Synergistic Targeting of Cell Membrane, Cytoplasm, and Nucleus of Cancer Cells Using Rod-Shaped Nanoparticles

Design of carriers for effective delivery and targeting of drugs to cellular and subcellular compartments is an unmet need in medicine. Here, we report pure drug nanoparticles comprising camptothecin (CPT), trastuzumab (TTZ), and doxorubicin (DOX) to enable cell-specific interactions, subcellular accumulation, and growth inhibition of breast cancer cells. CPT is formulated in the form of nanorods which are coated with TTZ. DOX is encapsulated in the TTZ corona around the CPT nanoparticle. Our results show that TTZ/DOX-coated CPT nanorods exhibit cell-specific internalization in BT-474 breast cancer cells, after which TTZ is recycled to the plasma membrane, leaving CPT nanorods in the perinuclear region and delivering DOX into the nucleus of the cells. The effects of CPT-TTZ-DOX nanoparticles on growth inhibition are synergistic (combination index = 0.17 ± 0.03) showing 10–10 000-fold lower inhibitory concentrations (IC₅₀) compared to those of individual drugs. The design of antibody-targeted pure drug nanoparticles offers a promising design strategy to facilitate intracellular delivery and therapeutic efficiency of anticancer drugs

Bovine insulin transport across short-term 3-day Caco-2 monolayers.

<p>Transport of active peptides was measured across the confluent Caco-2 monolayers at different loading concentrations. 0.05, 0.15, 0.6, and 1.0 mg of bovine insulin was loaded onto Caco-2 monolayer in apical chambers by dissolving the same in the culture medium. Apical-to-basolateral permeability was determined by analyzing the samples collected from basolateral chambers at different time points for up to 5 hrs. TEER values were also measured to account for the integrity of the monolayer during the experiment. Bovine insulin concentrations were measured by commercially available ELISA kits, as mentioned in the Methods section. (<b>a</b>) TEER values of Caco-2 monolayer at different loading concentrations of 0.05 (circles), 0.15 (triangles), 0.6 (diamonds), and 1.0 (crosses) mg/well of bovine insulin. (<b>b</b>) Cumulative amount of insulin transported (µg) to the basolateral chamber at different time-points at different loading concentrations of 0.05 (open circles), 0.15 (filled circles), 0.6 (filled squares), and 1.0 (open squares) mg/well. Data represent mean±SD (n = 3).</p

Permeability values under various conditions tested in this study.

<p>Data represent mean±SD (n = 3).</p

Transport of salmon Calcitonin (sCT) across Caco-2 monolayers.

<p>Permeation of salmon Calcitonin was measured at different loading concentrations. Apical-to-basolateral permeability was measured by analyzing the amount of sCT present in basolateral chamber at different time-points. TEER values were measured to validate integrity of the monolayers. sCT concentrations were measured by commercially available ELISA kits, as mentioned in the Methods section. (<b>a</b>) TEER values of Caco-2 monolayer at different sCT loading concentrations of 5.0 (circles) and 24.0 (triangles) µg/well. (<b>b</b>) Cumulative amount of calcitonin transported (µg) to the basolateral chamber at different time-points at different loading concentrations of 5.0 (open circles) and 24.0 (filled circles) µg/well. Data represent mean±SD (n = 3).</p

FITC-insulin transport across Caco-2 monolayers.

<p>(<b>a</b>) Time-course study of FITC-insulin transport (mg) at different loading concentrations. FITC-insulin was loaded in apical chambers at 0.05 (open circles), 0.15 (filled circles), 0.3 (squares), and 0.6 (triangles) mg/well respectively; and permeation was measured by measuring the fluorescence in samples collected from basolateral chamber at different time-points up to 5 hrs. (<b>b</b>) % FITC-insulin transport across Caco-2 monolayers. Data represent mean±SD (n = 3).</p

Sulforhodamine-B transport across Caco-2 monolayers.

<p>(<b>a</b>) Time-course study of sulforhodamine-B transport (mg) at different loading concentrations. Sulforhodamine-B was loaded in apical chambers at 0.05 (open circles), 0.15 (filled circles), 0.3 (squares), and 0.6 (triangles) mg/well respectively; and apical-to-basolateral permeation was measured by measuring the fluorescence in samples collected from basolateral chamber at different time-points up to 5 hrs. (<b>b</b>) % Sulforhodamine-B transport across Caco-2 monolayers over of 5 hrs of incubation. Data represent mean±SD (n = 3).</p

Transport of exenatide across Caco-2 monolayers.

