阅读说明：本技术 一种具有抗肿瘤效果的二氧化钛化合物 (Titanium dioxide compound with anti-tumor effect ) 是由 冒志磊 王淮燕 周军 张克良 秦志强 陈科锦 于 2021-09-08 设计创作，主要内容包括：本发明公开了一种具有抗肿瘤效果的二氧化钛化合物,属于生物制药技术领域,所述二氧化钛化合物的合成原料包括谷氨酰胺、钛酸丁酯以及铜离子,其中,所述谷氨酰胺的反应浓度为1mg/ml,所述钛酸丁酯的反应浓度为0.5mg/ml,所述谷氨酰胺和钛酸丁酯的反应浓度配比为2:1,所述铜离子的反应浓度为0.3μg/ml,所述谷氨酰胺和钛酸丁酯二者进行反应用于合成纳米二氧化钛,所述铜离子用于向纳米二氧化钛的晶格中进行掺杂对纳米二氧化钛进行修饰。通过进行扰乱相分离,抑制肿瘤细胞侵袭和转移能力的机制,为后续制药设计提供新思路,使得药物合成成本可控,能大幅度降低药物研发的费用,从而降低肿瘤患者家庭的巨额支出。(The invention discloses a titanium dioxide compound with an anti-tumor effect, which belongs to the technical field of biological pharmacy, wherein the synthetic raw materials of the titanium dioxide compound comprise glutamine, butyl titanate and copper ions, the reaction concentration of the glutamine is 1mg/ml, the reaction concentration of the butyl titanate is 0.5mg/ml, the reaction concentration ratio of the glutamine to the butyl titanate is 2:1, the reaction concentration of the copper ions is 0.3 mu g/ml, the glutamine and the butyl titanate are reacted to synthesize nano titanium dioxide, and the copper ions are used for doping crystal lattices of the nano titanium dioxide to modify the nano titanium dioxide. By means of a mechanism of disturbing phase separation and inhibiting tumor cell invasion and metastasis capacity, a new idea is provided for subsequent pharmaceutical design, so that the drug synthesis cost is controllable, the drug research and development cost can be greatly reduced, and the huge expenditure of families of tumor patients is reduced.)

1. The titanium dioxide compound with the anti-tumor effect is characterized in that raw materials for synthesizing the titanium dioxide compound comprise glutamine, butyl titanate and copper ions, wherein the reaction concentration of the glutamine is 1mg/ml, the reaction concentration of the butyl titanate is 0.5mg/ml, the reaction concentration ratio of the glutamine to the butyl titanate is 2:1, the reaction concentration of the copper ions is 0.3 mu g/ml, the glutamine and the butyl titanate are reacted to synthesize nano titanium dioxide, the copper ions are used for doping crystal lattices of the nano titanium dioxide to modify the nano titanium dioxide, and the formed titanium dioxide compound is glutamine-nano titanium dioxide;

the method comprises the steps of centrifugally precipitating nano-particles of glutamine-nano titanium dioxide modified by copper ions by using an ultra-high speed centrifuge, washing to remove un-synthesized raw materials, measuring the concentration of titanium elements by using ICP-MS (inductively coupled plasma-mass spectrometry), determining the compound yield of the glutamine-nano titanium dioxide, measuring the original particle size and the hydrated particle size of the glutamine-nano titanium dioxide compound by using a transmission electron microscope and a Malvern laser particle sizer, judging the dispersion degree and the uniformity of the particle size of the titanium dioxide compound, and measuring the concentration of copper ion impurities in the titanium dioxide compound by using an X-ray energy spectrometer.

2. The titanium dioxide compound with an antitumor effect according to claim 1, wherein the copper ion doping concentration is optimized to improve the synthesis efficiency of the titanium dioxide compound and the purity and yield of the product, under the condition of the previous synthesis, the copper ion concentration is 0.3 μ g/ml, the reaction concentrations of glutamine and butyl titanate are maintained unchanged, and the copper ion concentrations are set to be 0.5 μ g/ml, 1 μ g/ml, 2 μ g/ml, 5 μ g/ml and 10 μ g/ml, respectively.

3. The titanium dioxide compound having an antitumor effect according to claim 1, wherein a ratio of a reaction concentration of glutamine to a reaction concentration of butyl titanate is 1:1, the reaction concentration of glutamine is 1mg/ml, and the reaction concentration of butyl titanate is 1mg/ml, when the titanium dioxide compound is synthesized as a raw material.

