CN104415739A - Application of styrene-divinylbenzene medical resin - Google Patents

Abstract

The invention discloses an application of styrene-divinylbenzene medical resin. Micro pores with a pore size of 70-140 nanometers are densely distributed on a resin ball of the styrene-divinylbenzene medical resin, wherein a specific surface area of the resin ball is greater than 1000 square meters/gram, and the pore volume of the micro pores of the resin ball is greater than 6.000e-01cc/gram. The application of the styrene-divinylbenzene medical resin is characterized in that the resin is constructed as electron cloud field force, gp120 and gp41 amino acid conserved sequence target site adsorbents, wherein the electron cloud field force is formed by taking the pore size and the additional charge inside the resin as the principle thing; a protein film coats on the surface of the resin, and the specific amino acid short peptides of the loaded gp120 and gp41 amino acid conserved sequences are used as target probes.

Description

The application of styrene-divinylbenzene medical resin

Technical field

The invention belongs to medical macromolecular materials, specifically a kind of electron cloud field force intervenes the target absorption large aperture resin particle of HIV acceptor groups two dimension dynamic structure and the preparation and application of adsorption column.

Background technology

Human immunodeficiency syndrome be cause after a kind of HIV infection with immune system damage for main disease, AIDS is the disease of involving whole body multiple organ system, mucocutaneous, lymph node, eye, respiratory system, digestive system, nervous system, urinary system etc.Except immune system pathology, also comprise multisystem opportunistic infections (as virus, bacterium, fungi and protozoon) and malignant tumour (comprising Kaposi's sarcoma, malignant lymphoma and cervix cancer), constitute the clinical pathology change of AIDS complexity.The WHO report whole world in 2010 survival HIV carriers and AIDS patient totally 3,400 ten thousand, new infeetioa 2,700,000, annual dead 1,800,000 people.Have every day more than 7000 people's new infections, district from all parts of the world all has popular, but more than 97% in, low income country, especially attach most importance to Africa.China CDC estimates, ends to the end of the year 2011, China survival HIV carriers and AIDS patient about 780,000 people, annual new infections person 4.8 ten thousand people, dead 2.8 ten thousand people.Epidemic situation has covered national all provinces, autonomous regions and municipalities, and current China faces AIDS morbidity and dead peak period, and spreads to population by the people at highest risk such as drug abuse, unlicensed prostitute.

HIV belongs to the mankind slow virus group in Retroviridae lentivirus, be divided into 1 type and 2 types, Major Epidemic HIV-1 in current world wide, HIV-1 is 20 body cubic symmetry, diameter about 100 ~ 120nm spheric granules, be made up of core and coating two parts, core comprises two single-stranded RNA chains, core texture albumen and the necessary enzyme of virus replication, containing reverse transcriptase, integrase and protease.HIV-1 is optionally adsorbed on the CD4 acceptor of target cell, under the help of accessory receptor, enters host cell after infecting human body.Through cyclisation and integration, transcribe and translate, assemble, ripe and sprout, form ripe virion.HIV need enter cell by means of the acceptor on permissive cell surface, comprises the first acceptor and Co receptor.After HIV enters human body, in 24-48 hour, arrive regional nodes, within about about 5 days, viral composition can be detected in peripheral blood.Then produce viremia virusemia, cause acute infection.Because human immune system is mainly through for each strain specific antibodies of HIV albumen, the reaction of specific C D4+T lymphocyte immunity and CTL directly or secrete various cell factor (as TNF, interferon etc.), suppress virus replication, but the immune system of body can not remove virus completely, final formation chronic infection, enters incubation period.HIV virus is constantly bred in human body, patient CD4+T LC obviously declines, and many < 200/mm3, HIV plasma viral load obviously raises, showing as HIV related symptoms, various opportunistic infections and tumour, is the terminal stage after infected by HIV.

HIV causes patient to occur body dysfunction, is in progress as AIDS principal immune pathologic basis is that CD4+T lymphocyte quantity reduces.After infected by HIV, in body, CD4+T lymphocyte quantity constantly reduces, be divided into 3 stages: 1. acute infection period: CD4+T lymphocyte quantity is transient rapid minimizing in a short time, most of the infected is without special treatment, and CD4+T lymphocyte number can return to normal level or voluntarily close to normal level; 2. the symptomless infection phase: CD4+T lymphocyte quantity continues slowly to reduce, how between 800 ~ 350/mm3, this periods of months to ten several years phase not etc., does not on average continue about about 8 years; 3. the symptom phase is had: the minimizing faster again of CD4+T lymphocyte, how at below 350/mm3, part end-stage patients are down to below 200/mm3, and reduce fast.T helper cell 1 (Th1) cell is replaced by t helper cell 2 (Th2) cell, antigen presenting cell function is impaired, interleukin 2 produces and reduces and lose antigen reaction activity power, makes the various infection of patient HIV/AIDS Yi Fasheng.