<p>Apical-to-basolateral permeability of exenatide was measured at different apical loading. TEER values were determined to ensure monolayer integrity. (<b>a</b>) TEER values of Caco-2 monolayer following apical loading of different concentrations of exenatide at 0.3 (circles), 1.0 (triangles), 3.0 (diamonds), and 9.0 (crosses) µg/well. (<b>b</b>) Cumulative transport of exenatide (µg) to basolateral chamber during the experiment at different apical loading concentrations of 0.3 (filled circles), 1.0 (open circles), 3.0 (squares), and 9.0 (triangles) µg/well. Data represent mean±SD (n = 3).</p

MoS₂ Field-Effect Transistor for Next-Generation Label-Free Biosensors

Biosensors based on field-effect transistors (FETs) have attracted much attention, as they offer rapid, inexpensive, and label-free detection. While the low sensitivity of FET biosensors based on bulk 3D structures has been overcome by using 1D structures (nanotubes/nanowires), the latter face severe fabrication challenges, impairing their practical applications. In this paper, we introduce and demonstrate FET biosensors based on molybdenum disulfide (MoS₂), which provides extremely high sensitivity and at the same time offers easy patternability and device fabrication, due to its 2D atomically layered structure. A MoS₂-based pH sensor achieving sensitivity as high as 713 for a pH change by 1 unit along with efficient operation over a wide pH range (3–9) is demonstrated. Ultrasensitive and specific protein sensing is also achieved with a sensitivity of 196 even at 100 femtomolar concentration. While graphene is also a 2D material, we show here that it cannot compete with a MoS₂-based FET biosensor, which surpasses the sensitivity of that based on graphene by more than 74-fold. Moreover, we establish through theoretical analysis that MoS₂ is greatly advantageous for biosensor device scaling without compromising its sensitivity, which is beneficial for single molecular detection. Furthermore, MoS₂, with its highly flexible and transparent nature, can offer new opportunities in advanced diagnostics and medical prostheses. This unique fusion of desirable properties makes MoS₂ a highly potential candidate for next-generation low-cost biosensors

Skin Delivery of Hydrophilic Biomacromolecules Using Marine Sponge Spicules

We report the development of sponge <i>Haliclona</i> sp. spicules, referred to as SHS, and its topical application in skin delivery of hydrophilic biomacromolecules, a series of fluorescein isothiocyanate-dextrans (FDs). SHS are silicious oxeas which are sharp-edged and rod-shaped (∼120 μm in length and ∼7 μm in diameter). SHS can physically disrupt skin in a dose-dependent manner and retain within the skin over at least 72 h, which allows sustained skin penetration of hydrophilic biomacromolecules. The magnitude of enhancement of FD delivery into skin induced by SHS treatment was dependent on its molecular weight. Specifically, SHS topical application enhanced FD-10 (MW: 10 kDa) penetration into porcine skin <i>in vitro</i> by 33.09 ± 7.16-fold compared to control group (<i>p</i> < 0.01). SHS dramatically increased the accumulation of FD-10 into and across the dermis by 62.32 ± 13.48-fold compared to the control group (<i>p</i> < 0.01). <i>In vivo</i> experiments performed using BALB/c mice also confirmed the effectiveness of SHS topical application; the skin absorption of FD-10 with SHS topical application was 72.14 ± 48.75-fold (<i>p</i> < 0.05) and 15.39 ± 9.91-fold (<i>p</i> < 0.05) higher than those from the PBS and Dermaroller microneedling, respectively. Further, skin irritation study and transepidermal water loss (TEWL) measurement using guinea pig skin <i>in vivo</i> indicated that skin disruption induced by SHS treatment is self-limited and can be recovered with time and efficiently. SHS can offer a safe, effective, and sustained skin delivery of hydrophilic biomacromolecules and presents a promising platform technology for a wide range of cosmetic and medical applications

Synthesis of Oil-Laden Poly(ethylene glycol) Diacrylate Hydrogel Nanocapsules from Double Nanoemulsions

Multiple emulsions have received great interest due to their ability to be used as templates for the production of multicompartment particles for a variety of applications. However, scaling these complex droplets to nanoscale dimensions has been a challenge due to limitations on their fabrication methods. Here, we report the development of oil-in-water-in-oil (O₁/W/O₂) double nanoemulsions <i>via</i> a two-step high-energy method and their use as templates for complex nanogels comprised of inner oil droplets encapsulated within a hydrogel matrix. Using a combination of characterization methods, we determine how the properties of the nanogels are controlled by the size, stability, internal morphology, and chemical composition of the nanoemulsion templates from which they are formed. This allows for identification of compositional and emulsification parameters that can be used to optimize the size and oil encapsulation efficiency of the nanogels. Our templating method produces oil-laden nanogels with high oil encapsulation efficiencies and average diameters of 200–300 nm. In addition, we demonstrate the versatility of the system by varying the types of inner oil, the hydrogel chemistry, the amount of inner oil, and the hydrogel network cross-link density. These nontoxic oil-laden nanogels have potential applications in food, pharmaceutical, and cosmetic formulations