Nanotechnology-Based Strategy for Enhancing Therapeutic Efficacy in Pancreatic Cancer: Receptor-Targeted Drug Delivery by Somatostatin Analog.

A novel nanotechnology-based drug delivery system (DDS) targeted at pancreatic cancer cells was developed, characterized, and tested. The system consisted of liposomes as carriers, an anticancer drug (paclitaxel) as a chemotherapeutic agent, and a modified synthetic somatostatin analog, 5-pentacarbonyl-octreotide, a ligand for somatostatin receptor 2 (SSTR2), as a targeting moiety for pancreatic cancer. The cellular internalization, cytotoxicity, and antitumor activity of the DDS were tested in vitro using human pancreatic ductal adenocarcinoma (PDAC) cells with different expressions of the targeted SSTR2 receptors, and in vivo on immunodeficient mice bearing human PDAC xenografts. The targeted drug delivery system containing paclitaxel exhibited significantly enhanced cytotoxicity compared to non-targeted DDS, and this efficacy was directly related to the levels of SSTR2 expression. It was found that octreotide-targeted DDS proved exceptionally effective in suppressing the growth of PDAC tumors. This study underscores the potential of octreotide-targeted liposomal delivery systems to enhance the therapeutic outcomes for PDAC compared with non-targeted liposomal DDS and Paclitaxel-Cremophor® EL, suggesting a promising avenue for future cancer therapy innovations.

Personalized Versus Precision Nanomedicine for Treatment of Ovarian Cancer.

The response to treatment is substantially varied between individual patients with ovarian cancer. However, chemotherapy treatment plans rarely pay sufficient attention to the mentioned factors. Instead, standardized treatment protocols are usually employed for most ovarian cancer patients. Variations in an individual's sensitivity to drugs significantly limit the effectiveness of treatment in some patients and lead to severe toxicities in others. In the present investigation, a nanotechnology-based approach for personalized treatment of ovarian carcinoma (the most lethal type of gynecological cancer) constructed on the individual genetic profile of the patient's tumor is developed and validated. The expression of predefined genes and proteins is analyzed for each patient sample. Finally, a mixture of the complex nanocarrier-based targeted delivery system containing drug(s)/siRNA(s)/targeted peptide is selected from the pre-synthesized bank and tested in vivo on murine cancer model using cancer cells isolated from tumors of each patient. Based on the results of the present study, an innovative approach and protocol for personalized treatment of ovarian cancer are suggested and evaluated. The results of the present study clearly show the advantages and perspectives of the proposed individual treatment approach.

Inhibition of Mtorc1/2 and DNA-PK via CC-115 Synergizes with Carboplatin and Paclitaxel in Lung Squamous Cell Carcinoma.

Only a small percentage (<1%) of patients with late-stage lung squamous cell carcinoma (LUSC) are eligible for targeted therapy. Because PI3K/AKT/mTOR signaling, particularly Phosphatidylinositol 3-kinase CA (PIK3CA), is dysregulated in two-thirds of LUSC, and DNA damage response pathways are enriched in LUSC, we tested whether CC-115, a dual mTORC1/2 and DNA-PK inhibitor, sensitizes LUSC to chemotherapy. We demonstrate that CC-115 synergizes with carboplatin in six of 14 NSCLC cell lines, primarily PIK3CA-mutant LUSC. Synergy was more common in cell lines that had decreased basal levels of activated AKT and DNA-PK, evidenced by reduced P-S473-AKT, P-Th308-AKT, and P-S2056-DNA-PKcs. CC-115 sensitized LUSC to carboplatin by inhibiting chemotherapy-induced AKT activation and maintaining apoptosis, particularly in PIK3CA-mutant cells lacking wild-type (WT) TP53. In addition, pathway analysis revealed that enrichments in the IFNα and IFNγ pathways were significantly associated with synergy. In multiple LUSC patient-derived xenograft and cell line tumor models, CC-115 plus platinum-based doublet chemotherapy significantly inhibited tumor growth and increased overall survival as compared with either treatment alone at clinically relevant dosing schedules. IHC and immunoblot analysis of CC-115-treated tumors demonstrated decreased P-Th308-AKT, P-S473-AKT, P-S235/236-S6, and P-S2056-DNA-PKcs, showing direct pharmacodynamic evidence of inhibited PI3K/AKT/mTOR signaling cascades. Because PI3K pathway and DNA-PK inhibitors have shown toxicity in clinical trials, we assessed toxicity by examining weight and numerous organs in PRKDC-WT mice, which demonstrated that the combination treatment does not exacerbate the clinically accepted side effects of standard-of-care chemotherapy. This preclinical study provides strong support for the further investigation of CC-115 plus chemotherapy in LUSC.

