RESUMEN
Bone grafting and reconstruction are still challenging in clinical practice because of the limitations of bone autografts and the drawbacks of currently approved bone substitutes. We thus developed a gene-activated bone substitute based on octacalcium phosphate and naked plasmid DNA carrying the vascular endothelial growth factor gene. This advanced combined therapy medicinal product had no cytotoxic effects in vitro, slightly decreased bone marrow mesenchymal stromal cell (MSC) doubling time, and was characterized by a prolonged level of gene construct delivery in vivo in a luciferase bioimaging assay. In the model of critically sized cranial bone defects in rabbits, the gene-activated matrix increased bone tissue formation through angiogenesis induction. After preclinical studies, we conducted an open-label non-randomized clinical trial (NCT03076138). The primary study outcome was the proportion of patients with newly formed bone tissue within the surgical area as measured by computed tomography within 6 months after surgery. The main secondary outcomes included frequencies of adverse events (AEs) and serious adverse events (SAEs) as well as the surgical failure rate. After completing the clinical trial, the patients had dental implants placed in the bone grafting area, and trephine biopsy samples were collected. In total, 20 patients with alveolar ridge atrophy (n = 16) and jaw bone defects (n = 4) were enrolled in the study. There were no AEs or SAEs during the clinical trial or the follow-up period (30 months). In all patients, newly formed tissues with a bone density of 908.13 ± 114.40 HU were detected within the zone of bone grafting. There were no significant differences between the subgroups of patients with atrophy and bone defects: 915.28 ± 125.85 and 879.56 ± 48.36 HU, respectively (p = 0.60). Histological analysis showed that the bone grafting area comprised newly formed bone tissue with some fragments of the gene-activated bone substitute partially resorbed and integrated with bone, without fibrous tissue in between. The preclinical data and clinical trial results proved the feasibility, safety, and efficacy of the investigated material for jaw bone grafting, allowing us to bring the world's first gene-activated bone substitute from bench to bedside.
RESUMEN
Bone reconstruction techniques are mainly based on the use of tissue grafts and artificial scaffolds. The former presents well-known limitations, such as restricted graft availability and donor site morbidity, while the latter commonly results in poor graft integration and fixation in the bone, which leads to the unbalanced distribution of loads, impaired bone formation, increased pain perception, and risk of fracture, ultimately leading to recurrent surgeries. In the past decade, research efforts have been focused on the development of innovative bone substitutes that not only provide immediate mechanical support, but also ensure appropriate graft anchoring by, for example, promoting de novo bone tissue formation. From the countless studies that aimed in this direction, only few have made the big jump from the benchtop to the bedside, whilst most have perished along the challenging path of clinical translation. Herein, we describe some clinically successful cases of bone device development, including biological glues, stem cell-seeded scaffolds, and gene-functionalized bone substitutes. We also discuss the ventures that these technologies went through, the hindrances they faced and the common grounds among them, which might have been key for their success. The ultimate objective of this perspective article is to highlight the important aspects of the clinical translation of an innovative idea in the field of bone grafting, with the aim of commercially and clinically informing new research approaches in the sector.
RESUMEN
The stromal-derived factor-1 alpha (SDF-1α) and vascular endothelial growth factor (VEGF) play an important role in angiogenesis and exert a significant trophic function. SDF-1α is a chemoattractant for endothelial progenitor cells derived from bone marrow and promotes new blood vessel formation. VEGF regulates all types of vascular growth, stimulates angiogenesis, and is involved in the induction of lymphangiogenesis. The possibility of using these growth factors for regenerative medicine is currently under investigation. The angiogenic potential of a pBud-SDF-1α-VEGF165 bicistronic plasmid construct which simultaneously encodes VEGF165 and SDF-1α genes cDNA was evaluated in this study. The conditioned medium collected from HEK293T cells transfected with the pBud-SDF-1α-VEGF165 plasmid was shown to stimulate the formation of capillary-like structures by human umbilical vein-derived endothelial cells (HUVEC) on Matrigel and to increase the proliferative activity of these cells in vitro. Thus, the pBud-SDF-1α-VEGF165 plasmid exhibits angiogenic properties in cell cultures in vitro. As interest in the development of non-viral techniques for regenerative medicine increases, this plasmid which simultaneously expresses VEGF165 and SDF-1α may provide a platform for advanced methods of stimulating therapeutic angiogenesis.
