WO2021139697A1 - 一种化疗药物作用后的肿瘤细胞上清液的应用 - Google Patents
一种化疗药物作用后的肿瘤细胞上清液的应用 Download PDFInfo
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- A—HUMAN NECESSITIES
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- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/13—Tumour cells, irrespective of tissue of origin
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- the application belongs to the technical field of tumor cell supernatant, and specifically relates to an application of tumor cell supernatant after the action of a chemotherapeutic drug.
- Chemotherapy is the main treatment option for cancer, especially for patients who cannot get local treatment. Chemotherapy drugs can directly kill tumor cells through cytotoxicity. Therefore, although in clinical practice, some patients survive for a long time after pathological complete remission after chemotherapy, it is currently believed that chemotherapy drugs are not the only cause of cancer cell death. Perhaps the immune system may play an important role.
- the technical problem to be solved by this application is to provide an application of tumor cell supernatant after the action of chemotherapeutic drugs in response to the above-mentioned deficiencies of the prior art.
- the tumor cell supernatant after the action of the chemotherapeutic drugs can promote liver cancer cell differentiation and inhibit
- the transcription of liver tumor stem cell markers and inhibiting the proliferation of liver cancer cells can be used to prepare drugs that inhibit the proliferation of tumor cells and induce tumor cell differentiation.
- the technical scheme adopted in this application is: the application of a tumor cell supernatant after the action of a chemotherapeutic drug, and the supernatant of the tumor cell after the action of the chemotherapeutic drug is used to prepare a tumor cell inhibitor A drug that proliferates and induces the differentiation of the tumor cell;
- the chemotherapeutic drug includes all tumor chemotherapeutic drugs used in clinic;
- the tumor cell supernatant after the action of the chemotherapeutic drug does not contain the chemotherapeutic drug;
- the tumor cell is a malignant tumor cell ;
- the supernatant is one or a mixture of two or more of tumor cell secretions, cell lysates, and intercellular substance after the chemotherapeutic drug is temporarily acted on; the short-term effect is that the supernatant acts on the tumor cells. , According to the best control level of tumor cell proliferation and differentiation related indicators.
- the supernatant includes nucleic acids, protein factors, peptides, lipids and small molecular substances.
- the damaging effects of chemotherapeutic drugs on tumor cells include chemical damage and physical damage.
- the chemical damage is a drug; and the physical damage includes particles, radiation, electrotherapy, radiofrequency, laser or ultrasound.
- the supernatant is used alone or in combination to inhibit tumor cell proliferation and induce tumor cell differentiation.
- the tumor cells include all cell types diagnosed as tumors, and the cell types of the tumors include melanoma and glioma, as well as adrenal glands, bladder muscles, bones, bone marrow, brain, spine, chest, Neck, gallbladder, ganglia, gastrointestinal tract, stomach, colon, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary gland, skin, spleen, green pills, thymus, uterus Or cancer cells of the breast.
- the cell types of the tumors include melanoma and glioma, as well as adrenal glands, bladder muscles, bones, bone marrow, brain, spine, chest, Neck, gallbladder, ganglia, gastrointestinal tract, stomach, colon, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary gland, skin, spleen, green pills, thymus, uterus Or cancer cells of the breast
- This application proposes for the first time that the supernatant of malignant tumor cells after the temporary action of clinical chemotherapeutics has the effect of inhibiting the proliferation and inducing differentiation of untreated tumor cells.
- the supernatant can be used to prepare and inhibit the proliferation of tumor cells.
- Drugs that induce tumor cell differentiation have creatively discovered that the supernatant of malignant tumor cells after chemotherapeutic drugs has a cascading effect with tumor treatment, breaking the existing medical belief that chemotherapeutic drugs can only directly kill malignant tumor cells.
- This application proposes for the first time that the supernatant of malignant tumor cells after the temporary action of clinical chemotherapeutics has the effect of inhibiting the proliferation and inducing differentiation of untreated tumor cells.
- the supernatant can be used to prepare and inhibit the proliferation of tumor cells.
- Drugs that induce tumor cell differentiation have creatively discovered that the supernatant of malignant tumor cells after chemotherapeutic drugs has a cascading effect with tumor treatment, breaking the existing medical belief that chemotherapeutic drugs can only directly kill malignant tumor cells.
