CN109908143A - New application of Theo Buddhist nun sieve in preparation treatment acute myeloid leukemia drug - Google Patents
New application of Theo Buddhist nun sieve in preparation treatment acute myeloid leukemia drug Download PDFInfo
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- CN109908143A CN109908143A CN201811550290.1A CN201811550290A CN109908143A CN 109908143 A CN109908143 A CN 109908143A CN 201811550290 A CN201811550290 A CN 201811550290A CN 109908143 A CN109908143 A CN 109908143A
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Abstract
The present invention relates to medicinal application fields, more particularly to Theo Buddhist nun sieve, in the new application of preparation treatment acute myeloid leukemia drug, the new application is the purposes of Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate in the drug of preparation prevention and/or treatment acute myeloid leukemia.Theo Buddhist nun sieve inhibits acute myeloid leukemia cell growth by Src/Fyn/p38 and Erk/MEK signal path, induces its apoptosis and Colony forming.
Description
Technical field
The present invention relates to medicinal application fields more particularly to Theo Buddhist nun sieve in preparation treatment acute myeloid leukemia drug
New application.
Background technique
Acute myeloid leukemia (Acute Myeloid Leukemia, AML) is pernicious gram of a kind of hematopoietic stem/progenitor cells
Grand proliferative diseases, the heterogeneity with height, it is characterized in that leukaemia cell's differentiation is obstructed, initial cell is largely proliferated, from
And lead to the associated clinical symptoms such as anaemia, decrease of platelet, infection and histoorgan infiltration, the main means of AML treatment at present
It is combined chemotherapy and Allogeneic Hematopoietic Stem Cell Transplantation (Short NJ, Rytting ME, Cortes JE, Acute myeloid
leukaemia.[J].Lancet 2018 08 18;392 (10147)) (Short NJ, Rytting ME, Cortes JE,
Acute myeloid leukaemia.[J]. Lancet 2018 08 18;392(10147)).
In recent years, with the improvement of multiple medicine combined chemotherapy dosage, the progress of hematopoietic stem cell transplantation technology and novel targeted
The application of drug, AML entirety curative effect increases earlier above, but still has quite a few patient initial drug-resistant inevitably occur
And recurrence, and once there is recurrent intractable, prognosis is very poor.Newest U.S. SEER data show that AML5 survival rate is only at present
30%~40%, in addition, combined chemotherapy and Allogeneic Hematopoietic Stem Cell Transplantation will lead to serious complication, lead to threat to life
Infection, bleeding and GVHD, treatment related mortality it is higher, many patients be die of chemotherapy or transplanting complication rather than disease
Disease itself.The progress of the novel targeted therapeutic agent of AML is also slow, at present exploitation can clinical application be mainly FLT3 and
IDH1/IGH2 inhibitor, but it is suitable only for 40%AML patient, most of AML patient still lacks safely and effectively target therapeutic agent
(Luppi M,Fabbiano F,Visani G,et al.Novel Agents for Acute Myeloid
Leukemia.Cancers(Basel)2018Nov 09;10(11)).Therefore, the treatment of AML has become the one big of blood field
Problem needs to seek new treatment method.
Aurora B is the member for belonging to Aurora A family, participates in the mitosis of regulating cell, and research is found
Auror BAML high expression, research find Aurora B in AML high expression and drug resistance and poor prognosis in relation to (Ikezoe T,
Yang J,Nishioka C,et al.A novel treatment strategy targeting Aurora kinases
in acute myelogenous leukemia.[J].Mol.Cancer Ther.2007 Jun;6(6)).
Theo Buddhist nun sieve (CS2164) is the novel three-way road tumor inhibitor of target tumor microenvironment, mainly passes through inhibition
The accesses such as VEGFR2, Aurora B and CSF1R play antineoplastic action.Early-stage study shows that it includes non-to a variety of solid tumors
Small Cell Lung Cancer, liver cancer, colon cancer and lymthoma etc. have lethal effect, and Phase I clinical trial good tolerance as the result is shown
Property and toxic side effect it is small, and about Theo Buddhist nun sieve in acute myeloid leukemia application there is not yet relevant report.
