RESUMO
Anti-VEGF (vascular endothelial growth factor) drugs such as aflibercept (AFL) and bevacizumab (BVZ) inhibit pathological neo-angiogenesis and vascular permeability in retinal vascular diseases. As cytokines and growth factors are produced by Müller glial cells under stressful and pathological conditions, we evaluated the in vitro effect of AFL (Eylea®, 0.5 mg/mL) and BVZ (Avastin®, 0.5 mg/mL) on cell viability/metabolism, and cytokine/growth factor production by Müller cells (MIO-M1) under cobalt chloride (CoCl2)-induced hypoxia after 24h, 48h and 72h. Cell viability/metabolism were analyzed by Trypan Blue and MTT assays and cytokine/growth factors in supernatants by Luminex xMAP-based multiplex bead-based immunoassay. Cell viability increased with AFL at 48h and 72h and decreased with BVZ or hypoxia at 24h. BVZ-treated cells showed lower cell viability than AFL at all exposure times. Cell metabolism increased with AFL but decreased with BVZ (72h) and hypoxia (48h and72h). As expected, AFL and BVZ decreased VEGF levels. AFL increased PDGF-BB, IL-6 and TNF-α (24h) and BVZ increased PDGF-BB (72h). Hypoxia reduced IL-1ß, -6, -8, TNF-α and PDGF-BB at 24h, and its suppressive effect was more prominent than AFL (EGF, PDGF-BB, IL-1ß, IL-6, IL-8, and TNF-α) and BVZ (PDGF-BB and IL-6) effects. Hypoxia increased bFGF levels at 48h and 72h, even when combined with anti-VEGFs. However, the stimulatory effect of BVZ predominated over hypoxia for IL-8 and TNF-α (24h), as well as for IL-1ß (72h). Thus, AFL and BVZ exhibit distinct exposure times effects on MIO-M1 cells viability, metabolism, and cytokines/growth factors. Hypoxia and BVZ decreased MIO-M1 cell viability/metabolism, whereas AFL likely induced gliosis. Hypoxia resulted in immunosuppression, and BVZ stimulated inflammation in hypoxic MIO-M1 cells. These findings highlight the complexity of the cellular response as well as the interplay between anti-VEGF treatments and the hypoxic microenvironment.
Assuntos
Células Ependimogliais , Receptores de Fatores de Crescimento do Endotélio Vascular , Proteínas Recombinantes de Fusão , Fator A de Crescimento do Endotélio Vascular , Humanos , Bevacizumab/farmacologia , Bevacizumab/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Células Ependimogliais/metabolismo , Sobrevivência Celular , Becaplermina/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Interleucina-8/metabolismo , Interleucina-6/metabolismo , Fatores de Crescimento do Endotélio Vascular/metabolismo , Citocinas/metabolismo , Hipóxia/metabolismo , Neovascularização Patológica/patologia , Inflamação/patologiaRESUMO
Previously, we demonstrated that the expression of THBS1 is increased in esophageal squamous cell carcinoma (ESCC) tissues and is correlated with lymph node metastasis and poor prognosis, indicating that THBS1 might be a candidate oncogene in ESCC. In this study, we future studied the specific role of THBS1 in ESCC and its molecular mechanism. Silencing THBS1 expression resulted in inhibition of cell migration and cell invasion of ESCC cells, the decrease of colony formation and proliferation. Tube formation of human umbilical vein endothelial cells (HUVECs) in vitro was decreased when cultured with conditioned medium from THBS1-silenced cells. The expression of CD31, a marker for blood vessel endothelial cells, was decreased in tumor tissues derived from THBS1-silenced tumors in vivo. Silencing THBS1 leaded the decreased of hypoxia-inducible factor-1α (HIF-1α), HIF-1ß, and VEGFA protein. The expression of p-ERK and p-AKT were declined in HUVECs following incubation with conditioned medium from THBS1-silenced ESCC cells compared conditioned medium from control cells. Furthermore, the treatment with bevacizumab boosted the decrease of the p-ERK and p-AKT levels in HUVECs incubated with the conditioned medium from THBS1-silenced ESCC cells. THBS1 silencing combined with bevacizumab blocked VEGF, inhibited to the tube formation, colony formation and migration of HUVECs, which were superior to that of bevacizumab alone. We presumed that THBS1 can enhance HIF-1/VEGF signaling and subsequently induce angiogenesis by activating the AKT and ERK pathways in HUVECs, resulting in bevacizumab resistance. THBS1 would be a potential target in tumor antiangiogenesis therapies.
Assuntos
Neoplasias Esofágicas , Carcinoma de Células Escamosas do Esôfago , Humanos , Carcinoma de Células Escamosas do Esôfago/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Bevacizumab/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Esofágicas/patologia , Angiogênese , Meios de Cultivo Condicionados/farmacologia , Linhagem Celular Tumoral , Transdução de Sinais , Células Endoteliais da Veia Umbilical Humana/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismoRESUMO
Extracranial arteriovenous malformations (AVMs) are regarded as rare diseases and are prone to complications such as pain, bleeding, relentless growth, and high volume of shunted blood. Due to the high vascular pressure endothelial cells of AVMs are exposed to mechanical stress. To control symptoms and lesion growth pharmacological treatment strategies are urgently needed in addition to surgery and interventional radiology. AVM cells were isolated from three patients and exposed to cyclic mechanical stretching for 24 h. Thalidomide and bevacizumab, both VEGF inhibitors, were tested for their ability to prevent the formation of circular networks and proliferation of CD31+ endothelial AVM cells. Furthermore, the effect of thalidomide and bevacizumab on stretched endothelial AVM cells was evaluated. In response to mechanical stress, VEGF gene and protein expression increased in patient AVM endothelial cells. Thalidomide and bevacizumab reduced endothelial AVM cell proliferation. Bevacizumab inhibited circular network formation of endothelial AVM cells and lowered VEGF gene and protein expression, even though the cells were exposed to mechanical stress. With promising in vitro results, bevacizumab was used to treat three patients with unresectable AVMs or to prevent regrowth after incomplete resection. Bevacizumab controlled bleeding, pulsation, and pain over the follow up of eight months with no patient-reported side effects. Overall, mechanical stress increases VEGF expression in the microenvironment of AVM cells. The monoclonal VEGF antibody bevacizumab alleviates this effect, prevents circular network formation and proliferation of AVM endothelial cells in vitro. The clinical application of bevacizumab in AVM treatment demonstrates effective symptom control with no side effects.