Anti-hiv therapy therapeutic purpose: suppress copying of virus to greatest extent, Save and restore immunologic function, reduces case fatality rate and the diseases related incidence of disease of HIV, improves the quality of life of patient, reduce the propagation of AIDS.HAART (Highly Active Antiretroviral Therapy, HAART) is the most basic methods for the treatment of of AIDS, needs to take medicine throughout one's life.Current anti-retroviral (ARV) medicine has six large classes kind more than 30.Ucleosides reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), protease inhibitors (Pls), integrase inhibitor (raltegravir), fusion enzyme inhibitor (FIs) and CCR5 suppress neat (maraviroc).

Both at home and abroad about the research of aids prevention and treatment is carried out continual, regrettably all the failing for the treatment of of current all about prevention, within 2007, Merck & Co., Inc. announces vaccine development failure, within 2009, NIH declares vaccine research failure, being the failure of vaccine research on the one hand, is the exposure of a large amount of people at highest risk on the other hand.Simultaneously along with the carrying out of antiviral therapy, a large amount of antibody-resistant bacterium occurs, especially the toxic and side effect of antiviral therapy medicine, economic factor, patient compliance decline and treatment lack of standardization, antiviral failed, immuning failure is caused to occur in a large number to clinical failure, the failed initiation functional of serial therapy, there is clinical complication in patient, quality of life declines and disables, and causes a large amount of burden on societys.

For the failure of antiviral therapy, have and consider new methods for the treatment of, wherein blood purification treatment is constantly attempted, but HIV virus presents highly divergent isolate in human body environment, structure is changeable, simple adsorbing therapy is difficult to effectively remove virus, especially the removing being combined virus with CD4 is difficult to realize, be difficult to reach result for the treatment of, the application number that Wuhan University 2012.4.23 applies for is the patent of 201210118840.9, be improved the design of compatibility adsorbing therapy HIV, but, due to gp120 variability, functional group is changeable, affinity antibody is difficult to reach affine effect, especially albumen affinity antibody, passing through sterilization, in pathogenic microorganism inactivation process, albumen affinity receptor is very easy to deactivation, a small amount of remaining acceptor, because between each subunit of gp120 acceptor, molecule dynamic cooperating measure needs power, subunit group is needed to expose with the affine combination of conserved positions, also be single affine unapproachable.

Summary of the invention

The object of the invention is that the difficult point overcoming existing treating AIDS is with not enough, according to electron cloud field force, gp120 acceptor, the protein science feature of gp41 acceptor groups, for gp120, gp41 subunit amino acid conserved sequence is target spot, design height specific amino acid probe, specific binding HIV virus, according to gp120, gp41 subunit is in conjunction with the dynamic translation feature of two-dimensional structure after CD4 acceptor, the mechanics scope of two-dimensional space cooperating measure when going out gp120 specific binding CD4 according to steered molecular dynamics model inspection, the electron cloud field force that the target designing a kind of particular range combines intervenes the ethene-divinylbenzene material macroporous resin microspheres of HIV functional group dynamic structure conversion, microballoon itself is with hollow hole, specific designs pore size, siphon power is produced after meeting AIDS patient's environment, this siphon power can effectively adsorb HIV virion in human internal environment, particle surface wraps up protein envelope and does not destroy human normal structure, with this microballoon for adsorption column prepared by host material, oral granule removes AIDS virus, reach the object for the treatment of.

The medical macroporous absorbent resin of the present invention be using styrene-divinyl benzene resin ball as electron cloud field force, the medical polymeric adsorbent of gp120, gp41 amino acid consensus sequence target site adsorbent framework, its resin balls is gathered micropore, and pore diameter range is in 70-140 nanometer; Resin balls specific area be greater than 1000 square meters/gram; Resin balls Micropore volume is greater than 6.000e-01cc/ gram; Resin inside is the theme with aperture and additional charge, forms electron cloud field force, Surface coating protein film, is loaded into gp120, gp41 amino acid consensus sequence specific amino acid small peptide as target spot probe.Coated with resin can process as required with polarity, shows as negativity, keeps containing more positively charged chloro acid residue with V3 district, reduces the variation of viral high saltation zone, solves the removing of antiviral therapy resistance HIV Strain.

