Poly(2-oxazoline)-magnetite NanoFerrogels: Magnetic field responsive theranostic platform for cancer drug delivery and imaging.

Combining diagnosis and treatment approaches in one entity is the goal of theranostics for cancer therapy. Magnetic nanoparticles have been extensively used as contrast agents for nuclear magnetic resonance imaging as well as drug carriers and remote actuation agents. Poly(2-oxazoline)-based polymeric micelles, which have been shown to efficiently solubilize hydrophobic drugs and drug combinations, have high loading capacity (above 40% w/w) for paclitaxel. In this study, we report the development of novel theranostic system, NanoFerrogels, which is designed to capitalize on the magnetic nanoparticle properties as imaging agents and the poly(2-oxazoline)-based micelles as drug loading compartment. We developed six formulations with magnetic nanoparticle content of 0.3%-12% (w/w), with the z-average sizes of 85-130 nm and ξ-potential of 2.7-28.3 mV. The release profiles of paclitaxel from NanoFerrogels were notably dependent on the degree of dopamine grafting on poly(2-oxazoline)-based micelles. Paclitaxel loaded NanoFerrogels showed efficacy against three breast cancer lines which was comparable to free paclitaxel. They also showed improved tumor and lymph node accumulation and signal reduction in vivo (2.7% in tumor; 8.5% in lymph node) compared to clinically approved imaging agent ferumoxytol (FERAHEME®) 24 h after administration. NanoFerrogels responded to super-low frequency alternating current magnetic field (50 kA m-1, 50 Hz) which accelerated drug release from paclitaxel-loaded NanoFerrogels or caused death of cells loaded with NanoFerrogels. These proof-of-concept experiments demonstrate that NanoFerrogels have potential as remotely actuated theranostic platform for cancer diagnosis and treatment.

Redox control of enzymatic functions: The electronics of life's circuitry.

The field of redox biology has changed tremendously over the past 20 years. Formerly regarded as bi-products of the aerobic metabolism exclusively involved in tissue damage, reactive oxygen species (ROS) are now recognized as active participants of cell signaling events in health and in disease. In this sense, ROS and the more recently defined reactive nitrogen species (RNS) are, just like hormones and second messengers, acting as fundamental orchestrators of cell signaling pathways. The chemical modification of enzymes by ROS and RNS (that result in functional enzymatic alterations) accounts for a considerable fraction of the transient and persistent perturbations imposed by variations in oxidant levels. Upregulation of ROS and RNS in response to stress is a common cellular response that foments adaptation to a variety of physiologic alterations (hypoxia, hyperoxia, starvation, and cytokine production). Frequently, these are beneficial and increase the organisms' resistance against subsequent acute stress (preconditioning). Differently, the sustained ROS/RNS-dependent rerouting of signaling produces irreversible alterations in cellular functioning, often leading to pathogenic events. Thus, the duration and reversibility of protein oxidations define whether complex organisms remain "electronically" healthy. Among the 20 essential amino acids, four are particularly susceptible to oxidation: cysteine, methionine, tyrosine, and tryptophan. Here, we will critically review the mechanisms, implications, and repair systems involved in the redox modifications of these residues in proteins while analyzing well-characterized prototypic examples. Occasionally, we will discuss potential consequences of amino acid oxidation and speculate on the biologic necessity for such events in the context of adaptative redox signaling. © 2014 IUBMB Life, 66(3):167-181, 2014.

A cell-targeted photodynamic nanomedicine strategy for head and neck cancers.

Photodynamic therapy (PDT) holds great promise for the treatment of head and neck (H&N) carcinomas where repeated loco-regional therapy often becomes necessary due to the highly aggressive and recurrent nature of the cancers. While interstitial light delivery technologies are being refined for PDT of H&N and other cancers, a parallel clinically relevant research area is the formulation of photosensitizers in nanovehicles that allow systemic administration yet preferential enhanced uptake in the tumor. This approach can render dual-selectivity of PDT, by harnessing both the drug and the light delivery within the tumor. To this end, we report on a cell-targeted nanomedicine approach for the photosensitizer silicon phthalocyanine-4 (Pc 4), by packaging it within polymeric micelles that are surface-decorated with GE11-peptides to promote enhanced cell-selective binding and receptor-mediated internalization in EGFR-overexpressing H&N cancer cells. Using fluorescence spectroscopy and confocal microscopy, we demonstrate in vitro that the EGFR-targeted Pc 4-nanoformulation undergoes faster and higher uptake in EGFR-overexpressing H&N SCC-15 cells. We further demonstrate that this enhanced Pc 4 uptake results in significant cell-killing and drastically reduced post-PDT clonogenicity. Building on this in vitro data, we demonstrate that the EGFR-targeted Pc 4-nanoformulation results in significant intratumoral drug uptake and subsequent enhanced PDT response, in vivo, in SCC-15 xenografts in mice. Altogether our results show significant promise toward a cell-targeted photodynamic nanomedicine for effective treatment of H&N carcinomas.

