CN106084005A - The Al of targeting somatostatin receptor18f NOTA PEG6tATE and its preparation method and application - Google Patents
The Al of targeting somatostatin receptor18f NOTA PEG6tATE and its preparation method and application Download PDFInfo
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- 102000005157 Somatostatin Human genes 0.000 title abstract description 5
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
- A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
- A61K51/04—Organic compounds
- A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K19/00—Hybrid peptides, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes
Abstract
The invention discloses the Al of targeting somatostatin receptor18F‑NOTA‑PEG6TATE and its preparation method and application.The preparation method of this label is: somatostatin derivant TATE be attached by with PEG with bifunctional chelating agent NOTA, generates radioactive label precursor, then uses " one kettle way " to carry out positron radionuclide18F labelling, can obtain18Somatostatin derivant Al of F labelling18F‑NOTA‑PEG6‑TATE.The inventive method can be carried out in acetate buffer liquid system or acetonitrile reaction system, and the method has advantage easy and simple to handle, that mark rate, product purity are high.The positron label of targeting somatostatin receptor prepared by the present invention can be applicable in the preparation of PET/CT molecular probe or anti-tumor medicine, and the synthesis for clinical PET/CT molecular probe provides new thinking.
Description
Technical field
The invention belongs to radiolabeling art, be specifically related to the Al of targeting somatostatin receptor18F-NOTA-PEG6-
TATE and its preparation method and application.
Background technology
In recent years, radioisotope labeling receptor-binding peptides is one of study hotspot of cancer target diagnosis, wherein based on
The tumor imaging of somatostatin receptor (somatostatin receptor, SSTR) receives much concern.SSTR is as G-protein coupling
A member of receptor family, has 5 hypotypes, i.e. SSTR1～5 can be at small cell lung cancer, neuroendocrine tumor
The kinds of tumor cells surface high expresseds such as (neuroendocrine tumor, NET), meningioma and glioma.Based on this, more come
It can be maybe that diagnosing tumor novel targets carries that the most scholars think that searching has the somatostatin endogenous ligand of high-affinity to SSTR
For using for reference.Octreotide (octreotide, OC), is the ring octapeptide [D-Phe-that synthesizes after manually modified of natural somatostatin
Cys-Phe-D-Trp-Lys-Thr-Cys-Thr (ol)], it has high affinity to SSTR2, has slight affinity to SSTR5.
If 3 Phe are replaced by Tyr, the Thr (ol) of one of carbon tip is replaced by Thr, and octreotide can derive as TATE, its amino acid sequence
For: D-Phe-Cys-Tyr-D-Trp-Lys-Thr-Cys-Thr, the affinity of SSTR2 is increased by this derivant further, thus
Increase tumor cell internalization ratio, finally make peptide molecule SSTR high expressed tissue (pancreas, adrenal gland, hypophysis and
The tumor of SSTR high expressed) in have substantially high picked-up.
In American-European countries,111In-DTPA-otreotide (Octreoscan) is the gold mark of diagnosis NETs the most clinically
Standard, but because cyclotron produces111Costly, it extensively applies limited In, and compared with positron radionuclide,111In energy
Relatively low (171keV, 245keV) so that111The spatial resolution that In-DTPA-otreotide is ultimately imaged is relatively low.For positive electricity
For sub-PET imaging,68Ga、64Cu、18F etc. have been used for labelling octreotide derivant.Wherein68The octreotide derivant of Ga labelling
(68Ga-DOTATATE,68Ga-DOTATOC and68Ga-DOTANOC etc.) in neuroendocrine tumor images, there is great potential,
Use the most clinically and substantially increase.But because of68Ge/68The Ga generator daily output is relatively low, the most only can eluting 300-
700MBq, adds and lacks FDA approval68Ga labeled drug and68Ge/68Ga generator,68Ga PET imaging does not the most exist
Wide clinical application.
