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    • 专利摘要:
    The present invention relates to the use of biodegradable centroids for the release of radiation-sensitive anticancer agents for the manufacture of drugs which are intended to be used simultaneously, alone or over time for the treatment of gliomas. Use of the biodegradable centrosomes of the present invention results in patient survival time of at least 90 weeks, and therapeutically effective concentrations are maintained in the parenchymal space over this period. The core preferably contains 5-fluorouracil coated with poly (d, l-lactic acid-co-glycolic acid). The centrosome is implanted into the wall of the surgical site after excision of the tumor by in vivo injection. Radiation therapy targeting the tumor mass is irradiated at 60 Gy over about six weeks. The invention also relates to a process for preparing biodegradable centroids by emulsion-extraction and to suspensions containing the biodegradable centroids obtained using the method.
    公开号:KR20020000178A
    申请号:KR1020017014652
    申请日:2000-05-17
    公开日:2002-01-04
    发明作者:나탈리 페장뜨；쟝-피에르 베느와뜨；필리페 마네
    申请人:제라르드 레뒤끄；라보하또와훼 데 쁘호뒤뜨 에띠끄 에띠빠흠；
    IPC主号:
    专利说明:

    Use of biodegradable microspheres that release an anticancer agent for treating glioblastoma}
    [2] Gliomas belong to the group of rare diseases listed in the National Organization for Rare Disorders.
    [3] Malignant glioma is the primary tumor of the central nervous system, representing 13-22% of the cranial tumors, depending on the series. From a histological point of view, both types of malignant glioma are actually classified into undifferentiated astrocytoma and gliomas, with gliomas representing the most undifferentiated form of these tumors.
    [4] At present, there is no effective treatment for malignant glioma. Survival time for patients with gliomas does not exceed 1 year, even if chemotherapy and radiation therapy are combined with surgery.
    [5] Treatment of malignant glioma is mainly limited to three phenomena.
    [6] The first is the presence of a blood-brain barrier (BBB) that separates the central nervous system from the rest of the body. This BBS only passes small fat-soluble molecules. Other molecules must be administered at high doses to reach the central nervous system, which results in significant systemic side effects.
    [7] The second factor limiting the therapeutic efficacy of glioma is the invasiveness of these tumors. Because the brain is a highly functional organ, it is not possible to perform exclusive surgery on the brain in an oncological sense. The most complete resection possible is visually complete resection, so many tumor cells will infiltrate into the wall of the resection cavity. Many authorities have also found that 90% of malignant gliomas treated with radiation therapy recur within 2 cm of the initial tumor site.
    [8] The final factor limiting the therapeutic efficiency of glioma is a low therapeutic index. Tumor cells are hidden behind normal tissue, which is extremely susceptible to attack, for example by radiation therapy or certain anticancer agents.
    [9] The progress achieved in the treatment of glioma is insufficient [Kornblith PL, Walker M, Chemotherapy for malignant gliomas. J. Neurosurg, 68: 1-17, 1988; Shapiro WR, Green SB, Burger PC, Selker RG, VanGilder JC, Robertson JT, Mahaley SM, A randomized comparison of intra-arterial versus intravenous BCNU with or without intravenous 5-fluorouracil, for newly diagnosed patients with malignant glioma, J. Neurosurg . 76: 772-781, 1992.
    [10] Currently, conventional treatments for gliomas that are subsequently surgically resected are based on external radiation therapy. This cannot achieve a survival time of more than one year. Combination of radiation therapy with chemotherapy using 1- (2-chloroethyl) -3-cyclohexyl-1-nitrosouurea (BCNU) is effective only against undifferentiated astrocytoma. This contributes only moderately because it only increases the rate of survivors for 18 months without changing the survival time.
    [11] In addition, immunotherapy has not established itself in this field, and gene therapy has not yet proven itself.
    [12] Experiments have been conducted using subcutaneous reservoirs with various techniques aimed at increasing local concentrations of anticancer agents, such as osmotic rupture of the blood brain barrier, infusion into the cerebrospinal fluid, intracarotid infusion, and intratumoral administration. RJ and Brem H, Drug delivery to the central nervous system, Neurosurgery Quarterly, 2: 259-279, 1992]. These techniques cannot increase the survival time of patients and some techniques have proven very toxic.
    [13] Over the past few years, research in galenic preparations has developed implantable polymer systems that can reduce systemic side effects and protect the active substances from degeneration for a period of time and control local release. With the benefits of such implantable polymer systems, several teams have recently been urged to study their use in central nervous system pathology (Langer R, Polymer implants for drug delivery in the brain, J. Controlled Release, 16 :). 53-60, 1991]. In particular, such a system implanted into the tumor resection wall of malignant glioma slows tumor recurrence and prolongs patient survival. The isolated malignant cells persist around the cavity remaining after surgery, which causes 90% of recurrences and occurs within 2 cm of the surgical site. Within this site, the nerve tissue has a function and the blood brain barrier is still intact, limiting the action of conventional chemotherapy and radiation therapy.
    [14] Various implantable polymer systems have been developed and tested in animals that release active molecules. PCPP-SA (poly [1,3-bis (carboxyphenoxy) propane-co-sebacic acid]) and BCNU ( Biodegradable wafer systems have been developed despite careful results in clinical studies (Brem H, Polymers to treat brain tumors, Biomaterials 11: 699-701, 1990; Brem H, Mahaley MS, Vick NA, Black KL, Schold SC, Eller TW, Cozzens JW, Kenealy JN, Interstitial chemotherapy with drug polymer implants for the treatment of recurrent gliomas, J. Neurosurg 74: 441-446, 1991; Brem H, Walter KA, Langer R, Polymers as controlled drug delivery devices for the treatment of malignant brain tumors, Eur J Pharm Biopharm, 39 (1): 2-7, 1993; Brem H, Piantadosi S, Burger PC, Walker M, et al., Placebo-controlled trial of safety and efficacy of intraoperative controlled delivery by biodegradable polymers of chemotherapy for recurrent glioma, Lancet, 345: 1008-1-12, 1995].
