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Culture method for drug-resistant cell line of human oral squamous cell carcinoma CAL-27 and cell line obtained after culture

A technology of CAL-27 and drug-resistant cells, which is applied in the direction of tumor/cancer cells, microbial-based methods, animal cells, etc., can solve problems such as gene changes, and achieve the effect of stable traits

Inactive Publication Date: 2014-11-26
上海溥生生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The technical problem to be solved by the present invention is to overcome the defect that the human oral squamous cell carcinoma drug-resistant cell line screened by the current conventional method is easy to cause genetic changes due to human factors, and to use human oral squamous cell carcinoma CAL-27 cells as parent cells, select Tumor chemotherapeutic drugs are commonly used as screening drugs, and the real action process of drugs after administration in the human body is simulated in vitro, so as to establish a stable and effective screening system and method for drug-resistant cell lines

Method used

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  • Culture method for drug-resistant cell line of human oral squamous cell carcinoma CAL-27 and cell line obtained after culture
  • Culture method for drug-resistant cell line of human oral squamous cell carcinoma CAL-27 and cell line obtained after culture
  • Culture method for drug-resistant cell line of human oral squamous cell carcinoma CAL-27 and cell line obtained after culture

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1 Screening of human oral squamous cell carcinoma CAL-27 / 5-Fu drug-resistant cell lines

[0067] In this example, 5-fluorouracil (5-Fu) was used as a screening drug to induce and screen CAL-27 cells, establish CAL-27 / 5-Fu drug-resistant cell lines, and detect the characteristics of drug-resistant cells.

[0068] 1. Parental cell culture:

[0069] Human oral squamous cell carcinoma CAL-27 (ATCC NO. is CRL-2095) cells were grown in high-glucose DMEM medium containing 10% FBS, placed in 5% CO 2 , Cultured in a 37°C incubator.

[0070] 2. Parent cell IC 50 Determination of IC with 20 calculation

[0071] Drug: 5-Fluorouracil (5-Fu)

[0072] Dosage: Design 8 drug concentration gradients (0μg / ml, 0.02μg / ml, 0.04μg / ml, 0.08μg / ml, 0.16μg / ml, 0.32μg / ml, 0.64μg / ml, 1.28μg / ml), Measuring the IC of CAL-27 cells by MTT method 50 concentration. The full name of MTT is 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenyltetrazolium romide, and its Chinese chemical name is 3-...

Embodiment 2

[0108] Example 2 Screening of human oral squamous cell carcinoma CAL-27 / CDDP drug-resistant cell lines

[0109] CAL-27 cells were screened by cisplatin (CDDP) drug induction, CAL-27 / CDDP drug-resistant cell lines were established, and the characteristics of drug-resistant cells were detected.

[0110] One, parent cell culture: operation is the same as embodiment 1.

[0111] 2. Parent cell IC 50 Determination of IC with 20 calculation

[0112] Drug: Cisplatin (CDDP)

[0113] Dosage: with embodiment 1.

[0114] The IC of CAL-27 cells was calculated by the modified Cole's method 50 The concentration was 2.333 μg / ml, and the IC was calculated 20 The concentration of 0.75 μg / ml was used as the initial concentration for screening.

[0115] 3. Induction of drug-resistant cell lines in human oral squamous cell carcinoma

[0116] Human oral squamous cell carcinoma CAL-27 cells in good growth state and in the logarithmic growth phase were collected and counted by trypsinization....

Embodiment 3

[0134] Example 3 Screening of human oral squamous cell carcinoma CAL-27-HCPT drug-resistant cell line

[0135] CAL-27 cells were screened by hydroxycamptothecin (HCPT) drugs, CAL-27-CDDP drug-resistant cell lines were established, and the characteristics of drug-resistant cells were detected.

[0136] One, parent cell culture: operation is the same as embodiment 1

[0137] 2. Parent cell IC 50 Determination of IC with 20 calculation

[0138] Drug: Hydroxycamptothecin (HCPT)

[0139] Dosage: with embodiment 1.

[0140] The IC of CAL-27 cells was calculated by the modified Cole's method 50 The concentration is 0.101 μg / ml, and the IC is calculated 20 The concentration of 0.01 μg / ml was used as the initial concentration for screening.

[0141] 3. Induction of drug-resistant cell lines in human oral squamous cell carcinoma

[0142] Human oral squamous cell carcinoma CAL-27 cells in good growth state and in the logarithmic growth phase were collected and counted by trypsini...

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Abstract

The invention discloses a culture method for a drug-resistant cell line of human oral squamous cell carcinoma CAL-27 and the cell line obtained in the culture method. The method comprises the following steps: (1) cultivating CAL-27 parent cells in a cell culture dish, adding a screening drug when a fusion rate reaches 30 to 40% after the cells grow and adhere onto the wall of the dish, with the initial concentration of the drug being the IC20 value of the drug, and washing out dead cells and amplifying survival cells after continuous action of the drug for 24 to 72 h; and (2) when the cells are in a logarithmic phase again, repeating the operations of washing-out of dead cells and amplification of survival cells as in step (1), wherein the concentration of the screening drug for action is increased by 1 time, increase times of the concentration of the screening drug are reduced when the concentration of the screening drug is increased to a first fixed value, and cells are subjected to cryopreservation and then to resuscitation when the concentration of the screening drug is increased to a second fixed value so as to obtain the cell line. The obtained drug-resistant cell line has clinically biological characters of human oral squamous cell carcinoma and is an ideal model for research on a resistance mechanism of human oral squamous cell carcinoma cells.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a method for culturing a CAL-27 drug-resistant cell line of human oral cavity squamous cell carcinoma and the cell line obtained from the culture. Background technique [0002] In my country, carcinoma is the most common malignant tumor of the oral and maxillofacial region, and sarcoma is rare. Among cancers, squamous cell carcinoma is the most common, generally accounting for more than 80%; followed by glandular epithelial carcinoma (such as mucoepidermoid carcinoma, adenocarcinoma, adenoid cystic carcinoma, malignant pleomorphic adenoma, acinar carcinoma) cell carcinoma, etc.) and undifferentiated carcinoma. Squamous cell carcinoma often metastasizes to regional lymph nodes, and distant metastasis may occur in advanced stages. For squamous cell carcinoma, although many therapeutic drugs have been developed and have been effectively used clinically, however, drug resistance and mu...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/09A01K67/027C12R1/91
Inventor 王海峰洪谊姚远颋费倩岚谈竹君劳昕元
Owner 上海溥生生物科技有限公司