With radioactive metal nucleic68Ga and64Cu compares,18F application is the most extensive, because it has the relatively long half-life
(t1/2=110min), mental retardation (0.64MeV) and lack the excellent nucleic feature such as scattering.It addition,18F labelled compound is feasible
Delayed imaging, its initial specific activity is higher allows heavy dose to prepare, and18F labelled compound spatial resolution is higher, therefore with putting
Penetrating property18F labeling and growing chalone analog has more obvious advantage.
The most existing multiple radioactivity18The octreotide analog of F labelling is (such as 2-18F-FP-OC, Gluc-Lys ([18F]
FP)-TOCA, Gluc-S-Dpr ([18F] FBOA) TOCA and Cel-S-Dpr ([18F] FBOA) TOCA etc.) divide in neural
Secrete the imaging of tumor.But compound radioactivity preparation process is loaded down with trivial details, productivity is relatively low for these, and its routine clinical use is limited, and
Its preparation process is difficulty with automatization.Such as: Wester in 2003 et al. synthesizes glycosylated Gluc-Lys by prothetic group method
([18F] FP)-TOCA, first synthetic mesophase product18F-NFP, then pass through18The acylation of F, uses18F-NFP and Nα-(1-
deoxy-D-fructosyl)-Lys0-Tyr3-Lys5(Dde)-octreotate reacts, and synthesizes glycosylated Gluc-Lys
([18F] FP)-TOCA, wherein lysine (Lys) can be simultaneously connected with18F-NFP, glycosyl and octreotide derivant, when always synthesizing
Between about 3h, putting productivity is between 20%-30%.
Summary of the invention
PET imaging molecular probe that it is an object of the invention to provide a kind of targeting somatostatin receptor and preparation method thereof
And application.
The technical solution used in the present invention is:
The PET molecular probe of a kind of targeting somatostatin receptor, for Al18F-NOTA-PEG6-TATE, its structural formula is as follows:
PET molecular probe Al18F-NOTA-PEG6-TATE application in the tumor imaging of somatostatin receptor targeting.Bag
Include neuroendocrine tumour and non-neuroendocrine tumor (such as glioma etc.).
PET molecular probe Al18F-NOTA-PEG6-TATE divides in the nerve of the somatostatin receptor targeting of non-Bone tumour
Secrete the application of tumor imaging, or the application in cerebral glioma.
PET molecular probe Al18F-NOTA-PEG6-TATE application in neuroendocrine tumor molecular image diagnoses.
The preparation method of a kind of Small-molecule probe, comprises the following steps: filling18The reaction vessel of F ion adds
AlCl3And the micromolecular compound that bifunctional chelating agent NOTA connects, 90-105 DEG C is reacted 15-25 minute, is then peeled off pure
Change, prepare Al18The Small-molecule probe of F-NOTA labelling.
Preferably, micromolecular compound is selected from micromolecule polypeptide or vitamin.
Preferably, vitamin is folic acid.
Preferably, micromolecular compound is PEG6-TATE。
Preferably, reaction buffer is acetate solution or anhydrous acetonitrile.
Preferably, the concentration of acetate buffer be 0.1-0.5mol/L, pH be 3-6.5.
Preferably, the concentration of acetate buffer be 0.1-0.5mol/L, pH be 3.8-4.2.
Preferably, in acetate buffer, every 0.55 1.11GBq's18F ion adds the AlCl of 6nmol3And
54.5-136nmol NOTA-PEG6-TATE。
Preferably, in anhydrous acetonitrile, every 0.55 1.11GBq's18F ion adds the AlCl of 52-260nmol3And
26.4-264.5nmol NOTA-PEG6-TATE, and 26.4-264.5nmol glacial acetic acid.
The invention has the beneficial effects as follows:
The present invention establishes a kind of method of PET molecular probe preparing targeting somatostatin receptor, applies difunctional chela
The octreotide analog TATE that mixture NOTA modifies with hydrophilic group PEG prepares labelled precursor, is carried out by " one kettle way "18F marks
Note, say, that without intennediate purification step, two-step reaction is carried out in same container, is finally purified, obtain target
Product Al18F-NOTA-PEG6-TATE.This invention labeling method is easy, save time, and putting productivity is better than traditional prothetic group labelling method.
