SPIONs: Superparamagnetic iron oxide-based nanoparticles for the delivery of microRNAi-therapeutics in cancer.

Non-coding RNA (ncRNA)-based therapeutics that induce RNA interference (RNAi), such as microRNAs (miRNAs), have drawn considerable attention as a novel class of targeted cancer therapeutics because of their capacity to specifically target oncogenes/protooncogenes that regulate key signaling pathways involved in carcinogenesis, tumor growth and progression, metastasis, cell survival, proliferation, angiogenesis, and drug resistance. However, clinical translation of miRNA-based therapeutics, in particular, has been challenging due to the ineffective delivery of ncRNA molecules into tumors and their uptake into cancer cells. Recently, superparamagnetic iron oxide-based nanoparticles (SPIONs) have emerged as highly effective and efficient for the delivery of therapeutic RNAs to malignant tissues, as well as theranostic (therapy and diagnostic) applications, due to their excellent biocompatibility, magnetic responsiveness, broad functional surface modification, safety, and biodistribution profiles. This review highlights recent advances in the use of SPIONs for the delivery of ncRNA-based therapeutics with an emphasis on their synthesis and coating strategies. Moreover, the advantages and current limitations of SPIONs and their future perspectives are discussed.

BET inhibition induces vulnerability to MCL1 targeting through upregulation of fatty acid synthesis pathway in breast cancer.

Therapeutic options for treatment of basal-like breast cancers remain limited. Here, we demonstrate that bromodomain and extra-terminal (BET) inhibition induces an adaptive response leading to MCL1 protein-driven evasion of apoptosis in breast cancer cells. Consequently, co-targeting MCL1 and BET is highly synergistic in breast cancer models. The mechanism of adaptive response to BET inhibition involves the upregulation of lipid synthesis enzymes including the rate-limiting stearoyl-coenzyme A (CoA) desaturase. Changes in lipid synthesis pathway are associated with increases in cell motility and membrane fluidity as well as re-localization and activation of HER2/EGFR. In turn, the HER2/EGFR signaling results in the accumulation of and vulnerability to the inhibition of MCL1. Drug response and genomics analyses reveal that MCL1 copy-number alterations are associated with effective BET and MCL1 co-targeting. The high frequency of MCL1 chromosomal amplifications (>30%) in basal-like breast cancers suggests that BET and MCL1 co-targeting may have therapeutic utility in this aggressive subtype of breast cancer.

miRacle of microRNA-Driven Cancer Nanotherapeutics.

MicroRNAs (miRNAs) are non-protein-coding RNA molecules 20-25 nucleotides in length that can suppress the expression of genes involved in numerous physiological processes in cells. Accumulating evidence has shown that dysregulation of miRNA expression is related to the pathogenesis of various human diseases and cancers. Thus, stragegies involving either restoring the expression of tumor suppressor miRNAs or inhibiting overexpressed oncogenic miRNAs hold potential for targeted cancer therapies. However, delivery of miRNAs to tumor tissues is a challenging task. Recent advances in nanotechnology have enabled successful tumor-targeted delivery of miRNA therapeutics through newly designed nanoparticle-based carrier systems. As a result, miRNA therapeutics have entered human clinical trials with promising results, and they are expected to accelerate the transition of miRNAs from the bench to the bedside in the next decade. Here, we present recent perspectives and the newest developments, describing several engineered natural and synthetic novel miRNA nanocarrier formulations and their key in vivo applications and clinical trials.

RNAi-based therapeutics and tumor targeted delivery in cancer.

Over the past decade, non-coding RNA-based therapeutics have proven as a great potential for the development of targeted therapies for cancer and other diseases. The discovery of the critical function of microRNAs (miRNAs) has generated great excitement in developing miRNA-based therapies. The dysregulation of miRNAs contributes to the pathogenesis of various human diseases and cancers by modulating genes that are involved in critical cellular processes, including cell proliferation, differentiation, apoptosis, angiogenesis, metastasis, drug resistance, and tumorigenesis. miRNA (miRNA mimic, anti-miRNA/antagomir) and small interfering RNA (siRNA) can inhibit the expression of any cancer-related genes/mRNAs with high specificity through RNA interference (RNAi), thus representing a remarkable therapeutic tool for targeted therapies and precision medicine. siRNA and miRNA-based therapies have entered clinical trials and recently three novel siRNA-based therapeutics were approved by the Food and Drug Administration (FDA), indicating the beginning of a new era of targeted therapeutics. The successful clinical applications of miRNA and siRNA therapeutics rely on safe and effective nanodelivery strategies for targeting tumor cells or tumor microenvironment. For this purpose, promising nanodelivery/nanoparticle-based approaches have been developed using a variety of molecules for systemic administration and improved tumor targeted delivery with reduced side effects. In this review, we present an overview of RNAi-based therapeutics, the major pharmaceutical challenges, and the perspectives for the development of promising delivery systems for clinical translation. We also highlight the passive and active tumor targeting nanodelivery strategies and primarily focus on the current applications of nanoparticle-based delivery formulations for tumor targeted RNAi molecules and their recent advances in clinical trials in human cancers.