- Fig. 1 is a graph showing the expression of alpha-fetoprotein (AFP) mRNA in the supernatant of liver cancer cells after different concentrations of vincristine sulfate in Example 1 of the present application.
- AFP alpha-fetoprotein
- Example 2 is a graph showing the expression of alpha-fetoprotein (AFP) protein levels in the supernatant of liver cancer cells after different concentrations of vincristine sulfate in Example 1 of the present application act on liver cancer cells.
- AFP alpha-fetoprotein
- Fig. 3 is a graph showing the expression of alpha-fetoprotein (AFP) mRNA of liver cancer cells stimulated by the supernatant with a concentration of 4 ⁇ M vincristine sulfate aqueous solution in Example 1 of the present application for different times.
- AFP alpha-fetoprotein
- Example 4 is a graph showing the expression of alpha-fetoprotein (AFP) protein levels of liver cancer cells in the supernatant after stimulation with a 4 ⁇ M vincristine sulfate aqueous solution in Example 1 of the present application.
- AFP alpha-fetoprotein
- Figure 5 shows the liver cancer differentiation-related markers (HNF4 ⁇ and HNF6) and liver tumor stem cell markers (CD133 and CD133) of liver cancer cells after the supernatant stimulated by the 4 ⁇ M aqueous solution of vincristine sulfate in Example 1 of the present application.
- EpCAM and proliferation-related genes (PCNA and c-Myc) mRNA expression map.
- Figure 6 shows the liver cancer differentiation-related markers (HNF4 ⁇ and HNF6) and liver tumor stem cell markers (CD133 and CD133) of liver cancer cells after stimulation with a 4 ⁇ M aqueous solution of vincristine sulfate in Example 1 of the present application.
- EpCAM and proliferation-related genes (PCNA and c-Myc) protein expression map.
- the chemotherapeutic drug is vincristine sulfate (vincristine sulfate is referred to as VCR), and the supernatant after vincristine sulfate acts on liver cancer cells (
- the supernatant after vincristine sulfate acts on liver cancer cells is called TSN for short) can inhibit the proliferation of the tumor cells and induce the differentiation of the tumor cells; that is, the supernatant of each type of cancer cell only acts on the type that has not been treated by the drug.
- Cancer cells such as the supernatant of the liver cancer cell line SMMC7721 act on the liver cancer cell line SMMC7721; the supernatant of the liver cancer cell line Bel7402 act on the hepatoma cell line Bel7402; the supernatant of the liver cancer cell line Huh7 act on the liver cancer cell line Huh7; After VCR acts on liver cancer cells, the supernatant does not contain the chemotherapeutic drug VCR.
- the human liver cancer cell line SMMC7721 was cultured in DMEM medium containing 10% fetal bovine serum, 100U/mL penicillin and 100mg/mL streptomycin to make the cell abundance reach 80-90%, and then placed in a petri dish with a diameter of 60mm Or culture in a 6-well plate or a 12-well plate for 24 hours to obtain SMMC7721 cell liquid;
- the human liver cancer cell line Bel7402 was cultured in DMEM medium containing 10% fetal bovine serum, 100 U/mL penicillin and 100 mg/mL streptomycin to make the cell abundance reach 80-90%, and then placed in a culture dish with a diameter of 60 mm Or culture in a 6-well plate or a 12-well plate for 24 hours to obtain Bel7402 cell liquid;
- the human hepatoma cell line Huh7 was cultured in DMEM medium containing 10% fetal bovine serum, 100U/mL penicillin and 100mg/mL streptomycin to make the cell abundance reach 80-90%, and then cultured in a 60mm diameter Culture in a dish or a 6-well plate or a 12-well plate for 24 hours to obtain Huh7 cell liquid;
- the obtained SMMC7721 cell fluid, Bel7402 cell fluid, and Huh7 cell fluid were all stimulated with a VCR aqueous solution with a concentration of 4 ⁇ M, 6 ⁇ M, 8 ⁇ M or 10 ⁇ M for 30 minutes. After centrifugation, different concentrations of VCR were used to act on the supernatant of liver cancer cells.
- the TSN contains no VCR.
- the expression of alpha-fetoprotein (AFP) mRNA and the level of AFP protein in TSN are detected by qRT-PCR and Western blot (western blot), as can be seen from the figure, VCR can decrease AFP mRNA expression ( Figure 1) and AFP protein expression ( Figure 2) in a concentration-dependent manner, inhibit the proliferation of liver cancer cells, and induce their differentiation.
