CN105198790B - Promote the tetracyclic compound of double negative t cells in-vitro multiplication - Google Patents
Promote the tetracyclic compound of double negative t cells in-vitro multiplication Download PDFInfo
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- CN105198790B CN105198790B CN201510186837.4A CN201510186837A CN105198790B CN 105198790 B CN105198790 B CN 105198790B CN 201510186837 A CN201510186837 A CN 201510186837A CN 105198790 B CN105198790 B CN 105198790B
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
- C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
- C07D209/52—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring condensed with a ring other than six-membered
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Abstract
The invention discloses a kind of tetracyclic compound for promoting double negative t cells in-vitro multiplication, the tetracyclic compound is IMC 00239, and its molecular formula is：.The present invention relates to screened that CD3 can be promoted from compound library with high flux screening method+/CD4‑/CD8‑The tetracyclic compound of double negative t (DN T) cell proliferation in vitro, including its pharmaceutically acceptable salt and prodrug.Such compound is the inhibitor of Wnt signal paths, and DN T cell β Catenin degraded can be induced in testing in vitro, promotes cell cycle and mitosis.Such compound can be effectively promoted the structure of CAR DN T amplification in vitro systems, help the realization of CAR DN T cells factory.
Description
Technical field
The present invention relates to a kind of tetracyclic compound of promotion double negative t cells in-vitro multiplication of biomedicine field, especially
It is related to a kind of promotion CD3+/CD4-/CD8-The tetracyclic compound of double negative t (DN T) cell proliferation in vitro.
Background technology
Immunotherapy is considered as the third time revolution for the treatment of of cancer, wherein Chimeric antigen receptor T cell（CAR-T）Technology
To CD19+The remarkable clinical test results of B-lineage Acute Lymphocyte Leukemia have caused the extensive concern of medical field, are considered as
It is the therapy for being most hopeful to cure tumour.
However, current CAR-T treatments are to carry out CAR-T cell transformations with autologous patient T cell, this is just to the T of patient
Cell quantity, vigor have very high requirement with multiplication capacity.Found in clinical test practice, the T cell of the patient more than 80%
The requirement of CAR-T cells preparation is not reached, therefore Most patients can not be benefited from existing CAR-T treatments.In view of this
Problem, homologous Hetersomata CAR-T preparation is as the focus studied.
Current research strategy is by the φt cell receptor of the T cell of homologous xenogenic origin by gene Knockout（TCR）
After knockout, it is lost the characteristic of MHC limitations, be then purified into the T cell of TCR knockouts, then carry out the preparation of CAR-T cells,
It is allowed to play tumor-killing effect and avoids causing graft versus host disease（GVHD）.But run into practical operation very big
Technical bottleneck, main cause are that the carrier molecule amount such as CRISPR-Cas9 for gene Knockout is larger, import T cell and are stranded
Hardly possible, the efficiency of gene knockout are low.Also, if importing the carriers such as CRISPR-Cas9 using the mode of virus transfection, it can go out again
The resistance phenomenon of the secondary virus transfection of existing Chimeric antigen receptor, CAR-T cell yields are low, it is impossible to prepared on a large scale, should
It is poor with property.
Existing research finds a group CD3 be present in the peripheral blood of normal person+/CD4-/CD8-Double negative t (DN T) is thin
Born of the same parents, it is restricted without MHC, will not cause graft versus host disease, with reference to existing Chimeric antigen receptor technology, structure
Go out a kind of new Chimeric antigen receptor DN-T cells (CAR-DN-T), tumor patient can be fed back with allosome.But in view of DN T
Cell only accounts for and accounts for PMNC（PBMC）1-5%, it is difficult to reach CAR-DN-T treatment needed for cell concentration.
The content of the invention
It is an object of the invention to overcome problem above existing for prior art, there is provided one kind promotes double negative t cells body
The tetracyclic compound of outer propagation.
To realize above-mentioned technical purpose and the technique effect, the present invention is achieved through the following technical solutions：
A kind of tetracyclic compound for promoting double negative t cells in-vitro multiplication, the tetracyclic compound is IMC-00239, its
Molecular formula is：
。
Further, the tetracyclic compound for promoting double negative t cells in-vitro multiplication promotes DN T cell in-vitro multiplications
Application.
