RESUMEN
Currently, there is no gadolinium-based contrast agent available for conventional magnetic resonance imaging (MRI) detection of amyloidal beta (Aß) plaques in Alzheimer's disease (AD). Its timely finding would be vital for patient survival and quality of life. Curcumin (CUR), a common Indian spice effectively binds to Aß plaques which is a hallmark of AD. To address this binding, we have designed a novel nanoimaging agent (NIA) based on nature-derived poly(ß-l-malic acid) (PMLA) containing covalently attached gadolinium-DOTA(Gd-DOTA) and nature-derived CUR. The all-in-one agent recognizes and selectively binds to Aß plaques and is detected by MRI. It efficiently detected Aß plaques in human and mouse samples by an ex vivo staining. The method can be useful in clinic for safe and noninvasive diagnosis of AD.
Asunto(s)
Enfermedad de Alzheimer/diagnóstico , Medios de Contraste/química , Imagen por Resonancia Magnética , Malatos/química , Placa Amiloide/diagnóstico , Polímeros/química , Animales , Encéfalo/patología , Curcumina/análogos & derivados , Curcumina/química , Compuestos Heterocíclicos/química , Humanos , Ratones , Compuestos Organometálicos/químicaRESUMEN
Differential diagnosis of brain magnetic resonance imaging (MRI) enhancement(s) remains a significant problem, which may be difficult to resolve without biopsy, which can be often dangerous or even impossible. Such MRI enhancement(s) can result from metastasis of primary tumors such as lung or breast, radiation necrosis, infections, or a new primary brain tumor (glioma, meningioma). Neurological symptoms are often the same on initial presentation. To develop a more precise noninvasive MRI diagnostic method, we have engineered a new class of poly(ß-l-malic acid) polymeric nanoimaging agents (NIAs). The NIAs carrying attached MRI tracer are able to pass through the blood-brain barrier (BBB) and specifically target cancer cells for efficient imaging. A qualitative/quantitative "MRI virtual biopsy" method is based on a nanoconjugate carrying MRI contrast agent gadolinium-DOTA and antibodies recognizing tumor-specific markers and extravasating through the BBB. In newly developed double tumor xenogeneic mouse models of brain metastasis this noninvasive method allowed differential diagnosis of HER2- and EGFR-expressing brain tumors. After MRI diagnosis, breast and lung cancer brain metastases were successfully treated with similar tumor-targeted nanoconjugates carrying molecular inhibitors of EGFR or HER2 instead of imaging contrast agent. The treatment resulted in a significant increase in animal survival and markedly reduced immunostaining for several cancer stem cell markers. Novel NIAs could be useful for brain diagnostic MRI in the clinic without currently performed brain biopsies. This technology shows promise for differential MRI diagnosis and treatment of brain metastases and other pathologies when biopsies are difficult to perform.
Asunto(s)
Biopsia/métodos , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/patología , Encéfalo/patología , Imagen por Resonancia Magnética/métodos , Nanoconjugados , Nanomedicina/métodos , Animales , Secuencia de Bases , Barrera Hematoencefálica/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Transformación Celular Neoplásica , Diagnóstico Diferencial , Receptores ErbB/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Nanoconjugados/química , Metástasis de la Neoplasia , Oligonucleótidos Antisentido/química , Oligonucleótidos Antisentido/genética , Oligonucleótidos Antisentido/metabolismo , Receptor ErbB-2/metabolismo , Análisis de SupervivenciaRESUMEN
Multifunctional polymer nanoconjugates containing multiple components show great promise in cancer therapy, but in most cases complete analysis of each component is difficult. Polymalic acid (PMLA) based nanoconjugates have demonstrated successful brain and breast cancer treatment. They consist of multiple components including targeting antibodies, Morpholino antisense oligonucleotides (AONs), and endosome escape moieties. The component analysis of PMLA nanoconjugates is extremely difficult using conventional spectrometry and HPLC method. Taking advantage of the nature of polyester of PMLA, which can be cleaved by ammonium hydroxide, we describe a method to analyze the content of antibody and AON within nanoconjugates simultaneously using SEC-HPLC by selectively cleaving the PMLA backbone. The selected cleavage conditions only degrade PMLA without affecting the integrity and biological activity of the antibody. Although the amount of antibody could also be determined using the bicinchoninic acid (BCA) method, our selective cleavage method gives more reliable results and is more powerful. Our approach provides a new direction for the component analysis of polymer nanoconjugates and nanoparticles.
