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Composite medicine for targeted elimination of HIV/SIV

A drug and fusion protein technology, applied in the field of fusion proteins, can solve problems such as ineffectiveness

Inactive Publication Date: 2012-11-14
THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Thus, as far as the design of this new drug is concerned, the above-mentioned design scheme that tries to use CXCR4 as the target may not be effective for the clearance of other mutant subtypes

Method used

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  • Composite medicine for targeted elimination of HIV/SIV
  • Composite medicine for targeted elimination of HIV/SIV
  • Composite medicine for targeted elimination of HIV/SIV

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Example 1 Obtaining of Gene IL2 Fragment Encoding Recombination Targeting Fusion Protein

[0084] With the recombinant human IL2 gene containing C58S and C126S mutations (use PCR technology to change the DNA coding sequence of cysteine ​​Cysteine ​​at the 58th position of human IL2 into the coding sequence of serine Serine; change the cysteine ​​Cysteine ​​at the 126th position The DNA coding sequence is also changed to the coding sequence of serine Serine. In this way, through the expression of the recombinant plasmid, the derivative of IL2 is obtained, and its 58th and 126th positions are all serine. Such IL2 derivatives can only combine with its receptor The ability to inactivate IL2R without other biological activity.) as a template, the following primers were used to amplify the mutated IL2 gene fragment:

[0085] (1) Primer IL2L: gga ggt cga cca tat ggc acc tac ttc aag ttc, a total of 33mers, containing restriction enzymes SalI and NdeI cutting sites; (SEQ ID NO: ...

Embodiment 2

[0088] Example 2 Obtaining of Gene IL7 Fragment Encoding Recombination Targeting Fusion Protein

[0089] The IL7 gene used in the present invention comes from the human genome (see SEQ ID NO: 3 for its DNA coding sequence), and the codons therein are mainly encoded according to the frequency of codon usage preferred by eukaryotes. To express IL7 in prokaryotes requires Modify its codons. According to the characteristics of the codon usage frequency in the IL7 gene, the present invention uses the method of whole gene synthesis to transform the codon of IL7 so that it conforms to the codon preference of the host Escherichia coli, and the nucleotide sequence of the transformed IL7 is shown in SEQ ID NO: 4, see SEQ ID NO: 5 for the amino acid sequence. In addition, the coding sequence of Linker2 and the HindIII restriction site were fused to the 3' end of IL7 during synthesis, and placed in the vector pMD18-T. The primer sequences used in this embodiment are as follows:

[0090...

Embodiment 3

[0093] Example 3 Obtaining of Gene IL7-SON2 Fragment Encoding Recombination Targeting Fusion Protein

[0094] The present invention uses newIL7L containing restriction site EcoRI as the upstream primer (SEQ ID NO: 26), pSON1 (SEQ ID NO: 27), pSON2 (SEQ ID NO: 28), pSON3 (SEQ ID NO: 29), pSON4 (SEQ ID NO: 30), pSON5 (SEQ ID NO: 31) (including restriction site BamHI) as downstream primers, after 5 rounds of amplification, a fusion fragment containing IL7 coding sequence, SON2 coding sequence and Linker2 totaling 684bp was obtained. The results of agarose gel electrophoresis showed that the target product was obtained. Restriction sites EcoRI and BamHI were introduced into the upstream and downstream of the fusion fragment respectively. The fusion fragment was TA cloned into pGEM-T for sequencing. Sequencing results showed that the cloned fragment was correct, and the positive clone was named pT-IL7-SON2.

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Abstract

The invention relates to composite medicine for targeted elimination of HIV / SIV. Specifically, the invention relates to a fusion protein, which is composed of human cytokine polypeptide IL2, IL7, DNA binding protein SON2 rich in positive amino acids, as well as a co-receptor polypeptide CXCR4 able to achieve specific binding with gp120 and / or gp41 and neutralization of a free HIV virus, and serves as a non-viral vector. The non-viral vector fusion protein and a restrictive toxin gene expression recombinant DNA can be assembled to form the composite medicine. The invention also relates to application of the fusion protein and the composite medicine in elimination of HIV in AIDS (acquired immune deficiency syndrome) patients.

Description

technical field [0001] The present invention relates to a fusion protein and its use. Specifically, the fusion protein involved in the present invention is composed of human interleukin 2, interleukin 7, DNA binding protein SON rich in positive amino acids, and an auxiliary agent that specifically binds to gp120 and / or gp41 and neutralizes free HIV virus. Receptor CXCR4 polypeptide composition, as a non-viral vector. The non-viral vector fusion protein and the restricted expression toxin gene recombinant DNA are assembled into a compound drug, wherein the restricted expression toxin gene recombinant DNA is a toxin gene expression recombinant under the control of HIV 5'LTR; the toxin The gene is a toxin gene that encodes a toxin protein that has a killing effect on cells. The present invention also relates to the use of the fusion protein and the compound drug in eliminating HIV in AIDS patients. Background technique [0002] AIDS, caused by HIV infection, is the most seri...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62A61K48/00A61K47/42A61P31/18
Inventor 卢圣栋卢丽陈伟京李涛杜延平魏强涂新明秦川蒋虹丛喆路金芝佟巍
Owner THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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