RESUMO
The purpose of this exploratory study was to investigate the combination of a radiopharmaceutical, nanoparticles and ultrasound (US) enhanced delivery to develop a clinically viable therapeutic strategy for tumours overexpressing the epidermal growth factor receptor (EGFR). Molecularly targeted radionuclides have great potential for cancer therapy but are sometimes associated with insufficient delivery resulting in sub-cytotoxic amounts of radioactivity being delivered to the tumour. Liposome formulations are currently used in the clinic to reduce the side effects and improve the pharmacokinetic profile of chemotherapeutic drugs. However, in contrast to non-radioactive agents, loading and release of radiotherapeutics from liposomes can be challenging in the clinical setting. US-activated cavitation agents such as microbubbles (MBs) have been used to release therapeutics from liposomes to enhance the distribution/delivery in a target area. In an effort to harness the benefits of these techniques, the development of a liposome loaded radiopharmaceutical construct for enhanced delivery via acoustic cavitation was studied. The liposomal formulation was loaded with peptide, human epidermal growth factor (HEGF), coupled to a chelator for subsequent radiolabelling with 111Indium ([111In]In3+), in a manner designed to be compatible with preparation in a radiopharmacy. Liposomes were efficiently radiolabelled (57%) within 1 h, with release of ~12% of the radiopeptide following a 20 s exposure to US-mediated cavitation in vitro. In clonogenic studies this level of release resulted in cytotoxicity specifically in cells over-expressing the epidermal growth factor receptor (EGFR), with over 99% reduction in colony survival compared to controls. The formulation extended the circulation time and changed the biodistribution compared to the non-liposomal radiopeptide in vivo, although interestingly the biodistribution did not resemble that of liposome constructs currently used in the clinic. Cavitation of MBs co-injected with liposomes into tumours expressing high levels of EGFR resulted in a 2-fold enhancement in tumour uptake within 20 min. However, owing to the poor vascularisation of the tumour model used the same level of uptake was achieved without US after 24 h. By combining acoustic-cavitation-sensitive liposomes with radiopharmaceuticals this research represents a new concept in achieving targeted delivery of radiopharmaceuticals.
Assuntos
Radioisótopos de Índio , Lipossomos , Fator de Crescimento Epidérmico , Humanos , Distribuição TecidualRESUMO
Nanomedicines allow active targeting of cancer for diagnostic and therapeutic applications through incorporation of multiple functional components. Frequently, however, clinical translation is hindered by poor intratumoural delivery and distribution. The application of physical stimuli to promote tumour uptake is a viable route to overcome this limitation. In this study, ultrasound-mediated cavitation of microbubbles was investigated as a mean of enhancing the delivery of a liposome designed for chemo-radionuclide therapy targeted to EGFR overexpressing cancer. Method: Liposomes (111In-EGF-LP-Dox) were prepared by encapsulation of doxorubicin (Dox) and surface functionalisation with Indium-111 tagged epidermal growth factor. Human breast cancer cell lines with high and low EGFR expression (MDA-MB-468 and MCF7 respectively) were used to study selectivity of liposomal uptake, subcellular localisation of drug payload, cytotoxicity and DNA damage. Liposome extravasation following ultrasound-induced cavitation of microbubbles (SonoVue®) was studied using a tissue-mimicking phantom. In vivo stability, pharmacokinetic profile and biodistribution were evaluated following intravenous administration of 111In-labelled, EGF-functionalised liposomes to mice bearing subcutaneous MDA-MB-468 xenografts. Finally, the influence of ultrasound-mediated cavitation on the delivery of liposomes into tumours was studied. Results: Liposomes were loaded efficiently with Dox, surface decorated with 111In-EGF and showed selective uptake in MDA-MB-468 cells compared to MCF7. Following binding to EGFR, Dox was released into the intracellular space and 111In-EGF shuttled to the cell nucleus. DNA damage and cell kill were higher in MDA-MB-468 than MCF7 cells. Moreover, Dox and 111In were shown to have an additive cytotoxic effect in MDA-MB-468 cells. US-mediated cavitation increased the extravasation of liposomes in an in vitro gel phantom model. In vivo, the application of ultrasound with microbubbles increased tumour uptake by 66% (p<0.05) despite poor vascularisation of MDA-MB-468 xenografts (as shown by DCE-MRI). Conclusion:111In-EGF-LP-Dox designed for concurrent chemo-radionuclide therapy showed specificity for and cytotoxicity towards EGFR-overexpressing cancer cells. Delivery to tumours was enhanced by the use of ultrasound-mediated cavitation indicating that this approach has the potential to deliver cytotoxic levels of therapeutic radionuclide to solid tumours.
Assuntos
Antineoplásicos/administração & dosagem , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/radioterapia , Doxorrubicina/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Receptores ErbB/metabolismo , Radioisótopos de Índio/administração & dosagem , Animais , Antineoplásicos/química , Antineoplásicos/farmacocinética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Terapia Combinada , Doxorrubicina/química , Doxorrubicina/farmacocinética , Composição de Medicamentos , Sistemas de Liberação de Medicamentos/instrumentação , Receptores ErbB/genética , Feminino , Humanos , Radioisótopos de Índio/química , Radioisótopos de Índio/farmacocinética , Lipossomos/química , Camundongos , Camundongos Nus , Distribuição Tecidual , UltrassomRESUMO
Radiolabelled, drug-loaded nanoparticles may combine the theranostic properties of radionuclides, the controlled release of chemotherapy and cancer cell targeting. Here, we report the preparation of poly(lactic-co-glycolic acid) (PLGA) nanoparticles surface conjugated to DTPA-hEGF (DTPA = diethylenetriaminepentaacetic acid, hEGF = human epidermal growth factor) and encapsulating the ruthenium-based DNA replication inhibitor and radiosensitizer Ru(phen)2(tpphz)2+ (phen = 1,10-phenanthroline, tpphz = tetrapyridophenazine) Ru1. The functionalized PLGA surface incorporates the metal ion chelator DTPA for radiolabelling and the targeting ligand for EGF receptor (EGFR). Nanoparticles radiolabelled with 111In are taken up preferentially by EGFR-overexpressing oesophageal cancer cells, where they exhibit radiotoxicity through the generation of cellular DNA damage. Moreover, nanoparticle co-delivery of Ru1 alongside 111In results in decreased cell survival compared to single-agent formulations; an effect that occurs through DNA damage enhancement and an additive relationship between 111In and Ru1. Substantially decreased uptake and radiotoxicity of nanoparticles towards normal human fibroblasts and oesophageal cancer cells with normal EGFR levels is observed. This work demonstrates nanoparticle co-delivery of a therapeutic radionuclide plus a ruthenium-based radiosensitizer can achieve combinational and targeted therapeutic effects in cancer cells that overexpress EGFR.