Asunto(s)
Malatos/química , Nanoconjugados/química , Polímeros/química , Antineoplásicos/química , Sistemas de Liberación de Medicamentos , Cinética , Trastuzumab/químicaRESUMEN
Professor Black and colleagues have been working to improve the quality and sensitivity of imaging in the early detection of conditions from brain tumors to Alzheimer's disease to enhance treatment protocols and patient management. Professor Black et al introduced nanoparticles to improve MRI imaging. These nanoparticles consist of poly (b-L- malic acid (PMLA)) conjugates with monoclonal antibodies ((mAbs)) and Gd-DOTA. These are known as MRI nano-imaging agents (NIA). Most importantly, they can penetrate the endothelial blood-brain barrier (BBB) to reach brain tumors (primary or metastasis). This is effective in cases of brain tumors or breast cancer or other cancers such as lung cancer and gastric cancer having HER2 and/or EGFR positive crossing BBB. By the covalent conjugation of MR contrast (NIA), the MRI virtual biopsy can differentiate brain tumors from infections or other brain pathological conditions. The brain's intrinsic natural fluorescence such as NADH, FAD, lipopigments and porphyrin in the brain tissue can be identified by using time resolved fluorescence spectroscopy (TRFS) which is operated through the use of ultra-short laser. TRFS produces various color bands to differentiate the tumor from normal brain tissue in real time and registers the data on a 3D map. This is significant, as this will provide a greatly improved assessment methodology of tissue type. Consequently, this will potentially result in shorter operation times as well as more satisfactory tumor removal. In the detection of Alzheimer disease, amyloid plaque is deposited in retina tissue (including the RGC, RNFL and inner plexiform layer) which can produce a fluorescence effect by using curcumin as a contrast. This is then shown by human retina amyloid imaging device. Immunotherapies with glatiramer acetate (GA) have been shown to reduce amyloid deposits in brain and retinal AB deposits in mice. The study of advanced imaging technology and techniques including NIA, TRFS and the detection of amyloid plaque in Alzheimer disease are very important approaches to create a new era for diagnostic and therapeutic management of brain tumors and other cancers (HER2 and/or EGFR positive). This pioneering work by Professor Black, and colleagues, gives rise to a new hope for cancer patients for targeted therapy and for immunotherapies in Alzheimer's disease.
RESUMEN
Esterification of microbial poly(malic acid) is performed with either ethanol or 1-butanol to obtain polymalate conjugates capable to form nanoparticles (100-350 nm). Degradation under physiological conditions takes place with release of malic acid and the corresponding alcohol as unique degradation products. The anticancer drugs Temozolomide and Doxorubicin are encapsulated in nanoparticles with efficiency of 17 and 37%, respectively. In vitro drug release assays show that Temozolomide is almost completely discharged in a few hours whereas Doxorubicin is steadily released along several days. Drug-loaded nano-particles show remarkable effectiveness against cancer cells. Partially ethylated poly(malic acid) nano-particles are those showing the highest cellular uptake.
Asunto(s)
Antibióticos Antineoplásicos , Antineoplásicos Alquilantes , Dacarbazina/análogos & derivados , Doxorrubicina , Malatos , Nanocápsulas/química , Polímeros , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacocinética , Antibióticos Antineoplásicos/farmacología , Antineoplásicos Alquilantes/química , Antineoplásicos Alquilantes/farmacocinética , Antineoplásicos Alquilantes/farmacología , Línea Celular Tumoral , Dacarbazina/química , Dacarbazina/farmacocinética , Dacarbazina/farmacología , Preparaciones de Acción Retardada/síntesis química , Preparaciones de Acción Retardada/química , Preparaciones de Acción Retardada/farmacocinética , Preparaciones de Acción Retardada/farmacología , Doxorrubicina/química , Doxorrubicina/farmacocinética , Doxorrubicina/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Malatos/química , Malatos/farmacocinética , Malatos/farmacología , Nanocápsulas/ultraestructura , Polímeros/química , Polímeros/farmacocinética , Polímeros/farmacología , TemozolomidaRESUMEN
