Rhodamine B-Conjugated Fluorescent Block Copolymer Micelles for Efficient Chlorambucil Delivery and Intracellular Imaging.

The clinical development of the anticancer drug chlorambucil (CHL) is limited by its low solubility in water, poor bioavailability, and off-target toxicity. Besides, another constraint for monitoring intracellular drug delivery is the non-fluorescent nature of CHL. Nanocarriers based on block copolymers of poly(ethylene glycol)/poly(ethylene oxide) (PEG/PEO) and poly(ε-caprolactone) (PCL) are an elegant choice for drug delivery applications due to their high biocompatibility and inherent biodegradability properties. Here, we have designed and prepared block copolymer micelles (BCM) containing CHL (BCM-CHL) from a block copolymer having fluorescent probe rhodamine B (RhB) end-groups to achieve efficient drug delivery and intracellular imaging. For this purpose, the previously reported tetraphenylethylene (TPE)-containing poly(ethylene oxide)-b-poly(ε-caprolactone) [TPE-(PEO-b-PCL)2] triblock copolymer was conjugated with RhB by a feasible and effective post-polymerization modification method. In addition, the block copolymer was obtained by a facile and efficient synthetic strategy of one-pot block copolymerization. The amphiphilicity of the resulting block copolymer TPE-(PEO-b-PCL-RhB)2 led to the spontaneous formation of micelles (BCM) in aqueous media and successful encapsulation of the hydrophobic anticancer drug CHL (CHL-BCM). Dynamic light scattering and transmission electron microscopy analyses of BCM and CHL-BCM revealed a favorable size (10-100 nm) for passive targeting of tumor tissues via the enhanced permeability and retention effect. The fluorescence emission spectrum (λex 315 nm) of BCM demonstrated Förster resonance energy transfer between TPE aggregates (donor) and RhB (acceptor). On the other hand, CHL-BCM revealed TPE monomer emission, which may be attributed to the π-π stacking interaction between TPE and CHL molecules. The in vitro drug release profile showed that CHL-BCM exhibits drug release in a sustained manner over 48 h. A cytotoxicity study proved the biocompatibility of BCM, while CHL-BCM revealed significant toxicity to cervical (HeLa) cancer cells. The inherent fluorescence of RhB in the block copolymer offered an opportunity to directly monitor the cellular uptake of the micelles by confocal laser scanning microscopy imaging. These results demonstrate the potential of these block copolymers as drug nanocarriers and as bioimaging probes for theranostic applications.

Fluorescent ABC-Triblock Polymer Nanocarrier for Cisplatin Delivery to Cancer Cells.

Monitoring intracellular administration of non-luminescent anticancer drugs like cisplatin is a very challenging task in cancer research. Perylenebisimide (PBI) chromophore tagged fluorescent ABC-triblock polycaprolactone (PCL) nanoscaffold was engineered having carboxylic acid blocks for the chemical conjugation of cisplatin at the core and hydrophilic PEG blocks at the periphery. The amphiphilic ABC triblock Pt-prodrug was self-assembled into <200 nm nanoparticles and exhibited excellent shielding against drug detoxification by the glutathione (GSH) species in the cytosol. In vitro drug release studies confirmed that the Pt-prodrug was stable at extracellular conditions and the PCL block exclusively underwent lysosomal-enzymatic biodegradation at the intracellular level to release the cisplatin drug in the active-form for accomplishing more than 90% cell growth inhibition. Confocal microscopic imaging of the red-fluorescence signals from the perylene chromophores established the simultaneous monitoring and delivery aspects of the Pt-prodrug, and the proof-of-concept was successfully demonstrated in breast and cervical cancer cell lines.

AIE-Based Fluorescent Triblock Copolymer Micelles for Simultaneous Drug Delivery and Intracellular Imaging.

Fluorescent drug delivery systems have received increasing attention in cancer therapy because they combine drug delivery and bioimaging into a single platform. For example, polymers with aggregation-induced emission (AIE) fluorophores, such as tetraphenylethylene (TPE), have emerged as an elegant choice for drug delivery/bioimaging applications. In this work, we report one-pot sequential organocatalytic ring-opening polymerization of ε-caprolactone (CL) and ethylene oxide (EO) using TPE-(OH)2 as a difunctional initiator, in the presence of a t-BuP2/TEB Lewis pair (catalyst), in THF at room temperature. Two well-defined triblock copolymers with inverse block sequences, TPE-(PCL-b-PEO)2 and TPE-(PEO-b-PCL)2, were synthesized by altering the sequential addition of CL and EO. The physicochemical properties, including hydrodynamic diameter, morphology, and AIE properties of the synthesized amphiphilic triblock copolymers were investigated in aqueous media. The block copolymer micelles were loaded with anticancer drugs doxorubicin and curcumin to serve as drug delivery vehicles. In vitro studies revealed the accelerated drug release at lower pH (5.5), which mimics the tumor microenvironment, different from the physiological pH (7.4). In vitro cytotoxicity studies demonstrated that the neat block copolymer micelles are biocompatible, while drug-loaded micelles exhibited a significant cytotoxic effect in cancer cells. Cellular uptake, examined by confocal laser scanning microscopy, showed that the block copolymer micelles were rapidly internalized by the cells with simultaneous emission of TPE fluorophore. These results suggest that these triblock copolymers can be utilized for intracellular bioimaging.

STITCHLESS Percutaneous Endoscopic Cervical Discectomy: Are We Moving Towards Day Care Discectomy Procedure?

