Application of combination of PD-1 inhibitor and Peg-IFN-alpha in preparation of medicine for treating CHB which is not clinically cured

Abstract

The invention discloses application of a PD-1 inhibitor combined with Peg-IFN-alpha in preparation of a medicine for treating CHB which is not clinically cured, and belongs to the technical field of biological medicine. The PD-1 inhibitor is combined with Peg-IFN-alpha to treat a CHB patient which is not clinically cured, it is found that the PD-1 inhibitor can block a PD-1 / PD-L1 signal channel, relieve immunosuppression which is not clinically cured through Peg-IFN-alpha treatment, recover T cell activity and improve the HBV removing capacity of an immune system, and in the CHB treatment process, the good curative effect and safety are shown. By monitoring the expression level of target molecules related to nine interferon pathways in peripheral blood of a patient, it is found that the expression of ISG15 may be induced by adding a PD-1 inhibitor, and high expression of OAS1 may be a favorable factor for good treatment response of the combination of the PD-1 inhibitor and Peg-IFN-alpha.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to the use of a PD-1 inhibitor combined with Peg-IFN-α in the preparation of a drug for treating clinically incurable CHB. Background Art

[0002] Chronic hepatitis B (CHB) is one of the main causes of liver cirrhosis and liver cancer. The ideal goal of CHB treatment is to achieve clinical cure, that is, lasting clearance of hepatitis B surface antigen (HBsAg) and suppression of HBV DNA, with or without the appearance of hepatitis B virus surface antibody (HBsAb). This cure can largely block the occurrence of hepatitis B virus (HBV)-related liver cirrhosis and liver cancer, benefiting patients throughout their lives.

[0003] The existing antiviral drugs for CHB mainly include nucleoside (acid) analogs and pegylated interferon-α (Peg-IFN-α). Among them, Peg-IFN-α, as an immunomodulator, has become an important drug for the clinical treatment of CHB to achieve clinical cure by promoting the body's antiviral immune response. However, in actual clinical practice, only some patients can achieve clinical cure under Peg-IFN-α treatment, while most patients cannot achieve HBsAg clearance and need long-term NAs treatment. Therefore, it is urgent to solve the problem that Peg-IFN-α treatment of CHB fails to achieve clinical cure of hepatitis B.

[0004] Programmed death-1 (PD-1) is an immune checkpoint molecule expressed on T cells and belongs to the immunoglobulin superfamily. Its main function is to prevent overreaction of the immune system by inhibiting T cell activity. Summary of the invention

[0005] In order to overcome the shortcomings of the above-mentioned prior art, the purpose of the present invention is to provide a PD-1 inhibitor combined with Peg-IFN-α in the treatment of CHB that has not been clinically cured, so as to solve the technical problem that some CHB patients cannot achieve clinical cure after Peg-IFN-α treatment.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions: The first aspect of the present invention discloses the use of a PD-1 inhibitor combined with Peg-IFN-α in the preparation of a drug for treating CHB.

[0007] Preferably, the CHB is CHB that has not achieved HBsAg clearance after Peg-IFN-α treatment.

[0008] Preferably, the PD-1 inhibitor is sintilimab.

[0009] Preferably, the dosage of PD-1 inhibitor is 1 mg / kg, and the dosage of Peg-IFN-α is 180 μg / w.

[0010] Preferably, PD-1 inhibitors relieve the poor interferon response to Peg-IFN-α treatment by blocking the PD-1 / PD-L1 signaling pathway.

[0011] The second aspect of the present invention discloses the use of a PD-1 inhibitor in the preparation of a drug enhancer, wherein the drug enhancer is a enhancer for CHB where HBsAg clearance cannot be achieved after treatment with Peg-IFN-α.

[0012] The third aspect of the present invention discloses a combined therapeutic drug for treating clinically incurable CHB, comprising a PD-1 inhibitor and Peg-IFN-α.

[0013] Preferably, the PD-1 inhibitor is sintilimab.

[0014] Preferably, a pharmaceutically acceptable excipient is also included.