Assuntos
Malformações Arteriovenosas , Células Endoteliais , Humanos , Células Endoteliais/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Bevacizumab/farmacologia , Bevacizumab/uso terapêutico , Bevacizumab/metabolismo , Talidomida/metabolismo , Malformações Arteriovenosas/genética , Dor/metabolismoRESUMO
Previous work suggested that tenogenic differentiation of tendon stem/progenitor cells (TSPCs) was suppressed by upregulated expression of the angiogenic marker vascular endothelial growth factor (VEGF). The purpose of this study was to test the hypothesis that anti-VEGF antibody, bevacizumab, promotes in vitro tenogenic differentiation and maturation of two distinct types of TSPCs, tendon proper-derived cells (TDCs), and paratenon-derived cells (PDCs) originating from rat Achilles tendon. TDCs and PDCs were isolated from the tendon proper and the paratenon of rat Achilles tendons. TDCs and PDCs were cultured for 3 days on plates with or without VEGF. TDCs and PDCs were also cultured in collagen gel matrix, and the blocking effect of VEGF was examined by the addition of 100 ng/mL of bevacizumab. Effects of bevacizumab on tenogenic differentiation were assessed using real-time PCR, immunofluorescent staining, and western blotting. VEGF significantly attenuated expression of the Tnmd gene in both PDCs and TDCs (P<0.05). Expressions of the Scx, Tnmd, and Col1a1 genes were significantly upregulated by the addition of bevacizumab (P<0.05). Immunofluorescent staining showed that the percentage of tenomodulin-positive PDCs and TDCs was significantly higher with bevacizumab treatment than in control cultures (P<0.05). Western blotting showed that bevacizumab suppressed pVEGFR-2 protein expression in both PDCs and TDCs. Bevacizumab promoted the in vitro tenogenic differentiation and maturation of two distinct TSPCs derived from rat Achilles tendon. Since the previous studies demonstrated that TSPCs have a potential to contribute to tendon repair, attenuating VEGF levels in TSPCs by administration of bevacizumab is a novel candidate therapeutic option for promoting tendon repair.
Assuntos
Tendão do Calcâneo , Fator A de Crescimento do Endotélio Vascular , Ratos , Animais , Fator A de Crescimento do Endotélio Vascular/metabolismo , Bevacizumab/farmacologia , Bevacizumab/metabolismo , Diferenciação Celular , Células-TroncoRESUMO
Objective: To observe the effect of peritumoral electroacupuncture on the induction of vascular normalization in a mouse breast cancer model. Methods: A subcutaneous graft model of breast cancer was established with 4T1 breast cancer cell line in female BALB/c mice aged 6-8 weeks. The mice were randomly assigned to three groups, a tumor-bearing group (TG), peritumoral electroacupuncture tumor-bearing group (EATG), and bevacizumab tumor-bearing group (BTG), with 18 mice in each group. The TG mice did not receive any intervention, the EATG mice received peritumoral electroacupuncture for 30 minutes, and the BTG mice were intraperitoneally injected with bevacizumab at 10mg/kg. Immunofluorescence was performed to assess the expression of CD31/alpha smooth muscle actin (α-SMA) and hypoxia-inducible factor 1-alpha (HIF-1α) in the tumor tissue at various points of time, including before intervention and 3 days and 5 days after intervention. Then, 3 days after intervention, observation of morphological changes of the microvessels in the tumor tissue was performed through Hematoxylin and Eosin (HE) staining and scanning electron microscope. Results: There was no significant difference in the expression of CD31, α-SMA, and HIF-1α in the tumor tissues of all groups before experimental intervention ( P>0.05). On day 3 of the experimental interventions, the CD31 and HIF-1α expression levels in the tumor tissues of the EATG and BTG mice were significantly reduced ( P<0.01), while α-SMA expression levels were significantly increased ( P<0.01) in both groups. On day 5 of the experimental interventions, the CD31 and HIF-1α expression levels in the tumor tissues of the EATG and BTG mice were still significantly lower than those in the TG mice ( P<0.01), while the α-SMA expression level was significantly higher than that in the TG group ( P<0.05). On day 3 of the experimental interventions, H&E staining showed visible microvessels in the tumor tissues of all 3 groups. In addition, scanning electron microscopic observation showed that the tumor microvessel walls of the TG mice were rough and defective, and that obvious deformities appeared in the lumen. In contrast, the walls of the microvessels of the EATG and BTG mice were generally intact and there was no obvious deformities in the lumen. Conclusion: Peritumoral electroacupuncture may induce microvasculature normalization by decreasing microvascular density and increasing pericyte coverage of the neovasculature, thereby improving hypoxic microenvironment of breast cancer in mice.