Tumors with similar grade and morphology often respond differently to the same treatment because of variations in molecular profiling. To account for this diversity, personalized medicine is developed for silencing malignancy associated genes. Nano drugs fit these needs by targeting tumor and delivering antisense oligonucleotides for silencing of genes. As drugs for the treatment are often administered repeatedly, absence of toxicity and negligible immune response are desirable. In the example presented here, a nano medicine is synthesized from the biodegradable, non-toxic and non-immunogenic platform polymalic acid by controlled chemical ligation of antisense oligonucleotides and tumor targeting molecules. The synthesis and treatment is exemplified for human Her2-positive breast cancer using an experimental mouse model. The case can be translated towards synthesis and treatment of other tumors.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/tratamiento farmacológico , Malatos/administración & dosificación , Terapia Molecular Dirigida/métodos , Nanopartículas/administración & dosificación , Polímeros/administración & dosificación , Animales , Neoplasias de la Mama/enzimología , Femenino , Silenciador del Gen , Humanos , Malatos/síntesis química , Malatos/química , Malatos/metabolismo , Ratones , Ratones Desnudos , Nanopartículas/química , Oligonucleótidos Antisentido/administración & dosificación , Oligonucleótidos Antisentido/química , Oligonucleótidos Antisentido/genética , Physarum polycephalum/genética , Physarum polycephalum/metabolismo , Polímeros/síntesis química , Polímeros/química , Polímeros/metabolismo , Medicina de Precisión/métodos , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/biosíntesis , Receptor ErbB-2/genética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Engineered nanoparticles are widely used for delivery of drugs but frequently lack proof of safety for cancer patient's treatment. All-in-one covalent nanodrugs of the third generation have been synthesized based on a poly(ß-L-malic acid) (PMLA) platform, targeting human triple-negative breast cancer (TNBC). They significantly inhibited tumor growth in nude mice by blocking synthesis of epidermal growth factor receptor, and α4 and ß1 chains of laminin-411, the tumor vascular wall protein and angiogenesis marker. PMLA and nanodrug biocompatibility and toxicity at low and high dosages were evaluated in vitro and in vivo. The dual-action nanodrug and single-action precursor nanoconjugates were assessed under in vitro conditions and in vivo with multiple treatment regimens (6 and 12 treatments). The monitoring of TNBC treatment in vivo with different drugs included blood hematologic and immunologic analysis after multiple intravenous administrations. The present study demonstrates that the dual-action nanoconjugate is highly effective in preclinical TNBC treatment without side effects, supported by hematologic and immunologic assays data. PMLA-based nanodrugs of the Polycefin™ family passed multiple toxicity and efficacy tests in vitro and in vivo on preclinical level and may prove to be optimized and efficacious for the treatment of cancer patients in the future.
Asunto(s)
Antineoplásicos/administración & dosificación , Sistemas de Liberación de Medicamentos , Malatos/química , Polímeros/química , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Animales , Antineoplásicos/farmacología , Antineoplásicos/toxicidad , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Esquema de Medicación , Portadores de Fármacos/química , Evaluación Preclínica de Medicamentos , Receptores ErbB/antagonistas & inhibidores , Femenino , Humanos , Laminina/metabolismo , Ratones , Ratones Desnudos , Nanoconjugados , Nanopartículas , Poliésteres/química , Conejos , Neoplasias de la Mama Triple Negativas/patologíaRESUMEN
Breast cancer remains the second leading cause of cancer death among women in the United States. Breast cancer prognosis is particularly poor in case of tumors overexpressing the oncoprotein HER2/neu. A new nanobioconjugate of the Polycefin(TM) family of anti-cancer drugs based on biodegradable and non-toxic polymalic acid (PMLA) was engineered for a multi-pronged attack on HER2/neu-positive breast cancer cells. An antibody-cytokine fusion protein consisting of the immunostimulatory cytokine interleukin-2 (IL-2) genetically fused to an antibody specific for human HER2/neu [anti-HER2/neu IgG3-(IL-2)] was covalently attached to the PMLA backbone to target HER2/neu expressing tumors and ensure the delivery of IL-2 to the tumor microenvironment. Antisense oligonucleotides (AON) were conjugated to the nanodrug to inhibit the expression of vascular tumor protein laminin-411 in order to block tumor angiogenesis. It is shown that the nanobioconjugate was capable of specifically binding human HER2/neu and retained the biological activity of IL-2. We also showed the uptake of the nanobioconjugate into HER2/neu-positive breast cancer cells and enhanced tumor targeting in vivo. The nanobioconjugate exhibited marked anti-tumor activity manifested by significantly longer animal survival and significantly increased anti-HER2/neu immune response in immunocompetent mice bearing D2F2/E2 murine mammary tumors that express human HER2/neu. The combination of laminin-411 AON and antibody-cytokine fusion protein on a single polymeric platform results in a new nanobioconjugate that can act against cancer cells through inhibition of tumor growth and angiogenesis and the orchestration of an immune response against the tumor. The present Polycefin(TM) variant may be a promising agent for treating HER2/neu expressing tumors and demonstrates the versatility of the Polycefin(TM) nanobioconjugate platform.