    [15] Centrals that emit BCNU have been developed, but studies in animals have been relatively inspiring [Res. Al, Boisdron-Celle M, Benoit JP, Formulation of BCNU-loaded microspheres: influence of drug stability and solubility on the design of the microencapsulation procedure, J. Microencapsulation, 13: 41-51, 1996; T, Venier-Julienne MC, Benoit JP, Internal morphology of poly (D, L-lactide-co-glycolide) BNCU-loaded microspheres. Influence on drug stability, Eur. J. Pharm. Biopharm, 1998, 45, 31-39.
    [1] The present invention relates to the use of biodegradable centroids to release anticancer agents for treating gliomas.
    [16] Subject of the present invention is the use of an implantable biodegradable centroid that releases an anticancer agent for treating gliomas. Use of these centrosomes is combined with radiation therapy and surgery. After tumor resection, biodegradable centrosomes that release anticancer agents are implanted at the surgical site by in vivo infusion. The radiation therapy is then performed up to 7 days after the intervention.
    [17] By using these centrosomes, the inventors have succeeded entirely in doubling the survival time of patients with gliomas. Specifically, by using the core of the present invention, a survival time of 90 weeks or more can be achieved.
    [18] Accordingly, the present invention is directed to biodegradable centroids that release radiation-sensitive anticancer agents for the manufacture of drugs intended to be used simultaneously, alone or over time to treat gliomas and are implanted at the surgical site after resection of glioma. In the use of an anticancer agent, the centroids containing the anticancer agent are used to delay the release of the anticancer agent and to achieve a survival time of the treated patient for at least about 90 weeks, preferably for about 130 weeks, more preferably for 160 weeks. The present invention relates to the use of biodegradable centrosomes, which are coated with a polymer which maintains a therapeutically effective concentration in a parenchymal space over time.
    [19] The centroids used in the present invention preferably contain anticancer agents that exhibit hydrophilicity and / or do not interfere with the blood brain barrier. Advantageously, anticancer agents do not exhibit central neurotoxicity. This anticancer agent preferentially acts on dividing cells.
    [20] The anticancer agent consists of a radiosensitive anticancer compound or a mixture of anticancer compound containing one or more radiation sensitive anticancer compound, wherein the anticancer compound (s) is, for example, 5-fluorouracil (5-FU), carboplatin and cisplatin Taxanes such as Platin, Docetaxel and Paclitaxel, Gitatabine, VP16, Mitomycin, Indosuridine, Topoisomerase 1 inhibitors such as Irinotecan, Topotecan and Camptothecin, Nitroso such as BCNU, ACNU or MCNU Urea, methotrexate, bleomycin, adriamycin, cytosan and vincristine, immunoregulatory cytokines such as IL2, IL6, IL12 and IL13 and interferon.
    [21] The anticancer agent is preferably 5-FU.
    [22] 5-FU is a known long-acting antimitotic agent. It is a hydrophilic molecule whose activity is increased by topical administration because it very poorly interferes with the blood brain barrier. Bourke RS, West CR, Chheda G et al., Kinetics of entry and distribution of 5-fluorouracil in CSF and brain following intravenous injection in primate, Cancer Res, 33: 1735-1746, 1973; Gerosa MA, Dougherty DV, Wilson CB, Rosenblum ML, Improved treatment of a brain tumor model, Part 2: Sequential therapy with BCNU and 5-fluorouracil, J. Neurosurg. 58: 368, 1983; Kotsilimbas DG, Karpf R, Meredith S, Scheinberg LC, Evaluation of parenteral 5-FU on experimental brain tumors, Neurology, 16: 916-918, 1966; Levin VA, Edwards MS, Wara WM, Allen J, Ortega J, Vestnys P, 5-fluorouracil and 1- (2-chloroethyl) -3-cyclohexyl-1-nitrosourea (CCNU) followed by hydroxyurea, misonidazole and irradiation for brain stem gliomas: a pilot study of the brain tumor research center and the children cancer group, Neurosurgery, 14: 679-681, 1984; Oda Y, Tokuriki Y, Tsuda E, Handa H, Kieler J, Trial of anticancer pellet im malignant brain tumours, 5-FU and urokinase embedded in silastic. Proceeding the 6th European Congress of Neurosurgery, Acta neurochirurgica, Suppl. 28: 489-490, 1979; Penn RD, Kroin JS, Harris JE, Chiu KM, Braun DP, Chronic intratumoral chemotherapy of a rat tumor with cisplatin and fluorouracil, Appl. Neurophysio, 46: 240-244, 1983; Shapiro WR, Studies on the chemotherapy of experimental brain tumors: Evaluation of 1- (2-chloroethyl) -3-cyclohexyl-1-nitrosourea, vincristine and 5-fluorouracil, J. Nat. Cancer Institute, 46 (2), 359-368, 1971; Shapiro WR, Green SB, Burger PC, Selker RG, VanGilder JC, Robertson JT, Mahaley SM, A randomized comparison of intra-arterial versus intravenous BCNU with or without intravenous 5-fluorouracil, for newly diagnosed patients with malignant glioma, J. Neurosurg 76: 772-781, 1992; Soloway AH, Mark VH, Dukat EG et al., Chemotherapy of brain tumors. 1-Transplanted murine ependymoblastomas, Cancer Chemother Rep., 36: 1-4, 1964].
    [23] Thus, the activity of 5-FU is increased by sustained administration. 5-FU is an agent that acts by interfering with nucleic acid synthesis. Studies have shown that only 30-50% of the cells of rat malignant glioma (L9) divide in a predetermined period of time. In addition, the glioblastoma cell cycle is long (20 hours in L9 glioma and 3 to 7 days in human gliomas). From now on, 5-FU clearance in plasma is fast (30 min half-life) [Neuwelt EA, Barnett PA, Frenkel EP, Chemotherapeutic agent permeability to normal brain and delivery to avian sarcoma virus-induced brain tumors in the rodent: observation on problems drug delivery, Neurosurgery, 14: 154-160, 1984]. Thus, 5-FU cannot destroy many malignant cells by systemic administration or local infusion.
