RESUMO
L-lysine α-oxidase (LO) is an L-amino acid oxidase with antitumor, antimicrobial and antiviral properties. Pharmacokinetic (PK) studies were carried out by measuring LO concentration in plasma and tissue samples by enzyme immunoassay. L-lysine concentration in samples was measured spectrophotometrically using LO. After single i.v. injection of 1.0, 1.5, 3.0 mg/kg the circulating T1/2 of enzyme in mice varied from 51 to 74 min and the AUC0-inf values were 6.54 ± 0.46, 8.66 ± 0.59, 9.47 ± 1.45 µg/ml × h, respectively. LO was distributed in tissues and determined within 48 h after administration with maximal accumulation in liver and heart tissues. Mean time to reach the maximum concentration was highest for the liver-9 h, kidney-1 h and 15 min for the tissues of heart, spleen and brain. T1/2 of LO in tissues ranged from 7.75 ± 0.73 to 26.10 ± 2.60 h. In mice, plasma L-lysine decreased by 79% 15 min after LO administration in dose 1.6 mg/kg. The serum L-lysine levels remained very low from 1 to 9 h (< 25 µM, 17%), indicating an acute lack of L-lysine in animals for at least 9 h. Concentration of L-lysine in serum restored only 24 h after LO administration. The results of LO PK study show that it might be considered as a promising enzyme for further investigation as a potential anticancer agent.
Assuntos
Aminoácido Oxirredutases/farmacocinética , Trichoderma/enzimologia , Aminoácido Oxirredutases/administração & dosagem , Animais , Proteínas Fúngicas/administração & dosagem , Proteínas Fúngicas/farmacocinética , Lisina/sangue , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Distribuição TecidualRESUMO
Synergism effects of cisplatin and L-lysine-alpha-oxidase (LO), while sequential (no interval) administration of drugs depends on the tumor model and duration of treatment. Synergism is identified at intraperitoneal daily (during 3 days) administration of cisplatin to experimental animals in single doses of 1.5 or 3.0 mg/kg and intravenously 5-fold after 48 h administration of LO and also administered intravenously in cumulative doses of 300-600 E / kg discretely, the first dose--doubled. Synergism of cisplatin and LO is showed by significant (p < 0.05) therapeutic gain against cisplatin at such indicators as increased survival of mice with P388 tumor and increased inhibition of primary tumor melanoma B16.
Assuntos
Aminoácido Oxirredutases/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Cisplatino/farmacologia , Leucemia P388/tratamento farmacológico , Melanoma Experimental/tratamento farmacológico , Aminoácido Oxirredutases/administração & dosagem , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Cisplatino/administração & dosagem , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Análise de Sobrevida , Resultado do TratamentoRESUMO
In the United States, 5-12% of adults have at least one symptom of temporomandibular joint (TMJ) disorders, including TMJ osteoarthritis (TMJ-OA). However, there is no chondroprotective agent that is approved for clinical application. We showed that LOXL2 is elevated in the regenerative response during fracture healing in mice and has a critical role in chondrogenic differentiation. Indeed, LOXL2 is an anabolic effector that attenuates pro-inflammatory signaling in OA cartilage of the TMJ and knee joint, induces chondroprotective and regenerative responses, and attenuates NF-kB signaling. The specific goal of the study was to evaluate if adenoviral delivery of LOXL2 is anabolic to human and mouse TMJ condylar cartilage in vivo and evaluate the protective and anabolic effect on cartilage-specific factors. We employed two different models to assess TMJ-OA. In one model, clinical TMJ-OA cartilage from 5 different samples in TMJ-OA cartilage plugs were implanted subcutaneously in nude mice. Adenovirus LOXL2 -treated implants showed higher mRNA levels of LOXL2, ACAN, and other anabolic genes compared to the adenovirus-Empty-treated implants. Further characterization by RNA-seq analysis showed LOXL2 promotes proteoglycan networks and extracellular matrix in human TMJ-OA cartilage implants in vivo. In order to evaluate if LOXL2-induced functional and sex-linked differences, both male and female four-month-old chondrodysplasia (Cho/+) mice, which develop progressive TMJ-OA due to a point mutation in the Col11a1 gene, were subjected to intraperitoneal injection with Adv-RFP-LOXL2 every 2 weeks for 12 weeks. The data showed that adenovirus delivery of LOXL2 upregulated LOXL2 and aggrecan (Acan), whereas MMP13 expression was slightly downregulated. The fold change expression of Acan and Runx2 induced by Adv-RFP-LOXL2 was higher in females compared to males. Interestingly, Adv-RFP-LOXL2 injection significantly increased Rankl expression in male but there was no change in females, whereas VegfB gene expression was increased in females, but not in males, as compared to those injected with Adv-RFP-Empty in respective groups. Our findings indicate that LOXL2 can induce specifically the expression of Acan and other anabolic genes in two preclinical models in vivo. Further, LOXL2 has beneficial functions in human TMJ-OA cartilage implants and promotes gender-specific anabolic responses in Cho/+ mice with progressive TMJ-OA, suggesting its merit for further study as an anabolic therapy for TMJ-OA.