Anti-Tumor Effects of a Penetratin Peptide Targeting Transcription of E2F-1, 2 and 3a Is Enhanced When Used in Combination with Pemetrexed or Cisplatin.

BACKGROUND: We tested the antitumor effects of a modified E2F peptide substituting D-Arg for L-Arg, conjugated to penetratin (PEP) against solid tumor cell lines and the CCRF-leukemia cell line, alone and in combination with pemetrexed or with cisplatin. For in-vivo studies, the peptide was encapsulated in PEGylated liposomes (PL-PEP) to increase half-life and stability. METHODS: Prostate cancer (DU145 and PC3), breast cancer (MCF7, MDA-MB-468, and 4T1), lymphoma (CCRF-CEM), and non-small cell lung cancer (NSCLC) cell lines (H2009, H441, H1975, and H2228) were treated with D-Arg PEP in combination with cisplatin or pemetrexed. Western blot analysis was performed on the NSCLC for E2F-1, pRb, thymidylate synthase, and thymidine kinase. The H2009 cell line was selected for an in-vivo study. RESULTS: When the PEP was combined with cisplatin and tested against solid tumor cell lines and the CCRF-CEM leukemia cell line, there was a modest synergistic effect. A marked synergistic effect was seen when the combination of pemetrexed and the PEP was tested against the adenocarcinoma lung cancer cell lines. The addition of the PEP to pemetrexed enhanced the antitumor effects of pemetrexed in a xenograft of the H2009 in mice. CONCLUSIONS: The D-Arg PEP in combination with cisplatin caused synergistic cell kill against prostate, breast, lung cancers, and the CCRF-CEM cell line. Marked synergy resulted when the D-Arg PEP was used in combination with pemetrexed against the lung adenocarcinoma cell lines. A xenograft study using the PL-PEP in combination with pemetrexed showed enhanced anti-tumor effects compared to each drug alone.

Strategy to enhance lung cancer treatment by five essential elements: inhalation delivery, nanotechnology, tumor-receptor targeting, chemo- and gene therapy.

Non-Small Cell Lung Carcinoma (NSCLC), is the most common type of lung cancer (more than 80% of all cases). Small molecule Tyrosine Kinase (TK) Inhibitors acting on the Epidermal Growth Factor Receptors (EGFRs) are standard therapies for patients with NSCLC harboring EGFR-TK inhibitor-sensitizing mutations. However, fewer than 10 % of patients with NSCLC benefit from this therapy. Moreover, even the latest generation of EGFR inhibitors can cause severe systemic toxicities and are ineffective in preventing non-canonical EGFR signaling. In order to minimize and even overcome these limitations, we are proposing a novel multi-tier biotechnology treatment approach that includes: (1) suppression of all four types of EGFR-TKs by a pool of small interfering RNAs (siRNAs); (2) induction of cell death by an anticancer drug, (3) enhancing the efficiency of the treatment by the local inhalation delivery of therapeutic agents directly to the lungs (passive targeting), (4) active receptor-mediated targeting of the therapy specifically to cancer cells that in turn should minimize adverse side effects of treatment and (5) increasing the stability, solubility, and cellular penetration of siRNA and drug by using tumor targeted Nanostructured Lipid Carriers (NLC). Methods: NLCs targeted to NSCLC cells by a synthetic Luteinizing Hormone-Releasing Hormone (LHRH) decapeptide was used for the simultaneous delivery of paclitaxel (TAX) and a pool of siRNAs targeted to the four major forms of EGFR-TKs. LHRH-NLC-siRNAs-TAX nanoparticles were synthesized, characterized and tested in vitro using human lung cancer cells with different sensitivities to gefitinib (inhibitor of EGFR) and in vivo on an orthotopic NSCLC mouse model. Results: Proposed nanoparticle-based complex containing an anticancer drug, inhibitors of different types of EGFR-TKs and peptide targeted to the tumor-specific receptors (LHRH-NLC-siRNAs-TAX) demonstrated a favorable organ distribution and superior anticancer effect when compared with treatment by a single drug, inhibitor of one EGFR-TK and non-targeted therapy. Conclusions: The use of a multifunctional NLC-based delivery system substantially enhanced the efficiency of therapy for NSCLC and possibly will limit adverse side effects of the treatments. The results obtained have the potential to significantly impact the field of drug delivery and to improve the efficiency of therapy of lung and other types of cancer.