The NOTA-PEG of the present invention6-TATE can pass through Al18The success of F method labelling.Currently preferred reaction buffer is
Acetate solution or anhydrous acetonitrile.In acetate buffer reaction system, control every 0.55 1.11GBq's18In F fluorion
Add the AlCl of 6nmol3And 54.5-136nmolNOTA-PEG6-TATE.Under these conditions, putting of gained target product productivity
Between 70%-100%, in order to carry out heavy dose of labelling.Additionally, the present invention uses anhydrous acetonitrile to be that reaction dissolvent enters first
Row Al18F labelling NOTA-PEG6-TATE, it is thus achieved that success.Under anhydrous acetonitrile reaction system, control every 0.55 1.11GBq
's18F fluorion adds the AlCl of 52-260nmol3With 26.4-264.5nmol NOTA-PEG6-TATE, and 26.4-
264.5nmol glacial acetic acid, products therefrom putting productivity is between 64%-74%.It is 95% with top coal drawing after C18 column purification
Above, whole labelling and purge process can complete in 1 hour.
Lipid experiment records Log P=-2.45 ± 0.38, and Al is described18F-NOTA-PEG6-TATE water solublity
Relatively strong, this developer can stable existence 2 hours in PBS and calf serum；Cellular uptake and excretion experiment prove U87MG cell
Can quickly absorb and drain Al18F-NOTA-PEG6-TATE；This developer is pointed out in biodistribution experiment and Micro PET imaging
Higher at tumor uptake, and mainly pass through renal excretion.
The present invention selects people's glioblastoma multiforme U87MG cell to carry out cellular uptake and excretion experiment, with lotus U87MG tumor Mus
Carrying out biodistribution and Micro PET experiment, result proves Al18F-NOTA-PEG6-TATE is at U87MG tumor tissue and SSTR
In high expressed organ (adrenal gland and pancreas), picked-up is higher, illustrates on U87MG cell really containing SSTR, energy and Al18F-NOTA-
PEG6-TATE carries out specific binding.
Vivo biodistribution distribution experiments and Micro PET image results prompting double kidney picked-up are higher, and liver sausage picked-up is relatively low,
Illustrate that this developer is mainly drained by urinary system, draw this developer hydrophilic stronger conclusion base with lipid experiment
This is consistent.Additionally this developer absorbs relatively low in brain, and prompting developer is by blood brain barrier, and conventional clinically18F-
FDG can absorb higher by blood brain barrier in brain, therefore Al18F-NOTA-PEG6The imaging of the cerebral tumor is better than commonly using by-TATE
's18F-FDG。
Additionally, the present invention verifies developer Al18F-NOTA-PEG6-TATE and somatostatin receptor can carry out specificity knot
Close, therefore this developer can also be used for neuroendocrine tumor imaging, the god of the somatostatin receptor targeting of the most non-Bone tumour
Image through endocrine tumors.
Accompanying drawing explanation
Fig. 1 is Al18F-NOTA-PEG6The structural formula of-TATE；
Fig. 2 is the Al of (a) and (b) after purification after labelling18F-NOTA-PEG6The HPLC collection of illustrative plates of-TATE；
Fig. 3 is Al18F-NOTA-PEG6-TATE stability in PBS and hyclone；
Fig. 4 is U87MG picked-up (a) and eluting (b) Al18F-NOTA-PEG6The curve chart of-TATE；
Fig. 5 is injection Al18F-NOTA-PEG6-TATE is its biodistribution figure in lotus U87MG tumor Mus after 1 hour；
Fig. 6 is lotus U87MG tumor Mus, injection Al18F-NOTA-PEG6-the TATE normal imaging after 1 hour and 2 hours and
Injection precursor NOTA-PEG altogether6-TATE blocks imaging after 1 hour；
Fig. 7 is U87MG (A) and the SABC figure of KB (B) tumor tissue section SSTR expression.