Asunto(s)
Quimiocina CXCL12/genética , ADN/genética , Neovascularización Fisiológica/genética , Plásmidos , Factor A de Crecimiento Endotelial Vascular/genética , Células HEK293 , Humanos , Inmunofenotipificación , Técnicas In VitroRESUMEN
BACKGROUND: The effective treatment of chronic lower limb ischemia is one of the most challenging issues confronting vascular surgeons. Current pharmacological therapies play an auxiliary role and cannot prevent disease progression, and new treatment methods are needed. In 2011, a plasmid VEGF65-gene therapy drug was approved in Russia for the treatment of chronic lower limb ischemia ( ClinicalTrials.gov identifier: NCT03068585). The objective of this follow-up study was to evaluate the long-term safety and efficacy of gene therapy in patients with limb ischemia of atherosclerotic genesis. AIMS: To evaluate the long-term safety and efficacy of the therapeutic angiogenesis, 36 patients in the treatment group (pl- VEGF165) and 12 patients in the control group participated in a 5-year follow-up study. Planned examinations were carried out annually for 5 years after pl- VEGF165 administration. RESULTS: Differences in the frequency of major cardiovascular events (pl- VEGF165 5/36 versus control 2/12; p = 0.85), malignancies (pl- VEGF165 1/36 versus control 0/12; p = 0.38) and impaired vision (there was none in either group) over the 5-year follow-up period did not achieve statistical significance. The target limb salvage was 95% ( n = 36) and 67% ( n = 12) in the pl- VEGF165 and control groups, respectively. The pain-free walking distance value increased by 288% from 105.7 ± 16.5 m to 384 ± 39 m in the treatment group by the end of the fifth year, with a peak of 410.6 ± 86.1 m achieved by the end of the third year. The ankle-brachial index (ABI) increased from 0.47 ± 0.01 to 0.56 ± 0.02 by the end of the first year, with a subsequent slight decrease to 0.51 ± 0.02 by the fifth year. The maximum increment of transcutaneous oximetry test (tcoO2) by 36%, from 66.6 ± 3.7 mm Hg to 90.7 ± 4.9 mm Hg, was observed by the end of the second year. CONCLUSION: The therapeutic effect of angiogenesis induction by gene therapy persists for 5 years.
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Terapia Genética/métodos , Claudicación Intermitente/terapia , Isquemia/terapia , Extremidad Inferior/irrigación sanguínea , Enfermedad Arterial Periférica/terapia , Factor A de Crecimiento Endotelial Vascular/genética , Anciano , Índice Tobillo Braquial , Monitoreo de Gas Sanguíneo Transcutáneo , Tolerancia al Ejercicio , Femenino , Estudios de Seguimiento , Terapia Genética/efectos adversos , Humanos , Claudicación Intermitente/genética , Claudicación Intermitente/metabolismo , Claudicación Intermitente/fisiopatología , Isquemia/genética , Isquemia/metabolismo , Isquemia/fisiopatología , Recuperación del Miembro , Masculino , Persona de Mediana Edad , Neovascularización Fisiológica , Enfermedad Arterial Periférica/genética , Enfermedad Arterial Periférica/metabolismo , Enfermedad Arterial Periférica/fisiopatología , Recuperación de la Función , Federación de Rusia , Factores de Tiempo , Resultado del Tratamiento , Factor A de Crecimiento Endotelial Vascular/biosíntesis , Prueba de PasoRESUMEN
INTRODUCTION: The effective treatment of chronic lower limb ischemia is one of the most challenging issues confronting vascular surgeons. Current pharmacological therapies play an auxiliary role and cannot prevent disease progression, and new treatment methods are needed. pl-VEGF165, a gene therapy drug, was approved in Russia for the treatment of atherosclerotic peripheral arterial disease (PAD) after clinical studies in 2011. The study drug is an original gene construction in which pl-VEGF165 1.2 mg is the active substance. OBJECTIVE: This postmarketing surveillance study was undertaken to evaluate the safety (identification of uncommon side effects) and efficacy of gene therapy in patients in routine clinical practice. METHODS: In total, 210 patients with stage II-III