Multiple Therapeutic Agents Research Articles

Inside Cover: A Fragmenting Hybrid Approach for Targeted Delivery of Multiple Therapeutic Agents to the Malaria Parasite (ChemMedChem 3/2011)

Inside Cover: A Fragmenting Hybrid Approach for Targeted Delivery of Multiple Therapeutic Agents to the Malaria Parasite (ChemMedChem 3/2011)

A Fragmenting Hybrid Approach for Targeted Delivery of Multiple Therapeutic Agents to the Malaria Parasite

Artemisinin combination therapies (ACT) represent the current standard of care in the treatment of uncomplicated malaria. The widespread adoption of ACT has been motivated by a desire to minimize the emergence of drug resistance and to address the problem of recrudescence associated with artemisinin monotherapy.1–4 We set out to explore a single-molecule ‘fragmenting hybrid’ strategy in which an artemisinin-like peroxide is employed to deliver a partner drug, only upon activation by ferrous iron in the parasite. In principle, iron(II)-dependent drug delivery from a fragmenting hybrid could alleviate unwanted off-target bioactivities of the partner drug, which would be inactive in its hybrid form.

Drug delivery systems for differential release in combination therapy

Introduction: Combination therapy with multiple therapeutic agents has wide applicability in medical and surgical treatment, especially in the treatment of cancer. Thus, new drug delivery systems that can differentially release two or more drugs are desired. Utilizing new techniques to engineer the established drug delivery systems and synthesizing new materials and designing carriers with new structures are feasible ways to fabricate proper multi-agent delivery systems, which are critical to meet requirements in the clinic and improve therapeutic efficacy.Areas covered: This paper aims to give an overview about the multi-agent delivery systems developed in the last decade for differential release in combination therapy. Multi-agent delivery systems from nanoscale to bulk scale, such as liposomes, micelles, polymer conjugates, nano/micorparticles and hydrogels, developed over the last 10 years, have been collected and summarized. The characteristics of different delivery systems are described and discussed, including the structure of drug carriers, drug-loading techniques, release behaviors and consequent evaluation in biological assays.Expert opinion: The chemical structure of drug delivery systems is the key to controlling the release of therapeutic agents in combination therapy, and the differential release of multiple drugs could be realized by the successful design of a proper delivery system. Besides biological evaluation in vitro and in vivo, it is important to speed up practical application of the resulting delivery systems.

Continuous infusion of a standard combination solution in the management of hyperkalemia

Hyperkalemia, due to its effect on cardiac conductivity, is a potentially life-threatening electrolyte abnormality. Multiple therapeutic agents may be used alone or in combination for its prompt management. We report on the safety and efficacy of continuous infusion of a solution containing fixed concentrations of calcium gluconate, insulin, dextrose and sodium acetate (HyperK-Cocktail) for the treatment of hyperkalemia. This solution is prepared at our institution and is infused parenterally until the plasma potassium level stabilizes. Twenty-one consecutive hyperkalemic patients managed with HyperK-Cocktail on 23 occasions are reported. None of the subjects had intravenous extravasation injuries, hypernatremia, hypocalcemia, hypercalcemia or alkalosis during HyperK-Cocktail infusion. Transient hyperglycemia developed in nine subjects and hypoglycemia in one subject. The decrease in serum potassium was similar in the initial hour when compared to prior studies using a beta-agonist and/or insulin and glucose; a larger decrease was present from 2 to 8 h with the HyperK-Cocktail. The plasma potassium decreased by a mean of 1.0, 1.7, 2.1 and 2.1 mmol/L at 1, 2, 4 and 8 h, respectively. The mean serum potassium at hours 1-8 was significantly lower than the initial level. The results of our study demonstrated that HyperK-Cocktail is a safe and effective combination therapy for children with hyperkalemia.

