阅读说明：本技术 Obatoclax在制备HIV-1潜伏感染逆转剂中的应用 (Application of Obatoclax in preparation of HIV-1 latent infection reversal agent ) 是由 李琳 周晨亮 梁太珍 赖芳圆 郑腾羿 陈佩 李楚玉 刘叔文 于 2019-11-12 设计创作，主要内容包括：本发明提供Obatoclax及其药学上可接受的衍生物在制备HIV-1潜伏感染逆转剂中的应用,Obatoclax具有良好的激活HIV-1潜伏细胞库的活性,同时是Bcl-2家族蛋白的抑制剂,可防止HIV潜伏库的补充、促进再活化细胞的死亡,有望实现对HIV真正的“功能性治愈”。(The invention provides an application of Obatoclax and pharmaceutically acceptable derivatives thereof in preparation of HIV-1 latent infection reversal agents, wherein Obatoclax has good activity of activating HIV-1 latent cell banks, is an inhibitor of Bcl-2 family proteins, can prevent the supplement of HIV latent banks and promote the death of reactivated cells, and is expected to realize real 'functional cure' of HIV.)

The application of Obatoclax and pharmaceutically acceptable derivatives thereof in preparing HIV-1 latent infection reversal agents.

2. Use according to claim 1, characterized in that: the effective concentration of Obatoclax is 0.01-2.5 mu M.

3. Use according to claim 1, characterized in that: the pharmaceutically acceptable derivatives of Obatoclax are selected from pharmaceutically acceptable salts or pharmaceutically acceptable esters thereof.

4. Use according to claim 1, characterized in that: the HIV-1 latent infection reversal agent is in a dosage form of oral administration or non-oral administration.

5. Use according to claim 4, characterized in that: the oral administration dosage form is tablet, powder, granule, capsule, emulsion or syrup.

6. Use according to claim 4, characterized in that: the non-oral administration dosage form is injection.

Technical Field

The invention belongs to the technical field of medicines, relates to new application of an existing compound, and particularly relates to application of Obatoclax in preparation of an HIV-1 latent infection reversal agent.

Background

Aids is the most serious single etiological disease of humans caused by Human Immunodeficiency Virus (HIV), and is characterized by severe damage to the systemic immune system. By 2018, the number of people infected with HIV worldwide reaches 3690 ten thousand, and the health and survival of human beings are seriously threatened. In 1996, successful application of highly active antiretroviral therapy (HAART) or combined antiretroviral therapy (cART) converted AIDS from a lethal disease to a chronic controlled disease, greatly prolonging the survival time of AIDS patients. By 2018, 2170 million people worldwide have received cART treatment. However, cART has insurmountable limitations in development to date, such as toxic and side effects due to long-term administration, susceptibility to HIV drug resistance, and high treatment cost for patients. More importantly, the therapy cannot completely cure the patient, and once the drug is stopped, the latent HIV in the patient can rebound rapidly to cause reinfection, so that cART needs to be taken for the whole life, and the 'root chopping' cannot be realized.

It was found that when the virus titer in the plasma of a patient receiving cART treatment had decreased below the detection limit, there was still a low level of viral replication in the patient and the main reason why HIV could not be completely eliminated was the presence of a latent viral reservoir consisting of HIV-latently infected cells in the patient. HIV latency repository is primarily a group of HIV latently infected quiescent memory CD4⁺T cell composition, which is mainly characterized by: 1) after integration, the HIV provirus integrated in the host immune cell genome is subjected to gene silencing, so that the damage of a host immune system and the attack of a cART medicament are avoided, and once the conditions are suitable, the virus can recover the infectivity; 2) infected individuals carry a small number of latent infected cells, but the rate of attenuation is so low that complete elimination by cART therapy alone is not possible. Thus, the existence of a repository of HIV latent virus is a great obstacle to the current clinical treatment that does not completely eliminate HIV.