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Inmunoconjugados/uso terapéutico , Inmunoglobulina G/uso terapéutico , Interleucina-2/uso terapéutico , Malatos/uso terapéutico , Polímeros/uso terapéutico , Receptor ErbB-2/inmunología , Animales , Mama/efectos de los fármacos , Mama/inmunología , Mama/patología , Neoplasias de la Mama/inmunología , Neoplasias de la Mama/patología , Línea Celular Tumoral , Femenino , Humanos , Inmunización , Inmunoconjugados/inmunología , Inmunoconjugados/farmacocinética , Inmunoglobulina G/inmunología , Interleucina-2/inmunología , Interleucina-2/farmacocinética , Malatos/farmacocinética , Ratones , Ratones Desnudos , Polímeros/farmacocinética , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/farmacocinética , Proteínas Recombinantes de Fusión/uso terapéuticoRESUMEN
Anionic polymers are valuable components used in cosmetics and health sciences, especially in drug delivery, because of their chemical versatility and low toxicity. However, because of their highly negative charge they pose problems for penetration through hydrophobic barriers such as membranes. We have engineered anionic polymalic acid (PMLA) to penetrate biological membranes. PMLA copolymers of leucine ethyl ester (P/LOEt) or trileucine (P/LLL) show either pH-independent or pH-dependent activity for membrane penetration. We report here for the first time on the mechanisms which are different for those two copolymers. Formation of hydrophobic patches in either copolymer is detected by fluorescence techniques. The copolymers display distinctly different properties in solution and during membranolysis. P/LOEt copolymer binds to membrane as single molecules with high affinity, and induces leakage cooperatively through a mechanism known as "carpet" model, in which the polymer aligns at the surface throughout the entire process of membrane permeation. In contrast, P/LLL self-assembles to form an oligomer of 105 nm in a pH-dependent manner (pKa 5.5) and induces membrane leakage through a two-phase process: the concentration dependent first-phase of insertion of the oligomer into membrane followed by a concentration independent second-phase of rearrangement of the membrane-oligomer complex. The insertion of P/LLL is facilitated by hydrophobic interactions between trileucine side chains and lipids in the membrane core, resulting in transmembrane pores, through mechanism known as "barrel-stave" model. The understanding of the mechanism paves the way for future engineering of polymeric delivery systems with optimal cytoplasmic delivery efficiency and reduced systemic toxicity.
Asunto(s)
Malatos/farmacología , Polímeros/farmacología , Ésteres/química , Transferencia Resonante de Energía de Fluorescencia , Hidrodinámica , Concentración de Iones de Hidrógeno/efectos de los fármacos , Interacciones Hidrofóbicas e Hidrofílicas/efectos de los fármacos , Cinética , Liposomas/química , Malatos/síntesis química , Malatos/química , Membranas/anomalías , Microscopía Confocal , Oligopéptidos/química , Tamaño de la Partícula , Permeabilidad/efectos de los fármacos , Polímeros/síntesis química , Polímeros/química , Rodaminas/metabolismo , Soluciones , TemperaturaRESUMEN
Doxorubicin (DOX) is currently used in cancer chemotherapy to treat many tumors and shows improved delivery, reduced toxicity and higher treatment efficacy when being part of nanoscale delivery systems. However, a major drawback remains its toxicity to healthy tissue and the development of multi-drug resistance during prolonged treatment. This is why in our work we aimed to improve DOX delivery and reduce the toxicity by chemical conjugation with a new nanoplatform based on polymalic acid. For delivery into recipient cancer cells, DOX was conjugated via pH-sensitive hydrazone linkage along with polyethylene glycol (PEG) to a biodegradable, non-toxic and non-immunogenic nanoconjugate platform: poly(ß-l-malic acid) (PMLA). DOX-nanoconjugates were found stable under physiological conditions and shown to successfully inhibit in vitro cancer cell growth of several invasive breast carcinoma cell lines such as MDA-MB-231 and MDA-MB- 468 and of primary glioma cell lines such as U87MG and U251.