    [24] 5-FU is substantially active against rapidly regenerating tissues and exceptionally neurotoxic. 5-FU interferes with nucleic acid synthesis, and tissues that grow rapidly are particularly needed to ensure their proliferation and regeneration. This is of course not the case with brain tissue, and mitosis is rare in steady state and occurs only in glial populations. The toxic effect of 5-FU, which limits its administration via the common route, is virtually present in the blood and stomach. Rare neurological side effects of 5-FU have been disclosed, but their relatively unknown pathogenesis is presumably due to the exacerbation of the Krebs cycle by exacerbation of multifactorial (5-FU metabolites or preexisting thiamine deficiency. Aoki N, Reversible leukoencephalopathy caused by 5-fluorouracil derivatives, presenting as akinetic mutism, Surg Neurol, 25: 279-282, 1986: Moore DH, Fowler WC, Crumpler LS, 5-fluorouracil neurotoxicity, case report, Gynecol Oncology, 36: 152-154, 1990].
    [25] Finally, 5-FU shows radiation sensitivity. See Koutcher JA, Alfieri AA, Thaler H et al., Radiation enhancement by biochemical modulation and 5-FU, Int. J. Radit, Biol. Phys., 39: 1145-1152, 1997]. The excellence of the combination of 5-FU / radiation therapy in each of these separate treatments has already been demonstrated in 1960 for animal models and tumor cells in vitro. Bagshaw M, A possible role of potentiation in radiation therapy Amer J. Roentgenol, 85: 822-833, 1961; Vietti T, Eggerding F, Valeriote, F, Combined effect of X-radiation and 5-fluorouracil on survival of transplanted leukemic cells, J. Natl. Inst., 47: 865-870, 1971]. This synergistic effect is presumably due to the synchronization of tumor cell populations and the reduction of cell repair mechanisms by 5-FU. Combination of radiation therapy with antipyrimidine (5-FU or BrudR) has already been tried in humans [Goffman TE, Dachowski LJ, Bobo H et al., Long term follow-up on national cancer institute phase I / II study of glioblastoma multiforme treated with iododeoxyuridine and hyperfractionated irradiation, J. Clinical Oncology, 10: 264-268, 1992].
    [26] The lack of pronounced effect will be explained once again by the systemic route of administration of the drug.
    [27] If the anticancer agent is 5-FU, the concentration of the anticancer agent in the cerebrospinal fluid, which reflects the concentration in the parenchyma, is 3 to 20 ng / ml.
    [28] In order to limit the neurotoxicity of the anticancer agents contained in the core used in the present invention, neuroprotective compounds may be advantageously added to the anticancer agents. This neuroprotective compound is selected from peptide growth factors such as, for example, NGF or BDNF.
    [29] Biodegradable centroids used in the present invention are coated with a polymer that delays the release of the anticancer agent and maintains a therapeutically effective concentration in the parenchymal space for a period of at least three weeks, preferably at least four weeks.
    [30] The polymer is selected from ethylcellulose, polystyrene, poly (ε-caprolactone), poly (d, l-lactic acid) and poly (d, l-lactic acid-co-glycolic acid).
    [31] The polymer is preferably poly (d, l-lactic-co-glycolic acid), which is a biodegradable polymer that is acceptable for the formulation of sustained release galenose agents (unlike PCPP-SA, which is not approved for large scale clinical use). to be.
    [32] Poly (d, l-lactic acid-co-glycolic acid) is preferably 50:50 PLAGA (ie containing the same amount of lactic acid and glycolic acid), for example RG 506 (BI Chimie, France) with a weight average molecular weight of 72000, a polydispersity index of 1.8 and an intrinsic viscosity of 0.80 dl / g (0.1% polymer solution in chloroform at 25 ° C).
    [33] PLAGA is a hydrophobic copolymer and its degradation due to the hydrolysis reaction yields two conventional biological substrates, lactic acid and glycolic acid, and metabolize with CO 2 and H 2 O at the end of aerobic glycolysis. Long-term studies have shown that the respiratory pathway is the major pathway for the removal of these two substrates. The biodegradation rate of PLAGA depends on the respective ratios of lactic acid and glycolic acid. PLAGA exhibits complete biocompatibility and results in moderate foreign body reactions [Visscher GE, RL Robinson, HV Mauding, Fong JW, Pearson JE, Argentieri GJ, Biodegradation of and tissue reaction to 50:50 poly (DL-lactide-co -glycolide) microcapsules, J. Biomed. Mat. Res. 19: 345-365, 1985. PLAGA is a component of surgical sutures [Fraczza EJ, Schmidt EE, A new absorbable suture, J. Biomed. Mater. Res., 5: 43-58, 1971], is a subcutaneous implantable galenose preparation [Jalil R, Nixon JR, Biodegradable poly (lactic acid) and poly (lactide-co-glycolide) microcapsules: problems associated with preparative techniques and release properties (Review), J. Microencapsulation, 7: 297-325, 1990. 50:50 PLAGA centrosomes can be sterilized by γ radiation and once implanted into the rodent brain by stereoposition, it is fully biodegraded within two months, demonstrating that only a moderate response in the form of nonspecific glial astrocytic cells and histocytes occurs. Menei P, Daniel V, Montero-Menei C, Brouillard M, Pouplard-Barthelaix A, Benoit JP: Biodegradation and brain tissue reaction to poly (DL-lactide-co-glycolide) microspheres, Biomaterials 14: 470- 478, 1993; Menei P, Croue A, Daniel V, Pouplard-Barthelaix A, Benoit JP: Fate and biocompatibility of three types of microspheres implanted into the brain, J. Biomed Mat Res, 28, 1079-1085, 1994]. The latter results are later described in Kou JH, Emmett C, Shen P et al., Bioerosion and biocompatibility of poly (d, l-lactic-co-glycolic acid) implantsin brain, J Control Release, 43, 123- 130, 1997].
