CN101474170A - Application of isosulfocyanate compound in preparing medicament for treating leukemia - Google Patents
Application of isosulfocyanate compound in preparing medicament for treating leukemia Download PDFInfo
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- CN101474170A CN101474170A CNA2008102332218A CN200810233221A CN101474170A CN 101474170 A CN101474170 A CN 101474170A CN A2008102332218 A CNA2008102332218 A CN A2008102332218A CN 200810233221 A CN200810233221 A CN 200810233221A CN 101474170 A CN101474170 A CN 101474170A
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Abstract
The invention relates to a new use of isorhodanic ester compound in preparing pharmaceuticals for treatment of leukemia. The isorhodanic ester compound in preparing pharmaceuticals for treatment of leukemia has strong pharmacological action as well as characteristics of broad spectrum, special effect and low toxicity, thereby having good medical prospect. Additionally, the invention can be used for acute leukemia (including acute myelaid leukemia, acute T lymphatic leukemia and acute promyelocytic leukemia, and the like) and chronic leukemia.
Description
Technical field
The present invention relates to the purposes of isosulfocyanate compound, relate in particular to the application of isosulfocyanate compound in the leukemic medicine of preparation treatment.
Background technology
Isothiocyanate (isothiocyanates) extensively is present in various Cruciferae such as the plants such as broccoli, Brassica Oleracea Var.Acephala, brussels sprout, Brassica oleracea L. var. botrytis L., Radix Cochleariae officinalis and Nasturtium officinale.Crucifer is rich in thioglycoside (glucosinolates), and this material can produce the hydrolyzate of biologically active, i.e. isothiocyanate after plant myrosin or gastrointestinal microorganisms hydrolysis.
Isosulfocyanate compound mainly comprise isothiocyanic acid benzyl ester (BITC), isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN), and glycosides oil three eruciates (Erucin) etc. several.The molecular formula of isothiocyanic acid benzyl ester (BITC) is C
8H
7NS, structural formula is:
Leukemia is common blood system malignant tumor, accounts for the 6th of China adult mortality of malignant tumors, accounts for first of children malignant tumors mortality rate.The result shows according to Epidemiological study, and leukemic sickness rate is 4/100000ths, and the main age of onset of this disease wherein is the child more than 50% below 35 years old.Annual newly-increased, about 1,000,000 people of recurrence leukemia in the whole world.Annual newly-increased about 40,000 people of leukaemic in the whole nation die from about 30,000 people of leukemia every year.Because soaring year by year M ﹠ M is sought emphasis and focus that therapeutic strategy likely becomes leukemia research.Therefore, leukemia is that one of disease of keypoint control is demanded in the whole world urgently.In recent years, all adopt method inhibition leukaemia growths such as chemotherapy and radiation both at home and abroad, prolong patient's life-span.But the toxic and side effects of these class methods is big, and the relapse rate height not only brings great misery to patient, and has caused white elephant for its family, society.The sixties begin just bone marrow transplantation to be applied in the world leukemic treatment, and China has also carried out bone marrow transplantation and applied to the method for leukemia treating in nearly ten years, and has obtained good effect.But, because the medical expense costliness, and be subjected to joining the restriction in type source, still have many patients because can not get the suitable medical expense of joining the bone marrow of type or handing over not rise costliness, fail to be treated timely and effectively and cause death.
Summary of the invention
Purpose of the present invention just is to provide the application of isosulfocyanate compound in the leukemic medicine of preparation treatment.
Isosulfocyanate compound of the present invention can be isothiocyanic acid benzyl ester (BITC), isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN) or glycosides oil three eruciates (Erucin).
Isosulfocyanate compound of the present invention can directly be buied from market, also can make by following reaction: promptly adopt halogenated hydrocarbons and rhodanate generation nucleophilic substitution, make isothiocyanate (general formula is RSCN), reaction equation is:
RX+R’SCN→RSCN+R’X
Wherein R is a benzyl, and then RSCN is isothiocyanic acid benzyl ester (BITC); R is a phenethyl, and then RSCN is isothiocyanic acid phenylethylester (PEITC); R be acrylic then RSCN be isothiocyanic acid acrylic ester (AITC); R is a first sulfonyl butyl, and then RSCN is Sulforaphane (SFN); R is a first sulfo-butyl, and then RSCN is glycosides oil three eruciates.
Above-mentioned reaction adopts tetrabutyl ammonium bromide to cook phase transfer catalyst, carries out phase-transfer-catalyzed reactions by sodium rhodanate and chloric ether in aqueous solution, and preparation alkyl sulfocyanic ester intermediate heats up then, refluxes, and makes the sulfocyanic ester isomery become isothiocyanate.
Above-mentioned isosulfocyanate compound can be prepared into soft gelatin capsule by method well known in the art when being prepared into the leukemic medicine of treatment.
Above-mentioned soft gelatin capsule can be made every ball and contain 100,200 milligrams of two kinds of specifications of effective composition isosulfocyanate compound.
Above-mentioned leukemia comprises dissimilar acute leukemia (comprising acute myeloid leukaemia, acute T leukemic lymphoblastoid and acute promyelocytic leukemia etc.) and chronic leukemia.
Detailed directions, the consumption of above-mentioned soft gelatin capsule in the treatment leukemia is:
Oral, once a day, the effective dose of AITC, BITC, PEITC, SFN and Erucin by body weight for humans be respectively 10.0,7.5,7.0,16.5,10.5mg/kg.
In order to understand the present invention better, will the present invention be described by the pharmacological testing and the result thereof of various isosulfocyanate compounds below.
The purposes of isosulfocyanate compound in the leukemic medicine of preparation treatment, it is as follows at the leukemic mechanism of action:
One, the isothiocyanic acid benzyl ester (BITC) among the present invention, isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN) or glycosides oil three eruciates (Erucin) all can be induced the acute leukemia cells apoptosis, and have tangible dose-effect relationship and time-effect relationship
Acute leukemia cells (U937 or Jurkat cell) after above-mentioned five kinds of isothiocyanates are handled, adopts two stainings of AnnexinV/PI and flow cytometer to detect apoptosis respectively.Result of study shows, above-mentioned five kinds of isothiocyanates all can be induced acute leukemia cells generation apoptosis and be existed tangible dose-effect relationship (to see Fig. 1-1A, Fig. 1-2 A, Fig. 1-3A, Fig. 1-4A and Fig. 1-5A).Wherein acute T leukemic lymphoblastoid Jurkat cell is more responsive to isothiocyanic acid benzyl ester (BITC) and Lay abdomen sulfane (SFN), acute myeloid leukaemia U937 cell then to isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), and glycosides oil three eruciates (Erucin) more responsive.BITC, PEITC, AITC, SFN and Erucin induce the maximum effect dosage of acute leukemia cells generation apoptosis to be respectively 8 μ M, 8 μ M, 20 μ M, 40 μ M, reach 20 μ M.