Asunto(s)
Antibióticos Antineoplásicos , Neoplasias de la Mama/tratamiento farmacológico , Doxorrubicina , Sistemas de Liberación de Medicamentos , Glioma/tratamiento farmacológico , Malatos , Nanoconjugados/química , Polímeros , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacocinética , Antibióticos Antineoplásicos/farmacología , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Doxorrubicina/química , Doxorrubicina/farmacocinética , Doxorrubicina/farmacología , Femenino , Glioma/metabolismo , Glioma/patología , Humanos , Hidrazonas/química , Concentración de Iones de Hidrógeno , Malatos/química , Malatos/farmacocinética , Malatos/farmacología , Masculino , Polímeros/química , Polímeros/farmacocinética , Polímeros/farmacologíaRESUMEN
Treatment options for triple negative breast cancer (TNBC) are generally limited to cytotoxic chemotherapy. Recently, anti-epidermal growth factor receptor (EGFR) therapy has been introduced for TNBC patients. We engineered a novel nanobioconjugate based on a poly(ß-L-malic acid) (PMLA) nanoplatform for TNBC treatment. The nanobioconjugate carries anti-tumor nucleosome-specific monoclonal antibody (mAb) 2C5 to target breast cancer cells, anti-mouse transferrin receptor (TfR) antibody for drug delivery through the host endothelial system, and Morpholino antisense oligonucleotide (AON) to inhibit EGFR synthesis. The nanobioconjugates variants were: (1) P (BioPolymer) with AON, 2C5 and anti-TfR for tumor endothelial and cancer cell targeting, and EGFR suppression (P/AON/2C5/TfR), and (2) P with AON and 2C5 (P/AON/2C5). Controls included (3) P with 2C5 but without AON (P/2C5), (4) PBS, and (5) P with PEG and leucine ester (LOEt) for endosomal escape (P/mPEG/LOEt). Drugs were injected intravenously to MDA-MB-468 TNBC bearing mice. Tissue accumulation of injected nanobioconjugates labeled with Alexa Fluor 680 was examined by Xenogen IVIS 200 (live imaging) and confocal microscopy of tissue sections. Levels of EGFR, phosphorylated and total Akt in tumor samples were detected by western blotting. In vitro western blot showed that the leading nanobioconjugate P/AON/2C5/TfR inhibited EGFR synthesis significantly better than naked AON. In vivo imaging revealed that 2C5 increased drug-tumor accumulation. Significant tumor growth inhibition was observed in mice treated with the lead nanobioconjugate (1) [Pâ=â0.03 vs. controls; P<0.05 vs. nanobioconjugate variant (2)]. Lead nanobioconjugate (1) also showed stronger inhibition of EGFR expression and Akt phosphorylation than other treatments. Treatment of TNBC with the new nanobioconjugate results in tumor growth arrest by inhibiting EGFR and its downstream signaling intermediate, phosphorylated Akt. The nanobioconjugate represents a new generation of nanodrugs for treatment of TNBC.
Asunto(s)
Biopolímeros/uso terapéutico , Neoplasias de la Mama/terapia , Receptores ErbB/antagonistas & inhibidores , Malatos/química , Nanopartículas/uso terapéutico , Oligonucleótidos Antisentido/uso terapéutico , Polímeros/química , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Anticuerpos Monoclonales/uso terapéutico , Antineoplásicos/uso terapéutico , Western Blotting , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Receptores ErbB/genética , Receptores ErbB/metabolismo , Femenino , Humanos , Ratones , Ratones Desnudos , Receptor ErbB-2/metabolismo , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismo , Transducción de Señal , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Microbial poly(ß, l-malic acid) was modified with either l-leucine ethyl ester (L) or l-phenylalanine methyl ester (F) to produce amphiphylic copolymers. The degradation of these copolymers in aqueous buffer took place under physiological conditions in a few weeks by hydrolysis of the side chain ester group followed by cleavage of the main chain. Spherical nanoparticles with diameters ranging between 70 and 230 nm were prepared from these copolymers by the dialysis-precipitation method. No alteration of the cell viability was observed after incubation of these nanoparticles in different cell lines. Anticancer drugs temozolomide and doxorubicin were encapsulated in the nanoparticles. Temozolomide was released within several hours whereas doxorubicin took several weeks to be completely liberated.
RESUMEN
PMLA nanoparticles with diameters of 150-250 nm are prepared, and their hydrolytic degradation is studied under physiological conditions. Degradation occurs by hydrolysis of the side chain methyl ester followed by cleavage of the main-chain ester group with methanol and L-malic acid as the final degradation products. No alteration of the cell viability is found after 1 h of incubation, but toxicity increases significantly after 3 d, probably due to the noxious effect of the released methanol. Anticancer drugs temozolomide and doxorubicin are encapsulated in the NPs with 20-40% efficiency, and their release is monitored using in vitro essays. Temozolomide is fully liberated within several hours, whereas doxorubicin is steadily released from the particles over a period of 1 month.