    [34] The biodegradable centroids of the present invention have an average diameter of 48 ± 20 μm, preferably 46 ± 7 μm. They contain 15 to 35% by weight of anticancer agent, preferably 19 to 27% by weight of 5-FU, even more preferably 20% by weight of 5-FU, and 65 to 85% by weight of polymer.
    [35] In the present invention, 50:50 PLAGA centroid mediated 5-FU is particularly preferred.
    [36] In vitro, 50:50 PLAGA centromediate 5-FU may release 5-FU for 21 days. In vivo, when implanted subcutaneously in rabbits, these centrosomes can achieve 5-FU plato concentrations in plasma for 23 days. In vivo and in rodent brains, determination of 5-FU is visually observed in the centrosome until at least 19 days. In rabbits, after implantation in the cerebrum of the PLAGA-5-FU centroid (5-FU 7 mg / kg), no 5-FU trace was detected in the serum, suggesting that the drug was not substantially passed through the body circulation.
    [37] After cerebral transplantation of PLAGA-5-FU centrosomes in rodents, no overall signs of systemic toxicity or clinical or tissue neurotoxicity were observed with a total dose of 17 mg / kg of 5-FU. At a split dose of 24 Gy, the combination of 5-FU centrosome / cerebral radiation therapy is sufficiently resistant [Menei P, Vectorisation dans le SNC par implantation]. Vectorization in the CNS by stereotactic implantation of microspheres Sciences Pharmaceutiques [University doctorate in pharmaceutical sciences], [University of Angers], 1995. Finally, these centrosomes implanted into malignant glioma generated by stereoposition in rats (C6 glioma) significantly reduce mortality. Menei P, Boisdron-Celle M, Croue A, Guy G, Benoit JP , Effect of stereotactic implantation of biodegradable 5-fluorouracil-loaded microspheres in normal and C6-glioma bearing rats, Neurosurgery, 39: 117-124, 1996].
    [38] Advantageously, the centrosome is suspended in sterile solution and the suspension is injected into the wall of the surgical site after tumor resection.
    [39] Sterile solutions are preferably viscosity modifiers, for example sodium carboxymethylcellulose 1 to 1.5% w / v, preferably 1.25% w / v; Surfactants, for example polysorbates 0.5 to 1.5% w / v, preferably 1% w / v; And isotonic agents, for example mannitol 3.5 to 4.5% w / v, preferably 4% w / v.
    [40] The centrosome is preferably suspended immediately prior to injection. The suspension preferably contains 3 ml of the sterile solution described above and 700 to 800 mg of the biodegradable core.
    [41] After diagnosis of gliomas and confirmation of glioblastoma ablation, the suspension of the centrosome is implanted into the wall of the surgical site at a depth of at least 2 cm per cm 2, preferably 2 to 3 cm.
    [42] If the anticancer agent is 5-FU, the total dose of the injected suspension corresponds to an amount of 5-FU of 50-200 mg.
    [43] Radiation therapy focuses on tumor volume, the irradiated volume includes a preoperative tumor with a margin of at least 2 cm in all directions, and a total dose of 50-60 Gy is used.
    [44] Radiation therapy is preferably initiated 2 to 7 days after surgery. The overall dose of 50-60 Gy continues over a period of 4-8 weeks, for example at a rate of 5 fractions per week.
    [45] Radiation therapy is preferably performed at a total dose of 60 Gy for about 6 weeks, preferably at a rate of 5 fractions per week for 6.5 weeks.
    [46] After injecting the centrosome immediately after tumor resection, the fresh infusion of one or more centrosomes is done by stereoposition in case of tumor recurrence.
    [47] The centroids used in the present invention are described in variants of the method described in Boisdron-Celle M, Menei P, Benoit JP: Preparation of biodegradable 5-fluorouracil-loaded microspheres, J Pharm Pharmacol, 47, 108-114, 1995. According to the emulsification-extraction technique.
    [48] The present invention also relates to a method of making a core containing an anticancer agent coated with a polymer used in the present invention. The main step of this method is to prepare an organic phase in which the anticancer agent and the polymer are dispersed in an organic solvent. The organic phase and the aqueous phase are emulsified and then water is added to extract the organic solvent. Finally, the suspension of the obtained centrosome is filtered.
    [49] The process of the present invention is characterized, first of all, by the anticancer agent being dispersed in an organic solvent under vigorous stirring before the polymer is added.
    [50] According to a variant made in the prior art method, the active ingredient is ground in a planetary ball mill. The obtained crystal has a size of 15 to 50 µm. The size of crystals encapsulated and their degree of dispersion are in fact essential criteria for controlling the degree of encapsulation and release kinetics in vitro.
    [51] The active ingredient is then dispersed in an organic solvent of a round bottom tube, preferably dichloromethane, with stirring using a homogenizing rod before the polymer is added.
    [52] By homogenization, a homogeneous suspension is given, the difference between the different batches in one grinding batch is reduced, and the crystal size of the active ingredient is reduced.
    [53] The organic phase is prepared in solvent without auxiliary solvent. Without the cosolvent, precipitation of the polymer is slowed during the emulsifier, resulting in low porosity of the resulting particles.
    [54] The active ingredient dispersion is transferred to the first reactor.
    [55] The polymer is added at a mass ratio of 8 to 13%, preferably 11%. The organic phase obtained is kept at room temperature under constant stirring for 2-4 hours and then at 1-5 ° C., preferably at 2 ° C. for about 15 minutes. The organic phase is stirred at room temperature for a long time to ensure total solubility of the polymer in the solvent.
    [56] The water phase is preferably produced in a second reactor maintained at the same temperature as the organic phase, preferably at 2 ° C. As the temperature of the aqueous and organic phases decreases, the viscosity increases and the degree of encapsulation increases. The aqueous phase is for example a 10% aqueous PVA solution.