Use of Intravenous Immunoglobulin in the Treatment of Childhood Atopic Dermatitis

Atopic dermatitis (AD) is a chronically recurrent inflammatory skin disorder characterized by pruritus, a specific distribution, and a family history. It has recently been reported that the incidence of AD has increased in Korea.1,2 Pruritus, sleep loss, dietary restrictions, and psychosocial factors significantly decrease the quality of life for AD patients.3,4 Recently, the European Academy of Allergy and Clinical Immunology and the American Academy of Allergy, Asthma, and Immunology published the PRACTALL consensus report for the diagnosis and treatment of AD in children and adults.5 The report suggests a stepwise management that includes the addition of multiple therapeutic agents on the basis of the disease severity. The PRACTALL consensus report defines severe or recalcitrant AD as AD that cannot be controlled with topical treatment.6 In the 2009 Korean Work Group Report on the treatment of severe/recalcitrant AD, severe AD is defined as AD with a SCORAD index higher than 50 and that cannot be controlled with conventional treatment,7 while the 2008 Guideline of Atopic Dermatitis in Korean Children defines severe AD by a SCORAD index higher than 40.8 Specific criteria for the definition of recalcitrant and severe AD are necessary. For the management of severe AD, the PRACTALL consensus report recommends systemic therapy such as antimicrobial treatment, systemic corticosteroids, cyclosporin A, azathioprine, anti-histamines, phototherapy, and immunotherapy. Several reports, including the 2009 Korean Work Group Report, have described intravenous immunoglobulin (IVIg) treatment as one of various immunoregulatory treatments. Nevertheless, this treatment was not included in the PRACTALL report.7,9,10 IVIg treatment displays immunomodulatory and anti-inflammatory properties, and its effectiveness in several immune-mediated conditions such as Kawasaki disease and idiopathic thrombocytopenic purpura has been demonstrated.11 IVIg is considered a candidate for the treatment of AD because of its ability to downregulate T-cell function, particularly interleukin-4 production.12,13 A small number of observations on the efficiency of IVIg in AD have been reported, but prospective and randomized studies for its clinical efficiency in childhood AD are sparse. A randomized, placebo-controlled prospective study in childhood AD patients is therefore required.14 Jee et al.14 recently reported therapeutic effects of IVIg in childhood AD; however, this study involved moderate to severe AD patients, and it did not include severe AD patients because the disease severity might have affected the treatment results. Further randomized studies with strict criteria for recalcitrant/severe AD are warranted. In addition, the IVIg effective dose, the dosing interval for initiation and maintenance, the identification of biomarkers (e.g., ECP, ICAM-1, and IL-5/INF-gamma) to determine efficiency, and clear criteria for IVIg indications all require consideration. Currently, we lack evidence-based data supporting the use of IVIg and other immunomodulators in childhood AD. Before IVIg can be recommended, its cost-benefit ratio, course, duration, and adverse reactions compared with alternative therapeutic options must be determined. The effects of novel therapies such as IVIg for recalcitrant/severe AD patients should be verified through repeated research and numerous research discussions.

Synergistic anti-cancer effects via co-delivery of TNF-related apoptosis-inducing ligand (TRAIL/Apo2L) and doxorubicin using micellar nanoparticles

The use of small molecule drugs in cancer chemotherapy has mostly been limited by dose-dependent toxicity and development of drug resistance resulting from repeated administrations. To overcome such problems, efforts have been made to develop drug delivery systems that can bear multiple therapeutic agents in one system. The purpose of this study is to deliver human tumor necrosis factor (TNF)-related apoptosis-inducing ligand (Apo2L/TRAIL) and doxorubicin (Dox, an anti-cancer drug) with micellar nanoparticles self-assembled from a biodegradable cationic copolymer P(MDS-co-CES) to achieve synergistic cytotoxic effects in cancer cells. Exogenously expressed TRAIL using recombinant methods shows great potential in cancer therapy as it induces cell death selectively in cancer cells with limited toxicity to normal tissues. Dox-loaded nanoparticles and TRAIL formed stable nanocomplexes with a size of ∼ 225 nm and zeta potential of ∼ 70 mV. Effects of nanocomplexes on both wild type and TRAIL-resistant SW480 colorectal carcinoma cells were investigated. The assemblies of Dox and TRAIL with P(MDS-co-CES) nanoparticles were efficiently delivered to cancer cells. Receptor-blocking studies showed that the nanocomplexes entered cells via death receptor-mediated endocytosis. Synergism in cell death induction was analysed by the isobologram method to study drug interactions. Cytotoxicity of the nanocomplexes to non-cancerous cells was significantly lower than cancerous cells. Anti-proliferative effects of nanocomplexes were retained in remaining cancer cells in long-term cultures after treatment with the nanocomplexes. In summary, this Dox and TRAIL co-delivery system can be a promising candidate for cancer treatment.