The time-effect relationship result of study shows, these five kinds of isothiocyanate compounds cause that the maximum effect time of acute leukemia cells generation apoptosis also is inequalityly (to see Fig. 1-1B, Fig. 1-2 B, Fig. 1-3B, Fig. 1-4B and Fig. 1-5B).For example, just caused about 55% cell generation apoptosis in 6 hours behind the BITC processing Jurkat cell, reached maximum effect after 12 hours substantially and promptly caused about 70% cell generation apoptosis.PEITC has similar time dependence with Erucin, be that PEITC and Erucin processing U937 cell just causes about 60% and 40% cell generation apoptosis respectively after 3 hours, reached maximum effect after 6 hours substantially and promptly caused about 80% and 75% cell generation apoptosis respectively.SFN handles behind the Jurkat cell and just to cause nearly 50% cell generation apoptosis in 6 hours, and 12 hours and 24 hours just cause about 60% and 85% cell generation apoptosis respectively.And AITC handles behind the U937 cell and only to cause about 30% apoptosis in 6 hours, causes in 12 hours about 40% apoptosis to reach maximum effect and promptly to cause about 55% cell generation apoptosis in 24 hours.
Two, five kinds of isothiocyanate compounds among the present invention all can comprise Caspase-3 by apoptosis-induced protease (Caspase), Caspase-7, Caspase-8, the increase of activation/degraded of Caspase-9, and promote its degraded to increase to apoptotic effect substrate PARP (polyADP-ribose polymerase, poly polymerase)
Adopt Western Blot method to study above-mentioned five kinds of isothiocyanates apoptotic proteins enzyme (Caspase) in the acute leukemia cells is comprised Caspase-3, Caspase-7, Caspase-8, Caspase-9, and the influence of apoptotic effect substrate PARP expression.Result of study shows (sees Fig. 1-1C, Fig. 1-2 C, Fig. 1-3C, Fig. 1-4C, Fig. 1-5C, 1-1D, Fig. 1-2 D, Fig. 1-3D, Fig. 1-4D and Fig. 1-5D), above-mentioned five kinds of isothiocyanates all can comprise Caspase-3 by apoptosis-induced protease (Caspase), Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded of apoptotic effect substrate PARP increase, and there are tangible dose-effect relationship and time-effect relationship.
Three, BITC, PEITC, SFN reaches the expression that Erucin all can suppress anti-apoptotic albumen Mcl-1, and AITC is the expression of inducing cell cycle regulatory protein p21 then
Apoptosis is the programmed cell death mode that is subjected to series of genes control.The Bcl-2 family gene is the class protein of studying at most in the gene participating in apoptosis research, and it important role to these Bcl-2 family genes in the regulating cell apoptotic process.The Bcl-2 family gene mainly is divided into two classes: a class is anti-apoptotic albumen such as Bcl-2, Bcl-xL, Mcl-1, XIAP etc.; Another kind of then is short apoptosis protein such as Bax, Bad, Bim, Bid etc.Induce mechanism of action in the acute leukemia cells apoptosis in order to inquire into the Bcl-2 family gene at isosulfocyanate compound, we have studied above-mentioned five kinds of isothiocyanates and have handled the influence of the Bcl-2 family gene being expressed behind the acute leukemia cells.Result of study shows (sees Fig. 2-1A, Fig. 2-3A, Fig. 2-4A, Fig. 2-5A, 2-1B, Fig. 2-3B, Fig. 2-4B and Fig. 2-5B), BITC, PEITC, SFN, all the expression of anti-apoptotic proteins Mcl-1 be can suppress with Erucin, and tangible dose-effect relationship and time-effect relationship existed.AITC does not then influence the expression of Mcl-1.Result of study shows that also these five kinds of isothiocyanates all do not influence other Bcl-2 family gene such as XIAP, Bcl-2, Bcl-xL, Bax, Bad, the change that Bim etc. express.The above results shows that anti-apoptotic proteins Mcl-1 may be at BITC, PEITC, and SFN and Erucin induce in the acute leukemia cells apoptotic process and play an important role.
The inventor has also studied the influence that above-mentioned isosulfocyanate compound cell cycle modulin p21 expresses.P21 has the CDK inhibitor of extensive kinase inhibiting activity in the cell, it can combine with the cyclinD2/cdk4 complex, suppresses the proteic function of its phosphorylation Rb, thereby makes cell cycle arrest in the G1 phase, suppresses cell proliferation.P21 is also in close relations with apoptosis.In order to inquire into the influence that p21 expresses p21 after isosulfocyanate compound induces mechanism of action in the acute leukemia cells apoptosis, the inventor to study above-mentioned five kinds of isothiocyanates to handle acute leukemia cells.Result of study shows (sees that Fig. 2-2A and Fig. 2-2B), AITC can induce the expression of p21, and has tangible dose-effect relationship and time-effect relationship, and BITC that PEITC, SFN and Erucin then do not influence the change that p21 expresses.
Four, above-mentioned five kinds of isothiocyanates all can cause the reduction of Akt phosphorylation level, and the increase of JNK phosphorylation level
The PI3K/Akt/mTOR signal transduction pathway plays an important role in tumor development, and the activation of this path can suppress the apoptosis that multiple stimulation is brought out, and promotes the cell cycle progress, thereby promotes the existence and the propagation of cell.At present, the PI3K/Akt/mTOR signal transduction pathway has become the very promising target spot of prevention and treatment cancer.The former activated protein kinase of mitogenesis (Mitogen-activated protein kinases, MAPKs) the important pheron signaling system of a class of family's to be eukaryotic cell transducer cell external signal cause in cell cell effect.In mammalian cell, found at least 4 types MAPK approach at present, be extracellular signal adjusting protein kinase pathway (extra cellular regulated proteinkinases, ERK), c-Jun amino terminal kinases (c-JunN-terminal kinase, JNK), big silk splits plain activated protein kinase (BMK1) path for P38MAPK path and ERK5/.Wherein first three bar approach has become the focus of present research.MAPKs path and cell growth and apoptosis are closely related.On the whole, ERK is activated by somatomedin, and the propagation and the survival of pair cell play important effect.On the other hand, p38 and JNK path can cause apoptosis.
Induce mechanism of action in the acute leukemia cells apoptosis, the inventor to study after five kinds of isothiocyanates are handled acute leukemia cells influence in order to inquire into signal transduction pathways such as PI3K/Akt/mTOR and MAPK at isosulfocyanate compound to phosphorylations such as Akt and JNK (phosphorylation).Result of study shows (sees Fig. 2-1C, Fig. 2-2C, Fig. 2-3C, Fig. 2-4C, Fig. 2-5C, Fig. 2-1D, Fig. 2-2D, Fig. 2-3D, Fig. 2-4D and Fig. 2-5D), five kinds of isothiocyanates all can cause the reduction of Akt phosphorylation level, reach the increase of JNK phosphorylation level and have tangible dose-effect relationship and time-effect relationship.But five kinds of isothiocyanates do not influence the change of mTOR, ERK and p38 phosphorylation level, or cause some irregular variations.Above-mentioned result of study shows that PI3K/Akt and JNK signal path may be induced in the acute leukemia cells apoptosis at isosulfocyanate compound and be played an important role.