The beneficial effects of the invention are as follows:
1st, invention, which is related to be screened from compound library with high flux screening method, can promote CD3+/CD4-/CD8-Double negative t
The tetracyclic compound of (DN T) cell proliferation in vitro, including its pharmaceutically acceptable salt and prodrug.Such compound is Wnt letters
The inhibitor of number path, DN T cell β-Catenin degraded can be induced in testing in vitro, promote the cell cycle and have silk point
Split.Because DN T cells are restricted without MHC, graft versus host disease will not be caused（GVHD）, with reference to existing chimeric
Antigen receptor (CAR) technology, a kind of new Chimeric antigen receptor DN-T cells (CAR-DN-T) are constructed, can be fed back with allosome
Tumor patient, realize the industrialization of CAR-T cell factories.In view of DN T cells only account for and account for PMNC（PBMC）
1-5%, need to establish efficient external extensive amplification system could meet the cell concentration demand of CAR-DN-T treatments.Suchization
Compound can be effectively promoted the structure of CAR-DN-T amplification in vitro systems, help the realization of CAR-DN-T cell factories.
2nd, the compound IMC-00239 obtained by the high-flux cell propagation Screening Platform based on primary DN T cells
It is combined with φt cell receptor antibody and cell factor, external efficiently promotion DN T cell propagation can be done, its amplification ability is more than 4000
Times, 7 times are higher by than φt cell receptor antibody and cell factor treatment group, and be in dose-dependent effect.Under compound ≤30 μM
Without obvious cytotoxicity.Further study show that IMC-00239 mainly induces β-Catenin drops by suppressing Wnt signal paths
The propagation of solution regulation and control DN T cells.The CAR-DN-T cells of proliferation strong supports of the IMC-00239 to DN T cells
The foundation of amplification in vitro system, meet cell concentration needed for treatment.
Described above is only the general introduction of technical solution of the present invention, in order to better understand the technological means of the present invention,
And can be practiced according to the content of specification, below with presently preferred embodiments of the present invention and coordinate accompanying drawing describe in detail as after.
The embodiment of the present invention is shown in detail by following examples and its accompanying drawing.
Brief description of the drawings
Accompanying drawing described herein is used for providing a further understanding of the present invention, forms the part of the application, this hair
Bright schematic description and description is used to explain the present invention, does not form inappropriate limitation of the present invention.In the accompanying drawings：
Fig. 1 is the screening of high-flux cell propagation and CAR-T killing experiments flow charts based on primary DN T cells；
Fig. 2 is the propagation and its signal path figure one that IMC-00239 promotes DN T cells；
Fig. 3 is the propagation and its signal path figure two that IMC-00239 promotes DN T cells；
Fig. 4 is the propagation and its signal path figure three that IMC-00239 promotes DN T cells；
Fig. 5 is the propagation and its signal path figure four that IMC-00239 promotes DN T cells；
Fig. 6 is the facilitation schematic diagram that IMC-00239 breeds to CAR-DN-T；
Fig. 7 is that the CAR-DN-T cells of IMC-00239 compounds processing have Cytotoxicity in vitro functional schematic.
Embodiment
Below with reference to the accompanying drawings and in conjunction with the embodiments, the present invention is described in detail.