Roles of matrix metalloproteinases in cancer progression and their pharmacological targeting

Matrix metalloproteinases (MMPs) consist of a multigene family of zinc-dependent extracellular matrix (ECM) remodeling endopeptidases implicated in pathological processes, such as carcinogenesis. In this regard, their activity plays a pivotal role in tumor growth and the multistep processes of invasion and metastasis, including proteolytic degradation of ECM, alteration of the cell-cell and cell-ECM interactions, migration and angiogenesis. The underlying premise of the current minireview is that MMPs are able to proteolytically process substrates in the extracellular milieu and, in so doing, promote tumor progression. However, certain members of the MMP family exert contradicting roles at different stages during cancer progression, depending among other factors on the tumor stage, tumor site, enzyme localization and substrate profile. MMPs are therefore amenable to therapeutic intervention by synthetic and natural inhibitors, providing perspectives for future studies. Multiple therapeutic agents, called matrix metalloproteinase inhibitors (MMPIs) have been developed to target MMPs, attempting to control their enzymatic activity. Even though clinical trials with these compounds do not show the expected results in most cases, the field of MMPIs is ongoing. This minireview critically evaluates the role of MMPs in relation to cancer progression, and highlights the challenges, as well as future prospects, for the design, development and efficacy of MMPIs.

Safety and efficacy of maraviroc (MVC) combined with multiple different therapeutic agents in highly treatment‐experienced (TE) patients in Brazil

7‐11 November 2010, Tenth International Congress on Drug Therapy in HIV Infection, Glasgow, UK

Hidradenitis Suppurativa and Concomitant Pyoderma Gangrenosum

Hidradenitis suppurativa (HS) and pyoderma gangrenosum (PG) are both rare inflammatory skin conditions that are associated with systemic inflammatory diseases. We performed a retrospective medical chart review of patients with an overlap of HS and PG. We identified 11 cases of PG lesions presenting in patients with HS. Ten of the patients were women, and 9 were obese. All the patients developed HS lesions first, a median of 2.5 years (range, 0-15 years) preceding the appearance of PG lesions. All patients required multiple therapeutic agents because their diseases were often poorly responsive to standard therapies. Two patients received tumor necrosis factor inhibitors; 1 responded to treatment. One patient was treated with anakinra (interleukin-1 receptor antagonist) and had a 75% improvement of her lesions. We have identified a group of patients who have an overlap of PG and HS. Pyoderma gangrenosum can appear at any point after the development of HS and often has a severe, refractory course. We propose that PG and HS may represent variant manifestations of cytokine dysregulation by the innate immune system with common etiology. New therapeutic agents are eagerly sought, and further investigation with regard to interleukin 1 blockade is warranted.

Combinatorial Drug Conjugation Enables Nanoparticle Dual‐Drug Delivery

A new approach to loading multiple drugs onto the same drug-delivery nanocarrier in a precisely controllable manner, by covalently preconjugating multiple therapeutic agents through hydrolyzable linkers to form drug conjugates, is reported. In contrast to loading individual types of drugs separately, this drug-conjugates strategy enables the loading of multiple drugs onto the same carrier with a predefined stoichiometric ratio. The cleavable linkers allow the therapeutic activity of the individual drugs to be resumed after the drug conjugates are delivered into the target cells and unloaded from the delivery vehicle. As a proof of concept, the synthesis and characterization of paclitaxel-gemcitabine conjugates are demonstrated. The time-dependent hydrolysis kinetics and cytotoxicity of the combinatorial drug conjugates against human pancreatic cancer cells are examined. It is shown that the synthesized drug conjugates can be readily encapsulated into a lipid-coated polymeric drug-delivery nanoparticle, which significantly improves the cytotoxicity of the drug conjugates as compared to the free drug conjugates.