Five, isothiocyanate can act on dissimilar acute leukemia cellses, induces its apoptosis
Whether other acute leukemia cells is also had effect in order to inquire into isothiocyanate, the inventor has studied the influence of above-mentioned five kinds of isothiocyanates to three kinds of dissimilar acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) apoptosis.Result of study show (see Fig. 3-1A, Fig. 3-2A, Fig. 3-3A, Fig. 3-4A and Fig. 3-5A), though three kinds of acute leukemia cellses to the sensitivity difference of five kinds of isothiocyanates, isothiocyanate all can be induced three kinds of acute leukemia cells generation apoptosis.The Westernblot result of study shows (sees Fig. 3-1B, Fig. 3-2B, Fig. 3-3B, Fig. 3-4B and Fig. 3-5B), five kinds of isothiocyanates can induce all that apoptotic proteins enzyme (Caspase) comprises Caspase-3 in three kinds of acute leukemia cellses, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded of apoptotic effect substrate PARP.Above-mentioned five kinds of isothiocyanates can cause that also the downward modulation (downregulation) of anti-apoptotic proteins Mcl-1 in three kinds of acute leukemia cellses (sees Fig. 3-1C, Fig. 3-2C, Fig. 3-3C, Fig. 3-4C and Fig. 3-5C).At last, above-mentioned five kinds of isothiocyanates can cause that also the reduction of Akt phosphorylation level in three kinds of acute leukemia cellses and the increase of JNK phosphorylation level (see Fig. 3-1D, Fig. 3-2D, Fig. 3-3D, Fig. 3-4D and Fig. 3-5D).
Six, PI3K/Akt inhibitor LY294002 can strengthen above-mentioned isothiocyanate and induces the acute leukemia cells apoptosis
Play important effect in order to confirm that further the PI3K/Akt signal transduction pathway is induced in the acute leukemia cells apoptosis at above-mentioned isothiocyanate, the influence of acute leukemia cells apoptosis is induced in the inhibition that the inventor adopts the pharmacology inhibitor LY294002 of PI3K/Akt to study the PI3K/Akt path to above-mentioned isothiocyanate.Result of study shows (sees that Fig. 4-1A, Fig. 4-2A, Fig. 4-3A, Fig. 4-4A and Fig. 4-5A), after handling acute leukemia cells with LY294002 pretreatment and above-mentioned isothiocyanate, its apoptotic incidence rate obviously increases.Result of study also shows (sees Fig. 4-1B, Fig. 4-2B, Fig. 4-3B, Fig. 4-4B and Fig. 4-5B), pretreatment LY294002 can strengthen the inductive apoptotic proteins enzyme of above-mentioned isothiocyanate (Caspase) significantly and comprise Caspase-3, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded of apoptotic effect substrate PARP.Simultaneously, the pretreatment LY294002 expression that also can strengthen the downward modulation of the anti-apoptotic proteins Mcl-1 that above-mentioned isothiocyanate causes or p21 significantly increases and (sees Fig. 4-1C, Fig. 4-2C, Fig. 4-3C, Fig. 4-4C and Fig. 4-5C).Last result of study also shows (sees Fig. 4-1D, Fig. 4-2D, Fig. 4-3D, Fig. 4-4D and Fig. 4-5D), pretreatment LY294002 also can strengthen the inhibition of above-mentioned isothiocyanate to Akt activation (phosphorylation) significantly, and strengthens the activation of the inductive JNK of above-mentioned isothiocyanate.The above results shows that the PI3K/Akt path is induced in the acute leukemia cells apoptosis at above-mentioned isothiocyanate and played important effect.
Seven, the overexpression of Akt can be blocked above-mentioned isothiocyanate and induce the acute leukemia cells apoptosis
The inventor also sets up overexpression that Akt overexpression or dominant negative acute leukemia cells strain study Akt is induced the acute leukemia cells apoptosis to isothiocyanate influence by the means of molecular cloning.Result of study shows that (see that Fig. 5-1A, Fig. 5-2A, Fig. 5-3A, Fig. 5-4A and Fig. 5-5A), normal cell strain (pcDNA3.1) and Akt dominant cell strain (Akt-DN) are after above-mentioned isothiocyanate is handled, its apoptotic incidence rate obviously increases.But the cell strain of Akt overexpression is after above-mentioned isothiocyanate is handled, and its apoptotic incidence rate is starkly lower than pcDNA3.1 and Akt-DN.The result shows that the Akt overexpression can be blocked the inductive acute leukemia cells apoptosis of above-mentioned isothiocyanate.Result of study also shows (sees Fig. 5-1B, Fig. 5-2B, Fig. 5-3B, Fig. 5-4B and Fig. 5-5B), the Akt overexpression can also be blocked the inductive apoptotic proteins enzyme of above-mentioned isothiocyanate (Caspase) and comprise Caspase-3, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded of apoptotic effect substrate PARP.Simultaneously, the Akt overexpression can also be blocked the downward modulation of the anti-apoptotic proteins Mcl-1 that above-mentioned isothiocyanate causes, or the expression of p21 (is seen Fig. 5-1C, Fig. 5-2C, Fig. 5-3C, Fig. 5-4C and Fig. 5-5C).At last, and Westernblot result (see Fig. 5-1D, Fig. 5-2D, Fig. 5-3D, Fig. 5-4D and Fig. 5-5D) demonstration, the cell strain of Akt overexpression, its total Akt and Akt phosphorylation level are apparently higher than normal cell strain (pcDNA3.1).PcDNA3.1 and Akt-DN cell strain are after above-mentioned isothiocyanate is handled, and its Akt phosphorylation level obviously reduces, and the JNK phosphorylation level obviously increases.But its Akt phosphorylation level of the cell strain of Akt overexpression is not subjected to the influence of Akt isothiocyanate, and the JNK phosphorylation is subjected to blocking fully.The above results confirms further that the PI3K/Akt path is induced in the acute leukemia cells apoptosis at above-mentioned isothiocyanate and plays important effect.PI3K/Akt may become above-mentioned isothiocyanate prevention and treat leukemic very promising target spot.