[0015] Preferably, the combination therapy drug is in the form of tablets, pills, powders, suspensions, gels, emulsions, creams, granules, capsules, suppositories, injections or sprays.

[0016] Compared with the prior art, the present invention has the following beneficial effects: The application of the PD-1 inhibitor combined with Peg-IFN-α provided by the present invention in the preparation of a drug for treating CHB that has not been clinically cured, and the PD-1 inhibitor combined with Peg-IFN-α to treat CHB patients who have not been clinically cured, found that the PD-1 inhibitor can block the PD-1 / PD-L1 signaling pathway, relieve the immunosuppression of poor interferon response in Peg-IFN-α treatment, restore T cell activity, and improve the immune system's ability to clear HBV, and in the process of treating CHB, it showed good efficacy and safety. Further studies have found that the addition of PD-1 inhibitors may induce the expression of interferon-related factor ISG15, and the high expression of related factor OAS1 may be a favorable factor for the good response of PD-1 inhibitors combined with Peg-IFN treatment, which further illustrates that PD-1 inhibitors may have an interactive relationship with the interferon pathway. This scheme can provide new treatment ideas for CHB patients, especially in those patients who have not been clinically cured by conventional Peg-IFN-α treatment, to find a more effective clinical cure scheme. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The flowchart of the experimental design of the PD-1 inhibitor combined with Peg-IFN-α in the treatment of CHB that has not been clinically cured; Figure 2 is a graph showing the HBsAg changes and clearance curves of CHB patients who were not clinically cured after the PD-1 inhibitor combined with Peg-IFN-α of the present invention, and continued Peg-IFN-α alone treatment; wherein A is the HBsAg curve of the patient, the upper left is the HBsAg curve after 12 weeks of treatment with the PD-1 inhibitor combined with Peg-IFN-α, the upper right is the HBsAg curve after 24 weeks of treatment with the PD-1 inhibitor combined with Peg-IFN-α, the lower left is the HBsAg curve after 12 weeks of continued Peg-IFN-α alone treatment, and the lower right is the HBsAg curve after 24 weeks of continued Peg-IFN-α alone treatment. HBsAg curve at 24 weeks of IFN-α monotherapy, red columns represent patients with HBsAg clearance, blue columns represent patients who did not achieve HBsAg clearance, B is the HBsAg cumulative clearance curve, red lines represent patients in the PD-1 inhibitor combined with Peg-IFN-α treatment group, blue lines represent patients who continued to receive Peg-IFN-α monotherapy, C is the HBsAg dynamic change curve, red lines represent patients in the PD-1 inhibitor combined with Peg-IFN-α treatment group, blue lines represent patients who continued to receive Peg-IFN-α monotherapy; Figure 3The figure is a dynamic curve of transaminase in CHB patients who were not clinically cured after the treatment of the PD-1 inhibitor combined with Peg-IFN-α of the present invention; wherein A is alanine aminotransferase (ALT), B is aspartate aminotransferase (AST), C is a comparison of the dynamic changes of serum ALT levels between the HBsAg clearance group and the non-cleared patients at 24 weeks, and D is a comparison of the dynamic changes of serum AST levels between the HBsAg clearance group and the non-cleared patients at 24 weeks, wherein the red line represents the HBsAg clearance patients, and the blue line represents the HBsAg non-cleared patients. DETAILED DESCRIPTION

[0018] In order to enable those skilled in the art to understand the characteristics and effects of the present invention, the following is a general description and definition of the terms and expressions mentioned in the specification and claims. Unless otherwise specified, all technical and scientific terms used in the text are the common meanings understood by those skilled in the art for the present invention. In the event of a conflict, the definition in this specification shall prevail.

[0019] The theories or mechanisms described and disclosed herein, whether correct or incorrect, should not limit the scope of the present invention in any way, that is, the present invention can be implemented without being limited by any specific theory or mechanism.

[0020] In this article, all features such as values, quantities, contents and concentrations defined in the form of numerical ranges or percentage ranges are for simplicity and convenience only. Accordingly, the description of numerical ranges or percentage ranges should be considered to have included and specifically disclosed all possible secondary ranges and individual values ​​within the range (including integers and fractions).