Detailed description of the invention
Below in conjunction with specific embodiment, the present invention is further illustrated, but is not limited thereto.
Use material
All chemical reagent are all bought from the chemical company of specialty, for analytical grade, are not further purified during use.
NOTA-PEG6-TATE[p-SCN-Bn-NOTA-PEG6-(D)-Phe1-c(Cys2-Tyr3-(D)-Trp4-Lys5-Thr6-Cys7)-
Thr8] by this laboratory design, it is intended that certain biotech firm synthesizes, and chemical purity is more than 92%, and through RP-HPLC layer
Analysis method (reverse-phase high-performance liquid chromatography, RP-HPLC) and mass spectral analysis
Confirm.Positron radionuclide18F produces with the cyclotron PET/CTtrace (Siemens, Germany) at this center and obtains, and passes through
After activationLight Accell plus QMA anion-exchange column.Reversed phase extraction post C18Sep-Pak detached dowel from
Waters company buys (Milford, MA, USA), activates with ethanol and water successively before using.All radioactive compounds
Being all to be analyzed by RP-HPLC (Shimadzu, China), C18 post specification is 5 μm, 250 × 4.6mm, and during analysis, flow velocity is
1ml/min, flowing phase condition is as follows: A buffer is the water containing 0.1% trifluoroacetic acid, and B buffer is containing 0.1% trifluoroacetic acid
Acetonitrile, analysis gradient is 0-18min, 30%B-60%B, 18-20min, 60%B-30%B.
Acetate buffer configuration is as follows:
1) configuration of the acetate buffer of 0.5mol/L (i.e. 0.5M): with 20.5075g sodium acetate (molecular weight: 82.03)
Being dissolved in water, add 80.478mL glacial acetic acid (density 1.049g/mL), constant volume is 500mL, and the acetate being made into 0.5mol/L delays
Rush liquid；
2) configuration of the acetate buffer of 0.1mol/L (i.e. 0.1mM): take the acetate buffer of 60mL 0.5mol/L
It is settled to 300mL with water, is made into the acetate buffer of 0.1mol/L.
All data mean ± standard deviations obtained by experiment in following example represent, result non-matching t is examined
Testing, p is < statistically significant when 0.05.
Embodiment 1 radioactive label
NOTA-PEG6-TATE passes through Al18F chemical method is carried out18F labelling, based on two kinds of diverse reaction systems,
Acetate buffer (pH=4) and acetonitrile, be all to carry out in same reaction vessel, without the product of intermediate product in course of reaction
Raw.
1.1 based on acetate buffer
1ml is taken from this center accelerator18F aqueous solution, about 0.55 1.11GBq (15 30mCi), is passed through activation
After QMA post, then with the normal saline eluting of 0.2ml 0.9%18F ion, in 1.5ml EP pipe, is managed toward EP the most successively
The AlCl of middle addition 3 μ L 2mM3(6nmol) acetate buffer (0.1M, pH=4) and 54.5nmol NOTA-PEG6-TATE
Precursor solution.After sealing, reactant liquor is reacted 20 minutes at 100 DEG C, cooling, and dilute with 0.5ml PBS, then lead to
C18 post after overactivation, first with 5ml deionized water rinsing pillar to remove18F-and Al18F2+, then with the ethanol of 500 μ l 7:3/
Water mixed liquid eluting target compound, co-elute 3 times.The putting productivity of final products and top coal drawing are all by analytical type HPLC
Record, for experiment in vivo and vitro after being diluted by PBS.