Asunto(s)
Antineoplásicos/farmacología , Composición de Medicamentos/métodos , Malatos/síntesis química , Nanopartículas/química , Poliésteres/síntesis química , Polímeros/síntesis química , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Dacarbazina/análogos & derivados , Dacarbazina/farmacología , Doxorrubicina/farmacología , Emulsiones , Humanos , Concentración de Iones de Hidrógeno/efectos de los fármacos , Hidrólisis/efectos de los fármacos , Espectroscopía de Resonancia Magnética , Malatos/química , Nanopartículas/ultraestructura , Tamaño de la Partícula , Poliésteres/química , Polímeros/química , Polvos , Sonicación , Temozolomida , Factores de TiempoRESUMEN
Membranolytic macromolecules are promising vehicles for cytoplasmic drug delivery, but their efficiency and safety remains primary concerns. To address those concerns, membranolytic properties of various poly(ß-L-malic acid) (PMLA) copolymers were extensively investigated as a function of concentration and pH. PMLA, a naturally occurring biodegradable polymer, acquires membranolytic activities after substitution of pendent carboxylates with hydrophobic amino acid derivatives. Ruled by hydrophobization and charge neutralization, membranolysis of PMLA copolymers increased as a function of polymer molecular weight and demonstrated a maximum with 50% substitution of carboxylates. Charge neutralization was achieved either conditionally by pH-dependent protonation or permanently by masking carboxylates. Membranolysis of PMLA copolymers containing tripeptides of leucine, tryptophan and phenylalanine were pH-dependent in contrast to pH-independent copolymers of Leucine ethyl ester and Leu-Leu-Leu-NH(2) with permanent charge neutralization. PMLA and tripeptides seemed a unique combination for pH-dependent membranolysis. In contrast to nontoxic pH-dependent PMLA copolymers, pH-independent copolymers were found toxic at high concentration, which is ascribed to their nonspecific disruption of plasma membrane at physiological pH. pH-Dependent copolymers were membranolytically active only at acidic pH typical of maturating endosomes, and are thus devoid of cytotoxicity. The PMLA tripeptide copolymers are useful for safe and efficient cytoplasmic delivery routed through endosome.
Asunto(s)
Sistemas de Liberación de Medicamentos , Malatos/química , Péptidos/química , Polímeros/química , Membrana Celular/química , Membrana Celular/metabolismo , Portadores de Fármacos/síntesis química , Portadores de Fármacos/química , Endosomas/química , Endosomas/metabolismo , Concentración de Iones de Hidrógeno , Estructura Molecular , Peso Molecular , Polímeros/síntesis químicaRESUMEN
Biodegradable nanopolymers are believed to offer great potential in cancer therapy. Here, we report the characterization of a novel, targeted, nanobiopolymeric conjugate based on biodegradable, nontoxic, and nonimmunogenic PMLA [poly(ß-l-malic acid)]. The PMLA nanoplatform was synthesized for repetitive systemic treatments of HER2/neu-positive human breast tumors in a xenogeneic mouse model. Various moieties were covalently attached to PMLA, including a combination of morpholino antisense oligonucleotides (AON) directed against HER2/neu mRNA, to block new HER2/neu receptor synthesis; anti-HER2/neu antibody trastuzumab (Herceptin), to target breast cancer cells and inhibit receptor activity simultaneously; and transferrin receptor antibody, to target the tumor vasculature and mediate delivery of the nanobiopolymer through the host endothelial system. The results of the study showed that the lead drug tested significantly inhibited the growth of HER2/neu-positive breast cancer cells in vitro and in vivo by enhanced apoptosis and inhibition of HER2/neu receptor signaling with suppression of Akt phosphorylation. In vivo imaging analysis and confocal microscopy demonstrated selective accumulation of the nanodrug in tumor cells via an active delivery mechanism. Systemic treatment of human breast tumor-bearing nude mice resulted in more than 90% inhibition of tumor growth and tumor regression, as compared with partial (50%) tumor growth inhibition in mice treated with trastuzumab or AON, either free or attached to PMLA. Our findings offer a preclinical proof of concept for use of the PMLA nanoplatform for combination cancer therapy.