[0021] In this document, unless otherwise specified, “includes,” “including,” “contains,” “has,” or similar terms cover the meanings of “consisting of” and “mainly consisting of,” for example, “A includes a” covers the meanings of “A includes a and other” and “A only includes a.”

[0022] In this document, in order to make the description concise, not all possible combinations of various technical features in various embodiments or examples are described. Therefore, as long as there is no contradiction in the combination of these technical features, the various technical features in various embodiments or examples can be combined arbitrarily, and all possible combinations should be considered to be within the scope of this specification.

[0023] The present invention provides an application of a PD-1 inhibitor combined with Peg-IFN-α in treating clinically incurable CHB. Patients who have been treated with Peg-IFN-α for 48 weeks and whose HBsAg has not been cleared are regarded as clinically incurable CHB patients. According to the patients' treatment willingness, the patients are divided into a PD-1 inhibitor combined with Peg-IFN-α treatment group (experimental group) and a Peg-IFN-α continued treatment group (control group), and the two groups of patients are followed up for 24 weeks. At 12 and 24 weeks of follow-up, the HBsAg quantification, ALT, AST levels and drug-related adverse reactions of the two groups of patients were monitored, and the HBsAg clearance rate was calculated. The efficacy and safety of PD-1 inhibitors combined with Peg-IFN-α in the treatment of CHB that has not been clinically cured were evaluated; patients were evaluated face-to-face or followed up by phone at 1 day, 4 weeks, 12 weeks and 24 weeks after receiving PD-1 inhibitor treatment, and the adverse reactions and severity of the patients were recorded; patients who continued to receive Peg-IFN-α treatment were evaluated face-to-face or followed up by phone at 4 weeks, 12 weeks and 24 weeks of follow-up, and the adverse reactions and severity of the patients were recorded. At 12 and 24 weeks of follow-up, peripheral blood was collected from the two groups of patients, serum was separated, and the concentrations of 9 interferon pathway-related target molecules determined by single-cell sequencing were detected by ELISA to explore the potential mechanism of PD-1 inhibitors combined with Peg-IFN-α in the treatment of CHB patients who have not been clinically cured.

[0024] The following is a specific description of sintilimab injection as a PD-1 inhibitor, combined with Peg-IFN-α to treat patients with CHB who have not been clinically cured, to further illustrate the present invention. It should be understood that these embodiments are only used to illustrate the present invention and are not intended to limit the scope of the present invention. In addition, it should be understood that after reading the contents of the present invention, those skilled in the art can make various changes or modifications to the present invention, and other PD-1 inhibitors also fall within the scope defined by the claims attached to this application.

[0025] In the following examples, the Peg-IFN-α used is a polyethylene glycol interferon α-2b injection purchased from Xiamen Tebao Biotechnology Co., Ltd., with a trade name of Pegbin; the PD-1 inhibitor used is a sintilimab injection purchased from Innovent Biologics (Suzhou) Co., Ltd., with a trade name of Daboshu ; Other raw materials or reagents not specified are all conventional commercial products, and their specifications are conventional specifications in the field. The experimental methods in the following examples without specifying specific conditions are usually carried out under conventional conditions or under conditions recommended by the manufacturer. The instruments and equipment used are conventional instruments and equipment in the field.

[0026] This study was a single-center, non-randomized controlled clinical trial. Figure 1As shown in the figure, patients who still had not cleared HBsAg after 48 weeks of Peg-IFN-α treatment were considered as CHB patients who were not clinically cured. The inclusion criteria of patients were: 18 < age < 65 years old, HBsAg negative, HBsAg < 1500 IU / mL, HBV DNA < 20 IU / mL. The settings of each group during the experiment were: Experimental group: 35 patients with CHB who were not clinically cured were treated with Peg-IFN-α (the conventional adult dose is 180 μg / w) plus 1 mg / kg sintilimab injection. During the experiment, 2 patients were lost to follow-up, and finally 33 patients completed 24 weeks of follow-up.