Summary of the invention
In view of the deficiencies of the prior art and actual demand, the present invention provide the new medicine use of Buddhist nun Theo sieve.The present invention
A new approach and selection is provided for the treatment of acute myeloid leukemia.
To achieve this purpose, the present invention adopts the following technical scheme:
On the one hand, the present invention provides Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate prevent in preparation
And/or the purposes in the drug for the treatment of acute myeloid leukemia.
According to the present invention, the Drug inhibition Src/Fyn/p38 and Erk/MEK signal path.
According to the present invention, the phosphorylation level of the Drug inhibition VEGFR2.
According to the present invention, the Drug inhibition acute myeloid leukemia cell proliferation.
According to the present invention, the drug kills acute myeloid leukemia cell.
In the present invention, by utilizing Buddhist nun Theo sieve (CS2164) to various acute marrow series leukemia (AML) cell strain and original
It finds that it has the function of killing AML cell for the research that tumor formation in AML sample, animal body is tested, is in concentration and Time Dependent
Property, mechanism of action is related with by inhibiting the phosphorylation level of Src/Fyn/p38, Erk/MEK signal path and VEGFR2.
In the present invention, inventor has found that the action time of Buddhist nun Theo sieve is more than just to have apparent lethal effect, and medicine for 24 hours
Object concentration lethal effect when being lower than 2 μM is faint, and lethal effect is remarkably reinforced when higher than 2 μM.
In the present invention, the drug can by injection, injection, collunarium, eye drip, infiltration, absorption, physically or chemically mediate
Method imports body such as muscle, intradermal, subcutaneous, vein, mucosal tissue;Or machine is imported after other material mixings or package
Body.
When needs, one or more pharmaceutically acceptable carriers, the load can also be added in said medicine
Body includes excipient, diluent, carrier, flavoring agent, adhesive and filler of pharmaceutical field routine etc..
It can be made with the drug that Theo Buddhist nun sieve or its pharmaceutically acceptable salt, ester, solvate are active constituent preparation
At diversified forms such as injection, tablet, pulvis, granule, capsule, oral solutions, the drug of above-mentioned various dosage forms can be according to
It is prepared by the conventional method of pharmaceutical field.
Second aspect, the present invention provide a kind of drug prevented and/or treat acute myeloid leukemia, and active constituent is
Theo Buddhist nun sieve or its pharmaceutically acceptable salt, ester, solvate.
According to the present invention, the Drug inhibition Src/Fyn/p38 and Erk/MEK signal path.
According to the present invention, the phosphorylation level of the Drug inhibition VEGFR2.
According to the present invention, the Drug inhibition acute myeloid leukemia cell proliferation.
Preferably, the drug kills acute myeloid leukemia cell.
Preferably, the drug further includes pharmaceutically acceptable auxiliary material, the auxiliary material be excipient, diluent, carrier,
In flavoring agent, adhesive and filler any one or at least two combination.
The third aspect, the present invention provides Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate inhibit in preparation
Purposes in the drug of acute myeloid leukemia cell proliferation.
Fourth aspect, the present invention provides Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate are induced in preparation
Purposes in the drug of acute myeloid leukemia cell apoptosis.
5th aspect, the present invention provides Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate inhibit in preparation
Purposes in the drug of Src/Fyn/p38 and Erk/MEK signal path.
6th aspect, the present invention provides Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate inhibit in preparation
Purposes in the drug of the phosphorylation level of VEGFR2.
According to the present invention, the drug further includes pharmaceutically acceptable auxiliary material, the auxiliary material be excipient, diluent,
In carrier, flavoring agent, adhesive and filler any one or at least two combination.