chronic limb ischemia (according to the Fontaine classification modified by AV Pokrovsky) in 33 healthcare facilities in Russia and the Ukraine were enrolled in the study. The control group (n = 60) received conservative therapy without prostaglandins and prostacyclins, and the treatment group (n = 150) received treatment with pl-VEGF165 as two intramuscular injections for a total dose of 2.4 mg. Pain-free walking distance (PWD) (the primary efficacy criterion for Fontaine stages II-III), blood flow linear velocity (BFLV), and ankle-brachial index (ABI) were monitored for 6 months. The safety of pl-VEGF165 gene transfer in terms of the trial protocol was initially evaluated 6 months after the start of the study; adverse events (AEs) and serious adverse events (SAEs) were recorded during both routine visits and unscheduled requests for medical care. RESULTS: Overall, PWD increased by 177%, from 100.3 ± 6.9 to 277.1 ± 16.2 m (p = 0.0001), in the treatment group, whereas the mean value was unchanged in the control group (p = 0.218). Both BFLV and ABI values increased by 24% (p = 0.0001) in the treatment group but decreased in the control group. The greatest therapeutic effect was observed for stage III disease: PWD increased by 683% (p = 0.0001). No angiogenic therapy-related AEs or side effects were recorded, and target limb salvage was 96 and 97% in the treatment and control groups, respectively. The results obtained in this study are not significantly different from those observed in the phase IIb/III registration clinical study completed in 2011. CONCLUSION: pl-VEGF165 intramuscular gene transfer is an effective treatment for moderate to severe claudication due to chronic lower limb ischemia in routine clinical practice. ClinicalTrials.gov identifier: NCT02369809.
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Extremidades/fisiopatología , Isquemia/tratamiento farmacológico , Enfermedad Arterial Periférica/tratamiento farmacológico , Factor A de Crecimiento Endotelial Vascular/efectos adversos , Factor A de Crecimiento Endotelial Vascular/uso terapéutico , Anciano , Índice Tobillo Braquial/métodos , Femenino , Terapia Genética/métodos , Humanos , Masculino , Vigilancia de Productos Comercializados , Resultado del TratamientoRESUMEN
Effective treatment of chronic lower limb ischemia is one of the most challenging issues confronting vascular surgeons. There are a number of choices available to the vascular surgeon. Open or endovascular revascularization is the treatment of choice when applicable. Current pharmacological therapies play an auxiliary role and cannot prevent disease progression. Therefore, new methods of treatment are needed. We conducted a phase 2b/3 multicenter randomized controlled clinical trial of the intramuscular transfer of a plasmid DNA encoding vascular endothelial growth factor (VEGF) 165 with cytomegalovirus promotor (CMV) in patients with atherosclerotic lower limb ischemia. A total of 100 patients were enrolled in the study, that is, 75 patients were randomized into the test group and received 2 intramuscular injections of 1.2 mg of pCMV-vegf165, 14 days apart together with standard pharmacological treatment. In all, 25 patients were randomized into the control group and received standard treatment only. The following end points were evaluated within the first 6 months of the study and during a 1.5-year additional follow-up period: pain-free walking distance (PWD), ankle-brachial index (ABI), and blood flow velocity (BFV). The pCMV-vegf165 therapy appeared to be significantly more effective than standard treatment. The PWD increased in the test group by 110.4%, 167.2%, and 190.8% at 6 months, 1 year, and 2 years after treatment, respectively. The pCMV-vegf165 intramuscular transfer caused a statistically significant increase in ABI and BFV. There were no positive results in the control group. Thus, pCMV-vegf165 intramuscular gene transfer is an effective method of treatment of moderate to severe claudication due to chronic lower limb ischemia.