Correction to "Target-Driven Design of a Coumarinyl Chalcone Scaffold Based Novel EF2 Kinase Inhibitor Suppresses Breast Cancer Growth In Vivo".

[This corrects the article DOI: 10.1021/acsptsci.1c00030.].

Target-Driven Design of a Coumarinyl Chalcone Scaffold Based Novel EF2 Kinase Inhibitor Suppresses Breast Cancer Growth In Vivo.

Eukaryotic elongation factor 2 kinase (eEF-2K) is an unusual alpha kinase involved in protein synthesis through phosphorylation of elongation factor 2 (EF2). eEF-2K is highly overexpressed in breast cancer, and its activity is associated with significantly shortened patient survival and proven to be a potential molecular target in breast cancer. The crystal structure of eEF-2K remains unknown, and there is no potent, safe, and effective inhibitor available for clinical applications. We designed and synthesized several generations of potential inhibitors. The effect of the inhibitors at the binding pocket of eEF-2K was analyzed after developing a 3D target model by using a domain of another α-kinase called myosin heavy-chain kinase A (MHCKA) that closely resembles eEF-2K. In silico studies showed that compounds with a coumarin-chalcone core have high predicted binding affinities for eEF-2K. Using in vitro studies in highly aggressive and invasive (MDA-MB-436, MDA-MB-231, and BT20) and noninvazive (MCF-7) breast cancer cells, we identified a lead compound that was highly effective in inhibiting eEF-2K activity at submicromolar concentrations and at inhibiting cell proliferation by induction of apoptosis with no toxicity in normal breast epithelial cells. In vivo systemic administration of the lead compound encapsulated in single lipid-based liposomal nanoparticles twice a week significantly suppressed growth of MDA-MB-231 tumors in orthotopic breast cancer models in nude mice with no observed toxicity. In conclusion, our study provides a highly potent and in vivo effective novel small-molecule eEF-2K inhibitor that may be used as a molecularly targeted therapy breast cancer or other eEF-2K-dependent tumors.

Magnetically responsive, sorafenib loaded alginate microspheres for hepatocellular carcinoma treatment.

This study aimed to develop sorafenib loaded magnetic microspheres for the treatment of hepatocellular carcinoma. To achieve this goal, superparamagnetic iron oxide nanoparticles (SPIONs) were synthesised and encapsulated in alginate microspheres together with an antineoplastic agent, sorafenib. In the study, firstly SPIONs were synthesised and characterised by dynamic light scattering, energy-dispersive X-ray spectroscopy, and scanning electron microscopy. Then, alginate-SPIONs microspheres were developed, and further characterised by electron spin resonance spectrometer and vibrating sample magnetometer. Besides the magnetic properties of SPIONs, alginate microspheres with SPIONs were also found to have magnetic properties. The potential use of microspheres in hyperthermia treatment was then investigated and an increase of about 4°C in the environment was found out. Drug release studies and cytotoxicity tests were performed after sorafenib was encapsulated into the magnetic microspheres. According to release studies, sorafenib has been released from microspheres for 8 h. Cytotoxicity tests showed that alginate-SPION-sorafenib microspheres were highly effective against cancerous cells and promising for cancer therapy.

Silencing of survivin and cyclin B1 through siRNA-loaded arginine modified calcium phosphate nanoparticles for non-small-cell lung cancer therapy.