Eight, jnk inhibitor SP600125 can block the inductive acute leukemia cells apoptosis of above-mentioned isothiocyanate significantly
Above-mentioned result of study shows, can induce the activation of JNK signal path behind the above-mentioned isothiocyanate processing acute leukemia cells, and prompting JNK signal path may be induced in the acute leukemia cells apoptosis at above-mentioned isothiocyanate and be played an important role.Induce the mechanism of action in the acute leukemia cells apoptosis in order further to illustrate the JNK signal path at above-mentioned isothiocyanate, the influence of acute leukemia cells apoptosis is induced in the inhibition that the inventor adopts the pharmacology inhibitor SP600125 of JNK to study the JNK signal path to above-mentioned isothiocyanate.Result of study shows (sees that Fig. 6-1A, Fig. 6-2A, Fig. 6-3A, Fig. 6-4A and Fig. 6-5A), pretreatment SP600125 has obviously blocked the inductive acute leukemia cells apoptosis of above-mentioned isothiocyanate.Result of study also shows (sees Fig. 6-1B, Fig. 6-2B, Fig. 6-3B, Fig. 6-4B and Fig. 6-5B), pretreatment SP600125 has also obviously blocked the inductive apoptotic proteins enzyme of above-mentioned isothiocyanate (Caspase) and has comprised Caspase-3, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded of apoptotic effect substrate PARP.Pretreatment SP600125 has also obviously blocked the downward modulation of the anti-apoptotic proteins Mcl-1 that above-mentioned isothiocyanate causes or the expression of p21 and (has seen Fig. 6-1C, Fig. 6-2C, Fig. 6-3C, Fig. 6-4C and Fig. 6-5C).At last, the pretreatment SP600125 activation of also obviously having blocked the inductive JNK of above-mentioned isothiocyanate (is seen Fig. 6-1D, Fig. 6-2D, Fig. 6-3D, Fig. 6-4D and Fig. 6-5D).The above results confirms that further the JNK signal path is induced in the acute leukemia cells apoptosis at above-mentioned isothiocyanate and played an important role.
Nine, the overexpression of JNK can strengthen above-mentioned isothiocyanate and induces the acute leukemia cells apoptosis
The overexpression that JNK is studied in the acute leukemia cells strain that the inventor also sets up the JNK overexpression by the means of molecular cloning is induced the influence of acute leukemia cells apoptosis to above-mentioned isothiocyanate.Result of study shows that (see that Fig. 7-1A, Fig. 7-2A, Fig. 7-3A, Fig. 7-4A and Fig. 7-5A), normal cell strain (pcDNA3.1) is after above-mentioned low concentration isothiocyanate is handled, its apoptotic incidence rate is slight increase (about 25%) only.But the cell strain of JNK overexpression (JNK1 and JNK1/2) is after the isothiocyanate of identical low concentration is handled, and its apoptotic incidence rate obviously increases.Result of study also shows (sees Fig. 7-1B, Fig. 7-2B, Fig. 7-3B, Fig. 7-4B and Fig. 7-5B), the JNK overexpression can strengthen the inductive apoptotic proteins enzyme of isothiocyanate (Caspase) significantly and comprise Caspase-3, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded of apoptotic effect substrate PARP.The JNK overexpression also can strengthen the downward modulation of the Mcl-1 that above-mentioned isothiocyanate causes significantly and (see Fig. 7-1C, Fig. 7-3C, Fig. 7-4C and Fig. 7-5C).At last, the activation that also can strengthen the JNK that above-mentioned isothiocyanate causes significantly of JNK overexpression (is seen Fig. 7-1D, Fig. 7-2C, Fig. 7-3D, Fig. 7-4D and Fig. 7-5D).Above-mentioned result of study confirms further that the JNK signal path is induced in the acute leukemia cells apoptosis at above-mentioned isothiocyanate and plays an important role.JNK may become above-mentioned isothiocyanate prevention and treat leukemic very promising target spot.
According to above-mentioned result of study, beneficial effect of the present invention is:
1, the invention provides a new medical usage of isosulfocyanate compound, i.e. application in the leukemic medicine of preparation treatment;
2, with the leukemic medicine of isosulfocyanate compound preparation treatment provided by the invention, its pharmacological action is strong, has the characteristics of wide spectrum, specially good effect and low toxicity, has good prospect in medicine; And above-mentioned leukemia comprises dissimilar acute leukemia (comprising acute myeloid leukaemia, acute T leukemic lymphoblastoid and acute promyelocytic leukemia etc.) and chronic leukemia;
3, the isosulfocyanate compound among the present invention does not have obvious influence to normal cell, only acts on tumor cell.Five kinds of isothiocyanates all can be induced acute leukemia and chronic leukemia patient peripheral blood cells and myelomonocyte generation apoptosis (seeing embodiment 1, embodiment 2 among the present invention);
In a word, the above-mentioned treatment leukemia medicament that above-mentioned isosulfocyanate compound can be prepared into specially good effect, wide spectrum, low toxicity that studies show that.
Description of drawings
Fig. 1-1A detects the dose-effect relationship that isothiocyanic acid benzyl ester (BITC) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-1B detects the time-effect relationship that isothiocyanic acid benzyl ester (BITC) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-1C is the dose-effect relationship of isothiocyanic acid benzyl ester (BITC) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 1-1D is the time-effect relationship of isothiocyanic acid benzyl ester (BITC) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 2-1A is the dose-effect relationship that isothiocyanic acid benzyl ester (BITC) suppresses the expression of anti-apoptotic proteins Mcl-1; Fig. 2-1B is the time-effect relationship that isothiocyanic acid benzyl ester (BITC) suppresses the expression of anti-apoptotic proteins Mcl-1; Fig. 2-1C is that isothiocyanic acid benzyl ester (BITC) is handled behind the acute leukemia cells dose-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 2-1D is that isothiocyanic acid benzyl ester (BITC) is handled behind the acute leukemia cells time-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 3-1A is an isothiocyanic acid benzyl ester (BITC) to the influence of three kinds of dissimilar acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) apoptosis; To be isothiocyanic acid benzyl ester (BITC) comprise Caspase-3 to apoptotic proteins enzyme (Caspase) in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) to Fig. 3-1B, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded situation of apoptotic effect substrate PARP; Fig. 3-1C is an isothiocyanic acid benzyl ester (BITC) to the influence of anti-apoptotic proteins Mcl-1 in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 3-1D is an isothiocyanic acid benzyl ester (BITC) to the Akt phosphorylation level and the influence JNK phosphorylation level in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 4-1A utilizes the pharmacology inhibitor LY294002 of PI3K/Akt signal transduction pathway to confirm that the inhibition of PI3K/Akt path induces the influence of acute leukemia cells apoptosis to isothiocyanic acid benzyl ester (BITC); Fig. 4-1B is that pretreatment LY294002 comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid benzyl ester (BITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment LY294002 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid benzyl ester (BITC) Fig. 4-1C; Fig. 4-1D is that pretreatment LY294002 suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to isothiocyanic acid benzyl ester (BITC); Fig. 5-1A is Akt overexpression or dominant are induced the acute leukemia cells apoptosis to isothiocyanic acid benzyl ester (BITC) influence; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-1B is that the Akt overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid benzyl ester (BITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; To be the Akt overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid benzyl ester (BITC) Fig. 5-1C; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-1D is that the Akt overexpression suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to isothiocyanic acid benzyl ester (BITC); Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 6-1A utilizes the pharmacology inhibitor SP600125 of JNK signal transduction pathway to confirm that the