Assuntos
Neoplasias da Mama , Eletroacupuntura , Humanos , Camundongos , Feminino , Animais , Bevacizumab/metabolismo , Bevacizumab/farmacologia , Neoplasias da Mama/patologia , Xenoenxertos , Microvasos/metabolismo , Microvasos/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Microambiente TumoralRESUMO
BACKGROUND: Hemophilic arthropathy can cause recurrent hemarthroses and severe damage to the synovium and articular cartilage. Previous studies have shown that vascular endothelial growth factor (VEGF) plays an essential role in neoangiogenesis. Bevacizumab, a monoclonal VEGF inhibitor, is used clinically to prevent angiogenesis. However, its effects on hemophilic arthropathy are unknown. QUESTIONS/PURPOSES: Using a hemophilic arthropathy rabbit model, we asked: Does an intra-articular injection of bevacizumab (1) inhibit VEGF, (2) decrease signal intensity in dynamic contrast-enhanced MRI (DCE-MRI) as an assessment of capillary permeability and neoangiogenesis, (3) reduce cartilage damage, (4) reduce synovial changes, and (5) affect macroscopic changes during the development of hemophilic arthropathy? METHODS: Twenty-five male New Zealand rabbits were divided into four groups. Eight knees from four rabbits were used as the control group. We used an established animal model for hemophilic arthropathy in the remaining 21 rabbits. Animals were assigned randomly to three groups with seven rabbits in each group. One group was used to establish mild arthropathy, and the other two were used to establish severe arthropathy. Autologous blood from the rabbits' ears was injected into the right and left knees twice per week for 8 weeks to represent mild arthropathy and for 16 weeks to represent severe arthropathy. In the mild arthropathy group, bevacizumab was injected into the right knee once every 2 weeks. Bevacizumab was injected into the right knee of rabbits in one of the severe arthropathy groups once every 2 weeks for 16 weeks, and intra-articular bevacizumab injections were administered to the right knees of rabbits in the other severe arthropathy group once every 2 weeks after the eighth week. An equal volume of 0.9% saline was injected into the left knee of rabbits in all arthropathy groups. To explore the efficacy of bevacizumab, joint diameters were quantitatively measured, and cartilage and synovial changes were examined. Degeneration of articular cartilage was evaluated with the semiquantitative Osteoarthritis Research Society International grading system. Synovial damage was analyzed with a semiquantitative microscopic scoring system. In addition, we evaluated perfusion and angiogenesis using DCE-MRI (quantitative signal intensity changes). Immunohistochemical testing was used to measure VEGF levels (analyzed by Western blotting). RESULTS: Intra-articular bevacizumab treatment inhibited VEGF in our rabbit model of hemophilic arthropathy. VEGF protein expression levels were lower in the mild arthropathy group that received intra-articular bevacizumab (0.89 ± 0.45) than the mild arthropathy control group (1.41 ± 0.61) (mean difference -0.52 [95% CI -0.898 to -0.143]; p = 0.02). VEGF levels were lower in the severe arthropathy group that received treatment for 16 weeks (0.94 ± 0.27) than in the control knees (1.49 ± 0.36) (mean difference -0.55 [95% CI -0.935 to -0.161]; p = 0.01). In the severe arthropathy group, the Osteoarthritis Research Society International score indicating cartilage damage was lower in the group that received intra-articular bevacizumab treatment from the beginning than in the control group (median 17 [range 13 to 18] versus 18 [range 17 to 20]; difference of medians 1; p = 0.02). Additionally, the scores indicated synovial damage was lower in the group that received intra-articular bevacizumab treatment from the beginning than the control group (median 5 [range 4 to 9] versus 9 [range 8 to 12]; difference of medians 4; p = 0.02). The mean of mean values for signal intensity changes was higher in the nontreated severe groups than in the group of healthy knees. The signal intensity changes were higher in the severe arthropathy control groups (Groups BC and CC) (median 311.6 [range 301.4 to 361.2] and 315.1 [range 269.7 to 460.4]) than in the mild arthropathy control group (Group AC) (median 234.1 [range 212.5 to 304.2]; difference of medians 77.5 and 81, respectively; p = 0.02 and p = 0.04, respectively). In the severe arthropathy group, discoloration caused by hemosiderin deposition in the cartilage and synovium was more pronounced than in the mild arthropathy group. In the severe arthropathy group treated with intra-articular bevacizumab, joint diameters were smaller than in the control group (Group BT median 12.7 mm [range 12.3 to 14.0] versus Group BC median 14.0 mm [range 13.1 to 14.5]; difference of medians 1.3 mm; p = 0.02). CONCLUSION: Hemarthrosis damages the synovial tissues and cartilage in the knees of rabbits, regardless of whether they are treated with intra-articular bevacizumab. However, intra-articular injection of bevacizumab may reduce cartilage and synovial damage in rabbits when treatment is initiated early during the development of hemophilic arthropathy. CLINICAL RELEVANCE: If the findings in this study are replicated in larger-animal models that consider the limitations of our work, then a trial in humans might be appropriate to ascertain whether intra-articular injection of bevacizumab could reduce cartilage damage and synovial changes in patients with hemophilia whose hemarthroses cannot otherwise be controlled.