Prevention of paclitaxel-induced neuropathy by formulation approach.

Chemotherapy-induced peripheral neuropathy (CIPN) is a major adverse effect of paclitaxel. Several liposome-based products have been approved and demonstrated superior efficacy and safety profiles for other drugs. The first objective of this work was to evaluate the effect of liposome formulation of paclitaxel (L-PTX) on neurotoxicity in-vitro and in-vivo in comparison to the standard Taxol® formulation. The second aim was to investigate the effect of formulation on paclitaxel biodistribution following intravenous administration in an animal model. Free paclitaxel was toxic to cell of neuronal origin (IC50 = 18.4 µg/mL) at a lower concentration than to lung cancer cells (IC50 = 59.1 µg/mL), and L-PTX demonstrated a comparable toxicity in both cell lines (IC50 = 31.8 and 33.7 µg/mL). Administration of L-PTX at 2 mg/kg per dose for a total of 4 doses on day 0, 2, 4, and 6 to rats did not result in increased sensitivity in response to mechanical or thermal stimulation of hind paws, in comparison to Taxol® administration at the same dose level that resulted in neuropathy. Paclitaxel biodisposition was evaluated for two formulations in plasma, liver, lung, brain, spinal cord, skin and muscle of rats after single intravenous dose at 6 mg/kg. The exposure to paclitaxel in brain, spinal cord, muscle, and skin was lower in the L-PTX group compared to Taxol® group. PEGylated liposomes containing paclitaxel were successfully developed and demonstrated reduced neurotoxicity in-vitro in neuronal cells and prevented development of peripheral neuropathy in-vivo. This proof of concept study showed that formulation in nanoparticles is a promising approach for reducing (or preventing) neurotoxicity caused by cancer drugs.

Modeling and antitumor studies of a modified L-penetratin peptide targeting E2F in lung cancer and prostate cancer.

E2F1-3a overexpression due to amplification or to mutation or loss of the retinoblastoma gene, induces genes involved in DNA synthesis and leads to abnormal cellular proliferation, tumor growth, and invasion. Therefore, inhibiting the overexpression of one or more of these activating E2Fs is a recognized target in cancer therapeutics. In previous studies we identified by phage display, a novel 7-mer peptide (PEP) that bound tightly to an immobilized consensus E2F1 promoter sequence, and when conjugated to penetratin to increase its uptake into cells, was cytotoxic to several malignant cell lines and human prostate and small cell lung cancer xenografts. Based on molecular simulation studies that showed that the D-Arg penetratin peptide (D-Arg PEP) secondary structure is more stable than the L-Arg PEP, the L-Arg in the peptide was substituted with D-Arg. In vitro studies confirmed that it was more stable than the L- form and was more cytotoxic as compared to the L-Arg PEP when tested against the human castrate resistant cell line, DU145 and the human lung cancer H196 cell line. When encapsulated in PEGylated liposomes, the D-Arg-PEP potently inhibited growth of the DU145 xenograft in mice. Our findings validate D- Arg PEP, an inhibitor of E2F1and 3a transcription, as an improved second generation drug candidate for targeted molecular therapy of cancers with elevated levels of activated E2F(s).

Metastatic and triple-negative breast cancer: challenges and treatment options.

The major current conventional types of metastatic breast cancer (MBC) treatments include surgery, radiation, hormonal therapy, chemotherapy, or immunotherapy. Introducing biological drugs, targeted treatment and gene therapy can potentially reduce the mortality and improve the quality of life in patients with MBC. However, combination of several types of treatment is usually recommended. Triple negative breast cancer (TNBC) accounts for 10-20% of all cases of breast carcinoma and is characterized by the low expression of progesterone receptor (PR), estrogen receptor (ER), and human epidermal growth factor receptor 2 (HER2). Consequently, convenient treatments used for MBC that target these receptors are not effective for TNBC which therefore requires special treatment approaches. This review discusses the occurrence of MBC, the prognosis and predictive biomarkers of MBC, and focuses on the novel advanced tactics for treatment of MBC and TNBC. Nanotechnology-based combinatorial approach for the suppression of EGFR by siRNA and gifitinib is described.