Assuntos
Agrecanas/metabolismo , Aminoácido Oxirredutases/metabolismo , Cartilagem Articular/patologia , Osteoartrite/patologia , Transtornos da Articulação Temporomandibular/metabolismo , Adenoviridae/genética , Idoso , Aminoácido Oxirredutases/administração & dosagem , Aminoácido Oxirredutases/genética , Animais , Cartilagem Articular/metabolismo , Cartilagem Articular/transplante , Condrócitos/metabolismo , Colágeno/genética , Modelos Animais de Doenças , Feminino , Redes Reguladoras de Genes , Humanos , Masculino , Metabolismo/genética , Camundongos Mutantes , Camundongos Nus , Pessoa de Meia-Idade , Osteoartrite/metabolismo , Caracteres Sexuais , Transtornos da Articulação Temporomandibular/patologiaRESUMO
Evidences on the antileukemic effect of L-lysine alpha-oxidase from Trichoderma sp. are presented. It is inferred that enzyme needs detail studying as a potential chemotherapeutic drug both in experimental and clinical research.
Assuntos
Aminoácido Oxirredutases/uso terapêutico , Leucemia L1210/tratamento farmacológico , Leucemia P388/tratamento farmacológico , Leucemia Experimental/tratamento farmacológico , Fungos Mitospóricos/enzimologia , Trichoderma/enzimologia , Aminoácido Oxirredutases/administração & dosagem , Animais , Avaliação de Medicamentos , Masculino , Camundongos , Camundongos Endogâmicos , Fatores de TempoRESUMO
Maple syrup urine disease (MSUD) is a genetic disease caused by a deficiency of branched-chain keto acid dehydrogenase, a mitochondrial multienzyme complex responsible for the decarboxylation of leucine, isoleucine and valine. The complex consists of three subunits (E1, E2, and E3) and mutations in any subunit result in MSUD. No satisfactory treatment for MSUD is currently available. Here we report the successful use of retroviral gene transfer to restore leucine decarboxylation activity in fibroblasts derived from a MSUD patient containing a mutation in the E2 subunit. A full-length human E2 cDNA was inserted into a retroviral vector (MFG) and a stable CRIP producer line was generated. The amphotropic virus was then used to transduce mutant human fibroblasts. In untransduced mutant cells, 1-14C leucine decarboxylation activity was less than 2% that of the wild-type cells. Decarboxylation of 1-14C leucine in transduced mutant cells was restored to 93% of the wild-type level. Correct targeting of the expressed wild-type E2 protein to mitochondria was demonstrated by comparing the immunofluorescent pattern of E2 and a mitochondrial marker protein. Stable expression of enzyme activity has been obtained for at least 7 weeks. In contrast to most previous gene therapy attempts, which replace a single enzyme defect, the present results demonstrate complementation of a phenotype resulting from a gene defect whose product is a part of a multienzyme complex. Based on these results, studies can now be undertaken to investigate the feasibility of gene therapy to correct MSUD.