    [57] Two jacketed reactors are used and the coolant is circulated continuously in the two reactors. The temperature of the organic phase and the water phase is advantageously the same when the two phases are mixed together, preferably 2 ° C. By good temperature control, both effectively control the particle size, the dissociation rate of the active ingredient and the extraction rate of the solvent at the same time.
    [58] The organic phase is transferred from the first reactor to the second reactor. The water / organic volume ratio is 80/3 to 120/3, preferably 100/3.
    [59] The obtained emulsion is stirred for 3 minutes or more, preferably 3 to 6 minutes, more preferably 5 minutes. The choice of this period is directly related to the release kinetics, and especially directly over the "burst" effect over 24 to 48 hours.
    [60] In the absence of cosolvents combined with sufficient emulsification time, the surface or poorly coated active ingredients are dissolved, so that the release kinetics of the initial phase is well controlled.
    [61] To extract the organic solvent, water is added to the emulsion with an emulsion / water volume ratio of 1/4 to 1/2, preferably 1/3. The temperature of the water for extraction is 1 to 5 ° C, preferably 4 ° C.
    [62] The emulsification and extraction steps are done in the same reactor to limit time and save changes from one batch to another. The temperature of the extracting water is low to limit the excessive dissolution of the active ingredient.
    [63] The suspension of the obtained cores is mixed for a few minutes and then filtered under an inert atmosphere. Working in an inert atmosphere can limit the risk of contamination of the product.
    [64] It is advantageous that the centrosome obtainable according to the method described above is lyophilized.
    [65] 10 ml of sterile water is added to 2 to 5 g of the core powder (filter cake). The mixture is frozen at −40 ° C. and then introduced into the lyophilizer. Continue lyophilization for 18 hours. After the end of the operation, the secondary drying temperature should be kept below 10 ° C.
    [66] Cores should be stored at + 4 ° C even when dry.
    [67] The present invention also provides a sterile solution containing 1 to 1.5% m / v of viscosity modifier, 0.5 to 1.5% m / v of surfactant and 3.5 to 4.5% m / v of isotonic agent, and releasing anticancer agent and coated with a polymer. It relates to a suspension consisting of 200 to 300 mg / sterile solution, preferably 230 to 270 mg / ml biodegradable core obtained optionally according to the method.
    [68] The core is preferably composed of 15 to 35% by weight of the anticancer agent and 65 to 85% by weight of the polymer.
    [69] The polymer is preferably poly (d, l-lactic-co-glycolic acid) containing the same amount of lactic acid and glycolic acid.
    [70] The sterile solution preferably contains 1.25% w / v sodium carboxymethylcellulose, 1% w / v polysorbate 80 and 4% w / v mannitol.
    [71] The invention is illustrated without limitation by the following examples.
    [72] Example 1:
    [73] Emulsification of solvent technology according to a modification of the method described in Boisdron-Celle M, Menei P and Benoit JP (Preparation of biodegradable 5-fluorouracil-loaded microspheres, J Pharm Pharmacol, 47, 108-114, 1995). Extraction is used to prepare the centrosomes.
    [74] 5-FU Crushing:
    [75] Crush on planetary ball mills such as 7 (Fritsch). 8.5 g of 5-FU are introduced into a beaker containing seven beads. Grinding is continued at speed 7 for 10 minutes. The powder is recovered under a laminar flow hood. The obtained crystals are 15 to 50 mu m in size and classified into two parts: a particulate portion (less than 1 mu m in diameter) and a granulated portion (30 mu m or more).
    [76] Dispersion of 5-FU in Organic Solvents:
    [77] The milled 5-FU was dispersed in 45 ml of dichloromethane under stirring using a homogenizer such as an Ultra-Turrax machine at 13,500 rpm in a round bottom tube for 3 minutes.
    [78] Preparation of the organic phase:
    [79] The 5-FU dispersion is transferred to a 150 ml jacketed cooling reactor. PLAGA is added to bring the PLAGA / dichloromethane ratio to 11%. The organic phase is stirred using a paddle at 450 rpm for 4 hours at 20 ° C., followed by 15 minutes at 2 ° C. The temperature of the reactor is kept constant within 0.1 ° C with the aid of a cryostat.
    [80] Preparation of Emulsions:
    [81] 1500 ml of an autoclaved 10% PVA aqueous solution was prepared and maintained at 2 ° C. in a 6 L jacketed cooling reactor. The organic phase is then transferred to this reactor by opening the base valve of the first reactor. The organic phase is poured into the aqueous phase over 5-10 s. The water / organic volume ratio is 100/3.
    [82] The emulsion is stirred for 4 minutes 45 seconds.
    [83] extraction:
    [84] Once the emulsion is ready, 4.5 liters of extracting water is poured into the emulsion at 4 ° C. in an emulsion / water volume ratio of 1/3. Continue extraction for 2 minutes.
    [85] percolation:
    [86] The entire contents of the second reactor are transferred through the base to a stainless tank and then placed under nitrogen pressure. The suspension is filtered through a filter having a pore diameter of 3 μm.
    [87] After passing the entire suspension through the filter, the filter cake is washed twice with 3 liters of sterile water.
    [88] The encapsulation degree of the anticancer agent in the obtained centrosome is 20%. After sieving, desorption of dichloromethane is done in an oven for 48 hours. The core is then packaged and sterilized with a γ radiation of 19 kGy. After sterilization the degree of encapsulation is further monitored. The residual trace amount analysis of the solvent is performed. It is advantageous for a dichloromethane residual level of 0.5% to be detected. The sterility and in vitro release kinetics of the obtained centrosomes are monitored.
    [89] The obtained centrosome has an active ingredient content of 23 ± 3.5%.
    [90] Various batches were prepared according to the protocol described above, and the average particle diameter was calculated to be 48 ± 20 µm corresponding to 46 ± 6 µm (average of the average value of the batch produced) for all populations of batches.
    [91] The obtained core had an active ingredient content of 23 ± 3.5% and an average particle diameter of 48 ± 20 µm.