1.2 based on acetonitrile reaction system
1ml is taken from this center accelerator18F aqueous solution, about 0.55 1.11GBq (15 30mCi), is passed through activation
After QMA post, then with the 0.5mM K of 0.5ml2CO3Eluant solution, in the reaction bulb of 5ml, adds 0.5ml acetonitriles at 105 DEG C
Azeotropic water removing 3 times, is then sequentially added into the AlCl of 10 μ l 0.026mol/L (260nmol)3Solution and 200 μ l NOTA-PEG6-
TATE precursor solution (dissolving 0.5mg NOTA-TATE, i.e. 52.9nmol with 1ml acetonitrile) adds 30ul glacial acetic acid (52nmol) again,
Reactant liquor reacts 20 minutes at 105 DEG C, and cooling, purification process is ibid.
1.3 radioactive label results are as follows:
Al18F-NOTA-PEG6The structural formula of-TATE is shown in Fig. 1, whole radiolabeling procedures 100-105 DEG C of 60min it
Inside completing, after correction for attenuation, mark rate based on acetate reaction system is between 70%-100%, based on acetonitrile reaction body
System mark rate between 64%-74% (see Fig. 2.Fig. 2 is HPLC based on acetonitrile reaction system figure, and wherein 2-a is reaction
After, and 2-b is to have reacted to scheme through HPLC after purification).After using Sep-Pak C18 post detached dowel purification, HPLC detection,
No matter based on which kind of reaction system, putting purity and be all higher than 98%, specific activity is more than 16.9GBq/umol.
The invention demonstrates that NOTA-PEG6-TATE can pass through Al18F chelating chemical method labelling success, and this novel
Al18F chelating chemistry is more traditional18F prothetic group labelling method putting productivity is high.No matter based on acetate buffer or organic solvent
Acetonitrile, Al18F-NOTA-PEG6-TATE all can labelling success, simply putting of the latter productivity is lower slightly compared with the former.It addition, based on acetonitrile
The labeling method of system needs azeotropic water removing, and the time-consuming relatively acetate systems of total process is long.Can according to circumstances select any of the above-described body
System carries out Al18F chemical labeling.Final marked product can carry out isolated and purified without being purified with HPLC by C18 post.
Embodiment 2 lipid measures
Take the Al of 10 μ l (1.85MBq)18F-NOTA-PEG6-TATE reactant liquor, adds containing 0.5ml n-octyl alcohol and 0.5ml
In the EP pipe of the 1.5ml of phosphate buffer (PBS), at room temperature vortex 2min after sealing, centrifugal under the conditions of 1000r/min
5min.Take out organic facies with sample loading gun successively and each 50 μ l of aqueous phase are placed in V counter tube, measure counting with gamma counter.By public affairs
Formula LogP=lg (counting n-octyl alcohol/counting PBS) calculates the average LogP value of label, in triplicate.
Through measuring Al18F-NOTA-PEG6Log P=-2.45 ± 0.38 of-TATE, illustrates that this molecular probe is bright
Aobvious hydrophilic, therefore can speculate that this probe is drained by urinary system in vivo.
Embodiment 3 vitro stability is tested
Take 20 μ l's (about 3.7MBq)18F-NOTA-PEG6-TATE solution, is respectively placed in the PBS (pH=of 0.5mL
7.24,0.02mol/L) or in hyclone (FBS), it is placed at 37 DEG C and hatches, measure with HPLC respectively after 0.5,1,1.5 and 2h
Its radiochemical purity, to observe its vitro stability.But for hyclone, first use isopyknic dilution in acetonitrile, after sealing
At room temperature vortex 2min, under the conditions of 1000r/min, centrifugal 5min, takes supernatant HPLC and is analyzed.
Al18F-NOTA-PEG6-TATE vitro stability experimental result such as Fig. 3 institute in PBS and hyclone
Showing, as can be seen from Figure 3, this molecular probe hatches 2h top coal drawing still greater than 95% in 37 DEG C of PBS, simultaneously tire cattle
Serum is hatched 2h top coal drawing and is more than 80%, Al is described18F-NOTA-PEG6-TATE major part in vitro is stable existence.