Assuntos
Cartilagem Articular , Osteoartrite , Humanos , Coelhos , Masculino , Animais , Bevacizumab/farmacologia , Bevacizumab/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Hemartrose/tratamento farmacológico , Hemartrose/etiologia , Hemartrose/metabolismo , Membrana Sinovial/diagnóstico por imagem , Cartilagem Articular/diagnóstico por imagem , Injeções Intra-ArticularesRESUMO
BACKGROUND: This study was designed to test the hypothesis that blockade of vascular endothelial growth factor (VEGF) suppresses degenerative changes in articular cartilage from patients with osteoarthritis (OA). METHODS: Articular cartilage from eight OA patients was subjected to explant culture for 2 days in the presence or absence of 10 ng/ml recombinant interleukin (IL)-1ß. The blocking effect of VEGF was examined by the addition of 10 or 100 ng/ml of bevacizumab. The culture media were harvested, and markers for cartilage degradation were measured by sandwich enzyme-linked immunoassay. Total RNA was isolated from cartilage tissues, and gene expressions associated with the anabolic response were examined by the quantitative real-time polymerase chain reaction. RESULTS: Bevacizumab significantly reduced concentrations of matrix metalloproteinase (MMP)-2, MMP-3, and cartilage oligomeric matrix protein in the culture media with and without IL-1ß. Significant suppressive effects of bevacizumab on MMP-9 and MMP-13 were shown only in the presence of IL-1ß. Gene expression of Col2a1 was significantly increased by the addition of bevacizumab in the absence of IL-1ß. CONCLUSION: Bevacizumab inhibits catabolic reactions and stimulates anabolic function in articular cartilage derived from OA patients directly, suggesting a protective effect on articular cartilage from OA progression.
Assuntos
Cartilagem Articular , Osteoartrite do Joelho , Humanos , Cartilagem Articular/metabolismo , Bevacizumab/farmacologia , Bevacizumab/uso terapêutico , Bevacizumab/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Osteoartrite do Joelho/tratamento farmacológico , Osteoartrite do Joelho/metabolismo , Condrócitos/metabolismo , Interleucina-1beta/metabolismoRESUMO
The development of successful treatment regimens for breast cancer requires strong pre-clinical data generated in physiologically relevant pre-clinical models. Here we report the development of the chick embryo chorioallantoic membrane (CAM) model to study tumor growth and angiogenesis using breast cancer cell lines. MDA-MB-231 and MCF7 tumor cell lines were engrafted onto the chick embryo CAM to study tumor growth and treatment response. Tumor growth was evaluated through bioluminescence imaging and a significant increase in tumor size and vascularization was found over a 9-day period. We then evaluated the impact of anti-angiogenic drugs, axitinib and bevacizumab, on tumor growth and angiogenesis. Drug treatment significantly reduced tumor vascularization and size. Overall, our findings demonstrate that the chick embryo CAM is a clinically relevant model to monitor therapeutic response in breast cancer and can be used as a platform for drug screening to evaluate not only gross changes in tumor burden but physiological processes such as angiogenesis.
Assuntos
Neoplasias da Mama , Membrana Corioalantoide , Inibidores da Angiogênese/metabolismo , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Animais , Axitinibe , Bevacizumab/metabolismo , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Embrião de Galinha , Membrana Corioalantoide/metabolismo , Feminino , Humanos , Neovascularização Patológica/metabolismoRESUMO
Methamphetamine (METH) is an illicit amphetamine-like psychostimulant that is commonly abused. However, the modulation of METH-induced cardiac microvascular permeability is still not completely known. Previously, we discovered that the vascular endothelial growth factor (VEGF) regulated the cardiotoxicity produced by METH. In this work, we looked into the effect of METH exposure on cardiac microvascular permeability via the VEGF-PI3K-Akt-eNOS signaling pathway, as well as the efficacy of Bevacizumab treatment in reducing this effect. The findings revealed that METH exposure enhanced cardiac microvascular permeability while also activating the VEGF-PI3K-Akt-eNOS signaling pathway. Furthermore, treatment with Bevacizumab has been shown to be effective in reversing the METH-induced phenomena. Briefly stated, our research may provide fresh insight into the molecular underpinnings of METH-induced cardiac microvascular permeability, and it may also provide evidence for a relationship between METH misuse and Bevacizumab medication.
Assuntos
Metanfetamina , Fosfatidilinositol 3-Quinases , Bevacizumab/metabolismo , Bevacizumab/farmacologia , Permeabilidade Capilar , Metanfetamina/toxicidade , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: To evaluate the effectiveness of intravitreal bevacizumab treatment in patients with diabetic macular edema (DME) by assessing retinal changes using optical coherence tomography angiography (OCT-A). METHODS: This prospective study was performed in patients with treatment-naïve DME. The eyes of patients were imaged using a swept-source OCT system with a scan area of 6 × 6 mm. The DME patients with a central macular thickness (CMT) of ≥300 µm received nine bevacizumab injections within 12 months. The demographic, systemic, and ocular parameters, including the best-corrected visual acuity (BCVA), CMT, microaneurysm (MA) count, and foveal avascular zone (FAZ) area in both superficial capillary plexus (SCP) and deep capillary plexus (DCP), as well as vessel density in SCP, were assessed in the patients. In addition, the response (good or poor) of the DME eyes to bevacizumab treatment and the final visual acuity (BCVA of 75 letters) were analyzed. RESULTS: Seventy-seven eyes of DME patients were subjected to the final analysis. Bevacizumab treatment reduced CMT from 425.06 µm (±77.15) to 350.25 µm (±82.04) and improved BCVA by about 8.61 letters (from 64.73 to 73.34) in the patients. The mean number of MAs in SCP decreased from 3.51 ± 2.07 to 2.31 ± 1.15 (p < 0.001) and in DCP from 17.12 ± 11.56 to 12.21 ± 6.99 (p < 0.001), whereas the area of FAZ increased in SCP from 328.22 ± 131.38 to 399.70 ± 156.98 (p < 0.001) and in DCP from 571.13 ± 396.01 to 665.89 ± 412.77 (p = 0.001). The final BCVA letter score and CMT were statistically significant in both poor and good responders, as well as in BCVA < 75 and BCVA ≥ 75 groups. CONCLUSION: The fixed-regimen intravitreal bevacizumab therapy was effective in treating DME. Apart from noninvasive visualization of microvascular damage, OCT-A showed limited usefulness in predicting treatment response. Although the study showed that the number of MAs was significantly reduced during treatment, which is an OCT-A predictor of a good response to bevacizumab treatment at a 12-month visit, commonly observed artifacts may reduce the usefulness of OCT-A.