With the development of nanotechnology, various drug delivery systems including inorganic nanoparticles, liposomes, polymers, etc. have been developed over the past decade. Some of these nanoparticles are also forthcoming candidates for the successful delivery of small interfering RNA (siRNA) for targeted gene silencing. Upon its discovery, siRNA was perceived as a highly promising agent in the treatment of various diseases. However, it could not exhibit the expected clinical outcomes owing to the unfavorable challenges during delivery. One such challenge was identified as the lack of an effective carrier. Among the carriers, calcium phosphate (CaP) nanoparticles have attracted remarkable attention due to the superior biochemical properties and hold great promise for siRNA. It is well known that synthesis conditions influence the types of crystalline phases of CaPs as well as morphology. In this study, to address the influence of these parameters on the success of siRNA delivery, three different arginine (Arg) modified CaP nanoparticles having different chemical and morphological characteristics were synthesized as being the carriers of two specific siRNAs against survivin and cyclin B1. The functioning of CaP surfaces with Arg results in positive zeta potential on the surfaces. Functionalized nanoparticles have a higher loading capacity compared to unmodified particles, as they have a cationic surface that can be easily attached to negatively charged siRNAs. The gene silencing ability and the consequent in vitro antitumor activity of these CaP-Arg-siRNA complexes were investigated using A549 non-small-cell lung cancer cells. We found that high survivin and cyclin B1 expression is associated with worse survival in patients with lung cancer based on the Kaplan-Meier database. Considering the promoting role of survivin and cyclin B1 in cancer development and progression, CaP-Arg-siRNA mediated suppression of these genes resulted in a significant decrease in cell growth and induction of apoptosis. Our data suggest that all three CaP-Arg nanoparticles synthesized in this work can be used as safe and efficient nanocarriers for siRNA delivery, offering the opportunity to develop new therapeutic strategies for the treatment of lung cancer.

Designing siRNA-conjugated plant oil-based nanoparticles for gene silencing and cancer therapy.

In this study, the anticancer activities of two siRNA carriers were compared using a human lung adenocarcinoma epithelial cell line (A549). Firstly, poly(styrene)-graft-poly(linoleic acid) (PS-g-PLina) and poly(styrene)-graft-poly(linoleic acid)-graft-poly(ethylene glycol) (PS-g-PLina-g-PEG) graft copolymers were synthesized by free-radical polymerization. PS-PLina and PS-PLina-PEG nanoparticles (NPs) were prepared by solvent evaporation method and were then characterized. The size was found as 150 ± 10 nm for PS-PLina and 184 ± 6 nm for PS-PLina-PEG NPs. The NPs were functionalized with poly(l-lysine) (PLL) for c-myc siRNA conjugation. siRNA entrapment efficiencies were found in the range of 4-63% for PS-PLina-PLL and 6-42% for PS-PLina-PEG-PLL NPs. The short-term stability test was realised for 1 month. siRNA release profiles were also investigated. In vitro anticancer activity of siRNA-NPs was determined by MTT, flow cytometry, and fluorescence microscopy analyses. Obtained findings showed that both NPs systems were promising as siRNA delivery tool for lung cancer therapy.

Preparation and characterization of novel albumin-sericin nanoparticles as siRNA delivery vehicle for laryngeal cancer treatment.

Small interfering RNA (siRNA)-based gene silencing strategy has high potential on suppressing specific molecular targets, involved in cancer progression. However, the lack of an effective nanocarrier system that safely delivers siRNA to its target still limits the clinical applications of siRNA. This study aimed to develop albumin-sericin nanoparticles (Alb-Ser NPs) as a novel siRNA delivery system for laryngeal cancer treatment. Nanoparticle formulations composed of albumin and sericin at different ratios (1:1, 2:1, 1:2 w/w) were synthesized by desolvation method. The nanoparticles were modified with poly-L-lysine (PLL) for siRNA binding and decorated with hyaluronic acid (HA) to target laryngeal cancer cell line, Hep-2. HA/PLL/Alb-Ser NPs were individually loaded with siRNAs for casein kinase 2 (CK2), Absent, Small, or Homeotic-Like (ASH2L), and Cyclin D1 genes, which are overexpressed in Hep-2 cells. Downregulation of genes was confirmed by real-time PCR (RT-PCR). Size, morphological, and thermogravimetric characterizations revealed that Alb-Ser NPs having 2:1 (w/w) ratio are the most optimized formulation. Between 36.8 and 61.3% of siRNA entrapment efficiencies were achieved. HA/PLL-siRNA/Alb-Ser (2:1) NPs-mediated gene silencing resulted in a significant inhibition of cell growth and induction of apoptosis in cells. Our findings showed that HA/PLL/Alb-Ser (2:1) NPs were promising as a siRNA carrier.