Embodiment 4 cell is cultivated and model is set up
People's glioblastoma multiforme U87MG tumor cell line added with 10% hyclone, 100IU/mL penicillin and
The MEM culture medium of 100IU/ml streptomycin is cultivated, changes culture medium every other day.Cell is containing 5%CO2, temperature 37 DEG C
Humid air is cultivated.When separating with 0.05%EDTA and 0.01mol/L phosphate buffer (pH7.4) after cell confluent cultures ware
Cultivate for further cell.Every male nude mouse right shoulder subcutaneous vaccination 5 × 106Individual U87MG cell, treats that diameter of tumor reaches
To during 0.8 1cm for internal Micro PET experiment (about plantation after 4-5 week), all zooperies follow national standard and
Animal welfare principle.
Embodiment 5 cellular uptake and elution experiments
Every hole 1 × 10 is added in 24 orifice plates5Individual U87MG tumor cell, the most overnight to ensure that it is adherent, removes every other day
Culture medium, every hole adds the Al Han 185KBq18F-NOTA-PEG6The serum-free medium 0.5ml of-TATE, hatches 10 for 37 DEG C, and 30,
Wash 3 times, then with 0.25% trypsin/0.02%EDTA digestion with the PBS of 0.5ml/ hole frost after 60,90,180min
Cell, collects cell suspension γ-calculating instrument measurement count.Cellular uptake data use cell binding force after decay correction, i.e.
Percentage addition agent amount represents, experiment sets 3 Duplicate Samples, is repeated twice.
For cell elution experiments, 24 orifice plates add every hole 1 × 105Individual U87MG tumor cell, 37 DEG C with
185KBq/ hole Al18F-NOTA-PEG6-TATE is hatched 2h altogether and is made its abundant internalization.Then culture medium is removed, with frost PBS punching
Wash 3 times, add serum-free medium 37 DEG C and hatch 0,15,30,60,90,120min.3 are rinsed the most again with frost PBS liquid
Time, finally with 0.25% trypsin/0.02%EDTA peptic cell, collect cell suspension γ-calculating instrument measurement count.Carefully
Born of the same parents' excretion data represents by Cellular retention rate i.e. percentage addition agent amount after decay correction, and experiment sets 3 Duplicate Samples, repeats two
Secondary.Result is shown in Fig. 4.
From Fig. 4 result: U87MG cell can combine Al fast and efficiently18F-NOTA-PEG6-TATE, such as Fig. 4 a institute
Show, and along with the prolongation of incubation time, the cell binding force of U87MG is further enhanced, reaches when hatching 3h by developer
Peak, cell binding force peak is 1.77% ± 0.07%.In cell elution experiments, in initial 10min, Al18F-
NOTA-PEG6-TATE is the fastest in the intracellular excretion of U87MG, Cellular retention rate from 1.35% ± 0.10% being down to 0.41% ±
0.01%, this developer is slow in the intracellular excretion of U87MG afterwards, and 2h end Cellular retention 0.29% ± 0.07% is shown in Fig. 4 b.
Embodiment 6 vivo biodistribution distribution experiments
Vivo biodistribution experiment is to pass through tail vein injection by every lotus U87MG tumor Mus under 2% isoflurane anesthesia
The Al of 3.7MBq (100 μ Ci)18F-NOTA-PEG6-TATE, blocking experiment be by the tail above-mentioned tracer of vein co-injection and
Precursor solution NOTA-PEG6-TATE(10mg/kg).After injection developer, put to death animal after 1h, separate tumor and main dirty
Device is also weighed, and then measures radiocounting, the radioactive uptake per gram of tissue hundred of tumor and normal internal organs with γ-calculating instrument
Dispensing is penetrated dosage (%ID/g) and is represented.