    [92] Example 2:
    [93] Cores are prepared using the emulsification-extraction of the solvent technique of Example 1.
    [94] Crushing of 5-FU:
    [95] Follow the same procedure as in Example 1 to grind 4 g of 5-FU.
    [96] The obtained crystals are 15-50 탆 in size and are divided into two parts: a particulate portion (less than 1 탆 diameter) and a granulated portion (30 탆 or more).
    [97] Dispersion of 5-FU in Organic Solvents:
    [98] The milled 5-FU was dispersed in 40 ml of dichloromethane under stirring using a homogenizer such as an Ultra-Turax machine for 3.5 minutes at 13,500 rpm in a round bottom tube.
    [99] The organic phase and emulsion are prepared as in Example 1.
    [100] Extraction and filtration are performed as in Example 1.
    [101] Characteristics of the obtained centrosome:
    [102] 5-FU Content: 22%
    [103] Size: 46 ± 7㎛
    [104] Burst effect after 24 hours of radiation sterilization at 19 kGy: 40 ± 4%
    [105] Example 3:
    [106] A Phase I / II open pilot clinical study is conducted using the 50:50 PLAGA / 5-FU centrosome of Example 1.
    [107] The obtained core was subjected to sodium carboxymethylcellulose (Cooper) 1 to 1.5% w / v, polysorbate 80 1% w / v, mannitol 4% w / v, and a sufficient amount of injectable preparation water to obtain a total volume of 3 ml. It is immediately suspended in a solution containing.
    [108] The solution is previously sterilized by autoclave at 121 ° C. for 20 minutes and then radiation sterilized by gamma radiation in a dose of 5 kGy to 25 kGy, preferably 19 kGy.
    [109] The preparation of this suspension is precise because bubble formation should be avoided. Suspension is injected immediately after preparation because the core tends to settle on the dice and clog it.
    [110] The centrifugal suspension is visually implanted after glioma resection and implanted into the wall of the surgical site at a depth of 2-3 cm per cm 2 at a rate of 100 μl per injection.
    [111] 1 ml dice and 18 ga (1.3 × 45 mm) catheter ( Injection is performed to remove the metal mandrel to retain only an unbeveled foam-tipped plastic catheter for injecting the suspension into cerebral tissue. As many 1 ml of dice as needed are used. Injection of the suspension with the inclined needle introduces a hematoma risk and backflow of the centrosome. The catheter should be small enough in diameter to avoid traumatic cerebral tissue and large enough to prevent blockage by centro suspension.
    [112] Injection should be done very quietly and the catheter should remain in place for several minutes before being removed, to avoid backflow of the centrosome. Resorbable hemostatic compression bandage ( A 1 cm 2 fragment of) is used at the injection site.
    [113] Patients included in the study are 18 to 68 years old, have no history of tumor, have a Karnofsky index of 60 or higher, have clinical history and imaging to induce supporting gliomas, Diagnosis of gliomas is confirmed by their medical history investigation (performed according to WHO criteria: necrosis, vascular proliferation, nuclear polymorphism and mitosis activity) during these surgical procedures.
    [114] Patient exclusion criteria include: metabolic deficiency, pregnancy, and other previous cancer pathologies.
    [115] Three groups were initially envisioned to study the effects of increasing doses of 5-fluorouracil in chronological order of 70, 132 and 264 mg and started treatment of the following groups after observing treatment resistance by the tested group. It was.
    [116] Due to the occurrence of Grade II nervous system toxicity in patients receiving 5-FU 132 mg, the treatment phase expansion is stopped and an equal amount of 132 mg is administered to subsequent patients, according to the rules for discontinuing the protocol.
    [117] Conventional external radiation therapy (focus on tumor volume assessed for preoperative MRI at an energy of 10 μs) is initiated 2-7 days after surgery. A total dose of 60 Gy of 33 fractions of 1.8 Gy is applied at a rate of 5 fractions over 6.5 weeks. Volumes examined include preoperative tumors with margins of at least 2 cm in all directions.
    [118] Patients are clinically monitored by radiation: a 72 hour scan and clinical evaluation to confirm complete resection with the naked eye is done for D10, D20 and D30. MRI is performed on D10 and D30. Finally, analysis of 5-FU in blood and CSF is done for 72 hours D10, D20 and D30. Toxicity (neurologic, hematological, mucosal and cardiological) is assessed as a rating against criteria derived from the rating of WHO. After one month has elapsed, the patient is clinically monitored every two months and MRI is performed every three months.
    [119] Mild postoperative anemia, leukocytosis and mild lymphocytosis were observed in all patients.
    [120] Pharmacological studies have confirmed the sustained release of 5-FU in cerebrospinal fluid (CSF) at least 30 days, and transient passage of molecules into the body circulation at low levels. One month after transplantation, significant concentrations of 5-FU are present in CSF.
    [121] The release profile of 5-FU in CSF shows peaks at 10 and 20 days for doses of 70 and 132 mg, respectively. Levels of 5-FU in plasma were not detected on day 10 in half of the patients.
    [122] Systemic resistance is excellent in all treated patients. No change in chemistry or cell fidelity of CSF was observed. The dose could not continue to increase due to the appearance of cerebral edema during radiation therapy in patients treated with 132 mg.
    [123] Thus, eight patients were included in the study, including four men and four women with an average age of 48.5 years and a Karnovsky index of 90 or higher. Three first groups were administered 70 mg and five second groups were administered 132 mg.
    [124] Preliminary results on survival could not be explained statistically due to the small number of patients. However, it was very encouraging. At the final evaluation, in the first group treated with 70 mg, three patients died at 61, 114 and 125 weeks. Patients who died at week 114 should pay particular attention to the death of glioblastoma lung metastases. In the second group treated with 132 mg, three patients died at 31, 59 and 82 weeks, and two still showed remission at 159 and 172 weeks on the draft date of these preliminary results.