[0027] Control group: 34 CHB patients who were not clinically cured continued to receive Peg-IFN-α treatment (the conventional adult dose is 180 μg / w). During the experiment, 3 patients were lost to follow-up, and finally 31 patients completed the follow-up for 24 weeks.

[0028] Example 1: Evaluation of the efficacy and safety of Peg-IFN-α plus PD-1 inhibitors in the treatment of CHB patients who have not been clinically cured 1. Efficacy evaluation: In order to explore the HBsAg clearance of patients with uncured CHB after adding PD-1 inhibitors, quantitative detection (Architectassay; Abbott Diagnostics) was used to detect the HBsAg quantitative (detection limit of 0.05 IU / mL), ALT and AST levels of patients in the experimental group who were not clinically cured of CHB at 12 weeks and 24 weeks after adding sintilimab injection. During the treatment, the peripheral blood PD-1 level of patients was monitored by flow cytometry every 12 weeks. If the PD-1 level was higher than the upper limit of normal value, PD-1 inhibitor treatment could be given again.

[0029] At the same time, when patients in the control group who continued to use Peg-IFN-α were followed up for 12 and 24 weeks, a quantitative method (Architect assay; Abbott Diagnostics) was used to detect the HBsAg quantification of the patients (the detection limit was 0.05 IU / mL).

[0030] Table 1 Baseline characteristics of patients in the two groups

[0031] The baseline characteristics of the two groups of patients are shown in Table 1 , and the efficacy evaluation results are shown in Figures 2 and Figure 3Among the 33 patients who received Peg-IFN-α plus PD-1 inhibitors and completed 24 weeks of follow-up, 12.1% (4 / 33) of the patients had HBsAg clearance when PD-1 inhibitors were given and followed up for 12 weeks. At 24 weeks of follow-up, 30.3% (10 / 33) of the patients had HBsAg clearance ( Figure 2A The median time to HBsAg clearance was 12 (7, 20.5) weeks, of which the earliest HBsAg clearance occurred 2 weeks after PD-1 inhibitor injection. Among the patients who achieved HBsAg clearance, a total of 2 cases experienced HBsAg seroconversion. Twelve weeks after PD-1 inhibitor treatment, the median HBsAg level of 33 patients decreased from 0.95 log baseline to 10 IU / mL decreased to 0.37 log 10 IU / mL, but the overall HBsAg level at week 12 was not statistically different from the baseline. 24 weeks after treatment, the median HBsAg level of patients decreased to 0.03 log10 IU / mL, which was significantly different from the baseline ( P =0.023)( Figure 2C ).

[0032] Among the 31 patients who continued to receive Peg-IFN-α treatment, 9.7% (3 / 31) of the patients had HBsAg clearance at 12 weeks of follow-up, and 9.7% (3 / 31) of the patients had HBsAg clearance at 24 weeks of follow-up (A and B in Figure 2). Among the patients who achieved HBsAg clearance, HBsAg seroconversion occurred in 1 patient. When the 31 patients continued to receive Peg-IFN-α treatment and were followed up for 12 and 24 weeks, the overall HBsAg levels of the 31 patients were not statistically different from the baseline ( Figure 2C ).

[0033] At 12 weeks after the addition of PD-1 inhibitors, the overall ALT and AST levels of patients were significantly higher than those at baseline, but returned to the same level as baseline at 24 weeks ( Figure 3 A and B in the figure). The 33 patients were divided into two groups according to whether HBsAg clearance was achieved within 24 weeks. It was found that the ALT level of patients in the HBsAg clearance group was significantly higher than that of the baseline at 12 weeks and returned to the same level as the baseline at 24 weeks. Although the ALT level of patients who did not clear HBsAg was slightly higher than that of the baseline at 12 weeks, there was no significant statistical difference ( Figure 3 After comparing AST levels, it was found that regardless of whether HBsAg was cleared, the AST levels of patients increased significantly at 12 weeks, and the change trends of AST were almost the same ( Figure 3The results suggest that PD-1 inhibitors may activate immune activity in the liver and produce a controllable inflammatory response, which leads to an increase in transaminase levels around 12 weeks after treatment, and the increase in ALT at 12 weeks may be related to the clearance of HBsAg.

