Biomedicines, Volume 5, Issue 3 (September 2017) – 23 articles

The platelet-rich fibrin–like matrix (PRFM) is usually prepared onsite and immediately used for regenerative therapy. Nonetheless, to meet the clinical necessity of preserving the PRFM without quality deterioration, we developed a method for preparation of PRFMs from short-term-stored whole blood (WB) samples. In this study, to evaluate the practical expiration date of storage, we extended the storage time of WB samples from 2 to 7 days and assessed the quality of the resulting PRFMs. WB samples collected with acid-citrate-dextrose were stored with gentle agitation at ambient temperature. To prepare PRFMs, the stored WB samples were mixed with CaCl₂ in glass tubes and centrifuged. Fibrin fiber networks, CD41 and CD62P expression, and Platelet Derived Growth Factor-BB (PDGF-BB) levels were examined by scanning electron microscopy (SEM), flow cytometry, and an Enzyme-Linked ImmunoSorbent Assay (ELISA), respectively. Long-term storage had no significant effect on either blood cell counts or platelet functions tested. The resulting PRFMs were visually identical to freshly prepared ones. PDGF-BB levels did not markedly decrease in a time-dependent manner. However, fibrin fibers gradually became thinner after storage. Although the coagulation activity may diminish, we propose that PRFMs can be prepared—without evident loss of quality—from WB samples stored for up to 7 days by our previously developed method. Full article

To date, no curative therapy is available for the treatment of most chronic inflammatory diseases such as atopic dermatitis, rheumatoid arthritis, or autoimmune disorders. Current treatments require a lifetime supply for patients to alleviate clinical symptoms and are unable to stop the course of disease. In contrast, a new series of immunotherapeutic agents targeting the Fc γ receptor I (CD64) have emerged and demonstrated significant clinical potential to actually resolving chronic inflammation driven by M1-type dysregulated macrophages. This subpopulation plays a key role in the initiation and maintenance of a series of chronic diseases. The novel recombinant M1-specific immunotherapeutics offer the prospect of highly effective treatment strategies as they have been shown to selectively eliminate the disease-causing macrophage subpopulations. In this review, we provide a detailed summary of the data generated, together with the advantages and the clinical potential of CD64-based targeted therapies for the treatment of chronic inflammatory diseases. Full article

The impact of emerging contaminants in the presence of active pharmaceutical pollutants plays an important role in the persistence and activity of environmental bacteria. This manuscript focuses on the impact of amoxicillin functionalized iron oxide nanoparticles on bacterial growth, in the presence of dissolved organic carbon (humic acid). The impact of these emerging contaminants individually and collectively on the growth profiles of model gram positive and negative bacteria was tracked for 24 h. Results indicate exposure to subinhibitory concentrations of amoxicillin bound iron oxide nanoparticles, in the presence of humic acid, increase bacterial growth in Pseudomonas aeruginosa and Staphylococcus aureus. Accelerated bacterial growth was associated with an increase in iron ions, which have been shown to influence upregulation of cellular metabolism. Though iron oxide nanoparticles are often regarded as benign, this work demonstrates the distinguishable impact of amoxicillin bound iron oxide nanoparticles in the presence of dissolved organic carbon. The results indicate differential impacts of combined contaminants on bacterial growth, having potential implications for environmental and human health. Full article

Significant progress has been made in understanding the nature of molecular interactions on the cell membrane. To decipher such interactions, molecular scaffolds can be engineered as a tool to modulate these events as they occur on the cell membrane. To guarantee reliability, scaffolds that function as modulators of cell membrane events must be coupled to a targeting moiety with superior chemical versatility. In this regard, nucleic acid aptamers are a suitable class of targeting moieties. Aptamers are inherently chemical in nature, allowing extensive site-specific chemical modification to engineer sensing molecules. Aptamers can be easily selected using a simple laboratory-based in vitro evolution method enabling the design and development of aptamer-based functional molecular scaffolds against wide range of cell surface molecules. This article reviews the application of aptamers as monitors and modulators of molecular interactions on the mammalian cell surface with the aim of increasing our understanding of cell-surface receptor response to external stimuli. The information gained from these types of studies could eventually prove useful in engineering improved medical diagnostics and therapeutics. Full article