- AFP alpha-fetoprotein
- Con is a blank control. It can be seen from the figure that with different concentrations of VCR, the expression level of AFP mRNA is reduced to varying degrees; in Figure 2, GAPDHA (glyceraldehyde-3-phosphate dehydrogenase) protein is an internal control. , Con is the blank control.
- the upper image in Figure 2 is the immunoblotting diagram, and the lower image in Figure 2 is the numerical diagram of the expression level of AFP protein. It can be seen from Figure 1-2 that, as the concentration of the added VCR is different, different degrees It reduces the expression level of AFP mRNA and the expression of AFP egg level. When the VCR is 4 ⁇ M, the level of inhibition of the expression of AFP mRNA and AFP protein level is better.
- the TSN stimulated by a VCR aqueous solution at a concentration of 4 ⁇ M was used as a pretreatment condition for inhibiting the proliferation of tumor cells and inducing tumor cell differentiation.
- SMMC7721 cell fluid, Bel7402 cell fluid and Huh7 cell fluid were all stimulated with the above-mentioned 4 ⁇ M VCR aqueous solution to stimulate TSN for 5min, 10min, 15min, 20min and 25min, and the alpha-fetoprotein (AFP) was decreased in a time-dependent manner.
- Con is the blank control. It can be seen from the figure that the expression level of alpha-fetoprotein AFP mRNA is reduced to varying degrees with the action time; in Figure 4, the GAPDHA (glyceraldehyde-3-phosphate dehydrogenase) protein It is the internal control and Con is the blank control.
- the upper picture in Fig. 4 is the western blotting graph, and the lower picture in Fig. 4 is the numerical graph of the expression level of AFP protein. It can be seen from the graphs in Figs. Reduce the expression level of AFP mRNA and AFP egg level to a certain degree. When the action time is 15 minutes, the level of inhibition of AFP mRNA expression and AFP protein level expression is better.
- the TSN stimulated by a 4 ⁇ M aqueous solution of VCR stimulates liver cancer cells for 15 minutes as a treatment factor for inhibiting tumor cell proliferation and inducing tumor cell differentiation.
- the TSN stimulated by the above 4 ⁇ M VCR aqueous solution was used to stimulate SMMC7721, Bel7402, and Huh7 cells for 15 minutes, and then qRT-PCR and Western blot (immunoblotting) were used to detect the effects of TSN on SMMC7721, Bel7402, and Huh7.
- qRT-PCR and Western blot immunoblotting
- PCNA and c-Myc are cell differentiation-related markers for TSN.
- TSN inhibited SMMC7721, Bel7402 and Huh7 cell differentiation-related markers (HNF4 ⁇ and HNF6), liver tumor stem cell markers (CD133 and EpCAM) and proliferation-related genes (PCNA and c-Myc) ) MRNA expression.
- TSN has the effects of promoting the differentiation of liver cancer cells, inhibiting the transcription of liver cancer stem cell markers, and inhibiting the proliferation of liver cancer cells, and can be used to prepare drugs that inhibit the proliferation of tumor cells and induce tumor cell differentiation.
