Application of HOXA1 in preparation of antitumor drug

An anti-tumor drug, HOXA1 technology, applied in anti-tumor drugs, drug combinations, pharmaceutical formulations, etc., can solve problems such as unreported

Inactive Publication Date: 2017-08-15
ZHENJIANG NO 1 PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the role of HOXA1 in the activation, expansion and immunosuppressive function of MDSCs has not been reported so far.

Method used

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  • Application of HOXA1 in preparation of antitumor drug
  • Application of HOXA1 in preparation of antitumor drug
  • Application of HOXA1 in preparation of antitumor drug

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1: Detection of HOXA1 in MDSCs derived from tumor tissue of lung cancer patients

[0023] (1) Preparation of tumor and paracancerous tissue cell suspensions in patients with lung cancer

[0024] Weigh the fresh tissue and cut it to 1-2mm 3 . Then add tissue digestion solution (Sigma Company) at 10 mL / 0.27 g, digest in a water bath at 37 °C for 2 h, and pipette once every 10 min. After the digestion is over, use a sieve to filter out residual tissue clumps, centrifuge at 500g at 4°C for 5 min, and discard the supernatant to retain the cell pellet. Then erythrocytes were lysed by ACK, washed with PBS, centrifuged at 500 g for 5 min at 4°C, and the supernatant was discarded to retain the cell pellet.

[0025] (2) Induction of MDSCs in vitro

[0026] After Ficoll gradient centrifugation to separate healthy human peripheral blood PBMCs, according to 1 × 10 7 Seed per well in a 6-well plate, then add 40ng / mL GM-CSF (Peprotech) and 40ng / mL IL-1β (Peprotech) to sti...

Embodiment 2

[0047] Example 2: The influence of HOXA1 on the occurrence and development of tumors

[0048] (1) Cell culture in vitro: Lewis lung cancer cells were cultured in DMEM with 10% calf serum in 5% CO2 and 37°C in vitro, and the logarithmic growth phase was used for subsequent experiments.

[0049] (2) Establishment of lung cancer xenograft tumor model: male C57BL / 6 mice aged 6 to 8 weeks were taken, and the skin on the right abdominal wall of the mice was disinfected with medical alcohol in an ultra-clean bench, and each mouse was subcutaneously injected with 1x10 6 cells / 200 μL.

[0050] (3) The tumor-bearing mice were divided into two groups: Ctrl group and HOXA1 group. On the seventh day after tumor implantation, pcDNA3.1vector or pcDNA3.1-HOXA1 was locally injected into the tumor every three days for 2-4 weeks. During this period, the tumor size was measured every 2 days with the formula volume=0.5×length×width 2 to calculate the tumor volume. The mice were sacrificed 24 h...

Embodiment 3

[0052] Example 3: Regulation of HOXA1 on MDSCs immunosuppressive molecules during tumor development

[0053] (1) Sorting of MDSCs from tumor tissue of tumor-bearing mice

[0054] 1) Preparation of tumor tissue cell suspension of tumor-bearing mice: carefully strip the complete tumor tissue into a glass plate in an ultra-clean bench, weigh and cut the tumor tissue to 1-2mm 3 . Then add tissue digestion solution at 10mL / 0.27g, digest in a water bath at 37°C for 2h, and pipette once every 10min. After the digestion is over, use a sieve to filter out residual tissue clumps, centrifuge at 500g at 4°C for 5 min, and discard the supernatant to retain the cell pellet.

[0055] 2) Proportion of local MDSCs detected by flow cytometry

[0056] Take 1×10 6 Mouse tumor tissue cells in Ctrl group and HOXA1 group were resuspended in 1 mLEP tube in 100 μL PBS, and 0.25 μg of PE-anti-mouse-Gr1 and PE / CY5-anti-mouse-CD11b antibodies or corresponding isotype antibodies were added. Mix well ...

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Abstract

The invention belongs to the field of cytobiology and molecular biology and the field of clinical application, and relates to application of HOXA1. The invention discloses application of the HOXA1 in preparation of an antitumor drug, in particular application in preparation of a drug for suppressing the immunosuppressive function of myeloid-derived suppressor cells (MDSCs). The application shows that the HOXA1 can control the immunosuppressive effect of the MDSCs under a tumor condition; compared with the HOXA1 expression in a para-carcinoma tissue, the HOXA1 expression in the tumor tissue-derived MDSCs of a lung cancer patient is obviously down-regulated. A further experiment shows that by use of an HOXA1 overexpression carrier, in case of HOXA1 overexpression, growth of a tumor in a tumor-bearing mouse can be effectively delayed, and a Th1-CTL-mediated antitumor immune response can be promoted.

Description

technical field [0001] The invention relates to a new method of tumor immunotherapy targeting HOXA1 molecules, which belongs to the field of cell biology, molecular biology and clinical application, and specifically relates to the flow cytometry sorting of tumor tissue MDSCs of lung cancer patients, the construction of HOXA1 expression vector and its application in tumor treatment in the application. Background technique [0002] Tumor-induced immunosuppression is an important mechanism for tumor escape, which not only prevents natural tumor immune surveillance, but also limits tumor-targeted immunotherapy. The tumor microenvironment induces a large number of different immunosuppressive cells in the body, including regulatory T cells, tumor-associated macrophages, etc. These tumor-induced suppressor cells have an inhibitory effect on tumor-specific effector T cells, which is conducive to the further development and deterioration of tumors. In recent years, more and more st...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61P35/00A61P37/04
CPCA61K48/005
Inventor 王胜军田新宇田洁芮棵王运刚许化溪
Owner ZHENJIANG NO 1 PEOPLES HOSPITAL
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