Understanding and Overcoming Major Barriers in Cancer Nanomedicine

Cancer nanomedicine is currently under intense development for applications in cancer imaging, molecular diagnosis and targeted therapy [1–6]. The basic rationale is that nanometer-sized particles, such as quantum dots (QDs), colloidal gold and polymeric nanomicelles, have functional and structural properties that are not available from either discrete molecules or bulk materials [7–10]. When conjugated with targeting ligands such as monoclonal antibodies, peptides or small molecules, these nanoparticles can be used to target malignant tumor cells and the tumor microenvironment (e.g., tumor stroma and tumor vasculatures) with high specificity and affinity. In the ‘mesoscopic’ size range of 10 to 100 nm, nanoparticles also have large surface areas for conjugating to multiple diagnostic and therapeutic agents, opening up new possibilities in integrated cancer imaging and therapy. However, at present there are several fundamental problems and technical barriers that must be understood and overcome. These problems include nanoparticle surface opsonization (surface ‘fouling’ by nonspecific protein adsorption) [11–13], nanoparticle uptake and retention in reticuloendothelial organs (e.g., liver and spleen [14,15], difficulties in nanoparticle targeting and penetration in solid tumors [16,17], and the long-term fate and toxicity concerns of nanoparticles, especially nonbiodegradable nanomaterials containing toxic elements [18–20]. Thus, there is an urgent need to study these problems and to develop innovative strategies for overcoming them. In this article, we briefly discuss the major challenges and opportunities in developing cancer nanomedicine for molecular imaging, in vitro diagnostics and targeted therapy.

Abstract 5484: Simultaneous targeting of EGFR and MET pathways using nanotechnology improves the treatment outcomes of photodynamic therapy for pancreatic cancer

Abstract Pancreatic Cancer (PanCa) is difficult to treat due to its inherent resistance to most therapies. First line chemotherapy only has a 5% response rate that yields a dismal prognosis of 6-month median survival and <4% 5-year survival. Combination therapies only showed modest benefits and there are still vital unmet needs for alternative treatment that improve overall patient survival. Photodynamic Therapy (PDT), a light-regulated cytotoxic therapy, showed promise as a new treatment modality for PanCa which improved median survival from 6 to 9.5 months in a pilot clinical trial. This result triggered our preclinical development of PDT combination regimen targeting proliferative pathways to further enhance treatment outcome. Using nanotechnology, we designed and synthesized nano-constructs co-encapsulating multiple therapeutic agents that can simultaneously target EGFR and MET-regulated proliferative pathways in combination with cytotoxic PDT treatment. We tested these nano-constructs in orthotopic pancreatic cancer models for treatment efficacy and our major findings include: 1) The nanoscale multidrug delivery system allows simultaneous targeting of EGFR and MET-regulated proliferative pathways and offered the best reduction of local tumor burden and metastatic burden to lymph nodes, lung and liver. With only one time of treatment, the local tumor burden is reduced to 10% of that of non-treated, and significant reductions of organ metastasis were also achieved. 2) The nanoscale multidrug delivery system delivers PDT agent and MET inhibitor PHA-665752 with higher payload and reduces their systemic toxicity. The EGFR inhibitor Cetuximab is delivered intracellularly into cancer cells, which targets both intracellular and cell surface EGFR. Intracellular delivery of Cetuximab increased treatment efficacy compared to naked Cetuximab. We are continuing optimizing the scheduling of individual treatments to further improve treatment outcomes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5484.

Nanotechnology Applications in Surgical Oncology

Surgery is currently the most effective and widely used procedure in treating human cancers, and the single most important predictor of patient survival is a complete surgical resection. Major opportunities exist to develop new and innovative technologies that could help the surgeon to delineate tumor margins, to identify residual tumor cells and micrometastases, and to determine if the tumor has been completely removed. Here we discuss recent advances in nanotechnology and optical instrumentation, and how these advances can be integrated for applications in surgical oncology. A fundamental rationale is that nanometer-sized particles such as quantum dots and colloidal gold have functional and structural properties that are not available from either discrete molecules or bulk materials. When conjugated with targeting ligands such as monoclonal antibodies, peptides, or small molecules, these nanoparticles can be used to target malignant tumor cells and tumor microenvironments with high specificity and affinity. In the "mesoscopic" size range of 10-100 nm, nanoparticles also have large surface areas for conjugating to multiple diagnostic and therapeutic agents, opening new possibilities in integrated cancer imaging and therapy.