[0034] 2. Safety assessment: Adverse reactions in patients with CHB who were not clinically cured during treatment were obtained through face-to-face evaluation or telephone consultation. The adverse reactions in the experimental group (n=33) and the control group (n=31) during the 24 weeks of treatment were statistically collected, and the incidence of adverse reactions during the two treatment methods was compared. Specifically, face-to-face evaluation or telephone follow-up was performed on the experimental group patients who were not clinically cured of CHB on the first day, 4 weeks, 12 weeks, and 24 weeks, and the adverse reactions and severity of the patients in the experimental group were recorded. Face-to-face evaluation or telephone follow-up was performed on the experimental group patients who were not clinically cured on the first day, 4 weeks, 12 weeks, and 24 weeks, and the adverse reactions and severity of the patients in the experimental group were recorded. The results are shown in Table 2.

[0035] Table 2 Incidence of adverse reactions during treatment in patients with CHB who were not clinically cured

[0036] As can be seen from Table 2, during the period of continuing Peg-IFN-α treatment alone, the main adverse reactions observed were neutropenia, thrombocytopenia, fever, fatigue and anorexia. These adverse reactions are considered by researchers to be closely related to Peg-IFN-α treatment and have a high incidence. A few patients developed thyroid dysfunction, rash and hair loss during Peg-IFN-α treatment. Three patients had a sudden increase in ALT levels (ALT>5 ULN). During the treatment of PD-1 inhibitors combined with Peg-IFN-α, patients also experienced adverse reactions such as neutropenia, thrombocytopenia, fever, fatigue and anorexia. Three patients were observed to have oral ulcers during treatment, and this adverse reaction did not occur during Peg-IFN-α treatment. Therefore, it is believed that this adverse reaction may be related to PD-1 inhibitor treatment. It is worth mentioning that neutropenia and thrombocytopenia are long-term adverse reactions during Peg-IFN-α treatment, and these adverse reactions may accompany the entire Peg-IFN-α treatment period. Fever and fatigue often occur in the early stage of Peg-IFN-α treatment, especially in the first 12 weeks. As the treatment time increases, the above adverse reactions of patients can gradually improve. However, we found that during the treatment of PD-1 inhibitors combined with Peg-IFN-α, 12 (36.4%) patients had fever again, and 15 (45.5%) patients had fatigue again. This may be that the PD-1 inhibitor combined with Peg-IFN-α treatment aggravated or evoked the adverse reactions of Peg-IFN-α.

[0037] Example 2: ELISA method to determine the changes in the expression of interferon pathway-related molecules in the peripheral blood of patients treated with PD-1 inhibitors combined with Peg-IFN-α and those who continued to be treated with Peg-IFN-α Single-cell transcriptome sequencing was performed on peripheral blood PBMCs of two patients with CHB who were not clinically cured and received PD-1 inhibitors before and after treatment. The results showed that nine interferon pathway genes were significantly increased, namely IFIT1, MX1, IFI6, ISG15, OAS1, EIF2AK2, STAT1, IRF7 and IFITM3.

[0038] Then, at 12 and 24 weeks of follow-up, peripheral blood was collected from 33 patients who received PD-1 inhibitor combined with Peg-IFN-α treatment and 23 patients who continued Peg-IFN-α treatment alone, and serum was separated. The concentrations of 9 target molecules (IFIT1, MX1, IFI6, ISG15, OAS1, EIF2AK2, STAT1, IRF7 and IFITM3) of the patients were detected using a finished ELISA kit (Sino Best Biological Technology Co., Ltd, Shanghai). The 33 patients who received PD-1 inhibitor combined with Peg-IFN-α treatment were divided into a cured group and an uncured group according to whether HBsAg clearance was achieved during the observation period, in order to analyze the differences between the two groups at the baseline and after PD-1 inhibitor treatment. The test results are shown in Tables 3 to 5.