inhibition of JNK path induces the influence of acute leukemia cells apoptosis to isothiocyanic acid benzyl ester (BITC); Fig. 6-1B is that pretreatment SP600125 comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid benzyl ester (BITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment SP600125 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid benzyl ester (BITC) Fig. 6-1C; Fig. 6-1D is pretreatment SP600125 induces JNK to isothiocyanic acid benzyl ester (BITC) activatory influence; Fig. 7-1A is the JNK overexpression is induced the acute leukemia cells apoptosis to isothiocyanic acid benzyl ester (BITC) influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-1B is that the JNK overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid benzyl ester (BITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; To be the JNK overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid benzyl ester (BITC) Fig. 7-1C; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-1D is the JNK overexpression is induced JNK to isothiocyanic acid benzyl ester (BITC) activatory influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 8-1A handles apoptotic situation behind the mononuclear cell of 6 routine acute myeloid leukaemia (AML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid benzyl ester (BITC); Fig. 8-1B handles apoptotic situation behind the mononuclear cell of 3 routine acute lymphoblastic leukemia (ALL) patient's peripheral bloods or bone marrow for adopting two stainings of AnnexinV/PI and flow cytometer to detect isothiocyanic acid benzyl ester (BITC); Fig. 8-1C handles apoptotic situation behind the mononuclear cell of 3 routine chronic myelogenous leukemia (CML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid benzyl ester (BITC); Fig. 8-1D obtains hematopoietic stem cell (CD34 for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid benzyl ester (BITC) processing 4 routine normal person's bone marrow and separate
+) the apoptotic situation in back; Fig. 8-1E is for adopting the tumor incidence rate after isothiocyanic acid benzyl ester (BITC) and normal saline matched group are handled acute leukemia U937 cell NOD/SCID mice; Fig. 8-1F is for adopting the tumor weight after isothiocyanic acid benzyl ester (BITC) and normal saline matched group are handled acute leukemia U937 cell NOD/SCID mice;
Fig. 1-2 A detects the dose-effect relationship that isothiocyanic acid acrylic ester (AITC) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-2 B detects the time-effect relationship that isothiocyanic acid acrylic ester (AITC) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-2 C is the dose-effect relationship of isothiocyanic acid acrylic ester (AITC) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 1-2 D is the time-effect relationship of isothiocyanic acid acrylic ester (AITC) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 2-2A is the dose-effect relationship of the expression of isothiocyanic acid acrylic ester (AITC) inducing cell cycle regulatory protein p21; Fig. 2-2B is the dose-effect relationship of the expression of isothiocyanic acid acrylic ester (AITC) inducing cell cycle regulatory protein p21; Fig. 2-2C is that isothiocyanic acid acrylic ester (AITC) is handled behind the acute leukemia cells dose-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 2-2D is that isothiocyanic acid acrylic ester (AITC) is handled behind the acute leukemia cells time-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 3-2A is an isothiocyanic acid acrylic ester (AITC) to the influence of three kinds of dissimilar acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) apoptosis; To be isothiocyanic acid acrylic ester (AITC) comprise Caspase-3 to apoptotic proteins enzyme (Caspase) in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) to Fig. 3-2B, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded situation of apoptotic effect substrate PARP; Fig. 3-2C is an isothiocyanic acid acrylic ester (AITC) to the influence of anti-apoptotic proteins Mcl-1 in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 3-2D is an isothiocyanic acid acrylic ester (AITC) to the Akt phosphorylation level and the influence JNK phosphorylation level in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 4-2A utilizes the pharmacology inhibitor LY294002 of PI3K/Akt signal transduction pathway to confirm that the inhibition of PI3K/Akt path induces the influence of acute leukemia cells apoptosis to isothiocyanic acid acrylic ester (AITC); Fig. 4-2B is that pretreatment LY294002 comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid acrylic ester (AITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment LY294002 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid acrylic ester (AITC) Fig. 4-2C; Fig. 4-2D is that pretreatment LY294002 suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to isothiocyanic acid acrylic ester (AITC); Fig. 5-2A is Akt overexpression or dominant are induced the acute leukemia cells apoptosis to isothiocyanic acid acrylic ester (AITC) influence; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-2B is that the Akt overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid acrylic ester (AITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; To be the Akt overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid acrylic ester (AITC) Fig. 5-2C; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-2D is that the Akt overexpression suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to isothiocyanic acid acrylic ester (AITC); Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 6-2A utilizes the pharmacology inhibitor SP600125 of JNK signal transduction pathway to confirm that the inhibition of JNK path induces the influence of acute leukemia cells apoptosis to isothiocyanic acid acrylic ester (AITC); Fig. 6-2B is that pretreatment SP600125 comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid acrylic ester (AITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment SP600125 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid acrylic ester (AITC) Fig. 6-2C; Fig. 6-2D is pretreatment SP600125 induces JNK to isothiocyanic acid acrylic ester (AITC) activatory influence; Fig. 7-2A is the JNK overexpression is induced the acute leukemia cells apoptosis to isothiocyanic acid acrylic ester (AITC) influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-2B is that the JNK overexpression is to the inductive apoptotic proteins enzyme of isothiocyanic acid acrylic ester (AITC)
(Caspase) comprise Caspase-3, Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-2C is the JNK overexpression is induced JNK to isothiocyanic acid acrylic ester (AITC) activatory influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 8-2A handles apoptotic situation behind the mononuclear cell of 6 routine acute myeloid leukaemia (AML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid acrylic ester (AITC); Fig. 8-2B handles apoptotic situation behind the mononuclear cell of 3 routine acute lymphoblastic leukemia (ALL) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid acrylic ester (AITC); Fig. 8-2C handles apoptotic situation behind the mononuclear cell of 3 routine chronic myelogenous leukemia (CML) patient's peripheral bloods or bone marrow for adopting two stainings of AnnexinV/PI and flow cytometer to detect isothiocyanic acid acrylic ester (AITC); Fig. 8-2D obtains hematopoietic stem cell (CD34 for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid acrylic ester (AITC) processing 4 routine normal person's bone marrow and separate
+) the apoptotic situation in back;