Ophthalmologist Perceptions Regarding Treatment of Moderate-to-Severe Dry Eye: Results of a Physician Survey

To understand ophthalmologists' current perceptions and treatment of patients with moderate-to-severe dry eye disease (DED). Online survey. The online survey was sent to 7,882 ophthalmologists, including 51 corneal specialists, throughout the United States from October 9 to 21, 2008. The response rate was 3.1% (n = 245), typical for this type of survey. Only ophthalmologists who treated four or more moderate-to-severe DED patients per month (235 of 245 [96%]) were asked to complete the survey. Ninety-four percent of respondents agreed that more treatment options are needed for moderate-to-severe DED. Corneal specialists were more likely to strongly agree (63%) than general ophthalmologists (54%). Only 33% overall felt that current therapies were extremely or very effective for moderate DED and only 5% for severe disease. Ninety-two percent agreed that multiple therapeutic agents are needed to manage moderate-to-severe DED. The respondents prescribed or recommended a mean of 3.2 different treatments (standard deviation = 1.2) for moderate DED patients over the course of a year and 4.9 (standard deviation = 2.2) for patients with severe DED. The most highly ranked goals for treatment of moderate-to-severe DED patients were maintaining and protecting the ocular surface (ranked 1 or 2 x 74%) and lubricating and hydrating the ocular surface (ranked 1 or 2 x 67%). Corneal specialists ranked maintaining and protecting the ocular surface even more highly (ranked 1 or 2 x 82%). Results reflected the difficulty of treating moderate-to-severe DED, the importance of using multiple treatment approaches, the limitations of current treatment options, and the need for additional treatment options.

Novel Stably Transplantable Xenograft Models of Human Breast Cancer for Evaluation of Experimental Therapeutics.

Abstract In translational breast cancer research, our ability to evaluate clinical responses of human tumors to new therapeutic agents is severely limited experimentally. For example, it is not possible to evaluate the response of a single tumor to multiple candidate therapeutic agents. Conversely, the limited number of well-characterized in vivo preclinical human tumor models currently available precludes evaluation of multiple clinically relevant tumors with candidate therapeutic agents. These limitations severely impinge on our ability to develop and test novel therapeutic agents, particularly those that may target tumor-initiating “cancer stem cells”, which are relatively resistant to chemotherapy and radiation and may be responsible for disease recurrence and metastases.Historically, in vivo experimental therapeutic research has relied on either genetically engineered animal models, or “xenograft” transplantation models in which established human cancer cell lines are transplanted into immunocompromised host mice. However, each type of model has significant limitations. We sought to circumvent some of these limitations by propagating a cohort of human tumors as stably transplantable xenograft tissue lines grown in the absence of engineered or immortalized fibroblasts by transplanting clinical biopsies directly into the mammary fat pad of SCID/Beige immunocompromised mice (lacking B-cell, T-cell, and NK cell function) without intervening culture in vitro.Thus far, we have established 13 independent stably transplantable xenograft lines representing nine “triple-negative” (ER-PR-HER2-), two HER2+, and two ER+ breast cancers. Established xenograft lines show phenotypic similarity to the primary tumor with respect to histology and gene expression. Xenografts are being characterized genetically by whole genome sequencing as well as for the diversity of tumor-initiating cell types present. These models are proving useful for the evaluation of experimental therapeutics for their ability to inhibit tumor growth, and for their ability to target the subset cancer cells capable of regenerating tumors upon transplantation, with the goals of overcoming chemoresistance, preventing disease recurrence, and eliminating metastases. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 1159.