[0039] Table 3 PD-1 inhibitor combined with Peg-IFN-α treatment

[0040] Table 4 Clinical cure group and uncured group in the treatment of PD-1 inhibitor combined with Peg-IFN-α

[0041] Table 5 Expression levels of 9 target molecules in the Peg-IFN-α treatment group and comparison of the expression levels of 9 target molecules between the two groups at 24 weeks

[0042] As shown in Tables 3 and 4, the expression level of ISG15 increased significantly after PD-1 inhibitor treatment (36.73±7.99 pg / mL vs. 40.67±7.71 pg / mL, p =0.04, and the other 8 cytokines showed no significant changes. It was further found that the baseline of the CHB cured group (202.54±22.63 pg / mL vs. 156.78±38.03 pg / mL, p =0.002) and after PD-1 inhibitor treatment (204.14±22.09 pg / mL vs. 165.78±48.23 pg / mL, p=0.034) had significantly higher OAS1 levels than those in the uncured group. This result suggests that the addition of PD-1 inhibitors may induce the expression of ISG15, and the high expression of OAS1 may be a favorable factor for the good response to PD-1 inhibitors combined with Peg-IFN-α treatment. PD-1 inhibition may interact with the interferon pathway. As can be seen from Table 5, in patients who continued to receive Peg-IFN-α treatment alone, only the expression level of IFI6 was significantly increased after 24 weeks of treatment (1042.64±167.88 pg / mL vs.724.89±202.31 pg / mL, p =0.005), while the other 8 cytokines including ISG15 showed no significant changes. Further comparison of 9 cytokines in peripheral blood of 33 patients treated with PD-1 inhibitors and 23 patients who continued Peg-IFN-α treatment at 24 weeks of follow-up showed that the ISG15 level in the group treated with PD-1 inhibitors was significantly higher than that in the group treated with Peg-IFN-α (45.38±4.33 pg / mL vs. 36.50±6.30 pg / mL, p =0.005), which further indicated that the addition of PD-1 inhibitors may induce the expression of ISG15.

[0043] The above contents are only for explaining the technical idea of ​​the present invention and cannot be used to limit the protection scope of the present invention. Any changes made on the basis of the technical solution in accordance with the technical idea proposed by the present invention shall fall within the protection scope of the claims of the present invention.

Claims

1. Application of PD-1 inhibitor combined with Peg-IFN-α in the preparation of drugs for the treatment of CHB.

2. The use according to claim 1, characterized in that: The CHB is CHB in which HBsAg clearance has not been achieved after treatment with Peg-IFN-α.

3. The use according to claim 1 or 2, characterized in that: The PD-1 inhibitor is sintilimab.

4. The use according to claim 1 or 2, characterized in that: The dosage of PD-1 inhibitor was 1 mg / kg, and the dosage of Peg-IFN-α was 180 μg / w.

5. The use according to claim 1 or 2, characterized in that: PD-1 inhibitors relieve the poor response to interferon in Peg-IFN-α treatment by blocking the PD-1 / PD-L1 signaling pathway.

6. Use of a PD-1 inhibitor in the preparation of a drug enhancer, characterized in that: The drug enhancer is a enhancer for CHB where HBsAg clearance is still not achieved by Peg-IFN-α treatment of CHB.

7. A combined therapeutic drug for treating CHB that has not been clinically cured, characterized in that: Including PD-1 inhibitors and Peg-IFN-α.

8. A combined therapeutic drug for treating clinically incurable CHB according to claim 7, characterized in that: The PD-1 inhibitor is sintilimab.

9. The combined therapeutic drug for treating clinically incurable CHB according to claim 7, characterized in that: Pharmaceutically acceptable excipients are also included.

10. The combined therapeutic drug for treating clinically incurable CHB according to claim 7, characterized in that: The dosage form of the combined therapeutic drug is tablets, pills, powders, suspensions, gels, emulsions, creams, granules, capsules, suppositories, injections or sprays.

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