Result is shown in Fig. 5.After Fig. 5 result shows to inject developer 1h, U87MG tumor uptake value (the 3rd pillar in Fig. 5) is
2.43 ± 0.40%ID/g, higher than other normal internal organs, such as blood (1.22 ± 0.09%ID/g), muscle (0.86 ±
0.12%ID/g), brain (0.89 ± 0.09%ID/g) heart (1.68 ± 0.25%ID/g), liver (1.71 ± 0.14%ID/g)
And lungs (2.05 ± 0.15%ID/g), especially muscle and brain.Developer is higher in the picked-up of double kidneys, and up to 4.13 ± 0.02%
ID/g, and gastrointestinal picked-up is relatively low, and Al is described18F-NOTA-PEG6-TATE is to be drained by urinary system, rather than hepatobiliary excretion.Additionally
Visible substantially physiological accumulations in the pancreas expressed at SSTR and adrenal gland, uptake values be respectively 1.06 ± 0.18%ID/g and
3.88 ± 0.88%ID/g, points out Al18F-NOTA-PEG6-TATE and somatostatin receptor have high-affinity.In bone, picked-up is substantially
Increase, up to 67.18 ± 0.78%ID/g, Al is described18F-NOTA-PEG6The most easily there is the existing picture of defluorinate in-TATE.
Blocking experiment display tumor, pancreas and adrenal gland are to Al18F-NOTA-PEG6The specificity of-TATE absorbs in a large number
Can substantially lower in the presence of non-radioactive precursor, prove Al further18F-NOTA-PEG6-TATE has with somatostatin receptor
High-affinity.Additionally this developer absorbs relatively low in brain, and prompting developer is by blood brain barrier, and conventional clinically18F-FDG can absorb higher by blood brain barrier in brain, therefore Al18F-NOTA-PEG6The imaging of the cerebral tumor is better than often by-TATE
?18F-FDG。
And the picked-up of bone has no in the presence of excess precursor and is decreased obviously, illustrate that in bone, high picked-up is due to Al18F-NOTA-
PEG6-TATE there occurs defluorinate in vivo, and has SSTR high expressed in non-bone.Defluorinate at present is also inevitable, but because takes off
Fluorine be very trace, and in human body, itself is fluorine-containing, so there is not safety issue.The Al of the present invention18F-NOTA-
PEG6-TATE can be used for the tumor imaging of somatostatin receptor targeting；The targeting somatostatin receptor of the most non-Bone tumour
Neuroendocrine tumor images.
Embodiment 7Micro PET images
Micro PET acquisition and graphical analysis Siemens Inevon small animal position emission tomography (PET)/CT are carried out.Experimental group lotus glue
Matter tumor U87MG nude mice under 2% isoflurane anesthesia through the Al of tail vein injection 100 μ l about 3.7MBq (100 μ Ci)18F-NOTA-
PEG6-TATE, gathers still image after injection developer 60min and 120min.Blocking-up group is to inject aobvious altogether by tail vein
As agent 3.7MBq (100 μ Ci) and the precursor solution NOTA-PEG of 10mg/kg6-TATE, gathers quiet after injection developer 60min
State image.Laboratory animal is placed in ventricumbent position and is fixed, and persistently sucks different fluorine with the nose cup with connecting tube in videograph process
Alkane makes animal maintain under narcotism, and the body temperature water circulating-heating plate of laboratory animal makes it keep stable.Image is through with CT
After carrying out correction for attenuation and use 2D order subset expectation maximization method to rebuild.Every time after PET scan, use Inevon
Tumor and main organs is delineated on Research Workplace 4.1 software whole body Coronal image in attenuation corrected
Region of interest, measures radioactive uptake value, represents with %ID/g, and animal is put to death after terminating by PET imaging, it then follows animal welfare is former
Then.
Observable Al is imaged by Micro PET/CT18F-NOTA-PEG6The medicine that-TATE is internal in lotus U87MG tumor Mus
For dynamics and cancer target feature.As seen from Figure 6, U87MG tumor tuberosity shows clearly in Micro PET image,
Injection developer is the uptake values of tumor tissue respectively (3.8 ± 0.3) and (4.4 ± 0.4) %ID/g after 1 hour and 2 hours.In mistake
In the presence of amount non-radioactive precursor, tumor uptake is decreased obviously, and near background, is down to (0.33 ± 0.06) %ID/g, card
Real Al18F-NOTA-PEG6-TATE has obvious SSTR targeting to the U87MG tumor tissue that SSTR is positive.Otherwise similar to body
Interior distribution experiments, double kidneys absorb higher, liver and muscle picked-up is relatively low.Vivo biodistribution distribution experiments and Micro PET imaging
Result prompting double kidney picked-up is higher, and liver sausage picked-up is relatively low, illustrates that this developer is mainly drained by urinary system, joins with fat moisture
Coefficient experiment show that this stronger conclusion of developer hydrophilic is basically identical.