Combinatorial treatment of idiopathic pulmonary fibrosis using nanoparticles with prostaglandin E and siRNA(s).

Inhalation delivery of prostaglandin E (PGE2) in combination with selected siRNA(s) was proposed for the efficient treatment of idiopathic pulmonary fibrosis (IPF). Nanostructured lipid carriers (NLC) were used as a delivery system for PGE2 with and without siRNAs targeted to MMP3, CCL12, and HIF1Alpha mRNAs. The model of IPF was developed in SKH1 mice by intratracheal administration of bleomycin at a dose of 1.5U/kg. Results showed that NLC-PGE2 in combination with three siRNAs delivered locally to the lungs by inhalation markedly reduced mouse body mass, substantially limited hydroxyproline content in the lungs and disturbances of the mRNAs and protein expression, restricted lung tissue damage and prevented animal mortality. Our data provide evidence that IPF can be effectively treated by inhalation of the NLC-PGE2 in combination with siRNAs delivered locally into the lungs. This effect could not be achieved by using NLC containing just PGE2 or siRNA(s) alone.

Development of edge-activated liposomes for siRNA delivery to human basal epidermis for melanoma therapy.

Delivery of macromolecules such as siRNA into cells that reside in the basal epidermis of the skin is a major challenge due to the transport barriers that need to be overcome. siRNAs have potential therapeutic applications in various dermatological diseases such as psoriasis, atopic dermatitis, and cancer. Unfortunately, a low permeability of siRNA through the stratum corneum and epidermis has significantly limited its use for topical application. The objective of this study was to develop a topical siRNA delivery system that can permeate through the stratum corneum and viable epidermis and efficiently deposit therapeutic levels of siRNA to the basal epidermis/upper dermis where melanoma cells reside. To achieve this objective, a series of liposome compositions that contained various concentrations of edge activator in their structures were prepared and then complexed with siRNA at different ratios to generate a small library of liposome-siRNA complexes (lipoplexes) with different physicochemical properties. In this study we used melanoma as a disease model. Through use of quantitative imaging analysis, we identified the necessary design parameters for effective permeation of lipoplexes through the skin layers and deposition at the upper dermis. The ability of the formulated lipoplexes to internalize into melanoma cells, knockdown the expression of the BRAF protein and induce cell death in melanoma cells was studied by fluorescent microscopy, in-cell immunofluorescence assay and WST-1 cell proliferation assay. By providing direct quantitative and qualitative microscopy evidence, the results of this study demonstrate for the first time that the passive delivery of an edge-activated liposomal formulation can effectively carry siRNA through the stratum corneum and deposit it at the lower epidermis/upper dermis.

Stem cell-based gene therapy activated using magnetic hyperthermia to enhance the treatment of cancer.

Stem cell-based gene therapies, wherein stem cells are genetically engineered to express therapeutic molecules, have shown tremendous potential for cancer applications owing to their innate ability to home to tumors. However, traditional stem cell-based gene therapies are hampered by our current inability to control when the therapeutic genes are actually turned on, thereby resulting in detrimental side effects. Here, we report the novel application of magnetic core-shell nanoparticles for the dual purpose of delivering and activating a heat-inducible gene vector that encodes TNF-related apoptosis-inducing ligand (TRAIL) in adipose-derived mesenchymal stem cells (AD-MSCs). By combining the tumor tropism of the AD-MSCs with the spatiotemporal MCNP-based delivery and activation of TRAIL expression, this platform provides an attractive means with which to enhance our control over the activation of stem cell-based gene therapies. In particular, we found that these engineered AD-MSCs retained their innate ability to proliferate, differentiate, and, most importantly, home to tumors, making them ideal cellular carriers. Moreover, exposure of the engineered AD-MSCS to mild magnetic hyperthermia resulted in the selective expression of TRAIL from the engineered AD-MSCs and, as a result, induced significant ovarian cancer cell death in vitro and in vivo.

Biodegradable Janus nanoparticles for local pulmonary delivery of hydrophilic and hydrophobic molecules to the lungs.