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Carcinoma/tratamiento farmacológico , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Malatos/uso terapéutico , Polímeros/uso terapéutico , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/genética , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/química , Antineoplásicos/uso terapéutico , Biopolímeros/química , Biopolímeros/uso terapéutico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Carcinoma/genética , Carcinoma/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Femenino , Humanos , Inyecciones Intravenosas , Malatos/administración & dosificación , Malatos/química , Ratones , Ratones Desnudos , Modelos Biológicos , Nanopartículas/química , Nanopartículas/uso terapéutico , Polímeros/administración & dosificación , Polímeros/química , Receptor ErbB-2/metabolismo , Resultado del Tratamiento , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Nanotechnology is the design and assembly of submicroscopic devices called nanoparticles, which are 1-100 nm in diameter. Nanomedicine is the application of nanotechnology for the diagnosis and treatment of human disease. Disease-specific receptors on the surface of cells provide useful targets for nanoparticles. Because nanoparticles can be engineered from components that (1) recognize disease at the cellular level, (2) are visible on imaging studies, and (3) deliver therapeutic compounds, nanotechnology is well suited for the diagnosis and treatment of a variety of diseases. Nanotechnology will enable earlier detection and treatment of diseases that are best treated in their initial stages, such as cancer. Advances in nanotechnology will also spur the discovery of new methods for delivery of therapeutic compounds, including genes and proteins, to diseased tissue. A myriad of nanostructured drugs with effective site-targeting can be developed by combining a diverse selection of targeting, diagnostic, and therapeutic components. Incorporating immune target specificity with nanostructures introduces a new type of treatment modality, nano-immunochemotherapy, for patients with cancer. In this review, we will discuss the development and potential applications of nanoscale platforms in medical diagnosis and treatment. To impact the care of patients with neurological diseases, advances in nanotechnology will require accelerated translation to the fields of brain mapping, CNS imaging, and nanoneurosurgery. Advances in nanoplatform, nano-imaging, and nano-drug delivery will drive the future development of nanomedicine, personalized medicine, and targeted therapy. We believe that the formation of a science, technology, medicine law-healthcare policy (STML) hub/center, which encourages collaboration among universities, medical centers, US government, industry, patient advocacy groups, charitable foundations, and philanthropists, could significantly facilitate such advancements and contribute to the translation of nanotechnology across medical disciplines.
Asunto(s)
Antineoplásicos/uso terapéutico , Política de Salud/legislación & jurisprudencia , Política de Salud/tendencias , Nanomedicina/legislación & jurisprudencia , Nanomedicina/tendencias , Neoplasias/diagnóstico , Neoplasias/tratamiento farmacológico , Antineoplásicos/administración & dosificación , Sistemas de Liberación de Medicamentos/métodos , Sistemas de Liberación de Medicamentos/tendencias , Humanos , Imagenología Tridimensional/métodos , Nanomedicina/métodos , Nanoestructuras/uso terapéutico , Medicina de Precisión/métodos , Medicina de Precisión/tendenciasRESUMEN
Effective treatment of brain neurological disorders such as Alzheimer's disease, multiple sclerosis, or tumors should be possible with drug delivery through blood-brain barrier (BBB) or blood-brain tumor barrier (BTB) and targeting specific types of brain cells with drug release into the cell cytoplasm. A polymeric nanobioconjugate drug based on biodegradable, nontoxic, and nonimmunogenic polymalic acid as a universal delivery nanoplatform was used for design and synthesis of nanomedicine drug for i.v. treatment of brain tumors. The polymeric drug passes through the BTB and tumor cell membrane using tandem monoclonal antibodies targeting the BTB and tumor cells. The next step for polymeric drug action was inhibition of tumor angiogenesis by specifically blocking the synthesis of a tumor neovascular trimer protein, laminin-411, by attached antisense oligonucleotides (AONs). The AONs were released into the target cell cytoplasm via pH-activated trileucine, an endosomal escape moiety. Drug delivery to the brain tumor and the release mechanism were both studied for this nanobiopolymer. Introduction of a trileucine endosome escape unit resulted in significantly increased AON delivery to tumor cells, inhibition of laminin-411 synthesis in vitro and in vivo, specific accumulation in brain tumors, and suppression of intracranial glioma growth compared with pH-independent leucine ester. The availability of a systemically active polymeric drug delivery system that passes through the BTB, targets tumor cells, and inhibits glioma growth gives hope for a successful strategy of glioma treatment. This delivery system with drug release into the brain-specific cell type could be useful for treatment of various brain pathologies.
