Depleting tumor-associated Tregs via nanoparticle-mediated hyperthermia to enhance anti-CTLA-4 immunotherapy.

Aim: We aim to demonstrate that a local nanoparticle-mediated hyperthermia can effectively eliminate tumor-associated Tregs and thereby boost checkpoint blockade-based immunotherapy. Materials & methods: Photothermal therapy (PTT), mediated with systemically administered stealthy iron-oxide nanoparticles, was applied to treat BALB/c mice bearing 4T1 murine breast tumors. Flow cytometry was applied to evaluate both Treg and CD8+ T-cell population. Tumor growth following combination therapy of both PTT and anti-CTLA-4 was further evaluated. Results: Our data reveal that tumor-associated Tregs can be preferentially depleted via iron-oxide nanoparticles-mediated PTT. When combining PTT with anti-CTLA-4 immunotherapy, we demonstrate a significant inhibition of syngeneic 4T1 tumor growth. Conclusion: This study offers a novel strategy to overcome Treg-mediated immunosuppression and thereby to boost cancer immunotherapy.

Self-Assembled Au@Fe Core/Satellite Magnetic Nanoparticles for Versatile Biomolecule Functionalization.

Although the functionalization of magnetic nanoparticles (MNPs) with biomolecules has been widely explored for various biological applications, achieving efficient bioconjugations with a wide range of biomolecules through a single, universal, and versatile platform remains a challenge, which may significantly impact their applications' outcomes. Here, we report a novel MNP platform composed of Au@Fe core/satellite nanoparticles (CSNPs) for versatile and efficient bioconjugations. The engineering of the CSNPs is facilely formed through the self-assembly of ultrasmall gold nanoparticles (AuNPs, 2-3 nm in diameter) around MNPs with a polysiloxane-containing polymer coating. The formation of the hybrid magnetic nanostructure is revealed by absorption spectroscopy, dynamic light scattering (DLS), transmission electron microscopy (TEM), element analysis using atomic absorption spectroscopy, and vibrating sample magnetometer. The versatility of biomolecule loading to the CSNP is revealed through the bioconjugation of a wide range of relevant biomolecules, including streptavidin, antibodies, peptides, and oligonucleotides. Characterizations including DLS, TEM, lateral flow strip assay, fluorescence assay, giant magnetoresistive nanosensor array, high-performance liquid chromatography, and absorption spectrum are performed to further confirm the efficiency of various bioconjugations to the CSNP. In conclusion, this study demonstrates that the CSNP is a novel MNP-based platform that offers versatile and efficient surface functionalization with various biomolecules.

Sulforaphane enhances the anticancer activity of taxanes against triple negative breast cancer by killing cancer stem cells.

Triple negative breast cancer (TNBC) typically exhibits rapid progression, high mortality and faster relapse rates relative to other breast cancer subtypes. In this report we examine the combination of taxanes (paclitaxel or docetaxel) with a breast cancer stem cell (CSC)-targeting agent sulforaphane for use against TNBC. We demonstrate that paclitaxel or docetaxel treatment induces IL-6 secretion and results in expansion of CSCs in TNBC cell lines. Conversely, sulforaphane is capable of preferentially eliminating CSCs, by inhibiting NF-κB p65 subunit translocation, downregulating p52 and consequent downstream transcriptional activity. Sulforaphane also reverses taxane-induced aldehyde dehydrogenase-positive (ALDH+) cell enrichment, and dramatically reduces the size and number of primary and secondary mammospheres formed. In vivo in an advanced treatment orthotopic mouse xenograft model together with extreme limiting dilution analysis (ELDA), the combination of docetaxel and sulforaphane exhibits a greater reduction in primary tumor volume and significantly reduces secondary tumor formation relative to either treatment alone. These results suggest that treatment of TNBCs with cytotoxic chemotherapy would be greatly benefited by the addition of sulforaphane to prevent expansion of and eliminate breast CSCs.

Elimination of epithelial-like and mesenchymal-like breast cancer stem cells to inhibit metastasis following nanoparticle-mediated photothermal therapy.

Increasing evidence suggesting breast cancer stem cells (BCSCs) drive metastasis and evade traditional therapies underscores a critical need to exploit the untapped potential of nanotechnology to develop innovative therapies that will significantly improve patient survival. Photothermal therapy (PTT) to induce localized hyperthermia is one of few nanoparticle-based treatments to enter clinical trials in human cancer patients, and has recently gained attention for its ability to induce a systemic immune response targeting distal cancer cells in mouse models. Here, we first conduct classic cancer stem cell (CSC) assays, both in vitro and in immune-compromised mice, to demonstrate that PTT mediated by highly crystallized iron oxide nanoparticles effectively eliminates BCSCs in translational models of triple negative breast cancer. PTT in vitro preferentially targets epithelial-like ALDH + BCSCs, followed by mesenchymal-like CD44+/CD24- BCSCs, compared to bulk cancer cells. PTT inhibits BCSC self-renewal through reduction of mammosphere formation in primary and secondary generations. Secondary implantation in NOD/SCID mice reveals the ability of PTT to impede BCSC-driven tumor formation. Next, we explore the translational potential of PTT using metastatic and immune-competent mouse models. PTT to inhibit BCSCs significantly reduces metastasis to the lung and lymph nodes. In immune-competent BALB/c mice, PTT effectively eliminates ALDH + BCSCs. These results suggest the feasibility of incorporating PTT into standard clinical treatments such as surgery to enhance BCSC destruction and inhibit metastasis, and the potential of such combination therapy to improve long-term survival in patients with metastatic breast cancer.