A platelet-mimetic paradigm for metastasis-targeted nanomedicine platforms.

There is compelling evidence that, beyond their traditional role in hemostasis and thrombosis, platelets play a significant role in mediating hematologic mechanisms of tumor metastasis by directly and indirectly interacting with pro-metastatic cancer cells. With this rationale, we hypothesized that platelets can be an effective paradigm to develop nanomedicine platforms that utilize platelet-mimetic interaction mechanisms for targeted diagnosis and therapy of metastatic cancer cells. Here we report on our investigation of the development of nanoconstructs that interact with metastatic cancer cells via platelet-mimetic heteromultivalent ligand-receptor pathways. For our studies, pro-metastatic human breast cancer cell line MDA-MB-231 was studied for its surface expression of platelet-interactive receptors, in comparison to another low-metastatic human breast cancer cell line, MCF-7. Certain platelet-interactive receptors were found to be significantly overexpressed on the MDA-MB-231 cells, and these cells showed significantly enhanced binding interactions with active platelets compared to MCF-7 cells. Based upon these observations, two specific receptor interactions were selected, and corresponding ligands were engineered onto the surface of liposomes as model nanoconstructs, to enable platelet-mimetic binding to the cancer cells. Our model platelet-mimetic liposomal constructs showed enhanced targeting and attachment of MDA-MB-231 cells compared to the MCF-7 cells. These results demonstrate the promise of utilizing platelet-mimetic constructs in modifying nanovehicle constructs for metastasis-targeted drug as well as modifying surfaces for ex-vivo cell enrichment diagnostic technologies.

Optimization of a nanomedicine-based silicon phthalocyanine 4 photodynamic therapy (Pc 4-PDT) strategy for targeted treatment of EGFR-overexpressing cancers.

The current clinical mainstays for cancer treatment, namely, surgical resection, chemotherapy, and radiotherapy, can cause significant trauma, systemic toxicity, and functional/cosmetic debilitation of tissue, especially if repetitive treatment becomes necessary due to tumor recurrence. Hence there is significant clinical interest in alternate treatment strategies like photodynamic therapy (PDT) which can effectively and selectively eradicate tumors and can be safely repeated if needed. We have previously demonstrated that the second-generation photosensitizer Pc 4 (silicon phthalocyanine 4) can be formulated within polymeric micelles, and these micelles can be specifically targeted to EGFR-overexpressing cancer cells using GE11 peptide ligands, to enhance cell-specific Pc 4 delivery and internalization. In the current study, we report on the in vitro optimization of the EGFR-targeting, Pc 4 loading of the micellar nanoformulation, along with optimization of the corresponding photoirradiation conditions to maximize Pc 4 delivery, internalization, and subsequent PDT-induced cytotoxicity in EGFR-overexpressing cells in vitro. In our studies, absorption and fluorescence spectroscopy were used to monitor the cell-specific uptake of the GE11-decorated Pc 4-loaded micelles and the cytotoxic singlet oxygen production from the micelle-encapsulated Pc 4, to determine the optimum ligand density and Pc 4 loading. It was found that the micelle formulations bearing 10 mol % of GE11-modified polymer component resulted in the highest cellular uptake in EGFR-overexpressing A431 cells within the shortest incubation periods. Also, the loading of ∼ 50 µg of Pc 4 per mg of polymer in these micellar formulations resulted in the highest levels of singlet oxygen production. When formulations bearing these optimized parameters were tested in vitro on A431 cells for PDT effect, a formulation dose containing 400 nM Pc 4 and photoirradiation duration of 400 s at a fluence of 200 mJ/cm(2) yielded close to 100% cell death.

EGFR-mediated intracellular delivery of Pc 4 nanoformulation for targeted photodynamic therapy of cancer: in vitro studies.