4. The titanium dioxide compound having an antitumor effect according to claim 1, wherein a ratio of a reaction concentration of glutamine to a reaction concentration of butyl titanate is 1:2, the reaction concentration of glutamine is 0.5mg/ml, and the reaction concentration of butyl titanate is 1mg/ml, when the titanium dioxide compound is synthesized as a raw material.

5. The titanium dioxide compound having an antitumor effect according to claim 1, wherein a ratio of a reaction concentration of glutamine to a reaction concentration of butyl titanate is 2:1, the reaction concentration of glutamine is 0.7mg/ml, and the reaction concentration of butyl titanate is 0.35mg/ml in raw material synthesis.

6. The titanium dioxide compound having an antitumor effect according to claim 1, wherein the acute toxicity and accumulation of the titanium dioxide compound are analyzed, and the lethal dose of glutamine-nano titanium dioxide is estimated by half the lethal dose of nano titanium dioxide.

7. The titanium dioxide compound with anti-tumor effect according to claim 1, wherein the pharmacokinetics and the ability to reach tumor tissue of the titanium dioxide compound are analyzed, the maximum dose tail vein injection with a lethality rate of 0 is selected for pharmacokinetic study, blood and tissue samples within 48 hours are reserved at intervals of one hour for subsequent drug content measurement, a change curve is drawn according to the change of the titanium dioxide content in the blood and tissue, the apparent distribution volume is calculated, and the distribution of the drug and the degree of binding with the tissue are judged.

8. The titanium dioxide compound with the anti-tumor effect according to claim 1, wherein the effect of the titanium dioxide compound on inhibiting tumor formation and metastasis is analyzed, lentivirus transfection is constructed to construct tumor cells with stable and high luciferase expression, and the monitoring and evaluation of the growth and metastasis of the tumor cells are realized by using a visible light living body imaging technology.

9. The titanium dioxide compound with anti-tumor effect according to claim 1, wherein the mechanism of the titanium dioxide compound disturbing tumor cell phase separation to inhibit metastasis is analyzed to construct an in vitro microtubule polymerization model, and the interference effect of glutamine-nano titanium dioxide on the microtubule scaffold phase separation is determined.

Technical Field

The invention belongs to the technical field of biological pharmacy, and particularly relates to a titanium dioxide compound with an anti-tumor effect.

Background

Cancer cells have the property of "glutamine addiction" in that they enrich glutamine in large amounts for their rapid growth, and therefore, although glutamine is the most abundant amino acid in the body, a deficiency of glutamine often occurs in tumor-bearing organisms, and thus the supplemented glutamine is rapidly transported to tumor tissues. Although in vitro cell research shows that glutamine can promote tumor cell proliferation, in vivo research results contradict with the results, and the parenteral or enteral nutrition support containing glutamine does not obviously promote tumor growth of tumor-bearing host, probably because glutamine can promote positive nitrogen balance of host and improve body immunity, so that it is safe to supplement glutamine to tumor-bearing body, and the research related to glutamine metabolism is gradually becoming a new target and breakthrough for cancer treatment.

To this end, we propose a titanium dioxide compound having an antitumor effect.

Disclosure of Invention

In view of the deficiencies of the prior art, the present invention provides a titanium dioxide compound having an anti-tumor effect to solve the problems set forth in the background art described above.

In order to achieve the purpose, the invention provides the following technical scheme: a titanium dioxide compound with an anti-tumor effect is prepared from raw materials comprising glutamine, butyl titanate and copper ions, wherein the reaction concentration of the glutamine is 1mg/ml, the reaction concentration of the butyl titanate is 0.5mg/ml, the reaction concentration ratio of the glutamine to the butyl titanate is 2:1, the reaction concentration of the copper ions is 0.3 mu g/ml, the glutamine and the butyl titanate are reacted to synthesize nano titanium dioxide, the copper ions are used for doping crystal lattices of the nano titanium dioxide to modify the nano titanium dioxide, and the formed titanium dioxide compound is glutamine-nano titanium dioxide;

the method comprises the steps of centrifugally precipitating nano-particles of glutamine-nano titanium dioxide modified by copper ions by using an ultra-high speed centrifuge, washing to remove un-synthesized raw materials, measuring the concentration of titanium elements by using ICP-MS (inductively coupled plasma-mass spectrometry), determining the compound yield of the glutamine-nano titanium dioxide, measuring the original particle size and the hydrated particle size of the glutamine-nano titanium dioxide compound by using a transmission electron microscope and a Malvern laser particle sizer, judging the dispersion degree and the uniformity of the particle size of the titanium dioxide compound, and measuring the concentration of copper ion impurities in the titanium dioxide compound by using an X-ray energy spectrometer.