Cancer Nanotechnology - Prospects for Cancer Diagnostics and Therapy

Nanotechnology is a multidisciplinary field that brings together diverse fields of research and development such as engineering, biology, physics and chemistry. Formal definitions of nanotechnology refer to man-made devices, components and structures in the 1 – 100 nm range in at least one dimension. Advances in nanoscience are having a significant impact on many scientific fields, boosting the development of a variety of important technologies. Nanotechnology offers an unprecedented opportunity to interact with cancer cells in real time at the molecular and cellular scale. Because of their small size, nanoscale devices can readily interact with biomolecules on both the surface of cells and inside of cells. The concerted development of nanoscale devices, structures and components have provided essential breakthroughs in monitoring and fighting cancer at the earliest stages of the cancer process. Nanotechnology offers a wealth of tools that may provide researchers with new and innovative ways to diagnose and treat cancer - new imaging agents; systems for real-time assessments of therapeutic and surgical efficacy; multifunctional, targeted devices capable of bypassing biological barriers to deliver multiple therapeutic agents directly to cancer cells and tissues that play a critical role in cancer growth and metastasis; agents that can monitor predictive molecular changes allowing for preventive action against precancerous cells becoming malignant; minimizing costs for multiplex analysis. Nanotechnology, if properly integrated with conventional cancer research, may provide extraordinary prospects towards better diagnosis and effective therapy. Keywords: Nanotechnology, cancer, cancer therapy, cancer imaging, cancer molecular diagnostics, nanoparticles, nanosensors

CXCR4 inhibitor AMD3100 disrupts the interaction of multiple myeloma cells with the bone marrow microenvironment and enhances their sensitivity to therapy

AbstractThe interaction of multiple myeloma (MM) cells with their microenvironment in the bone marrow (BM) provides a protective environment and resistance to therapeutic agents. We hypothesized that disruption of the interaction of MM cells with their BM milieu would lead to their sensitization to therapeutic agents such as bortezomib, melphalan, doxorubicin, and dexamethasone. We report that the CXCR4 inhibitor AMD3100 induces disruption of the interaction of MM cells with the BM reflected by mobilization of MM cells into the circulation in vivo, with kinetics that differed from that of hematopoietic stem cells. AMD3100 enhanced sensitivity of MM cell to multiple therapeutic agents in vitro by disrupting adhesion of MM cells to bone marrow stromal cells (BMSCs). Moreover, AMD3100 increased mobilization of MM cells to the circulation in vivo, increased the ratio of apoptotic circulating MM cells, and enhanced the tumor reduction induced by bortezomib. Mechanistically, AMD3100 significantly inhibited Akt phosphorylation and enhanced poly(ADP-ribose) polymerase (PARP) cleavage as a result of bortezomib, in the presence of BMSCs in coculture. These experiments provide a proof of concept for the use of agents that disrupt interaction with the microenvironment for enhancement of efficacy of cytotoxic agents in cancer therapy.

Evolution of nonsurgical therapy for colorectal cancer

The management of colorectal cancer (CRC) has changed considerably in the past 15 years with the introduction of multiple novel active therapeutic agents. Chemotherapy regimens combining a fluoropyrimidine with either oxaliplatin or irinotecan are standard first-line and second-line therapy for advanced and metastatic disease. The first-line use of these combinations produces tumor response rates of approximately 50% and a median overall survival of approximately 20 months. Addition of bevacizumab to first-line treatment and addition of cetuximab to salvage therapy for patients who fail to respond to irinotecan have contributed to further increases in tumor response rates and enhanced progression-free survival. Such approaches have produced only marginal overall survival benefits, however, and entail considerable cost. Adjuvant chemotherapy, delivered after surgical resection of the primary tumor, increases cure rates by approximately 10% for stage III disease and approximately 3-4% for stage II disease. Encouraging reductions in local relapse rates have been observed in patients with early rectal cancer who have undergone chemoradiotherapy, and increasingly complex regimens are currently being explored in phase II clinical trials in an attempt to increase both the operability and long-term local control of CRC. The greater the therapeutic choice, the greater the cost (both financial and in terms of toxicity), thus the keener the clinical community becomes to develop biomarkers to select patient populations who will be most likely to benefit from a specific agent.

Mitotic trafficking of silicon microparticles.