Compared with prior art, the invention has the following beneficial effects:
(1) present invention passes through in vivo and in vitro the study found that Theo Buddhist nun sieve passes through Src/Fyn/p38 and Erk/MEK signal
Access inhibits acute myeloid leukemia cell growth, induces its apoptosis and Colony forming;
(2) present invention has the function of killing AML cell by primary cell level verification Theo Buddhist nun sieve, and to normally making
Hemocytoblast without overt toxicity, illustrate the application drug compared with chemotherapeutics with Small side effects, safety is good, can be significant
Related complication is treated in reduction, has more significant application prospect.
Detailed description of the invention
Fig. 1 is that Buddhist nun in Theo sieve has the obvious AML cell Proliferation that inhibits: Fig. 1 (A) and Fig. 1 (B) is shown respectively with CCK8 reagent
Box detects Buddhist nun Theo sieve and acts on the result acted on after 48h and 72h a variety of AML cell inhibitory effects;
Fig. 2 is that Buddhist nun in Theo sieve has the obvious result for inhibiting the experiment of AML cell Proliferation Colony forming: Fig. 2 (A) and Fig. 2
(B) histogram and typical Colony forming that Theo Buddhist nun sieve after acting on for 24 hours inhibits the Colony forming of Molm13 cell are respectively indicated
Figure;
Fig. 3 is the exercising result that Buddhist nun in Theo sieve induces AML cell strain apoptosis: Fig. 3 (A) and Fig. 3 (B) show use respectively
To the feelings for inducing a variety of AML cell strain apoptosis after the bis- dye method flow cytometer detection Theo Buddhist nuns sieve effect 48h and 72h of Annexin V/PI
Condition;
The Theo Fig. 4 Buddhist nun sieve induces the exercising result of AML cell strain and primary cell apoptosis: Fig. 4 (A) and Fig. 4 (B) difference table
It is shown as with Buddhist nun in Theo sieve after 48h to 42 primary AML samples and 10 normal haematopoetic (hematologic stem
Cell, HSC) lethal effect result;
Fig. 5 is the discussion for the mechanism of action that Theo Buddhist nun sieve kills AML cell: Fig. 5 (A) and Fig. 5 (B) is CS2164 effect
In U937 and Molm13 cell for 24 hours afterwards inhibit VEGFR2 phosphorylation level and Src/Fyn/p38 signal path expression water
It is flat;Fig. 5 (C) and Fig. 5 (D) acts on U937 and Molm13 cell for Buddhist nun Theo sieve and inhibits Erk/MEK signal path afterwards for 24 hours
Expression;
Fig. 6 is research of the Buddhist nun in Theo sieve to AML killing functions of immunocytes in vivo: Fig. 6 (A) is that Theo Buddhist nun sieve inhibits AML thin
The result of born of the same parents;Fig. 6 (B) is the comparing result that Buddhist nun in Theo sieve organizes mouse subcutaneous tumor weight and control group;Fig. 6 (C) is Buddhist nun Theo sieve
In vivo to the effect of mouse.
Specific embodiment
Further to illustrate technological means and its effect adopted by the present invention, below in conjunction with attached drawing and by specific real
Mode to further illustrate the technical scheme of the present invention is applied, but the present invention is not limited in scope of embodiments.
Implement process of the invention, condition, reagent, experimental method etc., is this in addition to what is specifically mentioned below
The universal knowledege and common knowledge in field, there are no special restrictions to content by the present invention.The reality of actual conditions is not specified in each embodiment
Proved recipe method, the condition as described in Sambrook et al. usually according to normal condition, or according to the normal condition proposed by manufacturer.
Unless otherwise indicated, the meaning of whole technical term and scientific words used in this specification with institute of the present invention
It is identical to belong to the meaning that those skilled in the art are commonly understood by.But if any conflict, it is subject to this specification comprising definition.
Experimental material: experimental material source is as follows:
Theo Buddhist nun sieve is provided by Shenzhen Weixin Biological Science and Technology Co., Ltd;
Experimental animal: it is purchased from Shanghai Lake.
