Charles J. Bowerman

Future of the Particle Replication in Nonwetting Templates (PRINT) Technology

Particle replication in nonwetting templates (PRINT) is a continuous, roll-to-roll, high-resolution molding technology which allows the design and synthesis of precisely defined micro- and nanoparticles. This technology adapts the lithographic techniques from the microelectronics industry and marries these with the roll-to-roll processes from the photographic film industry to enable researchers to have unprecedented control over particle size, shape, chemical composition, cargo, modulus, and surface properties. In addition, PRINT is a GMP-compliant (GMP=good manufacturing practice) platform amenable for particle fabrication on a large scale. Herein, we describe some of our most recent work involving the PRINT technology for application in the biomedical and material sciences.

Scalable Manufacture of Built-to-Order Nanomedicine: Spray-assisted Layer-by-Layer Functionalization of PRINT® Nanoparticles

Scalable methods, PRINT particle fabrication, and spray-assisted Layer-by-Layer deposition are combined to generate uniform and functional nanotechnologies with precise control over composition, size, shape, and surface functionality. A modular and tunable approach towards design of built-to-order nanoparticle systems, spray coating on PRINT particles is demonstrated to achieve technologies capable of targeted interactions with cancer cells for applications in drug delivery.

Particle Replication in Nonwetting Templates Nanoparticles with Tumor Selective Alkyl Silyl Ether Docetaxel Prodrug Reduces Toxicity

Delivery systems designed to have triggered release after passively targeting the tumor may improve small molecule chemotherapeutic delivery. Particle replication in nonwetting templates was used to prepare nanoparticles to passively target solid tumors in an A549 subcutaneous xenograft model. An acid labile prodrug was delivered to minimize systemic free docetaxel concentrations and improve tolerability without compromising efficacy.

Metronomic Docetaxel in PRINT Nanoparticles and EZH2 Silencing Have Synergistic Antitumor Effect in Ovarian Cancer

The purpose of this study was to investigate the antitumor effects of a combination of metronomic doses of a novel delivery vehicle, PLGA-PRINT nanoparticles containing docetaxel, and antiangiogenic mEZH2 siRNA incorporated into chitosan nanoparticles. In vivo dose-finding studies and therapeutic experiments were conducted in well-established orthotopic mouse models of epithelial ovarian cancer. Antitumor effects were determined on the basis of reduction in mean tumor weight and number of metastatic tumor nodules in the animals. The tumor tissues from these in vivo studies were stained to evaluate the proliferation index (Ki67), apoptosis index (cleaved caspase 3), and microvessel density (CD31). The lowest dose of metronomic regimen (0.5 mg/kg) resulted in significant reduction in tumor growth. The combination of PLGA-PRINT-docetaxel and CH-mEZH2 siRNA showed significant antitumor effects in the HeyA8 and SKOV3ip1 tumor models (P < 0.05). Individual as well as combination therapies showed significant antiangiogenic, antiproliferative, and proapoptotic effects, and combination therapy had additive effects. Metronomic delivery of PLGA-PRINT-docetaxel combined with CH-mEZH2 siRNA has significant antitumor activity in preclinical models of ovarian cancer. Mol Cancer Ther; 13(7); 1750–7. ©2014 AACR.

Efficacy and pharmacokinetics of a modified acid-labile docetaxel-PRINT® nanoparticle formulation against non-small-cell lung cancer brain metastases

AIM Particle Replication in Nonwetting Templates (PRINT(®)) PLGA nanoparticles of docetaxel and acid-labile C2-dimethyl-Si-Docetaxel were evaluated with small molecule docetaxel as treatments for non-small-cell lung cancer brain metastases. MATERIALS & METHODS Pharmacokinetics, survival, tumor growth and mice weight change were efficacy measures against intracranial A549 tumors in nude mice. Treatments were administered by intravenous injection. RESULTS Intracranial tumor concentrations of PRINT-docetaxel and PRINT-C2-docetaxel were 13- and sevenfold greater, respectively, than SM-docetaxel. C2-docetaxel conversion to docetaxel was threefold higher in intracranial tumor as compared with nontumor tissues. PRINT-C2-docetaxel increased median survival by 35% with less toxicity as compared with other treatments. CONCLUSION The decreased toxicity of the PRINT-C2-docetaxel improved treatment efficacy against non-small-cell lung cancer brain metastasis.