Multistage carriers were recently introduced by our laboratory, with the concurrent objectives of co-localized delivery of multiple therapeutic agents, the "theranostic" integration of bioactive moieties with imaging contrast, and the selective, potentially personalized bypassing of the multiplicity of biological barriers that adversely impact biodistribution of vascularly injected particulates. Mesoporous ("nanoporous") silicon microparticles were selected as primary carriers in multi-stage devices, with targets including vascular endothelia at pathological lesions. The objective of this study was to evaluate biocompatibility of mesoporous silicon microparticles with endothelial cells using in vitro assays with an emphasis on microparticle compatibility with mitotic events. We observed that vascular endothelial cells, following internalization of silicon microparticles, maintain cellular integrity, as demonstrated by cellular morphology, viability and intact mitotic trafficking of vesicles bearing silicon microparticles. The presence of gold or iron oxide nanoparticles within the porous matrix did not alter the cellular uptake of particles or the viability of endothelial cells subsequent to engulfment of microparticles. Endothelial cells maintained basal levels of IL-6 and IL-8 release in the presence of silicon microparticles. This is the first study that demonstrates polarized, ordered partitioning of endosomes based on tracking microparticles. The finding that mitotic sorting of endosomes is unencumbered by the presence of nanoporous silicon microparticles advocates the use of silicon microparticles for biomedical applications.

Inhibition of Human Lymphoma Cell Growth by the PKC-Beta Selective Inhibitor Enzastaurin (LY317615) in Combination with Multiple Therapeutic Agents.

Previous studies in our lab have shown that the PKC-beta inhibitor, enzastaurin (LY317615), when used to treat a panel of human diffuse large cell lymphoma (DLCL) lines, was able to induce cell death in vitro and substantially reduce tumor growth in xenograft assays. These findings support the hypothesis that activation of PKC contributes to tumor cell survival and proliferation, which has been implicated in the pathogenesis of human B cell lymphomas. Specifically, PKC-beta activation is increased in tumor cells from patients with poor prognosis DLCL, suggesting that PKC-beta may be a target for therapeutic intervention. In the present study, we have explored the interaction of enzastaurin with a panel of well characterized therapeutic agents to evaluate whether its anti-tumor activity can potentially be enhanced. Drugs were chosen for analysis based either on known single agent activity in lymphoma, or by preclinical evaluation indicating potential synergy with enzastaurin. For in vitro culture assays (48–72 hr treatment), the addition of gemcitabine, rapamycin, or bortezomib, increased the cytotoxicity of enzastaurin from 2 to 7 fold. This effect was evident with multiple human DLCL cell lines, (OCI-Ly3, 7, 10, 19, and SUDHL-4, and 6), as well as two independent primary DLCL cultures. For in vivo studies, subcutaneous transplantation of the DLCL cell line OCI-Ly19, (previously engineered to express luciferase which allows for real time in vivo imaging), or a primary DLCL isolate, into immune deficient NOD/SCID mice formed reproducible tumors. Recipient animals were separated into uniform cohorts when the tumors were of <=500 cubic mm in size. The animals were then simultaneously or sequentially treated with enzastaurin, (150 mg/kg b.i.d. via oral gavage) and a secondary drug, either gemcitabine, (2.5 or 5.0 mg/kg 1x/3 days IP), bortezomib, (0.4 mg/kg twice weekly IP), rapamycin, (1.0 mg/kg, daily IP), or rituxan, (5 mg/kg, weekly IP). Imaging and analysis of tumor volumes showed that addition of either rituxan or rapamycin provided no additional benefit in comparison to enzastaurin alone during the course of treatment. In contrast, the combination of either gemcitabine or bortezomib with enzastaurin demonstrated significantly reduced tumor volumes in comparison to enzastaurin alone (17% to 38% greater decrease with enzastaurin + gemcitabine, and 50% greater decrease in tumor volume with enzastaurin + bortezomib). These data suggest that the use of enzastaurin in combination with existing therapeutic drugs (gemcitabine or bortezomib) has the potential to limit tumor size/growth while lowering dose levels and thereby reducing potential side effects associated with traditional treatments.