/lysosome staining pre- and post-TUS application. Two animal models were used to assess the technique. Mice with dual tumors (PSMA expressing and PSMA negative) received PSMA-NB injection via the tail vein followed by TUS 1 hr. post injection (termed, targeted NB therapy or TNT). Twenty-four hours after treatment mice were euthanized and tumor cell apoptosis evaluated via TUNEL staining. Mice with single tumors (either PSMA + or -) were used for survival studies. Tumor size was measured for 80 days after four consecutive TNT treatments (every 3 days). To test the approach in a larger model, immunosuppressed rabbits with orthotopic human PSMA expressing tumors received PSMA-NB injection via the tail vein followed by TUS 30 min after injection. Tumor progression was assessed via US imaging and at the end point apoptosis was measured via TUNEL staining. ResultsIn vitro TNT studies using confocal microscopy showed that the internalized NBs and cellular compartments were disrupted after the TUS application, yet treated cells remained intact and viable. In vivo, PSMA-expressing tumors in mice receiving TNT treatment demonstrated a significantly greater extent of apoptosis (78.45 {+/-} 9.3%, p < 0.01) compared to the other groups. TNT treatment significantly inhibited the PSMA (+) tumor growth and overall survival significantly improved (median survival time increase by 103%, p < 0.001). A significant reduction in tumor progression compared to untreated control was also seen in the rabbit model in intraprostatic (90%) and in extraprostatic lesions (94%) (p = 0.069 and 0.003, respectively). ConclusionWe demonstrate for the first time the effect of PSMA-targeted nanobubble intracellular cavitation on cancer cell viability and tumor progression in two animal models. Data demonstrate that the targeted nanobubble therapy (TNT) approach relies primarily on mechanical disruption of intracellular vesicles and the resulting bioeffects appear to be more specific to target cancer cells expressing the PSMA receptor. The effect, while not lethal in vitro, resulted in significant tumor apoptosis in vivo in both a mouse and a rabbit model of PCa. While the mechanism of action of these effects is yet unclear, it is likely related to a locally-induced immune response, opening the door to future investigations in this area.

bioengineering↗

bioRxiv2021

Intracellular Vesicle Entrapment of Nanobubble Ultrasound Contrast Agents Targeted to PSMA Promotes Prolonged Enhancement and Stability In Vivo and In Vitro

Previous work has shown that active targeting of nanobubble (NB) ultrasound contrast agents to the prostate-specific membrane antigen (PSMA) significantly prolongs ultrasound signal enhancement in PSMA-expressing prostate cancer. However, the specific mechanism behind this effect is not well understood. Furthermore, prior studies were carried out using clinical ultrasound scanners in a single imaging plane. Because tumor heterogeneity can have a drastic effect on bubble kinetics and resulting contrast enhancement, a single region of interest in one imaging plane over time may not fully represent the contrast dynamics of the entire tumor. Accordingly, in the current work, we used high-frequency dynamic parametric contrast-enhanced ultrasound (DCE-US) imaging to gain a detailed understanding of NB kinetics in prostate tumors in mice. Specifically, we examined the differences in enhancement between the tumor periphery and tumor core in the same imaging plane. We also quantified intact nanobubble retention in the entire tumor volume. To better understand the mechanism behind prolonged tumor enhancement, intracellular retention and the acoustic activity of PSMA-NB were evaluated in cell culture. DCE-US US data suggest that both tumor wash-in and retention of PSMA-NB are delayed due to biomarker interaction and binding. The longer retention of PSMA-NB signal in tumor core supported target-driven bubble extravasation. In vitro studies demonstrated a higher level of internalization and prolonged-acoustic activity of internalized PSMA-NB. GC/MS analysis confirmed gas persistence in the cells after PSMA-NB internalization. The active-targeting of NB results in cellular internalization via receptor-mediated endocytosis, and the location with intracellular vesicles (late-stage endosomes/lysosomes) significantly prolongs gas retention within the cells. These features can enable background-free diagnostic imaging of the target cells/tissues, as well as highly focused ultrasound-modulated therapeutic interventions.

cancer biology↗