Synthesis and characterization of amino acid-functionalized calcium phosphate nanoparticles for siRNA delivery.

Small interfering RNAs (siRNA) are short nucleic acid fragments of about 20-27 nucleotides, which can inhibit the expression of specific genes. siRNA based RNAi technology has emerged as a promising method for the treatment of a variety of diseases. However, a major limitation in the therapeutic use of siRNA is its rapid degradation in plasma and cellular cytoplasm, resulting in short half-life. In addition, as siRNA molecules cannot penetrate into the cell efficiently, it is required to use a carrier system for its delivery. In this work, chemically and morphologically different calcium phosphate (CaP) nanoparticles, including spherical-like hydroxyapatite (HA-s), needle-like hydroxyapatite (HA-n) and calcium deficient hydroxyapatite (CDHA) nanoparticles were synthesized by the sol-gel technique and the effects of particle characteristics on the binding capacity of siRNA were investigated. In order to enhance the gene loading efficiency, the nanoparticles were functionalized with arginine and the morphological and their structural characteristics were analyzed. The addition of arginine did not significantly change the particle sizes; however, it provided a significantly increased binding of siRNA for all types of CaP nanoparticles, as revealed by spectrophotometric measurements analysis. Arginine functionalized HA-n nanoparticles showed the best binding behavior with siRNA among the other nanoparticles due to its high, positive zeta potential (+18.8mV) and high surface area of Ca++ rich "c" plane. MTT cytotoxicity assays demonstrated that all the nanoparticles tested herein were biocompatible. Our results suggest that high siRNA entrapment in each of the three modified non-toxic CaP nanoparticles make them promising candidates as a non-viral vector for delivering therapeutic siRNA molecules to treat cancer.

Downregulation of ABCE1 via siRNA affects the sensitivity of A549 cells against chemotherapeutic agents.

ATP-binding cassette E1 (ABCE1) is involved in several biological functions in cancer cells such as tumor proliferation, antiapoptotic pathway and chemoresistance mechanism. This work aimed to investigate the alterations in chemosensitivity of A549 lung cancer cells for 5-Fluorouracil (5-FU) and irinotecan by silencing ABCE1 using specific small interfering RNAs (siRNA). The cells were treated with low doses of drugs, alone and also their combinations with ABCE1 siRNA. Cytotoxicity, cell proliferation and apoptosis/necrosis evaluations were performed in order to examine the effects of the combined treatment. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to confirm the downregulation of ABCE1. We also investigated the levels of B cell lymphoma 2 (Bcl-2) and mammalian target of rapamycin (mTOR) after the treatments by RT-PCR. Downregulation of ABCE1 improved the anticancer effects of 5-FU in inducing cell viability/proliferation inhibition and apoptosis/necrosis, whereas interestingly, almost did not change or slightly reduced the anticancer effects of irinotecan. ABCE1 expression significantly decreased by transfecting the cells with ABCE1 siRNA. Moreover, Bcl-2 and mTOR levels changed after the single or combined therapy in parallel with the apoptotic and antiproliferation effect. In conclusion, the simultaneous treatment of lung cancer cells with ABCE1 siRNA and 5-FU exhibited synergistic or additive effects; however, ABCE1 siRNA and irinotecan had unexpected antagonistic effects. Our findings demonstrate that the strategy of downregulation of ABCE1 may be included in conventional 5-FU chemotherapy for lung cancer, minimizing the usage of 5-FU at high dosages.

A microbial biosensor based on Lactobacillus delbruecki sp. bacterial cells for simultaneous determination of lactic and pyruvic acid.

The aim of this study was mainly to develop a microbial biosensor for the simultaneous determination of lactic acid and pyruvic acid. In developing biosensor, lyophilised Lactobacillus delbruecki sp. bacterial cells were immobilised with polypyrrole on a platin electrode surface using electropolymerization method. Lactate concentration was determined based on the differences in amperometric responses at cathodic peak (+0.2V) of potassium ferricyanide, whereas pyruvate concentration was determined using the differences at anodic peak (+0.1V). The response of biosensor showed linearity between 0.1 and 1.0mM for both of two substrates. Optimisation studies were carried out for amount of microorganism, pyrrole concentration, pH and temperature. In the characterisation studies, substrate specificity, interference effect of some substances on the biosensor response, and storage stability were established.