Achyrocline satureioides (AS, family Asteraceae) is a plant widely used in traditional medicine for stomach, digestive, and gastrointestinal disorders during pregnancy. Studies regarding the indiscriminate use of plant infusions during pregnancy are limited. Recent reports have shown that chronic flavonoid supplementation induces toxicity in vivo and raises the mortality rates of healthy subjects. Therefore, we investigated whether supplementation of pregnant and lactating Wistar rats with two AS inflorescence extracts, consisting of an aqueous (AQ) extract similar to a tea (47 mg·kg⁻¹·day) and a hydroethanolic (HA) extract (35 mg·kg⁻¹·day⁻¹) with a higher flavonoid content, could induce redox-related side effects. Total reactive antioxidant potential (TRAP), thiobarbituric reactive species (TBARS), and total reduced thiol (SH) content were evaluated. Superoxide dismutase (SOD) and catalase (CAT) activities were additionally quantified. Our data suggest that both AQ and HA of AS inflorescence extracts may induce symptoms of toxicity in concentrations of (47 mg·kg⁻¹·day) and (35 mg·kg⁻¹·day⁻¹), respectively, in mothers regarding the delivery index and further decrease of neonatal survival. Of note, significant tissue-specific changes in maternal (liver, kidney, heart, and hippocampus) and pups (liver and kidney) biochemical oxidative parameters were observed. Our findings provide evidence that may support the need to control supplementation with the AQ of AS inflorescence extracts during gestation due to potential toxicity in vivo, which might be related, at least in part, to changes in tissue-specific redox homeostasis and enzymatic activity. Full article

Over the past decades there have been exciting and rapid developments of highly specific molecules to bind cancer antigens that are overexpressed on the surfaces of malignant cells. Nanomedicine aims to exploit these ligands to generate nanoscale platforms for targeted cancer therapy, and to do so with negligible off-target effects. Aptamers are structured nucleic acids that bind to defined molecular targets ranging from small molecules and proteins to whole cells or viruses. They are selected through an iterative process of amplification and enrichment called SELEX (systematic evolution of ligands by exponential enrichment), in which a combinatorial oligonucleotide library is exposed to the target of interest for several repetitive rounds. Nucleic acid ligands able to bind and internalize into malignant cells have been extensively used as tools for targeted delivery of therapeutic payloads both in vitro and in vivo. However, current cell targeting aptamer platforms suffer from limitations that have slowed their translation to the clinic. This is especially true for applications in which the cargo must reach the cytosol to exert its biological activity, as only a small percentage of the endocytosed cargo is typically able to translocate into the cytosol. Innovative technologies and selection strategies are required to enhance cytoplasmic delivery. In this review, we describe current selection methods used to generate aptamers that target cancer cells, and we highlight some of the factors that affect productive endosomal escape of cargoes. We also give an overview of the most promising strategies utilized to improve and monitor endosomal escape of therapeutic cargoes. The methods we highlight exploit tools and technologies that can potentially be incorporated in the SELEX process. Innovative selection protocols may identify aptamers with extended biological functionalities that allow effective cytosolic translocation of therapeutics. This in turn may facilitate successful translation of these platforms into clinical applications. Full article

Transcription factors of the nuclear factor κB (NF-κB) family are central coordinating regulators of the host defence responses to stress, injury and infection. Aberrant NF-κB activation also contributes to the pathogenesis of some of the most common current threats to global human health, including chronic inflammatory diseases, autoimmune disorders, diabetes, vascular diseases and the majority of cancers. Accordingly, the NF-κB pathway is widely considered an attractive therapeutic target in a broad range of malignant and non-malignant diseases. Yet, despite the aggressive efforts by the pharmaceutical industry to develop a specific NF-κB inhibitor, none has been clinically approved, due to the dose-limiting toxicities associated with the global suppression of NF-κB. In this review, we summarise the main strategies historically adopted to therapeutically target the NF-κB pathway with an emphasis on oncology, and some of the emerging strategies and newer agents being developed to pharmacologically inhibit this pathway. Full article