Further optimizing the technical scheme, when the copper ions are modified, the doping concentration of the copper ions is optimized, the synthesis efficiency of the titanium dioxide compound and the purity and yield of the product are improved, under the condition of early synthesis, the concentration of the copper ions is 0.3 mu g/ml, the reaction concentrations of glutamine and butyl titanate are kept unchanged, and the concentrations of the copper ions are respectively set to be 0.5 mu g/ml, 1 mu g/ml, 2 mu g/ml, 5 mu g/ml and 10 mu g/ml.

Further optimizing the technical scheme, when the titanium dioxide compound is synthesized from raw materials, the reaction concentration ratio of glutamine to butyl titanate is 1:1, the reaction concentration of glutamine is 1mg/ml, and the reaction concentration of butyl titanate is 1 mg/ml.

Further optimizing the technical scheme, when the titanium dioxide compound is synthesized from raw materials, the reaction concentration ratio of glutamine to butyl titanate is 1:2, the reaction concentration of glutamine is 0.5mg/ml, and the reaction concentration of butyl titanate is 1 mg/ml.

Further optimizing the technical scheme, when the titanium dioxide compound is synthesized from raw materials, the reaction concentration ratio of glutamine to butyl titanate is 2:1, the reaction concentration of glutamine is 0.7mg/ml, and the reaction concentration of butyl titanate is 0.35 mg/ml.

Further optimizing the technical scheme, analyzing the acute toxicity and the accumulation of the titanium dioxide compound, and estimating the lethal dose of the glutamine-nano titanium dioxide by half lethal dose of the nano titanium dioxide.

Further optimizing the technical scheme, analyzing the pharmacokinetics and the ability of the titanium dioxide compound to reach tumor tissues, selecting the maximum dose tail vein injection with the lethality rate of 0 to carry out pharmacokinetic research, reserving blood and tissue samples within 48 hours at intervals of one hour to carry out subsequent drug content determination, drawing a change curve according to the change of the titanium dioxide content in the blood and the tissue, calculating apparent distribution volume, and judging the distribution of the drug and the combination degree of the drug and the tissue.

The technical scheme is further optimized, the effect of the titanium dioxide compound on inhibiting tumor formation and metastasis is analyzed, slow virus transfection is constructed to construct tumor cells of stable and high-expression luciferase, and the monitoring and evaluation on the growth and metastasis of the tumor cells are realized by utilizing a visible light living body imaging technology.

Further optimizing the technical scheme, analyzing the mechanism of the titanium dioxide compound disturbing the tumor cell 'phase separation' to inhibit the transfer, constructing an in vitro microtubule polymerization model, and determining the interference effect of the glutamine-nanometer titanium dioxide on the microtubule skeleton phase separation of the cells.

Compared with the prior art, the invention provides a titanium dioxide compound with an anti-tumor effect, which has the following beneficial effects:

the titanium dioxide compound with the anti-tumor effect provides a new idea for subsequent pharmaceutical design by a mechanism of disturbing phase separation and inhibiting tumor cell invasion and metastasis capacity, converts the titanium dioxide compound into a medicine, enables the medicine synthesis cost to be controllable, can greatly reduce the research and development cost of the medicine, thereby reducing the huge expenditure of families of tumor patients, can effectively inhibit tumor metastasis in vivo, can effectively prolong the survival time of patients with advanced tumor, and has important social significance for the tumor patients and the families thereof.

Drawings

FIG. 1 is a schematic diagram of a process for synthesizing a titanium dioxide compound having an anti-tumor effect according to the present invention;

FIG. 2 is a graph showing the intermediate state and the completed state of a titanium dioxide compound having an antitumor effect according to the present invention.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

The first embodiment is as follows:

referring to fig. 1, a titanium dioxide compound with an anti-tumor effect is prepared from raw materials including glutamine, butyl titanate and copper ions, wherein the reaction concentration of glutamine is 1mg/ml, the reaction concentration of butyl titanate is 0.5mg/ml, the reaction concentration ratio of glutamine to butyl titanate is 2:1, the reaction concentration of copper ions is 0.3 μ g/ml, the glutamine and butyl titanate are reacted to synthesize nano titanium dioxide, the copper ions are used for doping crystal lattices of the nano titanium dioxide to modify the nano titanium dioxide, and the formed titanium dioxide compound is glutamine-nano titanium dioxide;

as shown in FIG. 2, photograph A in FIG. 2 shows that the copper ion-modified glutamine-nano titanium dioxide in the intermediate state is in the shape of a rod, and photograph B in FIG. 2 shows that the copper ion-modified glutamine-nano titanium dioxide in the complete state is in the shape of a sphere.