    [125] The median survival of patients is 98 weeks (this is 50.6 weeks in the literature for patients meeting the same criteria) [Devaux BC, O'Fallon JR, Kelly PJ, Resection, biopsy and survival in malignant glial neoplasms, J Neurosurg, 78: 767-775, 1993]. Five of the eight patients, or 62%, survived 18 months, while the literature found that the 18-month survival rate was 20% for patients who met the comprehensive criteria of this study. [Devaux BC, O'Fallon JR , Kelly PJ, Resection, biopsy and survival in malignant glial neoplasms, J Neurosurg, 78: 767-775, 1993].
    权利要求:
    Claims (37)
    [1" claim-type="Currently amended] In the use of biodegradable centrosomes, which release radiation-sensitive anticancer agents for the manufacture of drugs intended for use alone, or over time, for the treatment of gliomas, and which are to be implanted at the surgical site after resection of the glioma. The central body containing the anticancer agent may be used in the parenchymal space for the purpose of delaying the release of the anticancer agent and achieving the survival time of the treated patient for at least about 90 weeks, preferably about 130 weeks, more preferably 160 weeks. Use of a biodegradable centroid, characterized by coating with a polymer that maintains a therapeutically effective concentration over time.
    [2" claim-type="Currently amended] The use of biodegradable centroids according to claim 1, wherein the anticancer agent exhibits hydrophilicity and / or does not interfere with the blood brain barrier.
    [3" claim-type="Currently amended] Use according to claim 1 or 2, wherein the anticancer agent does not exhibit central neurotoxicity.
    [4" claim-type="Currently amended] The method according to any one of claims 1 to 3, wherein the anticancer agent consists of a radiation sensitive anticancer compound or a mixture of anticancer compound containing at least one radiation sensitive anticancer compound, wherein the anticancer compound (s) is for example 5- Platinum, such as fluorouracil (5-FU), carboplatin and cisplatin, taxanes such as docetaxel and paclitaxel, togitabine, VP16, mitomycin, indosuridine, irinotecan, topotecan and camptothecins such as camptothecin Merase 1 inhibitor, characterized in that it is selected from nitrosoureas such as BCNU, ACNU or MCNU, methotrexate, bleomycin, adriamycin, cytosan and vincristine, immunomodulatory cytokines such as IL2, IL6, IL12 and IL13 and interferon Use of biodegradable centroids.
    [5" claim-type="Currently amended] 5. The use of biodegradable core according to claim 4, wherein the anticancer agent is 5-fluorouracil.
    [6" claim-type="Currently amended] 6. The use of biodegradable centrosome according to claim 5, wherein the concentration of 5-FU in cerebrospinal fluid reflecting the concentration in the parenchyma is 3-20 ng / ml.
    [7" claim-type="Currently amended] Use of a biodegradable core according to any one of claims 1 to 6, characterized in that a neuroprotective compound selected from peptide growth factors such as NGF or BDNF is added to the anticancer agent.
    [8" claim-type="Currently amended] 8. The polymer according to claim 1, wherein the polymer coating the centroid that slows the release of the anticancer agent maintains a therapeutically effective concentration in the parenchymal space for at least 3 weeks, preferably at least 4 weeks. Use of biodegradable centrosomes.
    [9" claim-type="Currently amended] The polymer of claim 1, wherein the polymer is ethylcellulose, polystyrene, poly (ε-caprolactone), poly (d, l-lactic acid) and poly (d, l-lactic acid-co-glycolic acid The concentration of the biodegradable centrosome, characterized in that is selected from.
    [10" claim-type="Currently amended] Use according to any of the preceding claims, wherein the polymer coating the core is poly (d, l-lactic-co-glycolic acid).
    [11" claim-type="Currently amended] Use according to claim 10, wherein the polymer coating the core is poly (d, l-lactic-co-glycolic acid) containing the same amount of lactic acid and glycolic acid.
    [12" claim-type="Currently amended] 12. Use of a biodegradable core according to any one of the preceding claims, characterized in that the core has an average diameter of 48 ± 20 μm, preferably 46 ± 7 μm.
    [13" claim-type="Currently amended] 13. Use of a biodegradable core according to any one of claims 1 to 12, wherein the core contains 15 to 35% by weight of the anticancer agent and 65 to 85% by weight of the polymer.
    [14" claim-type="Currently amended] 14. Use of a biodegradable core according to claim 5 or 13, characterized in that the centroid contains 5-FU 19-27%, preferably 5-FU 20%.
    [15" claim-type="Currently amended] The method according to any one of claims 1 to 14, wherein the centrosome is suspended in sterile solution, and the obtained suspension is formed to be injected into the wall of the surgical site after resection of the glioma, wherein the sterile solution is 1 to 1.5% of a viscosity modifier. m / v, 0.5-1.5% m / v surfactant and 3.5-4.5% m / v isotonic agent.
    [16" claim-type="Currently amended] Use according to claim 15, wherein the sterile solution contains 1.25% w / v sodium carboxymethylcellulose, 1% w / v polysorbate 80 and 4% w / v mannitol.
    [17" claim-type="Currently amended] Use according to claim 15 or 16, characterized in that the suspension of the centrosome is injected into the wall of the surgical site at a depth of at least 2 cm, preferably 2 to 3 cm per cm 2.
    [18" claim-type="Currently amended] 6. Use of biodegradable centroids according to claim 5, characterized in that the total dose of 5-FU injected is 50-200 mg, preferably 130 mg.
    [19" claim-type="Currently amended] The method of claim 1, wherein the radiation therapy focuses on the tumor volume, the irradiated volume comprises a preoperative tumor having a margin of at least 2 cm in all directions, with a total dose of 50-60 Gy. Use of a biodegradable core, characterized in that used.
    [20" claim-type="Currently amended] 20. Use of the biodegradable centrosome according to any one of claims 1 to 19, characterized in that radiation therapy is preferably initiated 2 to 7 days after surgery.
    [21" claim-type="Currently amended] 21. The use of any one of claims 1 to 20, wherein the radiation therapy is performed at a total dose of 60 Gy for about 6 weeks.