Docetaxel-Loaded PLGA Nanoparticles Improve Efficacy in Taxane-Resistant Triple-Negative Breast Cancer

Novel treatment strategies, including nanomedicine, are needed for improving management of triple-negative breast cancer. Patients with triple-negative breast cancer, when considered as a group, have a worse outcome after chemotherapy than patients with breast cancers of other subtypes, a finding that reflects the intrinsically adverse prognosis associated with the disease. The aim of this study was to improve the efficacy of docetaxel by incorporation into a novel nanoparticle platform for the treatment of taxane-resistant triple-negative breast cancer. Rod-shaped nanoparticles encapsulating docetaxel were fabricated using an imprint lithography based technique referred to as Particle Replication in Nonwetting Templates (PRINT). These rod-shaped PLGA-docetaxel nanoparticles were tested in the C3(1)-T-antigen (C3Tag) genetically engineered mouse model (GEMM) of breast cancer that represents the basal-like subtype of triple-negative breast cancer and is resistant to therapeutics from the taxane family. This GEMM recapitulates the genetics of the human disease and is reflective of patient outcome and, therefore, better represents the clinical impact of new therapeutics. Pharmacokinetic analysis showed that delivery of these PLGA-docetaxel nanoparticles increased docetaxel circulation time and provided similar docetaxel exposure to tumor compared to the clinical formulation of docetaxel, Taxotere. These PLGA-docetaxel nanoparticles improved tumor growth inhibition and significantly increased median survival time. This study demonstrates the potential of nanotechnology to improve the therapeutic index of chemotherapies and rescue therapeutic efficacy to treat nonresponsive cancers.

Preparation and biological evaluation of synthetic and polymer-encapsulated congeners of the antitumor agent pactamycin: Insight into functional group effects and biological activity

The synthesis and biological analysis of a number of novel congeners of the aminocyclopentitol pactamycin is described. Specific attention was paid to the preparation of derivatives at crucial synthetic branch points of the parent structure, and biological assays revealed a number of insights into the source of pactamycins biological activity. Additionally, the encapsulation of pactamycin and select derivatives into the PRINT© nanoparticle technology was investigated as a proof-of-concept, and evidence of bioactivity modulation through nanoparticle delivery is demonstrated. This work has provided heretofore unrealized access to a large number of novel compounds for further evaluation.

A versatile acid-labile linker for antibody–drug conjugates

Antibody drug conjugates (ADC) couple therapeutic monoclonal antibodies (mAb) with potent toxins through a linker that is stable within systemic circulation, but cleaves within the target cells. In this report, silyl ether chemistry was used to couple the mAb trastuzumab with the chemotherapeutic, gemcitabine, to demonstrate the use of silyl ethers as a potential linker for ADCs.

Abstract 5403: Metronomic docetaxel in PRINT nanoparticles and EZH2 silencing have synergistic antitumor effect in ovarian cancer

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Purpose: The purpose of this study was to investigate the combined antitumor effects of metronomic docetaxel-containing PLGA-PRINT nanoparticles and antiangiogenic mEZH2 siRNA incorporated into chitosan nanoparticles (CH-mEZH2 siRNA). Method: In vivo dose-finding studies and therapeutic experiments with PLGA-PRINT-docetaxel and CH-mEZH2 siRNA were conducted in well-established orthotopic mouse models of ovarian cancer (HeyA8 and SKOV3ip1). Antitumor effects were quantified through mean tumor weight and tumor nodule counts. Tumor tissues from these studies were stained to evaluate the proliferation index (Ki67), apoptosis index (cleaved caspase-3), and microvessel density (CD31). Result: Dose-finding studies for PLGA-PRINT-docetaxel revealed all doses (0.5, 1.5, 2 mg/kg for metronomic and 20mg/kg for MTD) significantly reduced tumor burden (p<0.05 for MTD; p<0.01 for metronomic doses). The lowest dose (0.5 mg/kg) reduced tumor weight by ∼80% and was selected for subsequent therapeutic studies. We then tested whether combining PLGA-PRINT-docetaxel particles with CH-mEZH2 siRNA would enhance antitumor effect. CH-mEZH2 siRNA alone reduced tumor weight by ∼70% in both models (p<0.01 for HeyA8; p<0.05 for SKOV3ip1). PLGA-PRINT-docetaxel treatment reduced tumor weight by ∼80% (p<0.01). Combination therapy showed the greatest therapeutic benefit with ∼95% reduction in tumor weight in both models (p<0.001 in HeyA8; p<0.01 in SKOV3ip1). In both the models, the combination therapy resulted in the greatest reduction of tumor nodules. Individual as well as combination therapies showed a significant reduction in proliferation (p<0.001). Metronomic PLGA-PRINT-docetaxel and CH-mEZH2 siRNA monotherapy increased the apoptotic index significantly (p<0.05), whereas the combination therapy had an additive effect (p<0.001). CH-mEZH2 siRNA as well as metronomic PLGA-PRINT-docetaxel therapy resulted in significant reduction in MVD (p<0.01). Conclusion: Metronomic delivery of PLGA-PRINT-docetaxel combined with CH-mEZH2 siRNA is an attractive therapeutic strategy. Citation Format: Kshipra M. Gharpure, Kevin S. Chu, Charles Bowerman, Takahito Miyake, Sunila Pradeep, Lingegowda S. Mangala, Hee-Dong Han, Rajesha Rupaimoole, Sherry Y. Wu, Heather J. Dalton, Mary E. Napier, Gabriel Lopez-Berestein, Joseph M. DeSimone, Anil K. Sood. Metronomic docetaxel in PRINT nanoparticles and EZH2 silencing have synergistic antitumor effect in ovarian cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5403. doi:10.1158/1538-7445.AM2014-5403