Assuntos
Bevacizumab/farmacologia , Retinopatia Diabética/tratamento farmacológico , Idoso , Idoso de 80 Anos ou mais , Análise de Variância , Inibidores da Angiogênese/metabolismo , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Bevacizumab/metabolismo , Bevacizumab/uso terapêutico , Retinopatia Diabética/fisiopatologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Pesquisa Qualitativa , Estatísticas não Paramétricas , Tomografia de Coerência Óptica/métodos , Tomografia de Coerência Óptica/estatística & dados numéricosRESUMO
Tumoral hypoxia correlates with worse outcomes in glioblastoma (GBM). While bevacizumab is routinely used to treat recurrent GBM, it may exacerbate hypoxia. Evofosfamide is a hypoxia-targeting prodrug being tested for recurrent GBM. To characterize resistance to bevacizumab and identify those with recurrent GBM who may benefit from evofosfamide, we ascertained MRI features and hypoxia in patients with GBM progression receiving both agents. Thirty-three patients with recurrent GBM refractory to bevacizumab were enrolled. Patients underwent MR and 18F-FMISO PET imaging at baseline and 28 days. Tumor volumes were determined, MRI and 18F-FMISO PET-derived parameters calculated, and Spearman correlations between parameters assessed. Progression-free survival decreased significantly with hypoxic volume [hazard ratio (HR) = 1.67, 95% confidence interval (CI) 1.14 to 2.46, P = 0.009] and increased significantly with time to the maximum value of the residue (Tmax) (HR = 0.54, 95% CI 0.34 to 0.88, P = 0.01). Overall survival decreased significantly with hypoxic volume (HR = 1.71, 95% CI 1.12 to 12.61, p = 0.01), standardized relative cerebral blood volume (srCBV) (HR = 1.61, 95% CI 1.09 to 2.38, p = 0.02), and increased significantly with Tmax (HR = 0.31, 95% CI 0.15 to 0.62, p < 0.001). Decreases in hypoxic volume correlated with longer overall and progression-free survival, and increases correlated with shorter overall and progression-free survival. Hypoxic volume and volume ratio were positively correlated (rs = 0.77, P < 0.0001), as were hypoxia volume and T1 enhancing tumor volume (rs = 0.75, P < 0.0001). Hypoxia is a key biomarker in patients with bevacizumab-refractory GBM. Hypoxia and srCBV were inversely correlated with patient outcomes. These radiographic features may be useful in evaluating treatment and guiding treatment considerations.
Assuntos
Glioblastoma/metabolismo , Recidiva Local de Neoplasia/metabolismo , Hipóxia Tumoral/fisiologia , Adulto , Idoso , Bevacizumab/metabolismo , Bevacizumab/uso terapêutico , Biomarcadores Farmacológicos , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/patologia , Volume Sanguíneo Cerebral/fisiologia , Resistencia a Medicamentos Antineoplásicos/genética , Resistencia a Medicamentos Antineoplásicos/fisiologia , Feminino , Glioblastoma/diagnóstico por imagem , Glioblastoma/mortalidade , Humanos , Imageamento por Ressonância Magnética/métodos , Masculino , Pessoa de Meia-Idade , Misonidazol/análogos & derivados , Misonidazol/uso terapêutico , Tomografia por Emissão de Pósitrons/métodos , Intervalo Livre de Progressão , Adulto JovemRESUMO
BACKGROUND: Syringes containing anti-vascular endothelial growth factor drugs to treat retinal diseases are prepared in different ways by various parties with syringe selection, preparation, and storage conditions affecting the risk of injecting particles into the vitreous. This study examines particle loads from various syringes over time. METHODS: Four syringes were studied: two plastic transfer syringes lubricated with silicone oil or oleamide, a glass syringe with baked-on silicone, and a lubricant-free polymer syringe. Syringes were rinsed with water or filled with buffer and analyzed over time; particles were quantified by flow imaging. Particle formation in a bevacizumab formulation was also characterized. RESULTS: Insulin syringes consistently showed very high particle counts. Oleamide-lubricated syringes had substantially fewer particles, but showed appreciable increases over time (leading to visible particles). Baked-on silicone glass syringes and lubricant-free polymer syringes both showed low particle levels ≥10 µm. Lubricant-free syringes showed the lowest particle levels ≥1 µm and the lowest particle levels with bevacizumab agitation. CONCLUSION: Syringes have different intrinsic particle loads which can contribute to particle loads in the delivered drug. Oleamide-lubricated transfer syringes, commonly used for bevacizumab repackaging, have time-dependent particle loads and are associated with the formation of visible particles beyond 30 days of storage.