The aim of the present work is to synthesize, characterize, and test self-assembled anisotropic or Janus particles designed to load anticancer drugs for lung cancer treatment by inhalation. The particles were synthesized using binary mixtures of biodegradable and biocompatible materials. The particles did not demonstrate cyto- and genotoxic effects. Janus particles were internalized by cancer cells and accumulated both in the cytoplasm and nuclei. After inhalation delivery, nanoparticles accumulated preferentially in the lungs of mice and retained there for at least 24 h. Two drugs or other biologically active components with substantially different aqueous solubility can be simultaneously loaded in two-phases (polymer-lipid) of these nanoparticles. In the present proof-of-concept investigation, the particles were loaded with two anticancer drugs: doxorubicin and curcumin as model anticancer drugs with relatively high and low aqueous solubility, respectively. However, there are no obstacles for loading any hydrophobic or hydrophilic chemical agents. Nanoparticles with dual load were used for their local inhalation delivery directly to the lungs of mice with orthotopic model of human lung cancer. In vivo experiments showed that the selected nanoparticles with two anticancer drugs with different mechanisms of action prevented progression of lung tumors. It should be stressed that anticancer effects of the combined treatment with two anticancer drugs loaded in the same nanoparticle significantly exceeded the effect of either drug loaded in similar nanoparticles alone.

Delivery of antisense oligonucleotides using poly(alkylene oxide)-poly(propylacrylic acid) graft copolymers in conjunction with cationic liposomes.

The clinical application of gene silencing is hindered by poor stability and low delivery efficiency of naked oligonucleotides. Here, we present the in vitro and in vivo behaviors of a rationally designed, ternary, self-assembled nanoparticle complex, consisting of an anionic copolymer, cationic DOTAP liposome, and antisense oligonucleotide (AON). The multifunctional copolymers are based on backbone poly(propylacrylic acid) (PPAA), a pH-sensitive hydrophobic polymer, with grafted poly(alkylene oxides) (PAOs) varying in extent of grafting and PAO chemistry. The nanoparticle complexes with PPAA-g-PAO copolymers enhance antisense gene silencing effects in A2780 human ovarian cancer cells. A greater amount of AON is delivered to ovarian tumor xenografts using the ternary copolymer-stabilized delivery system, compared to a binary DOTAP/AON complex, following intraperitoneal injection in mice. Further, intratumoral injection of the nanoparticle complexes containing 1 mol% grafted PAO reduced tumoral bcl-2 expression by up to 60%. The data for complexes across the set of PAO polymers support a strong role for the hydrophilic-lipophilic balance of the graft copolymer in achieving serum stability and cellular uptake. Based upon these results, we anticipate that this novel nanoparticle delivery system can be extended to the delivery of plasmid DNA, siRNA, or aptamers for preclinical and clinical development.

Tumor-targeted responsive nanoparticle-based systems for magnetic resonance imaging and therapy.

PURPOSE: Design and synthesis of a tumor responsive nanoparticle-based system for imaging and treatment of various cancers. METHODS: Manganese oxide nanoparticles (Mn3O4 NPs) were synthesized and modified with LHRH targeting peptide or anti-melanoma antibodies (cancer targeting moieties) and a MMP2 cleavable peptide (a possible chemotactic factor). Nanostructured lipid carriers (NLCs) were used to entrap the BRAF inhibitor, vemurafenib, and enhance cytotoxicity of the drug. Size distribution, stability, drug entrapment, cytotoxicity and genotoxicity of synthesized nanoparticles were studied in vitro. Enhancement of MRI signal by nanoparticles and their body distribution were examined in vivo on mouse models of melanoma, ovarian and lung cancers. RESULTS: Uniform, stable cancer-targeted nanoparticles (PEGylated water-soluble Mn3O4 NPs and NLCs) were synthesized. No signs of cyto-,genotoxicity and DNA damage were detected for nanoparticles that do not contain an anticancer drug. Entrapment of vemurafenib into nanoparticles significantly enhanced drug toxicity in cancer cells with targeted V600E mutation. The developed nanoparticles containing LHRH and MMP2 peptides showed preferential accumulation in primary and metastatic tumors increasing the MRI signal in mice with melanoma, lung and ovarian cancers. CONCLUSIONS: The proposed nanoparticle-based systems provide the foundation for building an integrated MRI diagnostic and therapeutic approach for various types of cancer.