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Abstract
提供一种化疗药物作用后的肿瘤细胞上清液在制备抑制肿瘤的药物中的应用，所述抑制肿瘤是指抑制肿瘤细胞的增殖、诱导该肿瘤细胞分化。
Description
本申请要求于2020年01月07日提交的中国专利申请No.CN202010013864.2的优先权。在先申请的公开内容通过整体引用并入本申请。
本申请属于肿瘤细胞上清液技术领域，具体涉及一种化疗药物作用后的肿瘤细胞上清液的应用。
化疗是癌症的主要治疗选择，特别是对于无法获得局部治疗的患者。化疗药物可以通过细胞毒作用直接杀死肿瘤细胞。因此，虽然在临床实践中，一些患者在化疗后病理性完全缓解后长期存活，但目前认为化疗药物并不是癌细胞死亡的唯一原因，或许免疫系统可能起着重要作用。
到目前为止，研究人员主要集中在化疗药物对肿瘤及其直接靶点的直接杀伤或破坏作用。然而，用化疗药物短暂处理的癌细胞上清液随后对未治疗的肿瘤细胞的级联效应以及相关机制尚不清楚。
现有技术中只利用了化疗药物对肿瘤及其直接靶点的直接杀伤或破坏作用。
本申请所要解决的技术问题在于针对上述现有技术的不足，提供一种化疗药物作用后的肿瘤细胞上清液的应用，该化疗药物作用后的肿瘤细胞上清液具有促进肝癌细胞分化、抑制肝脏肿瘤干细胞标志物转录、抑制肝癌细胞增殖的作用，可用于制备抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的药物。
为解决上述技术问题，本申请采用的技术方案是：一种化疗药物作用后的肿瘤细胞上清液的应用，所述化疗药物作用后的肿瘤细胞的上清液用于制备抑制该肿瘤细胞的增殖、诱导该肿瘤细胞分化的药物；所述化疗药物包括临床中使用的所有肿瘤化疗药物；所述化疗药物作用后的肿瘤细胞上清液中不含有化疗药物；所述肿瘤细胞为恶性肿瘤细胞；
优选地，所述上清液为化疗药物短暂作用后的肿瘤细胞的分泌物、细胞裂解产物和细胞间质中的一种或者两种以上的混合物；短暂作用是上清液作用于肿瘤细胞后，根据肿瘤细胞的增殖和分化相关指标最佳控制水平确定。
优选地，所述上清液中包括核酸、蛋白因子、肽、脂类和小分子物质。
优选地，化疗药物对肿瘤细胞的损伤作用包括化学损伤和物理损伤，所述化学损伤为药物；所述物理损伤包括粒子、射线、电疗、射频、激光或超声。
优选地，所述上清液单独或联合应用于抑制肿瘤细胞的增殖、诱导肿瘤细胞分化。
优选地，所述肿瘤细胞包括所有被诊断为肿瘤的细胞类型，所述肿瘤的细胞类型包括为黑素瘤和神经胶质瘤，以及肾上腺、膀肌、骨、骨髓、脑、脊柱、胸、颈部、胆囊、神经节、胃肠道、胃、结肠、心脏、肾、肝、肺、肌肉、卵巢、胰、甲状旁腺、阴茎、前列腺、唾液腺、皮肤、脾、翠丸、胸腺、子宫或乳腺的癌细胞。
本申请与现有技术相比具有以下优点：
本申请首次提出了临床化疗药物短暂作用后的恶性肿瘤细胞的上清液对未治疗的肿瘤细胞具有抑制其增殖、诱导其分化的作用，可将上清液应用于制备抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的药物，创造性地发现了化疗药物作用后的恶性肿瘤细胞的上清液与肿瘤治疗存在级联效应，打破现有医学上认为化疗药物只有直接杀伤恶性肿瘤细胞的作用。
本申请与现有技术相比具有以下优点：
本申请首次提出了临床化疗药物短暂作用后的恶性肿瘤细胞的上清液对未治疗的肿瘤细胞具有抑制其增殖、诱导其分化的作用，可将上清液应用于制备抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的药物，创造性地发现了化疗药物作用后的恶性肿瘤细胞的上清液与肿瘤治疗存在级联效应，打破现有医学上认为化疗药物只有直接杀伤恶性肿瘤细胞的作用。
图1是本申请的实施例1的不同浓度的硫酸长春新碱作用于肝癌细胞后的上清液的甲胎蛋白（AFP）的mRNA的表达图。
图2是本申请的实施例1的不同浓度的硫酸长春新碱作用于肝癌细胞后的上清液的甲胎蛋白（AFP）的蛋白水平的表达图。
图3是本申请的实施例1的浓度为4μM的硫酸长春新碱水溶液刺激后上清液对肝癌细胞刺激不同时间的甲胎蛋白（AFP）的mRNA的表达图。
图4是本申请的实施例1的浓度为4μM的硫酸长春新碱水溶液刺激后上清液对肝癌细胞刺激不同时间的甲胎蛋白（AFP）的蛋白水平的表达图。