In photodynamic therapy (PDT), the light activation of a photosensitizer leads to the generation of reactive oxygen species that can trigger various mechanisms of cell death. Harnessing this process within cancer cells enables minimally invasive yet targeted cancer treatment. With this rationale, here we demonstrate tumor-targeted delivery of a highly hydrophobic photosensitizer Pc 4 loaded within biocompatible poly(ethylene glycol)-poly(ɛ-caprolactone) block co-polymer micelles. The micelles were surface-modified with epidermal growth factor receptor (EGFR)-targeting GE11 peptides for active targeting of EGFR-overexpressing cancer cells, in vitro. Pc 4-loaded EGFR-targeted micelles were incubated with EGFR-overexpressing A431 epidermoid carcinoma cells for various time periods, to determine Pc 4 uptake by epifluorescence microscopy. The cells were subsequently photoirradiated, and PDT-induced cell death for various incubation periods was determined by MTT assay and fluorescence Live/Dead assay. Our results indicate that active EGFR targeting of the Pc 4-loaded micelles accelerates intracellular uptake of the drug. Consequently, this enhances the PDT-induced cytotoxicity within shorter time periods. FROM THE CLINICAL EDITOR: Photodynamic cancer therapy using Pc 4, a light activated and highly hydrophobic photosensitizer is demonstrated in this paper in vitro. Pc 4 was delivered in block-copolymer micelles surface-modified with GE11 peptides targeting EGFR-overexpressing cancer cells.

Correction to "Luteinizing Hormone Releasing Hormone-Targeted Cisplatin-Loaded Magnetite Nanoclusters for Simultaneous MR Imaging and Chemotherapy of Ovarian Cancer".

[This corrects the article PMC10317193.].

Enhancing Chimeric Antigen Receptor T Cell Anti-tumor Function through Advanced Media Design.

Effective chimeric antigen receptor (CAR)-T cell therapy is dependent on optimal cell culture methods conducive to the activation and expansion of T cells ex vivo, as well as infection with CAR. Media formulations used in CAR-T cell manufacturing have not been optimized for gene delivery, cell expansion, and overall potency. Bioactive components and derivatives that support the generation of functionally-competent T cell progeny with long-lasting persistence are largely undefined. Current media formulations rely on fetal bovine serum (FBS) or human serum (HS), which suffer from a lack of consistency or supply issues. We recognize that components of blood cellular fractions that are absent in serum may have therapeutic value. Here we investigate whether a concentrated growth factor extract, purified from human transfusion grade whole blood fractions, and marketed as PhysiologixTM xeno-free (XF) hGFC (Phx), supports CAR-T cell expansion and function. We show that Phx supports T cell proliferation in clinical and research-grade media. We also show that Phx treatment enhances lentiviral-mediated gene expression across a wide range of multiplicity of infections (MOIs). We compared the ability of anti-GD-2 CAR-T cells expanded ex vivo in medium conditioned with either Phx or HS to clear tumor burden in a human xenograft model of neuroblastoma. We show that T cells expanded in Phx have superior engraftment and potency in vivo, as well as CAR-induced cytolytic activity in vitro. Metabolomic profiling revealed several factors unique to Phx that may have relevance for CAR-T cell preclinical discovery, process development, and manufacturing. In particular, we show that carnosine, a biogenic amine modestly enriched in Phx relative to HS, enhances lentiviral gene delivery in activated T cells. By limiting extracellular acidification, carnosine enhances the metabolic fitness of T cells, shifting their metabolic profile from an acidic, stressed state toward an oxidative, energetic state. These findings are very informative regarding potential derivatives to include in medium customized for gene delivery and overall potency for T cell adoptive immunotherapies.

Remote Actuation of Magnetic Nanoparticles For Cancer Cell Selective Treatment Through Cytoskeletal Disruption.

Motion of micron and sub-micron size magnetic particles in alternating magnetic fields can activate mechanosensitive cellular functions or physically destruct cancer cells. However, such effects are usually observed with relatively large magnetic particles (>250 nm) that would be difficult if at all possible to deliver to remote sites in the body to treat disease. Here we show a completely new mechanism of selective toxicity of superparamagnetic nanoparticles (SMNP) of 7 to 8 nm in diameter to cancer cells. These particles are coated by block copolymers, which facilitates their entry into the cells and clustering in the lysosomes, where they are then magneto-mechanically actuated by remotely applied alternating current (AC) magnetic fields of very low frequency (50 Hz). Such fields and treatments are safe for surrounding tissues but produce cytoskeletal disruption and subsequent death of cancer cells while leaving healthy cells intact.

Luteinizing Hormone Releasing Hormone-Targeted Cisplatin-Loaded Magnetite Nanoclusters for Simultaneous MR Imaging and Chemotherapy of Ovarian Cancer.