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Neoplasias Encefálicas/tratamiento farmacológico , Concentración de Iones de Hidrógeno , Malatos/uso terapéutico , Nanopartículas , Polímeros/uso terapéutico , Animales , Barrera Hematoencefálica , Neoplasias Encefálicas/patología , Endosomas/metabolismo , Infusiones Intravenosas , Malatos/administración & dosificación , Malatos/farmacocinética , Ratones , Ratones Desnudos , Polímeros/administración & dosificación , Polímeros/farmacocinéticaRESUMEN
PURPOSE: Temozolomide (TMZ) is a pro-drug releasing a DNA alkylating agent that is the most effective drug to treat glial tumors when combined with radiation. TMZ is toxic, and therapeutic dosages are limited by severe side effects. Targeted delivery is thus needed to improve efficiency and reduce non-tumor tissue toxicity. METHODS: Multifunctional targetable nanoconjugates of TMZ hydrazide were synthesized using poly(ß-L-malic acid) platform, which contained a targeting monoclonal antibody to transferrin receptor (TfR), trileucine (LLL), for pH-dependent endosomal membrane disruption, and PEG for protection. RESULTS: The water-soluble TMZ nanoconjugates had hydrodynamic diameters in the range of 6.5 to 14.8 nm and ζ potentials in the range of -6.3 to -17.7 mV. Fifty percent degradation in human plasma was observed in 40 h at 37°C. TMZ conjugated with polymer had a half-life of 5-7 h, compared with 1.8 h for free TMZ. The strongest reduction of human brain and breast cancer cell viability was obtained by versions of TMZ nanoconjugates containing LLL and anti-TfR antibody. TMZ-resistant cancer cell lines were sensitive to TMZ nanoconjugate treatment. CONCLUSIONS: TMZ-polymer nanoconjugates entered the tumor cells by receptor-mediated endocytosis, effectively reduced cancer cell viability, and can potentially be used for targeted tumor treatment.
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Antineoplásicos/administración & dosificación , Dacarbazina/análogos & derivados , Malatos/química , Nanopartículas , Polímeros/química , Línea Celular Tumoral , Dacarbazina/administración & dosificación , Humanos , Liposomas , Peso Molecular , TemozolomidaRESUMEN
New copolyesters derived from poly(ß,L-malic acid) have been designed to serve as nanoconjugate platforms in drug delivery. 25% and 50% methylated derivatives (coPMLA-Me(25)H(75) and coPMLA-Me(50)H(50)) with absolute molecular weights of 32 600 Da and 33 100 Da, hydrodynamic diameters of 3.0 nm and 5.2 nm and zeta potential of -15mV and -8.25mV, respectively, were found to destabilize membranes of liposomes at pH 5.0 and pH 7.5 at concentrations above 0.05mg/mL. The copolymers were soluble in PBS (half life of 40 hours) and in human plasma (half life of 15 hours) but they showed tendency to aggregate at high levels of methylation. Fluorescence-labeled copolymers were internalized into MDA-MB-231 breast cancer cells with increased efficiency for the higher methylated copolymer. Viability of cultured brain and breast cancer cell lines indicated moderate toxicity that increased with methylation. The conclusion of the present work is that partially methylated poly(ß,L-malic acid) copolyesters are suitable as nanoconjugate platforms for drug delivery.
RESUMEN
This study focused on the preparation and evaluation of nanoparticles made of alkyl esters of microbial poly(gamma-glutamic acid) (PGGA) to be used as drugs and proteins carrier and delivery systems. Racemic PGGA of bacterial origin was fully methylated or partially esterified to render non-water-soluble polymers. A set of co-polymers containing poly(glutamic acid) and ethyl, hexyl, dodecyl and octadecyl glutamate units with alkyl contents of 50 and 75% was prepared. Spherical nanoparticles with a diameter of 200-250 nm and a narrow distribution were generated from the alkylated polymers by the precipitation-dialysis method. These nanoparticles readily degraded hydrolytically upon incubation in simulated physiological medium at a rate dependent on the alkylation degree and the length of the alkyl group. All these nanoparticles were able to encapsulate efficiently erythromycin. Those made of carboxyl containing polyglutamates were also effective to load alpha-chymotrypsin. The release of such compounds from nanoparticles upon incubation proceeded essentially following the same profile that is followed in the hydrolysis of the corresponding substrate polymers. The loss of enzyme activity of the incubated protein diminished significantly upon encapsulation in these systems.