Nucleus-Targeted, Echogenic Polymersomes for Delivering a Cancer Stemness Inhibitor to Pancreatic Cancer Cells.

Chemotherapeutic agents for treating cancers show considerable side effects, toxicity, and drug resistance. To mitigate the problems, we designed nucleus-targeted, echogenic, stimuli-responsive polymeric vesicles (polymersomes) to transport and subsequently release the encapsulated anticancer drugs within the nuclei of pancreatic cancer cells. We synthesized an alkyne-dexamethasone derivative and conjugated it to N3-polyethylene glycol (PEG)-polylactic acid (PLA) copolymer employing the Cu2+ catalyzed "Click" reaction. We prepared polymersomes from the dexamethasone-PEG-PLA conjugate along with a synthesized stimuli-responsive polymer PEG-S-S-PLA. The dexamethasone group dilates the nuclear pore complexes and transports the vesicles to the nuclei. We designed the polymersomes to release the encapsulated drugs in the presence of a high concentration of reducing agents in the nuclei of pancreatic cancer cells. We observed that the nucleus-targeted, stimuli-responsive polymersomes released 70% of encapsulated contents in the nucleus-mimicking environment in 80 min. We encapsulated the cancer stemness inhibitor BBI608 in the vesicles and observed that the BBI608 encapsulated polymersomes reduced the viability of the BxPC3 cells to 43% in three-dimensional spheroid cultures. The polymersomes were prepared following a special protocol so that they scatter ultrasound, allowing imaging by a medical ultrasound scanner. Therefore, these echogenic, targeted, stimuli-responsive, and drug-encapsulated polymersomes have the potential for trackable, targeted carrier of chemotherapeutic drugs to cancer cell nuclei.

Tissue-Penetrating, Hypoxia-Responsive Echogenic Polymersomes For Drug Delivery To Solid Tumors.

Hypoxia in solid tumors facilitates the progression of the disease, develops resistance to chemo and radiotherapy, and contributes to relapse. Due to the lack of tumor penetration, most of the reported drug carriers are unable to reach the hypoxic niches of the solid tumors. We have developed tissue-penetrating, hypoxia-responsive echogenic polymersomes to deliver anticancer drugs to solid tumors. The polymersomes are composed of a hypoxia-responsive azobenzene conjugated and a tissue penetrating peptide functionalized polylactic acid-polyethylene glycol polymer. The drug-encapsulated, hypoxia-responsive polymersomes substantially decreased the viability of pancreatic cancer cells in spheroidal cultures. Under normoxic conditions, polymersomes were echogenic at diagnostic ultrasound frequencies but lose the echogenicity under hypoxia. In-vivo imaging studies with xenograft mouse model further confirmed the ability of the polymersomes to target, penetrate, and deliver the encapsulated contents in hypoxic pancreatic tumor tissues.

Peptide-targeted, stimuli-responsive polymersomes for delivering a cancer stemness inhibitor to cancer stem cell microtumors.

Often cancer relapses after an initial response to chemotherapy because of the tumor's heterogeneity and the presence of progenitor stem cells, which can renew. To overcome drug resistance, metastasis, and relapse in cancer, a promising approach is the inhibition of cancer stemness. In this study, the expression of the neuropilin-1 receptor in both pancreatic and prostate cancer stem cells was identified and targeted with a stimuli-responsive, polymeric nanocarrier to deliver a stemness inhibitor (napabucasin) to cancer stem cells. Reduction-sensitive amphiphilic block copolymers PEG1900-S-S-PLA6000 and the N3-PEG1900-PLA6000 were synthesized. The tumor penetrating iRGD peptide-hexynoic acid conjugate was linked to the N3-PEG1900-PLA6000 polymer via a Cu2+ catalyzed "Click" reaction. Subsequently, this peptide-polymer conjugate was incorporated into polymersomes for tumor targeting and tissue penetration. We prepared polymersomes containing 85% PEG1900-S-S-PLA6000, 10% iRGD-polymer conjugate, and 5% DPPE-lissamine rhodamine dye. The iRGD targeted polymersomes encapsulating the cancer stemness inhibitor napabucasin were internalized in both prostate and pancreatic cancer stem cells. The napabucasin encapsulated polymersomes significantly (p < .05) reduced the viability of both prostate and pancreatic cancer stem cells and decreased the stemness protein expression notch-1 and nanog compared to the control and vesicles without any drug. The napabucasin encapsulated polymersome formulations have the potential to lead to a new direction in prostate and pancreatic cancer therapy by penetrating deeply into the tumors, releasing the encapsulated stemness inhibitor, and killing cancer stem cells.

Acoustic Characterization of Echogenic Polymersomes Prepared From Amphiphilic Block Copolymers.