In view of DN T cells only account for PMNC（PBMC）1-5%, efficient external extensive expand need to be established
Increasing system could meet the cell concentration demand of CAR-DN-T treatments.We establish the high-flux cell based on primary DN T cells and increased
Screening Platform is grown, and the compound screened is demonstrate,proved in the enterprising step of CAR-DN-T cells, and confirms it not influenceing CAR-T's
Cellkilling capacity, idiographic flow is as shown in Figure 1.Pass through this platform, it has been found that a series of that DN T cells can be promoted to increase
The micromolecular compound grown, wherein, effect most strong is a tetracyclic compound（IMC-00239）, its molecular formula is：
。
The compound IMC- that we are obtained by the high-flux cell propagation Screening Platform based on primary DN T cells
00239 is combined with φt cell receptor antibody and cell factor, can do external efficiently promotion DN T cell propagation, and its amplification ability surpasses
4000 times are crossed, 7 times are higher by than φt cell receptor antibody and cell factor treatment group（0.5 μM, shown in reference picture 2）, and in dosage according to
Rely effect（PEC50=7.2, shown in reference picture 3）.Without obvious cytotoxicity under compound ≤30 μM, shown in reference picture 4.Enter
One step research finds that IMC-00239 mainly induces the increasing of β-Catenin degrading and regulating DN T cells by suppressing Wnt signal paths
Grow, shown in reference picture 5.The CAR-DN-T cell bodies of proliferation strong supports of the IMC-00239 to DN T cells extend out
The foundation of increasing system, meet cell concentration needed for treatment.
Embodiment
IMC-00239 does not influence CAR-DN-T cell killing function to CAR-DN-T rush amplification effect.
Slow virus of the DN T cells through Chimeric antigen receptor sub-elected from healthy donor's PMNC
After infection, respectively with φt cell receptor antibody and cell factor (Vehicle) and 0.5 μM of IMC-00239 joint φt cell receptor
Antibody and cell factor stimulate, and cultivate 14 days.IMC-00239 groups cell number is higher than about 7 times or so of Vehicle groups, the institute of reference picture 6
Show.By IMC-00239 groups and cell factor group CAR-DN-T carry out cellkilling capacity it was found that IMC-00239 to CAR-
DN-T cellkilling capacity has no significant effect, shown in reference picture 7.
The preferred embodiments of the present invention are the foregoing is only, are not intended to limit the invention, for the skill of this area
For art personnel, the present invention can have various modifications and variations.Within the spirit and principles of the invention, that is made any repaiies
Change, equivalent substitution, improvement etc., should be included in the scope of the protection.
Claims (1)
1. a kind of tetracyclic compound for promoting double negative t cells in-vitro multiplication, it is characterised in that the tetracyclic compound is IMC-
00239, its structural formula is：
。
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| CN201510186837.4A CN105198790B (en) | 2015-04-20 | 2015-04-20 | Promote the tetracyclic compound of double negative t cells in-vitro multiplication |
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| CN105198790B true CN105198790B (en) | 2018-02-16 |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1882342A (en) * | 2003-09-17 | 2006-12-20 | 艾科斯有限公司 | Use of CHK1 inhibitors to control cell proliferation |
| CN102695413A (en) * | 2009-11-10 | 2012-09-26 | 麦赛尔控股有限公司 | Stabilized formulations of fatty acids |
| CN104136458A (en) * | 2012-02-22 | 2014-11-05 | 宾夕法尼亚大学董事会 | Use of the cd2 signaling domain in second-generation chimeric antigen receptors |
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| WO2013049307A2 (en) * | 2011-09-30 | 2013-04-04 | University Of Miami | Enhanced immune memory development by aptamer targeted mtor inhibition of t cells |
| DK2981607T3 (en) * | 2013-04-03 | 2020-11-16 | Memorial Sloan Kettering Cancer Center | EFFICIENT GENERATION OF TUMOR-TARGETED T-CELLS DERIVED FROM PLURIPOTENT STEM CELLS |
| WO2015023551A1 (en) * | 2013-08-13 | 2015-02-19 | Bionumerik Pharmaceuticals, Inc. | Administration of karenitecin for the treatment of advanced ovarian cancer, including chemotherapy-resistant and/or the mucinous adenocarcinoma sub-types |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1882342A (en) * | 2003-09-17 | 2006-12-20 | 艾科斯有限公司 | Use of CHK1 inhibitors to control cell proliferation |
| CN102695413A (en) * | 2009-11-10 | 2012-09-26 | 麦赛尔控股有限公司 | Stabilized formulations of fatty acids |
| CN104136458A (en) * | 2012-02-22 | 2014-11-05 | 宾夕法尼亚大学董事会 | Use of the cd2 signaling domain in second-generation chimeric antigen receptors |
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