Then how does the HIV virus remain latent in the patient's body, and how does it evade the attack of antiviral drugs? After HIV is mediated by its envelope proteins, the CD4 receptor on the host target cell, and the chemokine receptor CCR5 or CXCR4 into the host cell, the viral replication cycle begins. Firstly, an HIV pre-integration compound entering a target cell through membrane fusion generates linear and circular double-stranded complementary DNA through the action of reverse transcriptase; subsequently, the linear double-stranded DNA and viral integrase enter the nucleus and are further integrated into the genome of the host immune cell; the integrated HIV proviral DNA, not only serves as a template for transcription of viral genes in infected cells, but is also critical for latent HIV integration.

CD4 due to genome and host memory of HIV⁺The genome of T cells is integrated together, so that how to remove HIV provirus latently infected in immune cells in a host in a targeted manner and prevent disease rebound becomes one of the difficulties of great headache of researchers. In addition, the latent HIV reservoir can escape the immune system surveillance and cART also cannot act on the integrated HIV provirus, which is the biggest obstacle to complete HIV eradication from infected subjects.

In recent years, researchers have generally considered that a method of "leading out of the snake" (Shock and Kill) is adopted, in which HIV latent infection reversing agents (LRAs) are first used to induce resting memory CD4⁺T cells are activated to release HIV, and then HIV virus is screened out in combination with cART. At the same time, CD4 appears with stationary period⁺Activation of T cells, natural or acquired HIV specific immune responses induced by cytopathic effects and/or the same, also enables the half-life of the population of cells to be significantly shortened and then rapidly killed, thereby achieving HIV "functional" cure, i.e., HIV virus is below the detection limit and patient does not experience disease rebound when antiviral drugs are discontinued.

Currently, there are six major groups of latent infection-reversing agents LRAs in clinical research, including: 1) positive transcription elongation factor b (P-TEFb) activator: based on Bromodomain and its external domain (BET) inhibitors, such as JQ1, 1-BET151, etc., which enhance the binding of Tat and PTEF-b to activate HIV latent infection virus; 2) histone Deacetylase (HDAC) inhibitors: vorinostat (saha), Givinostat (ITF2357), Panobinostat, and the like, which are unwinding agents for chromatin, promote binding of viral transcription factors to HIV Long Terminal Repeats (LTR) together with HDAC, and inhibit proviral transcription by promoting deacetylation of lysine residues on histones; 3) protein Kinase C (PKC) activators: prostratin, bryostatin-1, etc. which have very key action on the transcription initiation of latent HIV gene; 4) DNA methyltransferase (DNMT) inhibitors: decitabine, which inhibits the DNA methylation process of provirus; 5) histone Methyltransferase (HMT) inhibitors: EPZ5676, which inhibits the activity of cellular HMT and thus inhibits the methylation of proviral DNA; 6) cytokine immune function modulators: such as IL-2, IL-7, IL-15, GM-CSF, and the like.

Unfortunately, the current treatment of HIV latent infection with the above mentioned LRAs candidates is still only in clinical trial stage, and no breakthrough progress has been made, and they have more or less certain defects, which limits their wide clinical application. The main problems are as follows: 1) some epigenomic therapeutic drugs can also cause a plurality of side effects while activating viruses, for example, the mechanism of the HDAC inhibitor is more controversial, and nausea, vomiting, abnormal blood system, influence on normal gene function and the like are also reported in clinic; 2) PKC activators can affect the transcription factor signaling pathway for normal T cell activation, thereby complicating immune and chemical clearance of the body leading to a latent viral pool; 3) some medicines have poor action specificity, wide action, large toxic and side effects, and are not suitable for long-term application and the like. 4) Many of the above LRAs are still in clinical trials, and even some drugs only have been demonstrated to activate latent viruses in vitro cell experiments, which is a long way away from the wide clinical application.