Fig. 1-3A detects the dose-effect relationship that isothiocyanic acid phenylethylester (PEITC) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-3B detects the time-effect relationship that isothiocyanic acid phenylethylester (PEITC) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-3C is the dose-effect relationship of isothiocyanic acid phenylethylester (PEITC) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 1-3D is the time-effect relationship of isothiocyanic acid phenylethylester (PEITC) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 2-3A is the dose-effect relationship that isothiocyanic acid phenylethylester (PEITC) suppresses the expression of anti-apoptotic proteins Mcl-1; Fig. 2-3B is the time-effect relationship that isothiocyanic acid phenylethylester (PEITC) suppresses the expression of anti-apoptotic proteins Mcl-1; Fig. 2-3C is that isothiocyanic acid phenylethylester (PEITC) is handled behind the acute leukemia cells dose-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 2-3D is that isothiocyanic acid phenylethylester (PEITC) is handled behind the acute leukemia cells time-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 3-3A is an isothiocyanic acid phenylethylester (PEITC) to the influence of three kinds of dissimilar acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) apoptosis; To be isothiocyanic acid phenylethylester (PEITC) comprise Caspase-3 to apoptotic proteins enzyme (Caspase) in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) to Fig. 3-3B, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded situation of apoptotic effect substrate PARP; Fig. 3-3C is an isothiocyanic acid phenylethylester (PEITC) to the influence of anti-apoptotic proteins Mcl-1 in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 3-3D is an isothiocyanic acid phenylethylester (PEITC) to the Akt phosphorylation level and the influence JNK phosphorylation level in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 4-3A utilizes the pharmacology inhibitor LY294002 of PI3K/Akt signal transduction pathway to confirm that the inhibition of PI3K/Akt path induces the influence of acute leukemia cells apoptosis to isothiocyanic acid phenylethylester (PEITC); Fig. 4-3B is that pretreatment LY294002 comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid phenylethylester (PEITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment LY294002 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid phenylethylester (PEITC) Fig. 4-3C; Fig. 4-3D is that pretreatment LY294002 suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to isothiocyanic acid phenylethylester (PEITC); Fig. 5-3A is Akt overexpression or dominant are induced the acute leukemia cells apoptosis to isothiocyanic acid phenylethylester (PEITC) influence; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-3B is that the Akt overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid phenylethylester (PEITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; To be the Akt overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid phenylethylester (PEITC) Fig. 5-3C; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-3D is that the Akt overexpression suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to isothiocyanic acid phenylethylester (PEITC); Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 6-3A utilizes the pharmacology inhibitor SP600125 of JNK signal transduction pathway to confirm that the inhibition of JNK path induces the influence of acute leukemia cells apoptosis to isothiocyanic acid phenylethylester (PEITC); Fig. 6-3B is that pretreatment SP600125 comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid phenylethylester (PEITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment SP600125 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid phenylethylester (PEITC) Fig. 6-3C; Fig. 6-3D is pretreatment SP600125 induces JNK to isothiocyanic acid phenylethylester (PEITC) activatory influence; Fig. 7-3A is the JNK overexpression is induced the acute leukemia cells apoptosis to isothiocyanic acid phenylethylester (PEITC) influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-3B is that the JNK overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of isothiocyanic acid phenylethylester (PEITC) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; To be the JNK overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 isothiocyanic acid phenylethylester (PEITC) Fig. 7-3C; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-3D is the JNK overexpression is induced JNK to isothiocyanic acid phenylethylester (PEITC) activatory influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 8-3A handles apoptotic situation behind the mononuclear cell of 6 routine acute myeloid leukaemia (AML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid phenylethylester (PEITC); Fig. 8-3B handles apoptotic situation behind the mononuclear cell of 3 routine acute lymphoblastic leukemia (ALL) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid phenylethylester (PEITC); Fig. 8-3C handles apoptotic situation behind the mononuclear cell of 3 routine chronic myelogenous leukemia (CML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid phenylethylester (PEITC); Fig. 8-3D obtains hematopoietic stem cell (CD34 for adopting two stainings of Annexin V/PI and flow cytometer to detect isothiocyanic acid phenylethylester (PEITC) processing 4 routine normal person's bone marrow and separate
+) the apoptotic situation in back; Fig. 8-3E is for adopting the tumor incidence rate after isothiocyanic acid phenylethylester (PEITC) and normal saline matched group are handled acute leukemia U937 cell NOD/SCID mice; Fig. 8-3F is for adopting the tumor weight after isothiocyanic acid phenylethylester (PEITC) and normal saline matched group are handled acute leukemia U937 cell NOD/SCID mice;
Fig. 1-4A detects the dose-effect relationship that Lay abdomen sulfane (SFN) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-4B detects the time-effect relationship that Lay abdomen sulfane (SFN) is induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-4C is the dose-effect relationship of Lay abdomen sulfane (SFN) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 1-4D is the time-effect relationship of Lay abdomen sulfane (SFN) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 2-4A is the dose-effect relationship that Lay abdomen sulfane (SFN) suppresses the expression of anti-apoptotic proteins Mcl-1; Fig. 2-4B is the time-effect relationship that Lay abdomen sulfane (SFN) suppresses the expression of anti-apoptotic proteins Mcl-1; Fig. 2-4C is that Lay abdomen sulfane (SFN) is handled behind the acute leukemia cells dose-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 2-4D is that Lay abdomen sulfane (SFN) is handled behind the acute leukemia cells time-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 3-4A is a Lay abdomen sulfane (SFN) to the influence of three kinds of dissimilar acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) apoptosis; To be Lay abdomen sulfane (SFN) comprise Caspase-3 to apoptotic proteins enzyme (Caspase) in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) to Fig. 3-4B, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded situation of apoptotic effect substrate PARP; Fig. 3-4C is a Lay abdomen sulfane (SFN) to the influence of anti-apoptotic proteins Mcl-1 in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 3-4D is a Lay abdomen sulfane (SFN) to the Akt phosphorylation level and the influence JNK phosphorylation level in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 4-4A utilizes the pharmacology inhibitor LY294002 of PI3K/Akt signal transduction pathway to confirm that the inhibition of PI3K/Akt path induces the influence of acute leukemia cells apoptosis to Lay abdomen sulfane (SFN); Fig. 4-4B is that pretreatment LY294002 comprises Caspase-3 to the inductive apoptotic proteins enzyme of Lay abdomen sulfane (SFN) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment LY294002 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 Lay abdomen sulfane (SFN) Fig. 4-4C; Fig. 4-4D is that pretreatment LY294002 suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to Lay abdomen sulfane (SFN); Fig. 5-4A is Akt overexpression or dominant are induced the acute leukemia cells apoptosis to Lay abdomen sulfane (SFN) influence; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-4B is that the Akt overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of Lay abdomen sulfane (SFN) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; To be the Akt overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 Lay abdomen sulfane (SFN) Fig. 5-4C; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-4D is that the Akt overexpression suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to Lay abdomen sulfane (SFN); Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 6-4A utilizes the pharmacology inhibitor SP600125 of JNK signal transduction pathway to confirm that the inhibition of JNK path induces the influence of acute leukemia cells apoptosis to Lay abdomen sulfane (SFN); Fig. 6-4B is that pretreatment SP600125 comprises