Research finds: pl20 molecular structure feature and functional localization gpl20 molecule hold C to hold can be divided into successively T cell tropism major decision bunch, monocyte tropism major decision bunch, V3 ring, CD4 land, gpl20 and gp41 cutting area, corresponding circle of sensation and cross-film district from the N of protein peptide chain, gpl20 there are 2 domains with strong conserved sequence, play the effect of the binding site of the acceptors such as covering and hiding host cell surface CD4/CCR5/CXCR4; Simultaneously, the c-terminus of gpl20 also has 3 to have certain conservative but discontinuous sequence forms the binding domain of CD4 molecule jointly, between them, section has the amino acid sequence separation of the high frequency of mutation, thus makes immune response be difficult to play effective immune clearance effect for the binding site on gpl20; In addition, gpl20 surface also exists 5 region of variability (variable region, V district), wherein V3 district is the most important, research finds that the Neutralization and crystallization of HIV 1 is present on V3 ring, V3 ring is the critical function district of envelope protein, also be that HIV-I enters the most important link of host cell process simultaneously, V3 ring is atypical circulus, be made up of about 35 ~ 36 amino acid residues, respectively there are one and half Guang chloric acid at two ends, and form disulfide bond each other and connect, its secondary structure is irregular.Research shows that V3 district is containing more positively charged chloro acid residue, and its corresponding co-receptor CXCR4 and CCR5 contains more electronegative amino acid residue, in V3 district, the arginine of N mono-end can affect the interaction with CCR5 with the replacement of the hydrophobic amino acid phenylalanine near top, V3 district.Research also shows that V3 district has 6 amino acid residues to play an important role in the interaction process of gpl20 and cCR5, comprise arginine 303 ~ 11 third chloric acid 333 residue of the base portion being positioned at V3 ring, with the lysine 310 of D lamella being positioned at V3 ring both sides, isoleucine 312, arginine 318 and phenylalanine-3,4-quinone 20t ~.For this relative conserved sequence, only will merge with the polypeptide of gp120 specific binding and Fc fragment or ETA, and then formed only for the specific fusion protein polypeptide of gp120, formed in conjunction with target position, covalent bond.Form resin microsphere specific adsorption site, be that lasting electron cloud field force intervenes gp120 two-dimensional structure, destroy and CD4 receptors bind, reduce lymphocyte under fire with damage, keep HIV virus free state, reach the effect that resin high-efficient cleaning removes.

Gp120-CD4 is combined with Chemokines CC XCR4 or CXCR5 of target cells and forms CD4-gp120-CXCR4/CXCR5 tri-molecular complex, after compound is formed, gp41 exposes, play a part bridge wherein, utilize self hydrophobic effect mediate retroviral cyst membrane and cell membrane fusion, finally cause cell to be destroyed.Prepare the monoclonal antibody (mAb) of anti-HIV-1 gp41 improvement on synthesis gp41-5, the 4 strain mAb obtained can the new target instrument of the space conformation domain of specific bond gp41 core textures, form secondary target to combination, remove the HIV structure that gp120 can not intervene completely, formed and remove HIV dual structure, this is also a bright spot of the present invention, namely can remove the HIV be combined with cd4 cell.

Based on gpl20 functional structure and steered molecular dynamics, detect the required power size that gp120 changes in conjunction with cd4 cell time space two-dimensional structure, abundant utility tree lipid microspheres spinning electron cloud field force, intervene and change gpl20 two-dimensional structure, destroy gpl20 and CD4 receptor binding capacity, this is also specificity of the present invention and uniqueness characteristic.

Gpl20 core texture by X-ray crystal diffraction technology to itself and acceptor CD451] parsing of the crystal complex structure of antibody 17b draws, the core of gpl20 comprises inner domain and outer domains 2 main domains and so-called " bridge lamella " structure, in this crystal complex, gPl20 core texture does not comprise N mono-and C mono-end, V1/V2 and V3 and V4 ring structure, however, this composite structure shows, from gpl20 inner domain, some structural constituent of outer domains and bridge lamella all interacts with CD4 molecule, these interact gpl20 molecule stable at CD4 combination (CD4 mono-bound) conformational state. simultaneously, this composite structure is that analysis gpl20 antigen site and HIV mechanism provide architecture basics.

Steered molecular dynamics is based on the main movement pattern of gp120, main manifestations is inner domain, outer domains, rotation/distortion between bridge lamella and V3 ring, flexing/closedown, the combination of prolongation/compression movement or these motion modes, utilize the mutual transfer capability between the different conformation of essential sub-space overlap algorithm evaluation gp120, result shows, before CD4 combines, state (unliganded) gp120 is better than ability to removing CD4 composite form (CD4-free) and transforming to the ability that composite form (CD4-complexed) transforms, and CD4-freegp120 has the stronger ability to condition conversion before combining than CD4-complexed gp120. with the relation of gp120 dynamic structure and function, calculate the field force size that gp120 Different Dynamic cooperating measure needs, decision tree lipid microspheres electron cloud field force scope, restriction gp120 dynamic structure transfer capability, reach and intervene gp120 two-dimensional space conversion effect, control gp120 and CD4 combines.

Claims (2)

1. the application of styrene-divinylbenzene medical resin, its resin balls is gathered micropore, and pore diameter range is in 70-140 nanometer; Resin balls specific area be greater than 1000 square meters/gram; Resin balls Micropore volume is greater than 6.000e-01cc/ gram; It is characterized in that:

Using this resin as electron cloud field force, gp120, gp41 amino acid consensus sequence target site adsorbent framework, resin is inner based on aperture and additional charge, forms electron cloud field force; The coated protein film of resin surface, is loaded into gp120, gp41 amino acid consensus sequence specific amino acid small peptide as target spot probe.

2. styrene-divinylbenzene medical resin as claimed in claim 1, is characterized in that:

Described coated with resin can process as required with polarity, shows as negativity, keeps containing more positively charged chloro acid residue with V3 district, reduces the variation of viral high saltation zone, solves the removing of antiviral therapy resistance HIV Strain.