Aptamers are high affinity single-stranded DNA/RNA molecules, produced by a combinatorial procedure named SELEX (Systematic Evolution of Ligands by Exponential enrichment), that are emerging as promising diagnostic and therapeutic tools. Among selection strategies, procedures using living cells as complex targets (referred as “cell-SELEX”) have been developed as an effective mean to generate aptamers for heavily modified cell surface proteins, assuring the binding of the target in its native conformation. Here we give an up-to-date overview on cell-SELEX technology, discussing the most recent advances with a particular focus on cancer cell targeting. Examples of the different protocol applications and post-SELEX strategies will be briefly outlined. Full article

Hematopoietic cell transplant is a potentially curative procedure for many benign and malignant conditions. The efficacy of allogeneic transplant relies in part on the cytotoxicity of the conditioning regimen and the graft versus tumor effect mediated by alloreactive donor T cells; the same cells are also implicated in the development of graft versus host disease (GVHD). Selective identification and depletion of the T cells implicated in GVHD, while preserving the T cells responsible for graft versus tumor effect has been the focus of many research groups in the recent years. Here we briefly review the physiology of T cells in transplantation, and comment on a recent clinical trial published by Bleakly et al. using a novel way of graft engineered allograft via naïve T cell depletion. Full article

Widespread microRNA (miRNA) repression is a phenomenon observed in mammals after exposure to cigarette smoke and in many types of cancer. A comprehensive reduction in miRNA expression after treatment with the hormone estrogen has also previously been described. Here, we reveal a conserved association of miRNA downregulation after estrogen exposure in zebrafish, mouse, and human breast cancer cell line, with a high guanine content in the terminal loop sequences of their precursors, and offer a possible link between estrogen-related miRNA-adducts formation and carcinogenesis. We also show common gene expression patterns shared by breast cancer tumors and estrogen-treated zebrafish, suggesting that this organism can be used as a powerful model system for the study of human breast cancer. Full article

Oncolytic viruses would ideally be of use for systemic therapy to treat disseminated cancer. To do this safely, this may require multiple layers of cancer specificity. The pharmacology and specificity of oncolytic adenoviruses can be modified by (1) physical retargeting, (2) physical detargeting, (3) chemical shielding, or (4) by modifying the ability of viral early gene products to selectively activate in cancer versus normal cells. We explored the utility of these approaches with oncolytic adenovirus serotype 5 (Ad5) in immunocompetent Syrian hamsters bearing subcutaneous HaK tumors. After a single intravenous injection to reach the distant tumors, the physically hepatocyte-detargeted virus Ad5-hexon-BAP was more effective than conditionally replicating Ad5-dl1101/07 with mutations in its E1A protein. When these control or Ad5 treated animals were treated a second time by intratumoral injection, prior exposure to Ad5 did not affect tumor growth, suggesting that anti-Ad immunity neither prevented treatment nor amplified anti-tumor immune responses. Ad5-dl1101/07 was next chemically shielded with polyethylene glycol (PEG). While 5 kDa of PEG blunted pro-inflammatory IL-6 production induced by Ad5-dl1101/07, this shielding reduced Ad oncolytic activity. Full article

Synthetic nucleic acid ligands (aptamers) have emerged as effective delivery tools for many therapeutic oligonucleotide-based drugs, including small interfering RNAs (siRNAs). In this review, we summarize recent progress in the aptamer selection technology that has made possible the identification of cell-specific, cell-internalizing aptamers for the cell-targeted delivery of therapeutic oligonucleotides. In addition, we review the original, proof-of-concept aptamer-siRNA delivery studies and discuss recent advances in aptamer-siRNA conjugate designs for applications ranging from cancer therapy to the development of targeted antivirals. Challenges and prospects of aptamer-targeted siRNA drugs for clinical development are further highlighted. Full article

Plant-made biopharmaceuticals have long been considered a promising technology for providing inexpensive and efficacious medicines for developing countries, as well as for combating pandemic infectious diseases and for use in personalized medicine. Plant virus expression vectors produce high levels of pharmaceutical proteins within a very short time period. Recently, plant viruses have been employed as nanoparticles for novel forms of cancer treatment. This review provides a glimpse into the development of plant virus expression systems both for pharmaceutical production as well as for immunotherapy. Full article