Given the superior soft tissue contrasts obtained by MRI and the long residence times of magnetic nanoparticles (MNPs) in soft tissues, MNP-based theranostic systems are being developed for simultaneous imaging and treatment. However, development of such theranostic nanoformulations presents significant challenges of balancing the therapeutic and diagnostic functionalities in order to achieve optimum effect from both. Here we developed a simple theranostic nanoformulation based on magnetic nanoclusters (MNCs) stabilized by a bisphosphonate-modified poly(glutamic acid)-b-(ethylene glycol) block copolymer and complexed with cisplatin. The MNCs were decorated with luteinizing hormone releasing hormone (LHRH) to target LHRH receptors (LHRHr) overexpressed in ovarian cancer cells. The targeted MNCs significantly improved the uptake of the drug in cancer cells and decreased its IC50 compared to the nontargeted formulations. Also, the enhanced LHRHr-mediated uptake of the targeted MNCs resulted in enhancement in the T2-weighted negative contrast in cellular phantom gels. Taken together, the LHRH-conjugated MNCs show good potential as ovarian cancer theranostics.

Photodynamic nanomedicine in the treatment of solid tumors: perspectives and challenges.

Photodynamic therapy (PDT) is a promising treatment strategy where activation of photosensitizer drugs with specific wavelengths of light results in energy transfer cascades that ultimately yield cytotoxic reactive oxygen species which can render apoptotic and necrotic cell death. Without light the photosensitizer drugs are minimally toxic and the photoactivating light itself is non-ionizing. Therefore, harnessing this mechanism in tumors provides a safe and novel way to selectively eradicate tumor with reduced systemic toxicity and side effects on healthy tissues. For successful PDT of solid tumors, it is necessary to ensure tumor-selective delivery of the photosensitizers, as well as, the photoactivating light and to establish dosimetric correlation of light and drug parameters to PDT-induced tumor response. To this end, the nanomedicine approach provides a promising way towards enhanced control of photosensitizer biodistribution and tumor-selective delivery. In addition, refinement of nanoparticle designs can also allow incorporation of imaging agents, light delivery components and dosimetric components. This review aims at describing the current state-of-the-art regarding nanomedicine strategies in PDT, with a comprehensive narrative of the research that has been carried out in vitro and in vivo, with a discussion of the nanoformulation design aspects and a perspective on the promise and challenges of PDT regarding successful translation into clinical application.

EGF receptor-targeted nanocarriers for enhanced cancer treatment.

The 'nanomedicine' approach has revolutionized cancer therapy by enabling the packaging of therapeutic agents within engineered nanovehicles that can specifically accumulate within the tumor stroma and then be internalized within cancer cells, to render site-selective action while minimizing nonspecific uptake and harmful side effects. While the specific accumulation within the tumor stroma is rendered by the ability of the nanovehicles to passively permeate through the tumor's leaky vasculature, the cellular internalization is often achieved by exploiting receptor-mediated active endocytotic mechanisms using receptor-specific ligand decoration on the vehicle surface. To this end, a highly important receptor found in several cancers is the EGF receptor, which has been implicated in tumor aggression and proliferation. In this context, we provide a comprehensive review of the various approaches of ligand decorations on nanovehicles for active targeting to EGF receptors, and discuss their pros and cons towards optimizing the design of EGF receptor-targeted nanomedicine systems.

Delivery of the photosensitizer Pc 4 in PEG-PCL micelles for in vitro PDT studies.

The silicon phthalocyanine Pc 4 is a second-generation photosensitizer that has several properties superior to other photosensitizers currently approved by the FDA, and it has shown significant promise for photodynamic therapy (PDT) in several cancer cells in vitro and model tumor systems in vivo. However, because of the high hydrophobicity of Pc 4, its formulation for in vivo delivery and favorable biodistribution become challenging. To this end, we are studying encapsulation and delivery of Pc 4 in block copolymer micelles. Here, we report the development of biocompatible PEG-PCL micelle nanoparticles, encapsulation of Pc 4 within the micelle core by hydrophobic association with the PCL block, and in vitro PDT studies of the micelle-formulated Pc 4 in MCF-7c3 human breast cancer cells. Our studies demonstrate efficient encapsulation of Pc 4 in the micelles, intracellular uptake of the micelle-formulated Pc 4 in cells, and significant cytotoxic effect of the formulation upon photoirradiation. Quantitative estimation of the extent of Pc 4 loading in the micelles and the photocytotoxicity of the micelle-incorporated Pc 4 demonstrate the promise of our approach to develop a biocompatible nanomedicine platform for tumor-targeted delivery of Pc 4 for site-selective PDT.