The method comprises the steps of centrifugally precipitating nano-particles of glutamine-nano titanium dioxide modified by copper ions by using an ultra-high speed centrifuge, washing to remove un-synthesized raw materials, measuring the concentration of titanium elements by using ICP-MS (inductively coupled plasma-mass spectrometry), determining the compound yield of the glutamine-nano titanium dioxide, measuring the original particle size and the hydrated particle size of the glutamine-nano titanium dioxide compound by using a transmission electron microscope and a Malvern laser particle sizer, judging the dispersion degree and the uniformity of the particle size of the titanium dioxide compound, and measuring the concentration of copper ion impurities in the titanium dioxide compound by using an X-ray energy spectrometer.

Specifically, when the copper ions are modified, the doping concentration of the copper ions is optimized, the synthesis efficiency of the titanium dioxide compound and the purity and yield of the product are improved, under the condition of early synthesis, the concentration of the copper ions is 0.3 mu g/ml, the reaction concentrations of glutamine and butyl titanate are kept unchanged, and the concentrations of the copper ions are respectively set to be 0.5 mu g/ml, 1 mu g/ml, 2 mu g/ml, 5 mu g/ml and 10 mu g/ml.

Specifically, when the titanium dioxide compound is synthesized from raw materials, the reaction concentration ratio of glutamine to butyl titanate is 1:1, the reaction concentration of glutamine is 1mg/ml, and the reaction concentration of butyl titanate is 1 mg/ml.

Specifically, when the titanium dioxide compound is synthesized from raw materials, the reaction concentration ratio of glutamine to butyl titanate is 1:2, the reaction concentration of glutamine is 0.5mg/ml, and the reaction concentration of butyl titanate is 1 mg/ml.

Specifically, when the titanium dioxide compound is synthesized from raw materials, the reaction concentration ratio of glutamine to butyl titanate is 2:1, the reaction concentration of glutamine is 0.7mg/ml, and the reaction concentration of butyl titanate is 0.35 mg/ml.

Specifically, the acute toxicity and accumulation of the titanium dioxide compound are analyzed, and the lethal dose of glutamine-nano titanium dioxide is estimated by half of the lethal dose of nano titanium dioxide.

Specifically, the pharmacokinetics and the ability of the titanium dioxide compound to reach tumor tissues are analyzed, the maximum dose tail vein injection with the lethality rate of 0 is selected for pharmacokinetic research, blood and tissue samples within 48 hours are reserved at intervals of one hour for subsequent drug content determination, a change curve is drawn according to the change of the titanium dioxide content in the blood and the tissue, the apparent distribution volume is calculated, and the distribution of the drug and the binding degree with the tissue are judged.

Specifically, the effect of the titanium dioxide compound on inhibiting tumor formation and metastasis is analyzed, a lentivirus is constructed for transfection, tumor cells with stable and high-expression luciferase are constructed, and the monitoring and evaluation on the growth and metastasis of the tumor cells are realized by utilizing a visible light living body imaging technology.

Specifically, the mechanism of the titanium dioxide compound for disturbing the 'phase separation' of tumor cells to inhibit metastasis is analyzed, an in vitro microtubule polymerization model is constructed, and the interference effect of glutamine-nano titanium dioxide on the microtubule skeleton phase separation of cells is determined.

Example two:

when the acute toxicity and the accumulation of the titanium dioxide compound having an antitumor effect were analyzed based on the titanium dioxide compound having an antitumor effect described in example one, the lethal dose of glutamine-nano titanium dioxide was estimated from the half lethal dose of nano titanium dioxide. After the concentration is measured by ICP-MS, different concentrations of the venom are prepared, and the maximum dose, the median lethal dose and the absolute lethal dose of the nude mice with the death rate of 0 are determined by tail vein injection administration. The LD50 of the glutamine-nano titanium dioxide and the 95 percent confidence interval thereof are calculated by utilizing the improved karber method. The poisoning symptoms of the mice, and macroscopic and pathological changes of the internal organs were observed and recorded. The dose was designed based on LD50 by a dose escalation continuous toxicant exposure method, and the accumulation coefficient was calculated to evaluate the accumulation property.