Overwhelming evidence indicates that aspirin and related non-steroidal anti-inflammatory drugs (NSAIDs) have anti-tumour activity and the potential to prevent cancer, particularly colorectal cancer. However, the mechanisms underlying this effect remain hypothetical. Dysregulation of the nuclear factor-kappaB (NF-κB) transcription factor is a common event in many cancer types which contributes to tumour initiation and progression by driving expression of pro-proliferative/anti-apoptotic genes. In this review, we will focus on the current knowledge regarding NSAID effects on the NF-κB signalling pathway in pre-cancerous and cancerous lesions, and the evidence that these effects contribute to the anti-tumour activity of the agents. The nuclear organelle, the nucleolus, is emerging as a central regulator of transcription factor activity and cell growth and death. Nucleolar function is dysregulated in the majority of cancers which promotes cancer growth through direct and indirect mechanisms. Hence, this organelle is emerging as a promising target for novel therapeutic agents. Here, we will also discuss evidence for crosstalk between the NF-κB pathway and nucleoli, the role that this cross-talk has in the anti-tumour effects of NSAIDs and ways forward to exploit this crosstalk for therapeutic purpose. Full article

Angiogenesis is a critical process in numerous diseases, and intervention in neovascularization has therapeutic value in several disease settings, including ocular diseases, arthritis, and in tumor progression and metastatic spread. Various vascular targeting agents have been developed, including those that inhibit growth factor receptor tyrosine kinases, blocking antibodies that interfere with receptor signal transduction, and strategies that trap growth factor ligands. Limited anti-tumor efficacy studies have suggested that the targeted delivery of the human pro-apoptotic molecule Granzyme B to tumor cells has significant potential for cancer treatment. Here, we review biological vascular targeting agents, and describe a unique vascular targeting agent composed of Granzyme B and the VEGF receptor ligand VEGF₁₂₁. The fusion protein GrB/VEGF₁₂₁ demonstrates cytotoxicity at nanomolar or sub-nanomolar levels, excellent pharmacokinetic and efficacy profiles, and has significant therapeutic potential targeting tumor vasculature. Full article

Nucleic acid aptamers are short RNA- or DNA-based affinity reagents typically selected from combinatorial libraries to bind to a specific target such as a protein, a small molecule, whole cells or even animals. Aptamers have utility in the development of diagnostic, imaging and therapeutic applications due to their size, physico-chemical nature and ease of synthesis and modification to suit the application. A variety of oligonucleotide modifications have been used to enhance the stability of aptamers from nuclease degradation in vivo. The non-bridging oxygen atoms of the phosphodiester backbones of RNA and DNA aptamers can be substituted with one or two sulfur atoms, resulting in thioaptamers with phosphorothioate or phosphorodithioate linkages, respectively. Such thioaptamers are known to have increased binding affinity towards their target, as well as enhanced resistance to nuclease degradation. In this review, we discuss the development of phosphorothioate chemistry and thioaptamers, with a brief review of selection methods. Full article

The increasing occurrence of antibiotic-resistant Staphylococcus (S.) aureus tremendously limits the antibiotic-based treatment options; therefore, an open discussion of alternative treatment strategies is urgently needed. The use of naturally derived materials might become a more promising concept, not only as directly acting antimicrobials, but also for stimulation of the immune system. Costa Rican plant extracts were screened for their ability to enhance the antimicrobial activity of human blood-derived cells against S. aureus infections. We identified three plant extracts which significantly reduced the growth of S. aureus in the presence of human blood without directly acting as antibacterials: Byrsonima crassifolia acetone bark extract, Mandevilla veraguasensis acetone vine extract and Verbesina oerstediana acetone bark extract (VEOEBA). The effect of VEOEBA was studied in more detail, and revealed that VEOEBA increases the antimicrobial activity of neutrophils by enhancing the formation of neutrophil extracellular traps. Full article

After more than a decade of preclinical and clinical development, therapeutic infusion of mesenchymal stromal cells is now a leading investigational strategy for the treatment of acute graft-versus-host disease (GVHD). While their clinical use continues to expand, it is still unknown which of their immunomodulatory properties contributes most to their therapeutic activity. Herein we describe the proposed mechanisms, focusing on the inhibitory activity of mesenchymal stromal cells (MSCs) at immunologic checkpoints. A deeper understanding of the mechanism of action will allow us to design more effective treatment strategies. Full article