Assuntos
Inibidores da Angiogênese/metabolismo , Bevacizumab/metabolismo , Composição de Medicamentos/métodos , Material Particulado/metabolismo , Agregação Patológica de Proteínas/etiologia , Seringas , Embalagem de Medicamentos , Injeções Intravítreas , Lubrificantes , Agregação Patológica de Proteínas/diagnóstico , Agregação Patológica de Proteínas/metabolismo , Óleos de Silicone/química , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidoresRESUMO
Intravitreal injections are the standard procedure in the treatment of retinal pathologies, such as the administration of the anti-VEGF antibodies in age-related macular degeneration. The aim of this study is to evaluate the intraocular and blood pharmacokinetics after an intravitreal injection of 89Zr-labelled bevacizumab and 89Zr-labelled aflibercept in Sprague-Dawley rats using Positron Emission Tomography. First, both antibodies were radiolabelled to zirconium-89 with a maximum specific activity of 15 Mbq/mg for bevacizumab and 10 Mbq/mg for aflibercept. Four µL containing 1-1.2 Mq of 89Zr-labelled compound were injected into the vitreous through a 35 G needle. A microPET acquisition was carried out immediately after the injection and at different time points through a 12-day study and blood samples were obtained through the tail vein. Radiolabelling was successfully performed with a radiochemical purity after ultrafiltration above 95% for both agents. Both antibodies ocular curves followed a two-compartment model in which an intraocular elimination half-life of 16.44 h was found for 89Zr-bevacizumab and 4.51 h for 89Zr-aflibercept, considering the alpha phase as the elimination phase. Regarding the beta phase, a half-life of 3.23 days for 89Zr-bevacizumab and 4.69 days for 89Zr-aflibercept were observed. With regards to blood concentration, 89Zr-bevacizumab showed a blood half-life of 7.08 days, whereas 89Zr-aflibercept's was 3.18 days, by a one-compartment model with first-order absorption kinetics. In conclusion, this study shows for the first time the ocular and blood pharmacokinetic analysis after intravitreal injection of aflibercept and bevacizumab in rats.
Assuntos
Bevacizumab/metabolismo , Olho/metabolismo , Injeções Intravítreas/métodos , Tomografia por Emissão de Pósitrons/métodos , Receptores de Fatores de Crescimento do Endotélio Vascular/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Inibidores da Angiogênese/administração & dosagem , Inibidores da Angiogênese/sangue , Inibidores da Angiogênese/metabolismo , Animais , Bevacizumab/administração & dosagem , Bevacizumab/sangue , Olho/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-Dawley , Receptores de Fatores de Crescimento do Endotélio Vascular/administração & dosagem , Receptores de Fatores de Crescimento do Endotélio Vascular/sangue , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/sangueRESUMO
PURPOSE: Bevacizumab (BCZ) is a recombinant monoclonal antibody that inhibits the biological activity of the vascular endothelial growth factor, which has an important role in angiogenesis for tumoral growth and progression. In this way, our objective was to develop chitosan-coated lipid-core nanocapsules functionalized with BCZ by an organometallic complex using gold-III. METHODS: The formulation was produced and characterized in relation to physicochemical characteristics. Furthermore, the antitumoral and antiangiogenic activities were evaluated against C6 glioma cell line and chicken embryo chorioallantoic membrane (CAM), respectively. RESULTS: Final formulation showed nanometric size, narrow polydispersity, positive zeta potential and gold clusters size lower than 2 nm. BCZ in aqueous solution (0.01-0.10 µmol L-1) did not show cytotoxic activity in vitro against C6 glioma cell line; although, MLNC-Au-BCZ showed cytotoxicity with a median inhibition concentration of 30 nmol L-1 of BCZ. Moreover, MLNC-Au-BCZ demonstrated cellular internalization dependent on incubation time and BCZ concentration. BCZ solution did not induce significant apoptosis as compared to MLNC-Au-BCZ within 24 h of treatment. CAM assay evidenced potent antiangiogenic activity for MLNC-Au-BCZ, representing a decrease of 5.6 times in BCZ dose comparing to BCZ solution. CONCLUSION: MLNC-Au-BCZ is a promising product for the treatment of solid tumors.
Assuntos
Inibidores da Angiogênese/química , Bevacizumab/química , Quitosana/química , Glioma/tratamento farmacológico , Ouro/química , Lipídeos/química , Nanocápsulas/química , Inibidores da Angiogênese/farmacologia , Animais , Apoptose/efeitos dos fármacos , Bevacizumab/metabolismo , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Embrião de Galinha , Membrana Corioalantoide/efeitos dos fármacos , Complexos de Coordenação/química , Relação Dose-Resposta a Droga , Composição de Medicamentos/métodos , Hexoses/química , Humanos , Lectinas de Plantas/química , Polissorbatos/química , Proteínas de Soja/química , Propriedades de Superfície , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
The unstable response to bevacizumab is a big dilemma in the antiangiogenic therapy of high-grade glioma that appears to be linked to an increase in the post-treatment intratumor levels of hypoxia-inducible factor 1 α (HIF1α) and active AKT. Particularly, a selective phosphodiesterase IV (PDE4) inhibitor, rolipram is capable of inhibiting HIF1α and AKT in cancer cells. Here, the effect of bevacizumab alone and in presence of rolipram on therapeutic efficacy, intratumor hypoxia levels, angiogenesis, apoptosis and proliferation mechanisms were evaluated. BALB/c mice bearing C6 glioma were received bevacizumab and rolipram either alone or combined for 30 days (nâ¯=â¯11/group). At the last day of treatments, apoptosis, proliferation and microvessel density, in xenografts (3/group) were detected by TUNEL staining, Ki67 and CD31 markers, respectively. Relative expression of target proteins was measured using western blotting. Bevacizumab initially hindered the tumor progression but its antitumor effect was weakened later despite the vascular regression and apoptosis induction. Unpredictably, bevacizumab-treated tumors exhibited the highest cell proliferation coupled with PDE4A, HIF1α and AKT upregulation and p53 downregulation and reversed by co-treatment with rolipram. Unlike a similar antivascular pattern to bevacizumab, rolipram consistently led to a more tumor growth suppression and proapoptotic effect versus bevacizumab. Co-treatment maximally hampered the tumor progression and elongated survival along with the major vascular regression, hypoxia, apoptosis induction, p53 and caspase activities. In conclusion, superior and persistent therapeutic efficacy of co-treatment provides a new insight into antiangiogenic therapy of malignant gliomas, suggesting to be a potential substitute in selected patients.