图5是本申请的实施例1的浓度为4μM的硫酸长春新碱水溶液刺激后上清液对肝癌细胞刺激15min后的肝癌分化相关标志物（HNF4α和HNF6）、肝脏肿瘤干细胞标志物（CD133和EpCAM）和增殖相关基因（PCNA和c-Myc）的mRNA的表达图。
图6是本申请的实施例1的浓度为4μM的硫酸长春新碱水溶液刺激后上清液对肝癌细胞刺激15min后的肝癌分化相关标志物（HNF4α和HNF6）、肝脏肿瘤干细胞标志物（CD133和EpCAM）和增殖相关基因（PCNA和c-Myc）的蛋白水平的表达图。
本申请的实施方式
实施例1
本实施例的化疗药物作用后的肿瘤细胞上清液的应用，所述化疗药物为硫酸长春新碱（硫酸长春新碱简称为VCR），硫酸长春新碱作用于肝癌细胞后的上清液（硫酸长春新碱作用于肝癌细胞后的上清液简称为TSN）能够抑制该肿瘤细胞的增殖、诱导该肿瘤细胞分化；即每种癌细胞的上清液只作用于未经药物作用的该类癌细胞，如肝癌细胞系SMMC7721的上清液作用于肝癌细胞系SMMC7721；肝癌细胞系Bel7402的上清液作用于肝癌细胞系Bel7402；肝癌细胞系Huh7的上清液作用肝癌细胞系Huh7；所述VCR作用于肝癌细胞后的上清液中不含有化疗药物VCR。
将人肝癌细胞系SMMC7721培养在含有10％胎牛血清、100U/mL青霉素和100mg/mL的链霉素的DMEM培养基中使细胞丰度达到80-90％，然后在直径为60mm的培养皿或6孔板或12孔板中培养24h，得到SMMC7721细胞液；
将人肝癌细胞系Bel7402培养在含有10％胎牛血清、100U/mL青霉素和100mg/mL的链霉素的DMEM培养基中使细胞丰度达到80-90％，然后在直径为60mm的培养皿或6孔板或12孔板中培养24h，得到Bel7402细胞液；
将人肝癌细胞系Huh7培养在含有10％胎牛血清、100U/mL青霉素和100mg/mL的链霉素的DMEM培养基中，使细胞丰度达到80-90％，然后在直径为60mm的培养皿或6孔板或12孔板中培养24h，得到Huh7细胞液；
（1）用于刺激产生肝癌细胞后的上清液（TSN）的VCR的浓度优化：
将得到的SMMC7721细胞液、Bel7402细胞液和Huh7细胞液，均分别用浓度为4μM、6μM、8μM或10μM的VCR水溶液刺激30min，离心后，分别取不同浓度的VCR作用于肝癌细胞后的上清液（TSN），所述TSN中不含VCR，通过qRT-PCR和Western blot（免疫印迹法）检测TSN中的甲胎蛋白（AFP）的mRNA的表达和AFP蛋白水平的表达，由图可知，VCR可浓度依赖性地降低AFP的mRNA的表达（图1）和AFP蛋白水平的表达（图2），抑制肝癌细胞的增殖，诱导其分化。
图1中，Con为空白对照，由图可知，随着VCR的浓度不同，不同程度的降低AFP的mRNA的表达水平；图2中，GAPDHA（甘油醛-3-磷酸脱氢酶）蛋白为内参，Con为空白对照，图2中上图为免疫印迹图，图2中下图为AFP蛋白的表达量数值图，由图1-2由图可知，随着加入的VCR的浓度不同，不同程度的降低AFP的mRNA的表达水平和AFP的蛋水平的表达，在VCR在4μM时，对AFP的mRNA的表达和AFP蛋白水平的表达的抑制水平较好。
将浓度为4μM的VCR水溶液刺激后的TSN作为用于抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的前处理条件。
（2）用于抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的作用时间的确定：
将得到的SMMC7721细胞液、Bel7402细胞液和Huh7细胞液分别均用上述浓度为4μM的VCR水溶液刺激后的TSN作用5min、10min、15min、20min和25min，呈时间性依赖地降低甲胎蛋白（AFP）的mRNA的表达（图3）和AFP蛋白水平的表达（图4）。
图3中，Con为空白对照，由图可知，随着作用时间不同，不同程度的降低甲胎蛋白AFP的mRNA的表达水平；图4中，GAPDHA（甘油醛-3-磷酸脱氢酶）蛋白为内参，Con为空白对照，图4中上图为免疫印迹图，图4中下图为AFP蛋白的表达量数值图，由图3-4由图可知，随着TSN作用时间的不同，不同程度的降低AFP的mRNA的表达水平和AFP的蛋水平的表达，在作用时间为15min时，对AFP的mRNA的表达和AFP蛋白水平的表达的抑制水平较好。