    [22" claim-type="Currently amended] 22. Use of a biodegradable centrosome according to any one of claims 15 to 17 or 19 to 21, wherein the suspension of the centrosome is injected by one or more stereopositions in case of tumor recurrence.
    [23" claim-type="Currently amended] Dispersing the polymer and the anticancer agent in the organic solvent, mixing the organic phase obtained to obtain an emulsion with an aqueous phase, extracting the organic solvent by adding water, and then filtering the suspension of the obtained core, followed by polymerisation by emulsification-extraction. A method for producing a biodegradable centrosome containing an anticancer agent coated with an anticancer agent, wherein the anticancer agent is dispersed in an organic solvent under vigorous stirring before the polymer is added.
    [24" claim-type="Currently amended] 24. The method of claim 23, wherein the organic solvent is dichloromethane.
    [25" claim-type="Currently amended] The process for producing a biodegradable core according to claim 23 or 24, wherein the water phase and the organic phase are at the same temperature, preferably 2 ° C, when mixing.
    [26" claim-type="Currently amended] 26. The method of any of claims 23-25, wherein the organic phase contains 11% polymer.
    [27" claim-type="Currently amended] 27. The method of any of claims 23 to 26, wherein the water / organic phase ratio is 100/3.
    [28" claim-type="Currently amended] 28. The method of any of claims 23 to 27, wherein the emulsion consisting of an aqueous phase and an organic phase is mixed for at least 3 minutes.
    [29" claim-type="Currently amended] 29. The method according to any one of claims 23 to 28, wherein the water required to extract the organic solvent is added in a proportion of 1/3 the volume ratio of emulsion / water.
    [30" claim-type="Currently amended] 30. The method according to any one of claims 23 to 29, wherein the temperature of the water required to extract the organic solvent is 4 ° C.
    [31" claim-type="Currently amended] 31. The method according to any one of claims 23 to 30, wherein the anticancer agent is ground before being dispersed in an organic solvent such that the anticancer agent has a crystal size of 15 to 50 µm.
    [32" claim-type="Currently amended] 32. A process according to any one of claims 23 to 31, wherein after the extracting water is added, the suspension obtained is filtered under an inert atmosphere.
    [33" claim-type="Currently amended] 33. The method of any of claims 23 to 32, wherein the core is lyophilized.
    [34" claim-type="Currently amended] Aseptic solution containing 1 to 1.5% m / v of viscosity modifier, 0.5 to 1.5% m / v of surfactant and 3.5 to 4.5% m / v of isotonic agent, and biodegradable core 200 to 300 which release anticancer agent and are coated with polymer A suspension, which is composed of mg / ml of sterile solution.
    [35" claim-type="Currently amended] 35. The suspension of claim 34 wherein the core comprises 15-35 weight percent anticancer agent and 65-85 weight percent polymer.
    [36" claim-type="Currently amended] 36. A suspension according to claim 34 or 35, wherein the polymer is preferably poly (d, l-lactic-co-glycolic acid) containing the same amount of lactic acid and glycolic acid.
    [37" claim-type="Currently amended] 37. The method of any one of claims 34 to 36. A sterile solution containing 1.25% w / v sodium carboxymethylcellulose, 1% w / v polysorbate 80 and 4% w / v mannitol.
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    同族专利:
    公开号 | 公开日
    BR0010648A|2002-02-19|
    US6803052B2|2004-10-12|
    JP5230045B2|2013-07-10|
    ZA200109415B|2003-01-29|
    US20020051749A1|2002-05-02|
    FR2793684B1|2001-08-10|
    CA2388656C|2011-04-26|
    JP2002544219A|2002-12-24|
    SI1053746T1|2003-04-30|
    HU0201224A2|2002-11-28|
    IL146467A|2006-06-11|
    DK1053746T3|2003-03-31|
    IL146467D0|2002-07-25|
    CZ20014137A3|2002-06-12|
    ES2185544T3|2003-05-01|
    AU4764100A|2000-12-05|
    DE60000842D1|2003-01-09|
    NO20015501L|2002-01-09|
    FR2793684A1|2000-11-24|
    TWI229608B|2005-03-21|
    AT228353T|2002-12-15|
    CN1210020C|2005-07-13|
    US20030175356A1|2003-09-18|
    EP1053746A1|2000-11-22|
    BG106106A|2002-05-31|
    CN1356892A|2002-07-03|
    US7041241B2|2006-05-09|
    NO331686B1|2012-02-20|
    EP1228755A1|2002-08-07|
    SK16342001A3|2002-08-06|
    AR024008A1|2002-09-04|
    HK1047044B|2005-12-23|
    NZ515515A|2004-02-27|
    BG64936B1|2006-10-31|
    PT1053746E|2003-04-30|
    EA004943B1|2004-10-28|
    NO20015501D0|2001-11-09|
    MXPA01011919A|2002-05-06|
    WO2000069413A1|2000-11-23|
    PL352372A1|2003-08-25|
    DE60000842T2|2003-08-28|
    CA2388656A1|2000-11-23|
    PL201614B1|2009-04-30|
    EP1053746B1|2002-11-27|
    KR100649948B1|2006-11-24|
    AU775320B2|2004-07-29|
    HK1047044A1|2005-12-23|
    HU0201224A3|2004-05-28|
    EA200101206A1|2002-04-25|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    法律状态:
    1999-05-17|Priority to FR99/06,207
    1999-05-17|Priority to FR9906207A
    2000-05-17|Application filed by 제라르드 레뒤끄, 라보하또와훼 데 쁘호뒤뜨 에띠끄 에띠빠흠
    2002-01-04|Publication of KR20020000178A
    2006-11-24|Application granted
    2006-11-24|Publication of KR100649948B1
    优先权:
    申请号 | 申请日 | 专利标题
    FR99/06,207|1999-05-17|
    FR9906207A|FR2793684B1|1999-05-17|1999-05-17|Use of biodegradable microspheres releasing anti-cancer agent for the treatment of glioblastoma, process for preparing such microspheres and suspension containing them|
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