Assuntos
Bevacizumab/farmacologia , Glioblastoma/tratamento farmacológico , Rolipram/farmacologia , Inibidores da Angiogênese/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica , Apoptose/efeitos dos fármacos , Bevacizumab/metabolismo , Neoplasias Encefálicas/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Feminino , Glioblastoma/metabolismo , Glioma/tratamento farmacológico , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/tratamento farmacológico , Inibidores da Fosfodiesterase 4/farmacologia , Rolipram/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Promising results have been described for antibodies binding vascular endothelial growth factor (VEGF) in patients with corneal neovascularization. Whether veterinary patients would also benefit from this therapeutic approach has not been investigated yet. We examined binding properties of anti-human VEGF antibodies bevacizumab (Avastin®) and aflibercept (Zaltrap®) for canine, feline, and equine VEGF. METHODS: Human, equine, feline, and canine VEGF were analyzed for sequence similarity using the "Basic Local Alignment Search Tool" (BLAST). Western-blot analysis and ELISA were used to assess binding properties. RESULTS: BLAST analysis revealed a sequence homology of canine, feline, and equine VEGF to human VEGF-A of 93%, 92%, and 89%, respectively. Western-blot analysis showed immunoreactivity of bevacizumab with human, canine, and feline VEGF, but not with equine VEGF. Aflibercept recognized VEGF of all tested species. ELISA data indicated that bevacizumab and aflibercept bind canine VEGF in a dose-dependent manner. Feline VEGF was bound by bevacizumab and aflibercept in a dose-independent manner. ELISA study further confirmed the lack of bevacizumab binding to equine VEGF, and yielded also a dose-independent binding by aflibercept. CONCLUSIONS: Bevacizumab and aflibercept turned out to bind VEGF with species-specific differences. Further studies are required to investigate their efficacy and safety under clinical conditions.
Assuntos
Inibidores da Angiogênese/metabolismo , Bevacizumab/metabolismo , Gatos/metabolismo , Cães/metabolismo , Cavalos/metabolismo , Receptores de Fatores de Crescimento do Endotélio Vascular/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Western Blotting , Ensaio de Imunoadsorção Enzimática , Humanos , Ligação ProteicaRESUMO
Glioblastoma multiforme (GBM) is the most aggressive and common type of brain cancer. Five-year survival rates are below 12%, even with the most aggressive trimodal therapies. Poor blood-brain barrier (BBB) permeability of therapeutics is a major obstacle to efficacy. Intravenous administration of bevacizumab is the standard treatment for GBM. It has been recently demonstrated that a single intraarterial infusion of bevacizumab provides superior therapeutic outcomes in patients with recurrent GBM. Further GBM treatment benefits can be achieved through opening of the BBB before intraarterial infusion of bevacizumab. However, a rationale for intraarterial delivery and BBB opening when delivering antibodies is lacking. A method facilitating quantification of intraarterial delivery of bevacizumab is needed for more effective and personalized GBM treatment. Here, we demonstrate such a method using PET imaging of radiolabeled bevacizumab. Methods: Bevacizumab was conjugated with deferoxamine and subsequently radiolabeled with 89Zr. 89Zr-bevacizumab deferoxamine (89Zr-BVDFO) was prepared with a specific radioactivity of 81.4 ± 7.4 MBq/mg (2.2 ± 0.2 µCi/mg). Brain uptake of 89Zr-BVDFO on carotid artery and tail vein infusion with an intact BBB or with BBB opening with mannitol was initially monitored by dynamic PET, followed by whole-body PET/CT at 1 and 24 h after infusion. Th ex vivo biodistribution of 89Zr-BVDFO was also determined. Results: Intraarterial administration of 89Zr-BVDFO resulted in gradual accumulation of radioactivity in the ipsilateral hemisphere, with 9.16 ± 2.13 percentage injected dose/cm3 at the end of infusion. There was negligible signal observed in the contralateral hemisphere. BBB opening with mannitol before intraarterial infusion of 89Zr-BVDFO resulted in faster and higher uptake in the ipsilateral hemisphere (23.58 ± 4.46 percentage injected dose/cm3) and negligible uptake in the contralateral hemisphere. In contrast, intravenous infusion of 89Zr-BVDFO and subsequent BBB opening did not lead to uptake of radiotracer in the brain. The ex vivo biodistribution results validated the PET/CT studies. Conclusion: Our findings demonstrate that intraarterial delivery of bevacizumab into the brain across an osmotically opened BBB is effective, in contrast to the intravenous route.