综上，将浓度为4μM的VCR水溶液刺激后的TSN对肝癌细胞刺激15min，作为用于抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的处理因素。
（3）TSN的作用：
将上述浓度为4μM的VCR水溶液刺激后的TSN分别对SMMC7721细胞液、Bel7402细胞液和Huh7细胞液分别刺激15min后，通过qRT-PCR和Western blot（免疫印迹法）检测TSN对SMMC7721、Bel7402和Huh7细胞分化相关标志物（HNF4α和HNF6）、肝脏肿瘤干细胞标志物（CD133和EpCAM）和增殖相关基因（PCNA和c-Myc）表达的调节作用。
由图5可知，以Con为空白对照，TSN抑制了SMMC7721、Bel7402和Huh7细胞分化相关标志物（HNF4α和HNF6）、肝脏肿瘤干细胞标志物（CD133和EpCAM）和增殖相关基因（PCNA和c-Myc）的mRNA的表达。
由图6可知，以GAPDHA（甘油醛-3-磷酸脱氢酶）蛋白为内参，Con为空白对照，TSN抑制了SSMMC7721、Bel7402和Huh7细胞分化相关标志物（HNF4α和HNF6）、肝脏肿瘤干细胞标志物（CD133和EpCAM）和增殖相关基因（PCNA和c-Myc）的蛋白水平的表达。
说明TSN具有促进肝癌细胞分化、抑制肝脏肿瘤干细胞标志物转录、抑制肝癌细胞增殖的作用，可用于制备抑制肿瘤细胞的增殖、诱导肿瘤细胞分化的药物。
以上所述，仅是本申请的较佳实施例，并非对本申请作任何限制。凡是根据发明技术实质对以上实施例所作的任何简单修改、变更以及等效变化，均仍属于本申请技术方案的保护范围内。
Claims (5)
- 一种化疗药物作用后的肿瘤细胞上清液的应用，其特征在于，所述化疗药物作用后的肿瘤细胞的上清液用于制备抑制该肿瘤细胞的增殖、诱导该肿瘤细胞分化的药物；所述化疗药物包括临床中使用的所有肿瘤化疗药物；所述化疗药物作用后的肿瘤细胞上清液中不含有化疗药物；所述肿瘤细胞为恶性肿瘤细胞。
- 根据权利要求1所述的一种化疗药物作用后的肿瘤细胞上清液的应用，其特征在于，所述上清液为化疗药物短暂作用后的肿瘤细胞的分泌物、细胞裂解产物和细胞间质中的一种或者两种以上的混合物。
- 根据权利要求1或2所述的一种化疗药物作用后的肿瘤细胞上清液的应用，其特征在于，所述上清液中包括核酸、蛋白因子、肽、脂类和小分子物质。
- 根据权利要求1所述的一种化疗药物作用后的肿瘤细胞上清液的应用，其特征在于，化疗药物对肿瘤细胞的损伤作用包括化学损伤和物理损伤，所述化学损伤为药物；所述物理损伤包括粒子、射线、电疗、射频、激光或超声。
- 根据权利要求1所述的一种化疗药物作用后的肿瘤细胞上清液的应用，其特征在于，所述上清液单独或联合应用于抑制肿瘤细胞的增殖、诱导肿瘤细胞分化。
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| CN103861107A (zh) * | 2014-03-04 | 2014-06-18 | 肖文华 | 一种药物组合物及其用途 |
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| CN103861107A (zh) * | 2014-03-04 | 2014-06-18 | 肖文华 | 一种药物组合物及其用途 |
| CN105169379A (zh) * | 2015-10-28 | 2015-12-23 | 崔长友 | 一种用于治疗胃癌的肿瘤疫苗及其制备方法 |
| CN109498651A (zh) * | 2019-01-14 | 2019-03-22 | 华中科技大学同济医学院附属协和医院 | 一种抗肿瘤微颗粒的制备方法、药剂及应用 |
| CN111000867A (zh) * | 2020-01-07 | 2020-04-14 | 河北医科大学 | 一种化疗药物作用后的肿瘤细胞上清液的应用 |
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