Distribution in vivo experimental result is consistent, and Micro PET/CT imaging is further characterized by Al18F-NOTA-PEG6-TATE exists
Internal there occurs defluorinate phenomenon.In normal group, femur uptake values is higher, for (14.4 ± 2.7) %ID/g, and has no in blocking-up group
It is decreased obviously as (14.4 ± 1.8) %ID/g, illustrates that the high picked-up in bone is not due to SSTR high expressed in bone, but there occurs
Defluorinate.
Embodiment 8 immunohistochemical experiment
After imaging terminates, the neck that broken by lotus U87MG tumor Mus is put to death, and dissects out by tumor tuberosity, fixes with 10% formalin,
Paraffin embedding is also cut into slices to 4 μm, and dewax aquation, antigen retrieval, carries out SSTR2 antibody (H-50, SANTA) with DAB coloration method
Immunostaining.Meanwhile, negative control is carried out with non-neuroendocrine tumor-human oral cavity epithelial JEG-3 KB tumor tuberosity.
Showed by immune group result is at SSTR2 high expressed seen from U87MG tumor cell membrane surface, and KB tumor then becomes low table
Reaching, see Fig. 7, these results are consistent with Micro PET image results, and tumor uptake developer Al is described18F-NOTA-PEG6-
TATE is not as non-specific uptake, but caused by tumor cell surface high expressed SSTR2.
Above-described embodiment is the present invention preferably embodiment, but embodiments of the present invention are not by above-described embodiment
Limit, the change made under other any spirit without departing from the present invention and principle, modify, substitute, combine, simplify,
All should be the substitute mode of equivalence, within being included in protection scope of the present invention.
Claims (10)
1. a PET molecular probe for targeting somatostatin receptor, for Al18F-NOTA-PEG6-TATE, its structural formula is as follows:
。
2.PET molecular probe Al18F-NOTA-PEG6-TATE application in the tumor imaging of somatostatin receptor targeting.
3.PET molecular probe Al18F-NOTA-PEG6-TATE application in neuroendocrine tumor molecular image diagnoses.
4. the preparation method of a Small-molecule probe, it is characterised in that comprise the following steps: filling18The reaction vessel of F ion
Middle addition AlCl3And bifunctional chelating agent NOTA connect micromolecular compound, 90-105 DEG C react 15-25 minute, then
Isolated and purified, prepare Al18The Small-molecule probe of F-NOTA labelling.
Preparation method the most according to claim 4, it is characterised in that: micromolecular compound is raw selected from micromolecule polypeptide or dimension
Element.
Preparation method the most according to claim 5, it is characterised in that: micromolecular compound is PEG6-TATE。
Preparation method the most according to claim 4, it is characterised in that: reaction buffer is acetate solution or anhydrous second
Nitrile.
Preparation method the most according to claim 7, it is characterised in that: the concentration of acetate buffer is 0.1-0.5mol/
L, pH are 3-6.5.
Preparation method the most according to claim 6, it is characterised in that: in acetate buffer, every 0.55 1.11 GBq
's18F ion adds the AlCl of 6nmol3And the NOTA-PEG of 54.5-136nmol6-TATE 。
Preparation method the most according to claim 6, it is characterised in that: in anhydrous acetonitrile, every 0.55 1.11 GBq
's18F ion adds the AlCl of 52-260nmol3And 26.4-264.5nmol NOTA-PEG6-TATE, and 26.4-
264.5nmol glacial acetic acid.
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