Assuntos
Artérias/metabolismo , Bevacizumab/metabolismo , Barreira Hematoencefálica/metabolismo , Osmose , Tomografia por Emissão de Pósitrons , Radioisótopos/metabolismo , Zircônio/metabolismo , Animais , Bevacizumab/farmacocinética , Bevacizumab/uso terapêutico , Marcação por Isótopo , Camundongos , Radioisótopos/farmacocinética , Radioisótopos/uso terapêutico , Distribuição Tecidual , Zircônio/farmacocinética , Zircônio/uso terapêuticoRESUMO
PURPOSE: To compare functional and anatomic outcomes of treatment with intravitreal aflibercept versus bevacizumab and/or ranibizumab in patients with recurrent macular edema (ME) secondary to central retinal vein occlusion (CRVO). METHODS: Retrospective, comparative case series of patients with recurrent ME in the setting of CRVO. Patients with recurrent ME received treatment with aflibercept (Group 1, G1) or bevacizumab and/or ranibizumab (Group 2, G2). Primary outcome measures were best-corrected visual acuity (BCVA) and central foveal thickness (CFT). RESULTS: Of the 20 eyes (20 patients) with recurrent ME included in the study, 9 received aflibercept (G1) and 11 received bevacizumab and/or ranibizumab (G2). Median BCVA at recurrence of ME and at most recent follow-up was 20/60 (G1) and 20/80 (G2) and 20/40 (G1) and 20/50 (G2, P > 0.05 for all comparisons), respectively. Median CFT at recurrence of ME and at most recent follow-up was 492 µm (G1) and 448 µm (G2) and 291 µm (G1) and 295 µm (G2, P > 0.05 for all comparisons), respectively. Complete resolution of ME for at least 4 months was found in 78% (G1) and 55% (G2) of patients with a median injection free interval of 11 (G1) and 13 (G2) months (P > 0.05). CONCLUSIONS: In patients with recurrent ME secondary to CRVO, there was improvement in BCVA and CFT in all groups, although patients treated with aflibercept showed a trend toward better anatomical outcomes decreased need for recurrent injections.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Bevacizumab/metabolismo , Edema Macular/tratamento farmacológico , Ranibizumab/metabolismo , Receptores de Fatores de Crescimento do Endotélio Vascular/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Oclusão da Veia Retiniana/tratamento farmacológico , Idoso , Inibidores da Angiogênese/administração & dosagem , Bevacizumab/administração & dosagem , Feminino , Humanos , Injeções Intravítreas , Masculino , Ranibizumab/administração & dosagem , Receptores de Fatores de Crescimento do Endotélio Vascular/administração & dosagem , Proteínas Recombinantes de Fusão/administração & dosagemRESUMO
PURPOSE: This study aimed to evaluate the effects of bevacizumab, ranibizumab, and aflibercept on the microRNA (miRNA) expression in human retinal pigment epithelium cell (ARPE-19) culture model of oxidative stress. METHODS: Control cells were cultured in the hydrogen peroxide (H2O2)-free medium. In H2O2 group ARPE-19 cells were exposed to 600 µM H2O2 alone for 18 h. In study groups, cells were preincubated with bevacizumab, ranibizumab, and aflibercept (1.25-2.5, 0.5 and 2.0 mg/mL, respectively) for 3 h before H2O2 exposure. Another group of ARPE-19 cells were incubated with drugs for 3 h without H2O2 exposure. Cell viability and vascular endothelial growth factor (VEGF) levels were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and enzyme-linked immunosorbent assay. The expression levels of 1,152 miRNAs were determined by quantitative real-time PCR. RESULTS: Incubation with 600 µM H2O2 alone for 18 h decreased cell viability by â¼50%. Cell viability was greater in the anti-VEGF drug groups compared with the H2O2 group, but the differences were not significant (P > 0.05). VEGF levels were significantly lower in the anti-VEGF drug groups compared with the H2O2 group (P < 0.05 for all study groups), with no significant differences between the study groups (P > 0.05). Incubation with anti-VEGF drugs alone had no effect on miRNA expression in ARPE-19 cells. However, preincubation with bevacizumab, ranibizumab, and aflibercept significantly altered the profile of H2O2-modulated miRNA expression. CONCLUSIONS: Preincubation with anti-VEGF drugs can alter the miRNA expression profile in response to H2O2-induced oxidative stress, and these drugs may have epigenetic effects.
Assuntos
Inibidores da Angiogênese/farmacologia , Bevacizumab/metabolismo , MicroRNAs/genética , Ranibizumab/metabolismo , Receptores de Fatores de Crescimento do Endotélio Vascular/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Epitélio Pigmentado da Retina/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Perfilação da Expressão Gênica , Humanos , Modelos Biológicos , Estresse Oxidativo/efeitos dos fármacos , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Inadequate tumor uptake of the vascular endothelial growth factor antibody bevacizumab could explain lack of effect in diffuse intrinsic pontine glioma. Methods: By combining data from a PET imaging study using 89Zr-labeled bevacizumab and an autopsy study, a 1-on-1 analysis of multiregional in vivo and ex vivo 89Zr-bevacizumab uptake, tumor histology, and vascular morphology in a diffuse intrinsic pontine glioma patient was performed. Results: In vivo 89Zr-bevacizumab measurements showed heterogeneity between lesions. Additional ex vivo measurements and immunohistochemistry of cervicomedullary metastasis samples showed uptake to be highest in the area with marked microvascular proliferation. In the primary pontine tumor, all samples showed similar vascular morphology. Other histologic features were similar between the samples studied. Conclusion: In vivo 89Zr-bevacizumab PET serves to identify heterogeneous uptake between tumor lesions, whereas subcentimeter intralesional heterogeneity could be identified only by ex vivo measurements. 89Zr-bevacizumab uptake is enhanced by vascular proliferation, although our results suggest it is not the only determinant of intralesional uptake heterogeneity.