Polymersomes are a class of artificial vesicles prepared from amphiphilic polymers. Like lipid vesicles (liposomes), they too can encapsulate hydrophilic and hydrophobic drug molecules in the aqueous core and the hydrophobic bilayer respectively, but are more stable than liposomes. Although echogenic liposomes have been widely investigated for simultaneous ultrasound imaging and controlled drug delivery, the potential of the polymersomes remains unexplored. We prepared two different echogenic polymersomes from the amphiphilic copolymers polyethylene glycol-poly-DL-lactic acid (PEG-PLA) and polyethylene glycol-poly-L-lactic acid (PEG-PLLA), incorporating multiple freeze-dry cycles in the synthesis protocol to ensure their echogenicity. We investigated acoustic behavior with potential applications in biomedical imaging. We characterized the polymeric vesicles acoustically with three different excitation frequencies of 2.25, 5 and 10 MHz at 500 kPa. The polymersomes exhibited strong echogenicity at all three excitation frequencies (about 50- and 25-dB enhancements in fundamental and subharmonic, respectively, at 5-MHz excitation from 20 µg/mL polymers in solution). Unlike echogenic liposomes, they emitted strong subharmonic responses. The scattering results indicated their potential as contrast agents, which was also confirmed by clinical ultrasound imaging.

Prostate-Specific Membrane Antigen Targeted Polymersomes for Delivering Mocetinostat and Docetaxel to Prostate Cancer Cell Spheroids.

Prostate cancer cells overexpress the prostate-specific membrane antigen (PSMA) receptors on the surface. Targeting the PSMA receptor creates a unique opportunity for drug delivery. Docetaxel is a Food and Drug Administration-approved drug for treating metastatic and androgen-independent prostate cancer, and mocetinostat is a potent inhibitor of class I histone deacetylases. In this study, we prepared reduction-sensitive polymersomes presenting folic acid on the surface and encapsulating either docetaxel or mocetinostat. The presence of folic acid allowed efficient targeting of the PSMA receptor and subsequent internalization of the polymeric vesicles in cultured LNCaP prostate cancer cell spheroids. The intracellular reducing agents efficiently released docetaxel and mocetinostat from the polymersomes. The combination of the two drug-encapsulated polymersome formulations significantly (p < 0.05) decreased the viability of the LNCaP cells (compared to free drugs or control) in three-dimensional spheroid cultures. The calculated combination index value indicated a synergistic effect for the combination of mocetinostat and docetaxel. Thus, our PSMA-targeted drug-encapsulated polymersomes has the potential to lead to a new direction in prostate cancer therapy that decreases the toxicity and increases the efficacy of the drug delivery systems.

Biomarkers and Targeted Therapy in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) constitutes 90% of pancreatic cancers. PDAC is a complex and devastating disease with only 1%-3% survival rate in five years after the second stage. Treatment of PDAC is complicated due to the tumor microenvironment, changing cell behaviors to the mesenchymal type, altered drug delivery, and drug resistance. Considering that pancreatic cancer shows early invasion and metastasis, critical research is needed to explore different aspects of the disease, such as elaboration of biomarkers, specific signaling pathways, and gene aberration. In this review, we highlight the biomarkers, the fundamental signaling pathways, and their importance in targeted drug delivery for pancreatic cancers.

Sequential culture on biomimetic nanoclay scaffolds forms three-dimensional tumoroids.

In recent times, the limitation of two-dimensional cultures and complexity of in vivo models has paved the way for the development of three-dimensional models for studying cancer. Here we report the development of a new tumor model using PCL/HAPClay scaffolds seeded with a sequential culture of human mesenchymal stem cells (hMSCs) followed by human prostate cancer cells (HPCCs). This nanocomposite system is used as a test-bed for studying cancer metastasis and efficacy of anti-cancer drugs using a polymersome delivery method. A novel sequential cell culture system in three-dimensional in vitro bone model provides a unique bone mimetic environment. The hMSCs seeded scaffolds are seeded with prostate cancer cells after the hMSCs have differentiated into osteoblasts. Sequential culture on the scaffolds has shown formation of tumoroids or microtissue consisting of organized, densely packed round cells with hypoxic core regions similar to in vivo tumors. Such tumoroids are not observed on HPCC seeded scaffolds or when HPCCs sequentially cultured with human osteoblast cells. Clearly, the newly differentiated hMSCs play a vital role in the ability of cancer cells to grow into tumoroids and cause disease. The PCL/HAPclay scaffold system seeded with the sequential culture of hMSCs, and HPCCs presents a good model system for study of the interactions between prostate cancer cells and bone microenvironment. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1591-1602, 2016.