Inhalation treatment of lung cancer: the influence of composition, size and shape of nanocarriers on their lung accumulation and retention.

OBJECTIVE: Various nanoparticles have been designed and tested in order to select optimal carriers for the inhalation delivery of anticancer drugs to the lungs. METHODS: THE FOLLOWING NANOCARRIERS WERE STUDIED: micelles, liposomes, mesoporous silica nanoparticles (MSNs), poly propyleneimine (PPI) dendrimer-siRNA complexes nanoparticles, quantum dots (QDs), and poly (ethylene glycol) polymers. All particles were characterized using the following methods: dynamic light scattering, zeta potential, atomic force microscopy, in vitro cyto- and genotoxicity. In vivo organ distribution of all nanoparticles, retention in the lungs, and anticancer effects of liposomes loaded with doxorubicin were examined in nude mice after the pulmonary or intravenous delivery. RESULTS: Significant differences in lung uptake were found after the inhalation delivery of lipid-based and non-lipid-based nanoparticles. The accumulation of liposomes and micelles in lungs remained relatively high even 24 h after inhalation when compared with MSNs, QDs, and PPI dendrimers. There were notable differences between nanoparticle accumulation in the lungs and other organs 1 and 3 h after inhalation or intravenous administrations, but 24 h after intravenous injection all nanoparticles were mainly accumulated in the liver, kidneys, and spleen. Inhalation delivery of doxorubicin by liposomes significantly enhanced its anticancer effect and prevented severe adverse side effects of the treatment in mice bearing the orthotopic model of lung cancer. CONCLUSION: The results of the study demonstrate that lipid-based nanocarriers had considerably higher accumulation and longer retention time in the lungs when compared with non-lipid-based carriers after the inhalation delivery. These particles are most suitable for effective inhalation treatment of lung cancer.

A novel peptide that inhibits E2F transcription and regresses prostate tumor xenografts.

E2F-1, a key transcription factor necessary for cell growth, DNA repair and differentiation, is an attractive target for development of useful anticancer drugs in tumors that are E2F "oncogene addicted". A peptide, isolated from phage clones, based on its binding to an E2F-1 consensus sequence, was cytotoxic against a wide range of cancer cell lines. The peptide was coupled to penetratin (PEP) and tested against prostate cancer cell lines, and a fresh sample from a patient with metastatic cancer. As the PEP was found to be relatively unstable in serum, it was encapsulated in PEGylated liposomes for in vivo studies. The peptide was cytotoxic against prostate cell lines and a fresh sample from a patient with metastatic prostate cancer. Treatment of mice bearing the human Du-145 human prostate tumor with the PEP encapsulated in PEGylated liposomes (PL-PEP) caused tumor regression without significant toxicity. The liposome encapsulated PEP has promise as an antitumor agent, alone or in combination with inhibitors of DNA synthesis.

Targeted nanomedicine for suppression of CD44 and simultaneous cell death induction in ovarian cancer: an optimal delivery of siRNA and anticancer drug.

PURPOSE: The proposed project is aimed at enhancing the efficiency of epithelial ovarian cancer treatment and reducing adverse side effects of chemotherapy using nanotechnology. Overexpression of the CD44 membrane receptor results in tumor initiation, growth, cancer stem cells' specific behavior, development of drug resistance, and metastases. We hypothesize that a developed cancer-targeted delivery system that combines CD44 siRNA with paclitaxel would successfully deliver its payload inside cancer cells, effectively induce cell death, and prevent metastases. EXPERIMENTAL DESIGN: We synthesized, characterized, and tested a nanoscale-based drug delivery system (DDS) containing a modified polypropylenimine (PPI) dendrimer as a carrier; anticancer drug paclitaxel as a cell death inducer; a synthetic analog of luteinizing hormone-releasing hormone (LHRH) peptide as a tumor-targeting moiety; and siRNA targeted to CD44 mRNA. The proposed DDS was tested in vitro and in vivo using metastatic ovarian cancer cells isolated from patients with malignant ascites. RESULTS: We found that in contrast with cells isolated from primary tumors, CD44 was highly overexpressed in metastatic cancer cells. Treatment with the proposed tumor-targeted nanoscale-based nucleic acid and DDS led to the suppression of CD44 mRNA and protein, efficient induction of cell death, effective tumor shrinkage, and prevention of adverse side effects on healthy organs. CONCLUSION: We show a high therapeutic potential for combinatorial treatment of ovarian carcinoma with a novel DDS that effectively transports siRNA targeting to CD44 mRNA simultaneously with cytotoxic agents. Clin Cancer Res; 19(22); 6193-204. ©2013 AACR.