Caspase-3 to the inductive apoptotic proteins enzyme of Lay abdomen sulfane (SFN) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; To be pretreatment SP600125 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 Lay abdomen sulfane (SFN) Fig. 6-4C; Fig. 6-4D is pretreatment SP600125 induces JNK to Lay abdomen sulfane (SFN) activatory influence; Fig. 7-4A is the JNK overexpression is induced the acute leukemia cells apoptosis to Lay abdomen sulfane (SFN) influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-4B is that the JNK overexpression comprises Caspase-3 to the inductive apoptotic proteins enzyme of Lay abdomen sulfane (SFN) (Caspase), Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; To be the JNK overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 Lay abdomen sulfane (SFN) Fig. 7-4C; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-4D is the JNK overexpression is induced JNK to Lay abdomen sulfane (SFN) activatory influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 8-4A handles apoptotic situation behind the mononuclear cell of 6 routine acute myeloid leukaemia (AML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect Lay abdomen sulfane (SFN); Fig. 8-4B handles apoptotic situation behind the mononuclear cell of 3 routine acute lymphoblastic leukemia (ALL) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect Lay abdomen sulfane (SFN); Fig. 8-4C handles apoptotic situation behind the mononuclear cell of 3 routine chronic myelogenous leukemia (CML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect Lay abdomen sulfane (SFN); Fig. 8-4D obtains hematopoietic stem cell (CD34 for adopting two stainings of Annexin V/PI and flow cytometer to detect Lay abdomen sulfane (SFN) processing 4 routine normal person's bone marrow and separate
+) the apoptotic situation in back;
Fig. 1-5A detects the dose-effect relationship that glycosides oil three eruciates (Erucin) are induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-5B detects the time-effect relationship that glycosides oil three eruciates (Erucin) are induced acute leukemia cells generation apoptosis for adopting two stainings of AnnexinV/PI and flow cytometer; Fig. 1-5C is the dose-effect relationships of glycosides oil three eruciates (Erucin) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 1-5D is the time-effect relationships of glycosides oil three eruciates (Erucin) to the influence of apoptotic proteins enzyme (Caspase) in the acute leukemia cells and apoptotic effect substrate PARP expression; Fig. 2-5A is the dose-effect relationship that glycosides oil three eruciates (Erucin) suppress the expression of anti-apoptotic proteins Mcl-1; Fig. 2-5B is the time-effect relationship that glycosides oil three eruciates (Erucin) suppress the expression of anti-apoptotic proteins Mcl-1; Fig. 2-5C is that glycosides oil three eruciates (Erucin) are handled behind the acute leukemia cells dose-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 2-5D is that glycosides oil three eruciates (Erucin) are handled behind the acute leukemia cells time-effect relationship to the influence of phosphorylations such as Akt and JNK (phosphorylation); Fig. 3-5A is glycosides oil three eruciates (Erucin) to the influence of three kinds of dissimilar acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) apoptosis; To be glycosides oil three eruciates (Erucin) comprise Caspase-3 to apoptotic proteins enzyme (Caspase) in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell) to Fig. 3-5B, Caspase-7, Caspase-8, activation/degraded of Caspase-9, and the degraded situation of apoptotic effect substrate PARP; Fig. 3-5C is glycosides oil three eruciates (Erucin) to the influence of anti-apoptotic proteins Mcl-1 in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 3-5D is glycosides oil three eruciates (Erucin) to the Akt phosphorylation level and the influence JNK phosphorylation level in three kinds of acute leukemia cellses (acute myeloid leukaemia U937 cell, acute T leukemic lymphoblastoid Jurkat cell, and acute promyelocytic leukemia HL60 cell); Fig. 4-5A utilizes the pharmacology inhibitor LY294002 of PI3K/Akt signal transduction pathway to confirm that the inhibition of PI3K/Akt path induces the influence of acute leukemia cells apoptosis to glycosides oil three eruciates (Erucin); Fig. 4-5B is that pretreatment LY294002 comprises Caspase-3, Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP to glycosides oil three eruciates (Erucin) inductive apoptotic proteins enzymes (Caspase); To be pretreatment LY294002 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 glycosides oil three eruciates (Erucin) Fig. 4-5C; Fig. 4-5D is that pretreatment LY294002 suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to glycosides oil three eruciates (Erucin); Fig. 5-5A is Akt overexpression or dominant are induced the acute leukemia cells apoptosis to glycosides oil three eruciates (Erucin) influence; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-5B is that the Akt overexpression comprises Caspase-3, Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP to glycosides oil three eruciates (Erucin) inductive apoptotic proteins enzymes (Caspase); Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; To be the Akt overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 glycosides oil three eruciates (Erucin) Fig. 5-5C; Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 5-5D is that the Akt overexpression suppresses the activation (phosphorylation) of Akt and the activatory influence of inducing JNK to glycosides oil three eruciates (Erucin); Wherein pcDNA3.1 is the normal cell strain, and Akt-DN is an Akt dominant cell strain, and Akt is the cell strain of overexpression; Fig. 6-5A utilizes the pharmacology inhibitor SP600125 of JNK signal transduction pathway to confirm that the inhibition of JNK path induces the influence of acute leukemia cells apoptosis to glycosides oil three eruciates (Erucin); Fig. 6-5B is that pretreatment SP600125 comprises Caspase-3, Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP to glycosides oil three eruciates (Erucin) inductive apoptotic proteins enzymes (Caspase); To be pretreatment SP600125 induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 glycosides oil three eruciates (Erucin) Fig. 6-5C; Fig. 6-5D is the activatory influence that pretreatment SP600125 induces JNK to glycosides oil three eruciates (Erucin); Fig. 7-5A is the JNK overexpression is induced the acute leukemia cells apoptosis to glycosides oil three eruciates (Erucin) influence; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-5B is that the JNK overexpression comprises Caspase-3, Caspase-7, Caspase-8, the influence of the degraded of the activation/degraded of Caspase-9 and apoptotic effect substrate PARP to glycosides oil three eruciates (Erucin) inductive apoptotic proteins enzymes (Caspase); Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; To be the JNK overexpression induce the downward modulation of anti-apoptotic proteins Mcl-1 or increase the influence of the expression of p21 glycosides oil three eruciates (Erucin) Fig. 7-5C; Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 7-5D is the activatory influence that the JNK overexpression is induced JNK to glycosides oil three eruciates (Erucin); Wherein pcDNA3.1 is the normal cell strain, and JNK1 and JNK1/2 are the cell strain of overexpression; Fig. 8-5A handles apoptotic situation behind the mononuclear cell of 6 routine acute myeloid leukaemia (AML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect glycosides oil three eruciates (Erucin); Fig. 8-5B handles apoptotic situation behind the mononuclear cell of 3 routine acute lymphoblastic leukemia (ALL) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect glycosides oil three eruciates (Erucin); Fig. 8-5C handles apoptotic situation behind the mononuclear cell of 3 routine chronic myelogenous leukemia (CML) patient's peripheral bloods or bone marrow for adopting two stainings of Annexin V/PI and flow cytometer to detect glycosides oil three eruciates (Erucin); Fig. 8-5D obtains hematopoietic stem cell (CD34 for adopting two stainings of Annexin V/PI and flow cytometer detection glycosides oil three eruciates (Erucin) to handle 4 routine normal person's bone marrow and separate
+) the apoptotic situation in back;
Among the figure: control all represents matched group.