Example three:

based on the titanium dioxide compound with the anti-tumor effect, in the first embodiment, when the pharmacokinetics and the ability of the titanium dioxide compound to reach tumor tissues are analyzed, the maximum dose tail vein injection with the lethality rate of 0 is selected for pharmacokinetic research, blood and tissue samples within 48 hours are reserved at intervals of one hour for subsequent drug content measurement, a change curve is drawn according to the change of the titanium dioxide content in the blood and the tissue, the apparent distribution volume is calculated, and the distribution of the drug and the binding degree of the drug and the tissue are judged. The half-life, clearance rate and elimination kinetics of the glutamine-nanometer titanium dioxide are calculated according to the content of the titanium dioxide in the plasma. Culturing tumor cells in vitro, inoculating the tumor cells to the middle and rear parts of armpits of nude mice under the skin, selecting the nude mice with better tumor formation, injecting glutamine-nano titanium dioxide into tail veins, and detecting the distribution of the glutamine-nano titanium dioxide in tumor tissues by utilizing ICP-MS after 48 hours. Urine and feces of nude mice are collected by a metabolism cage, main excretion modes and excretion amounts of glutamine-nano titanium dioxide are judged, and accumulation conditions of glutamine-nano titanium dioxide in vivo are evaluated.

Example four:

based on the titanium dioxide compound with the anti-tumor effect, when the effect of the titanium dioxide compound on inhibiting tumor formation and metastasis is analyzed, lentivirus transfection is constructed to construct tumor cells with stable and high-expression luciferase, and the growth and metastasis of the tumor cells are monitored and evaluated by using a visible light living body imaging technology. The visible light living body imaging technology is utilized to observe the tumor forming capability of the glutamine-nanometer titanium dioxide on the treated tumor cells, and the angiogenesis condition in the tumor and the invasion condition to the basement membrane tissue are observed after pathological sections are cut. The visible light living body imaging technology is combined with a flow cytometer to evaluate the tumor growth, invasion and metastasis inhibition conditions of the glutamine-nanometer titanium dioxide on tumor-bearing nude mice. Injecting tumor cells into a nude mouse body through a tail vein to construct a tumor metastasis model, injecting glutamine-nano titanium dioxide, observing the formation condition of a tumor systemic metastasis focus of the nude mouse, and recording the size and the life cycle of the tumor. Constructing a primary tumor nude mouse transplantation tumor model, injecting glutamine-nano titanium dioxide, and observing the growth of the tumor and the invasion and metastasis conditions to the periphery.

Example five:

when analyzing a mechanism of disturbing tumor cells to inhibit metastasis by 'phase separation' based on the titanium dioxide compound with the anti-tumor effect described in the first embodiment, an in vitro microtubule polymerization model is constructed, microtubule-associated protein tau, glutamine-nano titanium dioxide, tau protein + glutamine-nano titanium dioxide are added respectively, the change of a polymerization curve is recorded, and the influence of tau protein and glutamine-nano titanium dioxide on microtubule polymerization is judged; and observing the solution in which tau protein and glutamine-nano titanium dioxide are added in different time periods in the polymerization process of the microtubule by using an optical microscope to observe the influence on the phase separation droplet-shaped structure. The method comprises the steps of constructing a tubulin and tau protein overexpression vector by using lentivirus, observing a dynamic process of the two which participates in microtubule phase separation in a living cell by using a laser confocal microscope, interfering the expression of the tubulin and the tau protein by using siRNA respectively, constructing IDR structural domain mutant protein participating in the phase separation, adding glutamine-nano titanium dioxide, observing the change of fluorescence under each condition by using the laser confocal microscope, and determining the interference effect of the glutamine-nano titanium dioxide on the microtubule skeleton phase separation of the cell.

The invention has the beneficial effects that: the titanium dioxide compound with the anti-tumor effect provides a new idea for subsequent pharmaceutical design by a mechanism of disturbing phase separation and inhibiting tumor cell invasion and metastasis capacity, converts the titanium dioxide compound into a medicine, enables the medicine synthesis cost to be controllable, can greatly reduce the research and development cost of the medicine, thereby reducing the huge expenditure of families of tumor patients, can effectively inhibit tumor metastasis in vivo, can effectively prolong the survival time of patients with advanced tumor, and has important social significance for the tumor patients and the families thereof.

In the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.

Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.