Tumor regression by combined immunotherapy and hyperthermia using magnetic nanoparticles in an experimental subcutaneous murine melanoma

Abstract

Immunotherapy (IT) has become an accepted therapeutic modality. We previously reported that intracellular hyperthermia (IH) using magnetic nanoparticles induces antitumor immunity. We undertook these studies in order to study the combined effects of IT and IH on melanoma. Magnetite cationic liposomes (MCLs) have a positive surface charge and generate heat in an alternating magnetic field (AMF) due to hysteresis loss. MCLs were injected into a B16 melanoma nodule in C57BL/6 mice, which were subjected to AMF for 30 min. The temperature at the tumor reached 43°C and was maintained by controlling the magnetic field intensity. At 24 h after IH, interleukin-2 (IL-2) or granulocyte macrophage-colony stimulating factor (GM-CSF) was injected directly into the melanoma. Mice were divided into six groups: group I (control), group II (IH), group III (IL-2), group IV (GM-CSF), group V (IH + + + +IL-2), and group VI (IH + + + +GM-CSF). yperthermia has been used for many years to treat a wide variety of tumors both in experimental animals and patients.1) The most commonly used heating method in clinical settings is capacitive heating using a radiofrequency (RF) electric field.2) However, specifically heating tumors by capacitive heating using an RF electric field is difficult, because the heating characteristics are influenced by various factors, such as tumor size, position of electrodes, and adhesion of electrodes at uneven sites. From a clinical point of view, a simple heat mediator is preferable for superficially seated tumors such as cutaneous melanoma. Magnetic nanoparticles have been applied to generate hyperthermia in an attempt to overcome these disadvantages.3, 4) These magnetic nanoparticles generate heat in an alternating magnetic field (AMF) due to hysteresis loss. 5)We have developed "magnetite cationic liposomes" (MCLs) for intracellular hyperthermia (IH).6, 7) MCLs were developed to show improved adsorption and accumulation in tumor cells and have a ten-fold higher affinity for tumor cells than for neutrally charged magnetoliposomes due to electrostatic interaction with the negatively charged cell membrane.6) In our in vitro experiments, 55% of MCLs were incorporated into cells and the intracellular magnetic nanoparticles could generate heat under AMF.6) We have also demonstrated the efficacy of IH using MCLs against T-9 rat glioma in an in vivo study. 8)We previously reported that our IH system induced antitumor immunity. 9) Hyperthermia is known to induce heat shock proteins (HSPs).10) Because expression of HSP70 protects cells from heat-induced apoptosis, 11) HSP70 expression has been considered to be a complicating factor in hyperthermia. On the other hand, recent reports have shown the importance of HSPs, such as HSP70, HSP90 and glucose-regulated protein 96, in immune reactions.12, 13) HSP-mediated antitumor immunity was reported to cause a vaccine effect due to HSP-peptide complexes purified from human melanoma cells.14) With regard to the mechanism of antitumor immunity induced by IH, we demonstrated th...

Intratumoral injection of immature dendritic cells enhances antitumor effect of hyperthermia using magnetic nanoparticles

“…We previously studied the H-2K b amount on B16 cells by flow cytometry. 39 Flow cytometric analysis using the anti-mouse H-2K b antibody revealed that B16 cells used in this study expressed a much lower amount of MHC class I antigen when compared with EL4 cells. In this study, we used poorly immunogenic B16 cells to assess the preclinical feasibility of a novel cancer therapy.…”

“…We previously reported that MCL-induced hyperthermia treatment at 43°C for 30 min induced necrotic cell death in B16 melanoma. 39 In this study, apoptosis detection assay by the terminal deoxynucreotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) method was performed, but no significant number of apoptotic cells was observed (data not shown). On the other hand, conventional hyperthermia systems are applied to induce apoptosis, not necrotic cell death.…”

“…We previously demonstrated the augmentation of MHC class I antigen presentation via HSP70 expression after hyperthermia. 17,39 The T-9 rat glioma cell surface presentation of MHC class I antigen increased in tandem with increased HSP70 expression and the immunogenicity of these cells was enhanced by hyperthermia, 17 which indicates a mechanism of antitumor immunity induced by hyperthermia. In addition, DCs have been reported to activate NK cells by secretion of cytokines (IL-12 and IL-18) and costimulatory molecules.…”

“…In a previous study, we reported the infiltration of CD8-positive T cells and Mac-3-positive macrophage/DCs into a tumor and the activation of cytotoxicity against a melanoma nodule after hyperthrmia. 39 Our hyperthermia system using MCLs and AMF induced antitumor immunity, but other cancer therapies, such as chemotherapy and radiotherapy, often decrease leukocytes and surgery also causes attenuation of immunity via hyposthenia. Our hyperthermia system is able to heat the tumor specifically by means of the MCLs (Fig.…”

Chains of Magnetosomes Extracted from AMB-1 Magnetotactic Bacteria for Application in Alternative Magnetic Field Cancer Therapy

“…In this case, the destruction or elimination of tumors would occur by increasing the tumor temperature typically within the range of 37-45 C for hyperthermia (1,2,5) or above 45 C for thermoablation, (3). In previous work, the heat has been induced using chemically synthesized nanoparticles, mainly in the form superparamagnetic iron oxide nanoparticles (SPION), which were either mixed in solution or mixed with cells or administered to a living organism (1)(2)(3)(4)(5)(6)(7)(8). The anti-tumoral activity of these heated nanoparticles has been evaluated both on animal models and clinically on humans (1)(2)(3)(4)(5)(6)(7)(8).…”