The specific embodiment
Below in conjunction with accompanying drawing, by concrete test the present invention is described, but content of the present invention is not limited to this fully.
Gather 6 routine acute myeloid leukaemias (AML), 3 routine acute lymphoblastic leukemia (ALL) and 3 routine chronic myelogenous leukemia (CML) patient's peripheral blood cells or bone marrow, separate obtaining mononuclear cell.Gather 4 routine normal person's bone marrow simultaneously and separate and obtain hematopoietic stem cell (CD34
+).Adopt two stainings of AnnexinV/PI and flow cytometer to detect among the present invention five kinds of isosulfocyanate compounds and handle apoptotic generation behind the cell, five kinds of compound concentrations are respectively: BITC 8 μ M, AITC 20 μ M, PEITC 8 μ M, SFN 40 μ M, Erucin 20 μ M; Be 24 hours action time.Result of study is seen Fig. 8-1A, Fig. 8-2A, Fig. 8-3A, Fig. 8-4A, Fig. 8-5A, Fig. 8-1B, Fig. 8-2B, Fig. 8-3B, Fig. 8-4B, Fig. 8-5B, Fig. 8-1C, Fig. 8-2C, Fig. 8-3C, Fig. 8-4C and Fig. 8-5C, the result shows that five kinds of isothiocyanates all can induce AML, ALL, with CML cell generation apoptosis,, normal cell (do not see Fig. 8-1D, Fig. 8-2D, Fig. 8-3D, Fig. 8-4D and Fig. 8-5D) but being had obvious influence.The result shows that isothiocyanate has leukemia resisting action widely.
Select 60 of 4-5 NOD/SCID mices in age in week for use, female, and set up acute leukemia U937 cell NOD/SCID mice xenotransplantation tumor model.The right back butt of NOD/SCID mice inoculation 0.2ml acute leukemia U937 cell suspension down (contains cell about 2 * 10
6Individual), be divided into matched group, BITC processed group and PEITC processed group at random by the mice body weight in 3 days behind the inoculating cell.Control group mice is through intraperitoneal injection of saline; The processed group mice is respectively through lumbar injection BITC and PEITC, and dosage is 12 μ mol/ days, injects weekly 3 times, totally 4 weeks.The result shows that BITC and PEITC processed group tumor incidence rate are respectively 42.5% and 45%, (sees Fig. 8-1E and Fig. 8-3E) and normal saline matched group tumor incidence rate is 87.5%; BITC and PEITC processed group tumor weight are 1.33 ± 1.90 and 1.30 ± 1.87, and matched group is that Fig. 8-1F and Fig. 8-3F) (are seen in 6.93 ± 2.98 (P＜0.01).BITC and PEITC have no side effect to mice.Results suggest, BITC and PEITC have the obvious suppression effect to the growth of people's acute leukemia cells NOD/SCID mice xenotransplantation tumor.
By well known to a person skilled in the art that the soft gelatin capsule method prepares the isothiocyanate soft gelatin capsule; Can make the soft gelatin capsule that contains 50% isothiocyanic acid benzyl ester (BITC), isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN) or glycosides oil three eruciates (Erucin) by actual needs in the wherein said soft gelatin capsule, also can strengthen or reduce the content of isothiocyanic acid benzyl ester (BITC), isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN) or glycosides oil three eruciates (Erucin).Use commercially available isosulfocyanate compound, get 1500 gram isothiocyanic acid benzyl esters (BITC), isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN) or glycosides oil three eruciates (Erucin), 418 gram soybean oils, 400 gram gelatin, 400 gram purified water, 70 gram Cera Flavas, 10 gram red ferric oxide, 10 gram iron oxide blacks, 2 gram ethyl hydroxybenzoates, 200 gram glycerol are prepared into soft gelatin capsule by known preparation soft gelatin capsule technology, that is: adopting soybean oil and Cera Flava is that substrate prepares the capsule content; Gelatin adds the suitable quantity of water swelling; Red ferric oxide, iron oxide black add water makes dissolving; Ethyl hydroxybenzoate adds small amount of ethanol makes dissolving standby.Gelatin solution makes dissolving fully in 80 ℃ of heating, adds red ferric oxide, iron oxide black solution, adds glycerol and residue purified water, ethyl hydroxybenzoate liquid, stirs; Other gets soybean oil and Cera Flava is heated to fusion, stir, add monomer isothiocyanate monomer medicine [isothiocyanic acid benzyl ester (BITC), isothiocyanic acid phenylethylester (PEITC), isothiocyanic acid acrylic ester (AITC), Lay abdomen sulfane (SFN) or glycosides oil three eruciates (Erucin)], stir once more, milling treatment of colloid obtains uniform soft capsule content, with the rotation rolling capsule machine suppress soft gelatin capsule.
Claims (2)
1, the application of isosulfocyanate compound in the leukemic medicine of preparation treatment.
2, application according to claim 1, wherein said isosulfocyanate compound are isothiocyanic acid benzyl ester, isothiocyanic acid phenylethylester, isothiocyanic acid acrylic ester, Lay abdomen sulfane or glycosides oil three eruciates.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101822663A (en) * | 2010-05-07 | 2010-09-08 | 天津医科大学总医院 | Application of isothiocyanate to preparation of medicine for preventing and treating medicine-resistance tumor |
| CN103159691A (en) * | 2011-12-19 | 2013-06-19 | 天津市国际生物医药联合研究院 | Preparing method and application of isothiocyanate compound |
| CN106955284A (en) * | 2017-02-27 | 2017-07-18 | 新乡医学院 | Suppress a kind of medicine that leukemic stem cells update and bred by regulating and controlling Shh signal paths |
| CN107007578A (en) * | 2017-02-27 | 2017-08-04 | 新乡医学院 | Application of the sulforaphane in treatment leukemia medicament is prepared |
-
2008
- 2008-12-04 CN CNA2008102332218A patent/CN101474170A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101822663A (en) * | 2010-05-07 | 2010-09-08 | 天津医科大学总医院 | Application of isothiocyanate to preparation of medicine for preventing and treating medicine-resistance tumor |
| CN103159691A (en) * | 2011-12-19 | 2013-06-19 | 天津市国际生物医药联合研究院 | Preparing method and application of isothiocyanate compound |
| CN103159691B (en) * | 2011-12-19 | 2017-05-03 | 天津市国际生物医药联合研究院 | Preparing method and application of isothiocyanate compound |
| CN106955284A (en) * | 2017-02-27 | 2017-07-18 | 新乡医学院 | Suppress a kind of medicine that leukemic stem cells update and bred by regulating and controlling Shh signal paths |
| CN107007578A (en) * | 2017-02-27 | 2017-08-04 | 新乡医学院 | Application of the sulforaphane in treatment leukemia medicament is prepared |
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