Embodiment 1 studies the effect that Buddhist nun in Theo sieve increases strain Proliferation Ability to AML cell
Take 2 × 105Logarithmic growth phase AML cell strain (including HL60, K562, Molm13, NB4, Thp1, U937,
MV411, OCI-AML3) 24 orifice plates are inoculated in, control group, various concentration Theo Buddhist nun sieve group (0.5,1,2,4,8,16 μ is respectively set
M) act on 48h and 72h after, using CCK8 kit detection different experiments group AML cell proliferative conditions, result such as Fig. 1 and
It is specific as follows shown in table 1:
1 Theo Buddhist nun sieve of table acts on the IC50 value after 48h and 72h to a variety of AML cell strain inhibited proliferations respectively
It can be seen that the increase with Theo sieve Buddhist nun's concentration from Fig. 1 (A)-Fig. 1 (B), under the survival rate of AML cell is significant
Drop, can be seen that as time increases, the survival rate of AML cell also declines, it is seen then that Theo sieve from the result of 48h and 72h
Time and concentration dependent is presented in Buddhist nun;
As it can be seen from table 1 Theo Buddhist nun sieve inhibits a variety of AML cell Proliferations, and the IC50 of 72h after acting on 48h and 72h
Value is lower than the IC50 value of 48h.Embodiment 2 is further tested by Colony forming experiment (Colony formation unit, CFU)
Demonstrate,prove the effect that Buddhist nun in Theo sieve inhibits AML cell Proliferation
In the present embodiment, verifying Buddhist nun Theo sieve inhibits AML cel l proliferation, the specific steps are as follows:
1) 2 × 10 are taken5Control group and Theo sieve Buddhist nun's group is arranged in 24 orifice plates in the Molm13 cell inoculation of logarithmic growth phase
(2,4 μM) effects are for 24 hours;
2) it takes control group respectively and 500, Molm3 cell of Theo Buddhist nun sieve group (2,4 μM) is inoculated in methylcellulose culture
In, in 37 DEG C and 5%CO2Colony Board Lot is calculated with inverted microscope after humidified incubator culture 10 days.All cultures are equal
In 6 orifice plates (3 hole of multiple cropping), to be greater than 50 cells as colony, three wells is averaged;
3) it takes pictures after adding 500 μ l MTT reagents to be further cultured for 4h, as a result as shown in Figure 2.
From Fig. 2 (A)-Fig. 2 (B) as can be seen that the colony forming unit of 2 μM of concentration groups is only the 61% of control group, and 4 μM
The colony forming unit of group is only control group 5% or so, and Buddhist nun in Theo sieve is prompted to have the obvious Colony forming energy for inhibiting AML cell
Power, and be in concentration dependent.
Embodiment 3 induces the apoptotic effect of AML cell with bis- dye method flow cytometer detection Theo Buddhist nuns sieve of Annexin V/PI
Take 2 × 105Logarithmic growth phase AML cell strain (including HL60, K562, Molm13, NB4, Thp1, U937,
MV411, OCI-AML3) 24 orifice plates are inoculated in, control group, (1,2,4,8 μM) of various concentration Theo Buddhist nun sieve group effect is respectively set
After 48h and 72h, using the apoptosis situation of Annexin V/PI kit detection different experiments group AML cell, as a result such as Fig. 3 institute
Show.
From Fig. 3 (A)-Fig. 3 (B) as can be seen that Buddhist nun in Theo sieve has in various degree apoptosis-induced to a variety of AML cell strains
Effect, be in time-and concentration-dependent.
Effect of the embodiment 4 from primary cell level verification Theo Buddhist nun sieve to AML cell and normal haematopoetic
The collection of clinical case sample: collecting initial or recurrent intractable AML Bone Marrow of Patients sample and normal hematopoiesis is dry thin
Born of the same parents (HSC) extract mononuclearcell with lymphocyte separation medium.
After (0.5,1,2,4,8,16 μM) of the various concentration Theo Buddhist nun sieve primary AML cell of processing and normal HSC 48h,
Annexin V/PI method streaming technology detects primary AML cell and the apoptosis ratio of normal HSC, as a result as shown in Fig. 4 and table 2,
It is specific as follows:
Table 2
From Fig. 4 (A)-Fig. 4 (B) as can be seen that Buddhist nun in Theo sieve has killing to the primary AML sample standard deviation of 42 separate sources
Effect, and the higher apoptosis ratio of concentration is bigger, and it derives from normal candidate stem cell without obvious lethal effect to 10.
