Next to surgical resection, tumor ablation is a commonly used intervention in the treatment of solid tumors. Tumor ablation methods include thermal therapies, photodynamic therapy, and reactive oxygen species (ROS) producing agents. Thermal therapies induce tumor cell death via thermal energy and include radiofrequency, microwave, high intensity focused ultrasound, and cryoablation. Photodynamic therapy and ROS producing agents cause increased oxidative stress in tumor cells leading to apoptosis. While these therapies are safe and viable alternatives when resection of malignancies is not feasible, they do have associated limitations that prevent their widespread use in clinical applications. To improve the efficacy of these treatments, nanoparticles are being studied in combination with nonsurgical ablation regimens. In addition to better thermal effect on tumor ablation, nanoparticles can deliver anticancer therapeutics that show synergistic anti-tumor effect in the presence of heat and can also be imaged to achieve precision in therapy. Understanding the molecular mechanism of nanoparticle-mediated tumor ablation could further help engineer nanoparticles of appropriate composition and properties to synergize the ablation effect. This review aims to explore the various types of nonsurgical tumor ablation methods currently used in cancer treatment and potential improvements by nanotechnology applications.

To address efficacy issues of cancer diagnosis and chemotherapy, we have developed a magnetic nanoparticle (MNP) formulation with combined drug delivery and imaging properties that can potentially be used in image-guided drug therapy. Our MNP consists of an iron-oxide magnetic core coated with oleic acid (OA) and stabilized with an amphiphilic block copolymer. Previously, we reported that our MNP formulation can provide prolonged contrast for tumor magnetic resonance imaging and can be loaded with hydrophobic anticancer agents for sustained drug delivery. In this study, we developed MNPs with optical imaging properties using new near-infrared dyes to quantitatively determine their long-term biodistribution and tumor localization with and without an external magnetic field in mice with xenograft breast tumors. MNPs localized slowly in the tumor, reaching a peak 48 h post injection before slowly declining over the next 11 days. One-hour exposure of the tumor to a magnetic field further enhanced MNP localization to tumors. Our MNPs can be developed with combined drug delivery and multimodal imaging properties to improve cancer diagnosis, provide sustained treatment, and monitor therapeutic effects in tumors over time.

Pharmaceutical intervention often requires therapeutics and/or their carriers to enter cells via endocytosis. Therefore, endocytic aberrancies resulting from disease represent a key, yet often overlooked, parameter in designing therapeutic strategies. In the case of lysosomal storage diseases (LSDs), characterized by lysosomal accumulation of undegraded substances, common clinical interventions rely on endocytosis of recombinant enzymes. However, the lysosomal defect in these diseases can affect endocytosis, as we recently demonstrated for clathrin-mediated uptake in patient fibroblasts with type A Niemann-Pick disease (NPD), a disorder characterized by acid sphingomylinase (ASM) deficiency and subsequent sphingomyelin storage. Using similar cells, we have examined if this is also the case for clathrin-independent pathways, including caveolae-mediated endocytosis and macropinocytosis. We observed impaired caveolin-1 enrichment at ligand-binding sites in NPD vs. wild type fibroblasts, corresponding with altered uptake of ligands and fluid-phase markers by both pathways. Similarly, aberrant lysosomal storage of sphingomyelin induced by pharmacological means also diminished uptake. Partial degradation of the lysosomal storage by untargeted recombinant ASM lead to partial uptake enhancement, while both parameters were restored to wild type levels by ASM delivery using model polymer nanocarriers specifically targeted to intercellular adhesion molecule-1 (anti-ICAM NCs). Carriers also restored caveolin-1 enrichment at ligand-binding sites and uptake through the caveolar and macropinocytic routes. These results demonstrate a link between lysosomal storage in NPD and alterations in clathrin-independent endocytosis, which could apply to other LSDs. Hence, this study shall guide the design of therapeutic approaches using viable endocytic pathways.