Antitumor and modeling studies of a penetratin-peptide that targets E2F-1 in small cell lung cancer.

E2F-1, a key transcription factor necessary for cell growth, DNA repair, and differentiation, is an attractive target for development of anticancer drugs in tumors that are E2F "oncogene addicted". We identified a peptide isolated from phage clones that bound tightly to the E2F-1 promoter consensus sequence. The peptide was coupled to penetratin to enhance cellular uptake. Modeling of the penetratin-peptide (PEP) binding to the DNA E2F-1 promoter demonstrated favorable interactions that also involved the participation of most of the penetratin sequence. The penetratin-peptide (PEP) demonstrated potent in vitro cytotoxic effects against a range of cancer cell lines, particularly against Burkitt lymphoma cells and small cell lung cancer (SCLC) cells. Further studies in the H-69 SCLC cell line showed that the PEP inhibited transcription of E2F-1 and also several important E2F-regulated enzymes involved in DNA synthesis, namely, thymidylate synthase, thymidine kinase, and ribonucleotide reductase. As the PEP was found to be relatively unstable in serum, it was encapsulated in PEGylated liposomes for in vivo studies. Treatment of mice bearing the human small cell lung carcinoma H-69 with the PEP encapsulated in PEGylated liposomes (PL-PEP) caused tumor regression without significant toxicity. The liposome encapsulated PEP has promise as an antitumor agent, alone or in combination with inhibitors of DNA synthesis.

Nanostructured lipid carriers as multifunctional nanomedicine platform for pulmonary co-delivery of anticancer drugs and siRNA.

We developed, synthesized, and tested a multifunctional nanostructured lipid nanocarrier-based system (NLCS) for efficient delivery of an anticancer drug and siRNA directly into the lungs by inhalation. The system contains: (1) nanostructured lipid carriers (NLC); (2) anticancer drug (doxorubicin or paclitaxel); (3) siRNA targeted to MRP1 mRNA as a suppressor of pump drug resistance; (4) siRNA targeted to BCL2 mRNA as a suppressor of nonpump cellular resistance and (5) a modified synthetic analog of luteinizing hormone-releasing hormone (LHRH) as a targeting moiety specific to the receptors that are overexpressed in the plasma membrane of lung cancer cells. The NLCS was tested in vitro using human lung cancer cells and in vivo utilizing mouse orthotopic model of human lung cancer. After inhalation, the proposed NLCS effectively delivered its payload into lung cancer cells leaving healthy lung tissues intact and also significantly decreasing the exposure of healthy organs when compared with intravenous injection. The NLCS showed enhanced antitumor activity when compared with intravenous treatment. The data obtained demonstrated high efficiency of proposed NLCS for tumor-targeted local delivery by inhalation of anticancer drugs and mixture of siRNAs specifically to lung cancer cells and, as a result, efficient suppression of tumor growth and prevention of adverse side effects on healthy organs.

Genotoxicity of different nanocarriers: possible modifications for the delivery of nucleic acids.

The prevention of cyto- and genotoxicity of nanocarriers is an important task in nanomedicine. In the present investigation, we, at the first time using similar experimental conditions, compared genotoxicity of nanocarriers with different composition, architecture, size, molecular weight and charge. Poly(ethylene glycol) polymers, neutral and cationic liposomes, micelles, poly(amindo amine) and poly(propyleneimine) dendrimers, quantum dots, mesoporous silica, and supermagnetic iron oxide (SPIO) nanoparticles were studied. All nanoparticles were used in non-cytotoxic concentrations. However, even in these concentrations, positively charged cationic liposomes, dendrimers, and SPIO nanoparticles induced genotoxicity leading to the significant formation of micronuclei in cells. Negatively charged and neutral nanocarriers were not genotoxic. A strong positive correlation was found between the number of formed micronuclei and the positive charge of nanocarriers. We proposed modifications of both types of dendrimers and SPIO nanoparticles that substantially decreased their genotoxicity and allowed for an efficient intracellular delivery of nucleic acids.