B-lymphocytoma-2 (B-cell lymphoma-2) is abbreviated as Bcl-2, plays an important role in maintaining normal metabolism of a human body, and is one of the most widely researched apoptosis regulating genes. The Bcl-2 family is largely classified by functional classification into anti-apoptotic proteins (e.g., MCL-1, Bcl-2, Bcl-w, Bcl-xl, and A1) and pro-apoptotic proteins (Bak, Bok, and BH3-only proteins). Among them, Bax causes a decrease in membrane potential and outflow of cytochrome C and Apoptosis Inducing Factor (AIF), thereby inducing apoptosis, and Bcl-2 binds to Bax to form a dimer, thereby inhibiting apoptosis. The existing research shows that Bcl-2 has certain promotion effect on maintaining the steady proliferation of an infected cell line with HIV and the spread of acute infection of primary CD4T cells, and causes obstruction to HIV treatment. Therefore, inhibition of Bcl-2 prevents the recruitment of HIV latency pools and directly promotes the death of reactivated cells, which is beneficial for HIV therapy.

On the basis, if the Bcl-2 inhibitor which further meets the activation effect can be found, the effects of three effects of inventory reduction, activation and killing are integrated, and the clinical research value of functional cure of HIV is great.

Obatoclax (also known as GX15-070, Obatoclax Mesylate) is an inhibitor of Bcl-2 family proteins, and its Mesylate has the chemical structural formula shown in formula I:

obatoclax is generally used for treating lung cancer and has entered clinical phase 3 trial studies on advanced small cell lung cancer, but pharmacological activity of Obatoclax in treatment of other diseases has not been reported yet.

Disclosure of Invention

The invention aims to solve the problems and provides a novel HIV-1 latent infection reversing agent which can activate HIV latent infection cells, promote the HIV latent infection cells to release HIV viruses, inhibit Bcl-2, prevent the supplement of HIV latent pool and hopefully realize 'functional treatment' on HIV.

The technical scheme adopted by the invention is as follows:

the application of Obatoclax and pharmaceutically acceptable derivatives thereof in preparing HIV-1 latent infection reversal agents.

Furthermore, the effective concentration of the Obatoclax is 0.01-2.5 mu M.

Further, the pharmaceutically acceptable derivatives of Obatoclax are selected from pharmaceutically acceptable salts or pharmaceutically acceptable esters thereof.

Further, the HIV-1 latent infection reversal agent is in a pharmaceutical dosage form of oral administration or non-oral administration.

Further, the oral administration dosage form is tablet, powder, granule, capsule, emulsion or syrup.

Further, the non-oral administration dosage form is injection.

Compared with the prior art, the invention applies Obatoclax and pharmaceutically acceptable derivatives thereof to the preparation of HIV-1 latent infection reversal agents, and experiments prove that Obatoclax has good activity of activating HIV-1 latent cell banks; meanwhile, Obatoclax is an inhibitor of Bcl-2 family protein, can prevent the supplement of HIV latent pool and promote the death of reactivated cells, and is expected to realize the real 'functional cure' of HIV.

Drawings

FIG. 1 is a graph showing the effect of ObatoClax on P24 protein expression in a J-Lat 10.6 latently infected cell line;

FIG. 2 is a graph showing the effect of ObatoClax on P24 protein expression in a cell line latently infected with ACH 2;

FIG. 3 is a graph showing the effect of ObatoClax on HIV-1p24 protein expression in ACH2 cells, a model of HIV-1 latently infected cells;

FIG. 4 shows the cytotoxicity of Obatoclax on J-Lat 10.6 cells;

FIG. 5 is a graph showing the cytotoxicity of Obatoclax against ACH2 cells;

FIG. 6 shows the cytotoxicity of Obatoclax on J-Lat A2 cells.

Detailed Description

The inventor finds that Obatocrax has a novel pharmacological effect of activating latent HIV, and the technical scheme of the invention is described by combining specific experiments.