Application of Gold nanoparticles and Cold Atmospheric plasma as a targeted therapeutic adjunct has been widely investigated separately in cancer therapy. Gold nanoparticles, with their biocompatibility, lower cytotoxicity and superior efficacy, are becoming substantially more significant in modern cancer therapy. Likewise, cold atmospheric plasma, with rich reactive species including reactive oxygen species (ROS) and reactive nitrogen species (RNS), is being explored to selectively target and kill cancer cells, making them a promising anticancer agent. Recent scientific studies have shown that there is a potential synergy between these two aspects. Induction of apoptosis/necrosis due to oxidative stress may be a probable mechanism of their cytotoxic effect. The synergetic effect of the two therapeutic approaches could be tantamount to maximized targeted efficacy on the treatment of diseases like cancer. Full article

Chondroitin-sulfate proteoglycan 4 (CSPG4) is a transmembrane glycoprotein overexpressed on malignant cells in several cancer types with only limited expression on normal cells. CSPG4 is implicated in several signaling pathways believed to drive cancer progression, particularly proliferation, motility and metastatic spread. Expression may serve as a prognostic marker for survival and risk of relapse in treatment-resistant malignancies including melanoma, triple negative breast cancer, rhabdomyosarcoma and acute lymphoblastic leukemia. This tumor-associated overexpression of CSPG4 points towards a highly promising therapeutic target for antibody-guided cancer therapy. Monoclonal αCSPG4 antibodies have been shown to inhibit cancer progression by blocking ligand access to the CSPG4 extracellular binding sites. Moreover, CSPG4-directed antibody conjugates have been shown to be selectively internalized by CSPG4-expressing cancer cells via endocytosis. CSPG4-directed immunotherapy may be approached in several ways, including: (1) antibody-based fusion proteins for the selective delivery of a pro-apoptotic factors such as tumor necrosis factor-related apoptosis-inducing ligand to agonistic death receptors 4 and 5 on the cell surface; and (2) CSPG4-specific immunotoxins which bind selectively to diseased cells expressing CSPG4, are internalized by them and induce arrest of biosynthesis, closely followed by initiation of apoptotic signaling. Here we review various methods of exploiting tumor-associated CSPG4 expression to improve targeted cancer therapy. Full article

Some of the most promising small molecule toxins used to generate antibody drug conjugates (ADCs) include anti-mitotic agents (e.g., auristatin and its derivatives) which are designed to attack cancerous cells at their most vulnerable state during mitosis. We were interested in identifying a human cystostatic protein eventually showing comparable activities and allowing the generation of corresponding targeted fully human cytolytic fusion proteins. Recently, we identified the human microtubule associated protein tau (MAP tau), which binds specifically to tubulin and modulates the stability of microtubules, thereby blocking mitosis and presumably vesicular transport. By binding and stabilizing polymerized microtubule filaments, MAP tau-based fusion proteins skew microtubule dynamics towards cell cycle arrest and apoptosis. This biological activity makes rapidly proliferating cells (e.g., cancer and inflammatory cells) an excellent target for MAP tau-based targeted treatments. Their superior selectivity for proliferating cells confers additional selectivity towards upregulated tumor-associated antigens at their surface, thereby preventing off-target related toxicity against normal cells bearing tumor-associated antigens at physiologically normal to low levels. In this review, we highlight recent findings on MAP tau-based targeted cytolytic fusion proteins reported in preclinical immunotherapeutic studies. Full article

Graft versus host disease (GVHD) represents a major complication of allogeneic hematopoietic stem cell transplantation (allo HCT). Graft cellular manipulation has been used to mitigate the risk of GVHD. The αβ T cells are considered the primary culprit for causing GVHD therefore depletion of this T cell subset emerged as a promising cellular manipulation strategy to overcome the human leukocyte antigen (HLA) barrier of haploidentical (haplo) HCT. This approach is also being investigated in HLA-matched HCT. In several studies, αβ T cell depletion HCT has been performed without pharmacologic GVHD prophylaxis, thus unleashing favorable effect of donor’s natural killer cells (NK) and γδ T cells. This article will discuss the evolution of this method in clinical practice and the clinical outcome as described in different clinical trials. Full article