From table 2 it can be seen that Theo Buddhist nun sieve to the lethal effect of primary AML cell may with NPM1, ASXL1 and
TET2 mutation is related, while outer marrow infiltration may also influence the fragmentation effect of Theo Buddhist nun sieve;And white blood cell count(WBC), the LDH of patient
The factors such as horizontal, morbid state and FLT3-ITD mutation have no effect on the killing efficiency of CS2164.
Embodiment 5 kills the mechanism of action of AML cell with WB (western blot) method research Theo Buddhist nun sieve
0,0.5,1,2,4 μM of Theo sieve Buddhist nun's group is respectively set in U937 the and Molm13 cell of logarithmic growth phase, effect
It receives cell afterwards for 24 hours, then extracts albumen for western blot and detect VEGFR2 and downstream Src/Fyn/p38 and Erk/MEK letter
The expression of number access, as a result as shown in Figure 5.
From fig. 5, it can be seen that Fig. 5 (A) and Fig. 5 (C) shows that Buddhist nun in Theo sieve acts on U937 cell and inhibits afterwards for 24 hours
The phosphorylation level of VEGFR2 and the expression for inhibiting Src/Fyn/p38 and Erk/MEK signal path;Fig. 5 (B) and Fig. 5 (D)
Show Theo Buddhist nun sieve act on Molm13 cell for 24 hours afterwards inhibit VEGFR2 phosphorylation level and inhibit Src/Fyn/p38 and
Erk/MEK signal path.
Embodiment 6 has the function of killing AML from animal level verification Theo Buddhist nun sieve in vivo
1) nude mice lotus knurl model is constructed using AML cell strain (U937 and Molm13)
SPF grades of 4-6 weeks nude mice age sizes, half male and half female, operations all to mouse carry out in sterile laminar flow room.It will
U937 or Molm13 cell is suspended in 0.2ml and contains in the culture medium of 0.5%FBS (containing 5 × 10 in every 0.2ml6It is a thin
Born of the same parents), it is subcutaneous to be inoculated in mouse right fore, long to 75-150mm to gross tumor volume3, internal medication experiment can be started.
2) mouse experiment in vivo further verifies Theo Buddhist nun sieve and has the function of killing AML cell (U937 and Molm13)
Control group and Theo Buddhist nun sieve group (40mg/kg/d) is respectively set, the week is administered, and continuous 3 weeks, every two heavenly prison or jail
Survey the weight of mouse and the size of tumour
3) mouse is directly euthanized after medication, takes knurl for calculated weight and taken pictures, as a result as shown in Figure 6.
From Fig. 6 (A) as can be seen that Buddhist nun in Theo sieve inhibits AML cell (U937 and Molm13) intracorporal growth;Fig. 6 (B) can
It is weighs significantly less than control group to find out that Theo Buddhist nun sieve organizes mouse subcutaneous tumors, has the function of inhibiting knurl growth;Fig. 6 (C) can
To find out that Theo Buddhist nun sieve is acted in vivo to mouse without apparent toxic side effect, do not change the changes of weight of mouse.
In conclusion the present invention by vivo and in vitro the study found that Theo Buddhist nun sieve by inhibit Src/Fyn/p38 and
The phosphorylation level killing acute myeloid leukemia cell of Erk/MEK signal path and inhibition VEGFR2 induce its apoptosis and collection
It falls to be formed.
The Applicant declares that the present invention is explained by the above embodiments method detailed of the invention, but the present invention not office
Be limited to above-mentioned method detailed, that is, do not mean that the invention must rely on the above detailed methods to implement.Technical field
Technical staff it will be clearly understood that any improvement in the present invention, equivalence replacement and auxiliary element to each raw material of product of the present invention
Addition, selection of concrete mode etc., all of which fall within the scope of protection and disclosure of the present invention.