Facile Fabrication of Near-Infrared-Resonant and Magnetic Resonance Imaging-Capable Nanomediators for Photothermal Therapy.

Although many techniques exist for fabricating near-infrared (NIR)-resonant and magnetic resonance imaging (MRI)-capable nanomediators for photothermal cancer therapy, preparing them in an efficient and scalable process remains a significant challenge. In this report, we exploit one-step siloxane chemistry to facilely conjugate NIR-absorbing satellites onto a well-developed polysiloxane-containing polymer-coated iron oxide nanoparticle (IONP) core to generate dual functional core-satellite nanomediators for photothermal therapy. An advantage of this nanocomposite design is the variety of potential satellites that can be simply attached to impart NIR resonance, which we demonstrate using NIR-resonant gold sulfide nanoparticles (Au2SNPs) and the NIR dye IR820 as two example satellites. The core-satellite nanomediators are fully characterized by using absorption spectra, dynamic light scattering, ζ potential measurements, and transmission electron microscopy. The enhanced photothermal effect under the irradiation of NIR laser light is identified through in vitro solutions and in vivo mice studies. The MRI capabilities as contrast agents are demonstrated in mice. Our data suggest that polysiloxane-containing polymer-coated IONPs can be used as a versatile platform to build such dual functional nanomediators for translatable, MRI-guided photothermal cancer therapy.

Thiol-reactive amphiphilic block copolymer for coating gold nanoparticles with neutral and functionable surfaces.

Nanoparticles designed for biomedical applications are often coated with polymers containing reactive functional groups, such as -COOH and -NH2, to conjugate targeting ligands or drugs. However, introducing highly charged surfaces promotes binding of the nanoparticles to biomolecules in biological systems through ionic interactions, causing the nanoparticles to aggregate in biological environments and consequently undergo strong non-specific binding to off-target cells and tissues. Developing a unique polymer with neutral surfaces that can be further functionalized directly would be critical to develop suitable nanomaterials for nanomedicine. Here, we report a thiol-reactive amphiphilic block copolymer poly(ethylene oxide)-block-poly(pyridyldisulfide ethylmeth acrylate) (PEO-b-PPDSM) for coating gold nanoparticles (AuNPs). The resultant polymer-coated AuNPs have almost neutral surfaces with slightly negative zeta potentials from -10 to 0 mV over a wide pH range from 2 to 12. Although the zeta potential is close to zero we show that the PEO-b-PPDSM copolymer-coated AuNPs have both good stability in various physiological conditions and reduced non-specific adsorption of proteins/biomolecules. Because of the multiple pyridyldisulfide groups on the PPDSM block, these individually dispersed nanocomplexes with an overall hydrodynamic size around 43.8 nm can be directly functionalized via disulfide-thiol exchange chemistry.

Noninvasive fluorescence resonance energy transfer imaging of in vivo premature drug release from polymeric nanoparticles.

Understanding in vivo drug release kinetics is critical for the development of nanoparticle-based delivery systems. In this study, we developed a fluorescence resonance energy transfer (FRET) imaging approach to noninvasively monitor in vitro and in vivo cargo release from polymeric nanoparticles. The FRET donor dye (DiO or DiD) and acceptor dye (DiI or DiR) were individually encapsulated into poly(ethylene oxide)-b-polystyrene (PEO-PS) nanoparticles. When DiO (donor) nanoparticles and DiI (acceptor) nanoparticles were coincubated with cancer cells for 2 h, increased FRET signals were observed from cell membranes, suggesting rapid release of DiO and DiI to cell membranes. Similarly, increased FRET ratios were detected in nude mice after intravenous coadministration of DiD (donor) nanoparticles and DiR (acceptor) nanoparticles. In contrast, another group of nude mice i.v. administrated with DiD/DiR coloaded nanoparticles showed decreased FRET ratios. Based on the difference in FRET ratios between the two groups, in vivo DiD/DiR release half-life from PEO-PS nanoparticles was determined to be 9.2 min. In addition, it was observed that the presence of cell membranes facilitated burst release of lipophilic cargos while incorporation of oleic acid-coated iron oxide into PEO-PS nanoparticles slowed the release of DiD/DiR to cell membranes. The developed in vitro and in vivo FRET imaging techniques can be used to screening stable nanoformulations for lipophilic drug delivery.

Activation of an IL6 inflammatory loop mediates trastuzumab resistance in HER2+ breast cancer by expanding the cancer stem cell population.

Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain elusive. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrate that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of the cancer stem cell (CSC) population. Long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies demonstrate that trastuzumab resistance may be mediated by an IL6 inflammatory loop and suggest that blocking this loop may provide alternative strategy to overcome trastuzumab resistance.

Applications of human pharmacokinetic prediction in first-in-human dose estimation.

Quantitative estimations of first-in-human (FIH) doses are critical for phase I clinical trials in drug development. Human pharmacokinetic (PK) prediction methods have been developed to project the human clearance (CL) and bioavailability with reasonable accuracy, which facilitates estimation of a safe yet efficacious FIH dose. However, the FIH dose estimation is still very challenging and complex. The aim of this article is to review the common approaches for FIH dose estimation with an emphasis on PK-guided estimation. We discuss 5 methods for FIH dose estimation, 17 approaches for the prediction of human CL, 6 methods for the prediction of bioavailability, and 3 tools for the prediction of PK profiles. This review may serve as a practical protocol for PK- or pharmacokinetic/pharmacodynamic-guided estimation of the FIH dose.