BACKGROUND: STITCHLESS percutaneous endoscopic cervical discectomy s[PECD] is safe, precise, targeted, and a complete endoscopic procedure to treat soft cervical disc herniation with unilateral radiculopathy. It allows direct visualization of herniated fragment and its removal, inspection of decompressed nerve root in an awake and aware patient. It reduces the risk related to general anesthesia and to the neurological structures. However, all the patients treated with PECD can be candidates for anterior cervical discectomy and fusion (ACDF). ACDF requires a longer period of stay, expense, and more risk to neurological structures and ultimately loss of the disc space by fusion. MATERIALS AND METHODS: Twenty consecutively treated patients by sPECD over a period of 2 years with soft cervical disc herniation and unilateral radiculopathy were included in the study. PECD enables removal of offending fragment under vision and irrigation and ablation of inflammation with few complications. All patients were followed for minimum of 6 months with visual analog score (VAS) and neck disability index (NDI). RESULTS: All treated patients had a good outcome in terms of pain relief (VAS) and functional recovery (NDI). One patient had episodes of cough lying in the supine position and another patient had transient hoarseness of voice, (both recovered). CONCLUSION: Potential benefits of sPECD include safety as it is done under local anesthesia, smaller incision, short hospitalization, fewer complications, avoidance of fusion, preservation of segmental motion, preventing the adjacent segment degeneration, and avoidance of the risk related to the hardware (nonunion and pseudarthrosis). sPECD is an effective treatment modality for soft cervical disc herniation.

Fluorescent-Tagged Biodegradable Polycaprolactone Block Copolymer FRET Probe for Intracellular Bioimaging in Cancer Cells.

The present investigation reports a new fluorophore-tagged biodegradable polycaprolactone (PCL) block copolymer FRET-probe for intracellular imaging in cancer therapy. A hydroxyl functionalized π-conjugated oligo-phenylenevinylene (OPV) chromophore was tailor-made, and it was incorporated in a t-butyl ester substituted polycaprolactone block copolymer via ring opening polymerization. This blue-luminescent OPV-PCL triblock self-assembled as <200 nm spherical nanoparticles (FRET donor), and it encapsulated water insoluble Nile red (NR, FRET acceptor) to yield an OPV-NR FRET probe. Selective photo excitation of the OPV chromophore in block nanoassemblies enabled the excitation energy transfer from the OPV to NR and facilitated the efficient FRET process in aqueous medium. Time-correlated fluorescent decay dynamics and detailed photophysical studies were carried out to estimate the Förster distance, donor-acceptor distance, and the excitation energy transfer efficiency. These parameters confirmed the occurrence of the FRET process within the confined nanoparticle environment. The PCL chains in the FRET probe were susceptible to enzymatic biodegradation in intracellular environments, and the degradation process controlled the FRET on/off mechanism. Cytotoxicity studies revealed that the FRET probe was biocompatible and nontoxic to cells, and the FRET-probe was found to be readily taken up by the cancer cells, and it was internalized in the cytoplasm and peri-nuclear environment. Selective photoexcitation of the OPV chromophore in a confocal microscope exhibited dual emission from the FRET probe. The cancer cells exhibited blue luminescence (self-emission) with respect to the OPV chromophore (in the blue channel) and bright red-luminescence from the NR dye followed by the FRET process at the cellular level (in the red channel). The dual luminescence characteristics, biodegradation and biocompatibility, make the newly designed PCL-OPV-NR FRET probe an excellent biomedical nanodevice for bioimaging applications, and the proof-of-concept was established in cervical (HeLa) and breast cancer (MCF 7) cell lines.

Dual Functional Nanocarrier for Cellular Imaging and Drug Delivery in Cancer Cells Based on π-Conjugated Core and Biodegradable Polymer Arms.

Multipurpose polymer nanoscaffolds for cellular imaging and delivery of anticancer drug are urgently required for the cancer therapy. The present investigation reports a new polymer drug delivery concept based on biodegradable polycaprolactone (PCL) and highly luminescent π-conjugated fluorophore as dual functional nanocarrier for cellular imaging and delivery vehicles for anticancer drug to cancer cells. To accomplish this goal, a new substituted caprolactone monomer was designed, and it was subjected to ring opening polymerization using a blue luminescent bishydroxyloligo-phenylenevinylene (OPV) fluorophore as an initiator. A series of A-B-A triblock copolymer building blocks with a fixed OPV π-core and variable chain biodegradable PCL arm length were tailor-made. These triblocks self-assembled in organic solvents to produce well-defined helical nanofibers, whereas in water they produced spherical nanoparticles (size ∼150 nm) with blue luminescence. The hydrophobic pocket of the polymer nanoparticle was found to be an efficient host for loading water insoluble anticancer drug such as doxorubicin (DOX). The photophysical studies revealed that there was no cross-talking between the OPV and DOX chromophores, and their optical purity was retained in the nanoparticle assembly for cellular imaging. In vitro studies revealed that the biodegradable PCL arm was susceptible to enzymatic cleavage at the intracellular lysosomal esterase under physiological conditions to release the loaded drugs. The nascent nanoparticles were found to be nontoxic to cancer cells, whereas the DOX-loaded nanoparticles accomplished more than 80% killing in HeLa cells. Confocal microscopic analysis confirmed the cell penetrating ability of the blue luminescent polymer nanoparticles and their accumulation preferably in the cytoplasm. The DOX loaded red luminescent polymer nanoparticles were also taken up by the cells, and the drug was found to be accumulated at the perinuclear environment. The new nanocarrier approach reported in the present manuscript accomplishes both cellular imaging and delivering drugs to intracellular compartments in a single polymer system. The present investigation is one of the first examples to demonstrate the dual functional biodegradable luminescence nanocarrier concept in the literature, and the studies established this proof-of-concept in cellular imaging and drug delivery in cancer cells.