Many cellular activities and pharmaceutical interventions involve endocytosis and delivery to lysosomes for processing. Hence, lysosomal processing defects can cause cell and tissue damage, as in lysosomal storage diseases (LSDs) characterized by lysosomal accumulation of undegraded materials. This storage causes endocytic and trafficking alterations, which exacerbate disease and hinder treatment. However, there have been no systematic studies comparing different endocytic routes or LSDs. Here, we used genetic and pharmacological models of four LSDs (type A Niemann-Pick, type C Niemann-Pick, Fabry, and Gaucher diseases) and evaluated the pinocytic and receptor-mediated activity of the clathrin-, caveolae-, and macropinocytic routes. Bulk pinocytosis was diminished in all diseases, suggesting a generic endocytic alteration linked to lysosomal storage. Fluid-phase (dextran) and ligand (transferrin) uptake via the clathrin route were lower for all LSDs. Fluid-phase and ligand (cholera toxin B) uptake via the caveolar route were both affected, but less acutely in Fabry or Gaucher diseases. EGF-induced macropinocytosis was altered in Niemann-Pick cells, not other LSDs. Intracellular trafficking of ligands was also distorted in LSD vs. wild-type cells. The extent of these endocytic alterations paralleled the level of cholesterol storage in disease cell lines. Confirming this, pharmacological induction of cholesterol storage in wild-type cells disrupted endocytosis, and model therapeutics restored uptake in proportion to their efficacy in attenuating storage. This suggests a proportional and reversible relationship between endocytosis and lipid (cholesterol) storage. By analogy, the accumulation of biological material in other diseases, or foreign material from drugs or their carriers, may cause similar deficits, warranting further investigation.

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Quantum dots (QDs) are well-established as photoluminescent nanoparticle probes for in vitro or in vivo imaging, sensing, and even drug delivery. A critical component of this research is the need to reliably conjugate peptides, proteins, oligonucleotides, and other biomolecules to QDs in a controlled manner. In this chapter, we describe the conjugation of peptides to CdSe/ZnS QDs using a combination of polyhistidine self-assembly and hydrazone ligation. The former is a high-affinity interaction with the inorganic surface of the QD; the latter is a highly efficient and chemoselective reaction that occurs between 4-formylbenzoyl (4FB) and 2-hydrazinonicotinoyl (HYNIC) moieties. Two methods are presented for modifying peptides with these functional groups: (1) solid phase peptide synthesis; and (2) solution phase modification of pre-synthesized, commercial peptides. We further describe the aniline-catalyzed ligation of 4FB- and HYNIC-modified peptides, in the presence of a fluorescent label on the latter peptide, as well as subsequent assembly of the ligated peptide to water-soluble QDs. Many technical elements of these protocols can be extended to labeling peptides with other small molecule reagents. Overall, the bioconjugate chemistry is robust, selective, and modular, thereby potentiating the controlled conjugation of QDs with a diverse array of biomolecules for various applications.

Targeting of drug carriers to endocytic cell-receptors facilitates intracellular drug delivery. Carrier size and number of targeting moieties (valency) influence cell binding and uptake. However, how these parameters influence receptor-mediated cell-signaling (the link between binding and uptake) remains uncharacterized. We studied this using polymer carriers of different sizes and valencies, targeted to endothelial intercellular adhesion molecule-1 (ICAM-1), a marker overexpressed in many pathologies. Unexpectedly, induction of cell-signals (ceramide and PKC enrichment and activation) and uptake, were independent of: carrier avidity, total number of carriers bound per cell, cumulative cell-surface area occupied by carriers, number of targeting antibodies at the carrier-cell contact, and cumulative receptor engagement by all bound carriers. Instead, “valency density” (number of antibodies per carrier surface area) ruled signaling, and carrier size independently influenced uptake. These results are key to understanding the interplay between carrier design parameters and receptor-mediated signaling conducive to endocytosis, paramount for intracellular drug delivery.

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