Claims (10)
1. Theo Buddhist nun sieve or its pharmaceutically acceptable salt, ester, solvate are in preparation prevention and/or the treatment white blood of acute myeloid
Purposes in the drug of disease.
2. purposes according to claim 1, which is characterized in that the Drug inhibition Src/Fyn/p38 and Erk/MEK signal
Access;
Preferably, the phosphorylation level of the Drug inhibition VEGFR2.
3. purposes according to claim 1 or 2, which is characterized in that the Drug inhibition acute myeloid leukemia cell increases
It grows;
Preferably, the drug kills acute myeloid leukemia cell;
Preferably, the drug further includes pharmaceutically acceptable auxiliary material;
Preferably, the auxiliary material be excipient, diluent, carrier, flavoring agent, adhesive and filler in any one or extremely
Few two kinds of combination.
4. the drug of a kind of prevention and/or treatment acute myeloid leukemia, active constituent is Buddhist nun Theo sieve or it pharmaceutically may be used
Salt, ester, the solvate of receiving.
5. drug according to claim 4, which is characterized in that the Drug inhibition Src/Fyn/p38 and Erk/MEK signal
Access;
Preferably, the phosphorylation level of the Drug inhibition VEGFR2.
6. drug according to claim 4 or 5, which is characterized in that the Drug inhibition acute myeloid leukemia cell increases
It grows;
Preferably, the drug kills acute myeloid leukemia cell;
Preferably, the drug further includes pharmaceutically acceptable auxiliary material;
Preferably, the auxiliary material be excipient, diluent, carrier, flavoring agent, adhesive and filler in any one or extremely
Few two kinds of combination.
7. Theo Buddhist nun sieve or its pharmaceutically acceptable salt, ester, solvate inhibit acute myeloid leukemia cell to increase in preparation
The purposes in drug grown.
8. Theo Buddhist nun sieve or its pharmaceutically acceptable salt, ester, solvate wither in preparation induction acute myeloid leukemia cell
The purposes in drug died.
9. Theo Buddhist nun sieve or its pharmaceutically acceptable salt, ester, solvate inhibit Src/Fyn/p38 and Erk/MEK in preparation
Purposes in the drug of signal path;
Preferably, Buddhist nun Theo sieve or its pharmaceutically acceptable salt, ester, solvate inhibit the phosphorylation water of VEGFR2 in preparation
Purposes in flat drug.
10. the purposes according to any one of claim 7-9, which is characterized in that the drug further includes that can pharmaceutically connect
The auxiliary material received;
Preferably, the auxiliary material be excipient, diluent, carrier, flavoring agent, adhesive and filler in any one or extremely
Few two kinds of combination.
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CN111214475A (en) * | 2020-02-25 | 2020-06-02 | 厦门大学附属第一医院 | Combined pharmaceutical composition for resisting double-hit lymphoma and application thereof |
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CN114762685A (en) * | 2021-01-12 | 2022-07-19 | 陈璋辉 | Application of tegaserod maleate in treating acute myeloid leukemia and colorectal cancer |
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CN111214475A (en) * | 2020-02-25 | 2020-06-02 | 厦门大学附属第一医院 | Combined pharmaceutical composition for resisting double-hit lymphoma and application thereof |
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WO2022007745A1 (en) * | 2020-07-08 | 2022-01-13 | 深圳微芯生物科技股份有限公司 | Use of chiauranib and drug combination thereof in treatment of non-hodgkin's lymphoma |
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WO2023169384A1 (en) * | 2022-03-09 | 2023-09-14 | 深圳微芯生物科技股份有限公司 | Use of chiauranib in combination with anti-metabolism chemotherapeutic drug in treatment of breast cancer |
CN115177620A (en) * | 2022-07-18 | 2022-10-14 | 厦门大学附属第一医院 | Application of seolonide or pharmaceutically acceptable salt thereof in preparation of medicine for preventing or treating follicular lymphoma |
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