Peripheral blood marker combination for predicting nasopharynx cancer immune curative effect and dynamically monitoring nasopharynx cancer immune curative effect

Abstract

The invention discloses a peripheral blood marker combination for predicting nasopharyngeal carcinoma immune curative effect and dynamically monitoring, and belongs to the technical field of tumor immunotherapy, and the peripheral blood marker combination is characterized in that before immunotherapy, the immune therapy curative effect is predicted at the molecular levels of CD3, CD4, CD8, CD45RA, CCR7 and PD-1 in peripheral blood; after immunotherapy, the molecular levels of CD3, CD4, CD8, CD45RA, CCR7, CD69 and CXCR5 in peripheral blood are subjected to dynamic monitoring of the curative effect of immunotherapy. In the invention, the immunotherapy curative effect is predicted by detecting the molecular levels of CD3, CD4, CD8, CD45RA, CCR7 and PD-1 in peripheral blood of nasopharyngeal carcinoma patients before immunotherapy, and the patients with potential benefits are screened for immunotherapy; immunotherapy curative effect dynamic monitoring is carried out by detecting the molecular levels of CD3, CD4, CD8, CD45RA, CCR7, CD69 and CXCR5 in peripheral blood after nasopharynx cancer patients are subjected to immunotherapy, and then a subsequent treatment scheme is guided. The clinical large-scale popularization and application are worthy.

Description

Technical Field

[0001] The present invention relates to the technical field of tumor immunotherapy, and particularly to a combination of peripheral blood markers for predicting the immune efficacy of nasopharyngeal carcinoma and dynamic monitoring. Background Art

[0002] Immunotherapy represented by PD1 / PDL1 inhibitors has become the main treatment method for nasopharyngeal carcinoma. Currently, common markers related to immunotherapy in clinical practice include the expression level of PD-L1, tumor mutation burden (TMB), microsatellite instability (MSI) / mismatch repair deficiency (dMMR), etc. However, the predictive performance of the above markers in nasopharyngeal carcinoma is not good at present, and there is no unified judgment standard or industry specification yet. For example, the expression level of PD-L1 in nasopharyngeal carcinoma is as high as 90%, and multiple previous studies have reported that the expression level of PD-L1 cannot distinguish sensitive patients from drug-resistant patients; while MSI / dMMR is only applicable to specific tumors such as colorectal cancer and is not applicable to nasopharyngeal carcinoma.

[0003] The mechanism of action of immunotherapy is different from that of surgery, radiotherapy, and chemotherapy. It mainly activates T cells and mobilizes T cells to carry out anti-tumor immunity as the main effective way. Therefore, in recent years, experts and scholars in the field have proposed that the functional state of immune cells in the peripheral blood circulation of the body is closely related to the outcome of tumor treatment, and the phenotypic characteristics of immune cells in peripheral blood can be used as potential markers for predicting immunotherapy. In addition, compared with tumor tissue markers, peripheral blood immune markers have the advantages of less invasiveness, high safety, and dynamic monitoring, so their roles and predictive values have attracted more and more clinical attention.

[0004] Currently, in melanoma, lung cancer and other cancers, studies have confirmed that peripheral blood markers can effectively predict the efficacy of immunotherapy. For example, in patients with non-small cell lung cancer receiving PD-1 / PD-L1 inhibitor treatment, a higher proportion (>40%) of well-differentiated CD4+ T cells (CD27-CD28low / −) with a higher peripheral blood baseline can predict the objective response rate, with 100% specificity and 70% sensitivity. However, there is currently no effective peripheral blood marker for predicting the immune efficacy of nasopharyngeal carcinoma. For this reason, a combination of peripheral blood markers for predicting the immune efficacy of nasopharyngeal carcinoma and dynamic monitoring is proposed.

[0005] The above information disclosed in this background art is only used to increase the understanding of the background art of the present invention. Therefore, it may include prior art that is not known to those of ordinary skill in the art. Summary of the Invention

[0006] The present invention aims to at least solve one of the technical problems existing in the prior art. For this reason, an object of the present invention is to propose a combination of peripheral blood markers for predicting the immune efficacy of nasopharyngeal carcinoma and dynamic monitoring.

[0007] To achieve the above object, the present invention provides the following technical solutions:

[0008] A peripheral blood biomarker combination for predicting the immune efficacy of nasopharyngeal carcinoma and dynamic monitoring, comprising:

[0009] Predicting the immune efficacy by the molecular levels of CD3, CD4, CD8, CD45RA, CCR7, and PD-1 in peripheral blood before immunotherapy;

[0010] Dynamically monitoring the immune efficacy by the molecular levels of CD3, CD4, CD8, CD45RA, CCR7, CD69, and CXCR5 in peripheral blood after immunotherapy.

[0011] A method for predicting the immune efficacy of nasopharyngeal carcinoma and dynamic monitoring based on the phenotypic characteristics of peripheral blood immune cells:

[0012] Performing spectral flow cytometry detection on the peripheral blood of the enrolled patients before treatment in a prospective clinical trial, and performing grouped comparative analysis according to the efficacy (treatment response group R vs. treatment-resistant group NR). A total of 8 major cell populations including T cells, B cells, monocytes, and dendritic cells were identified. Subsequently, the T cells were subdivided into subsets to screen for T cell subsets in which the cell ratio was significantly correlated with the efficacy.

[0013] Subsequently, the previous results were verified through two separate nasopharyngeal carcinoma cohorts of recurrence and metastasis (validation set 1) and locally advanced (validation set 2). A combination of 6 indicators, namely CD3, CD4, CD8, CD45RA, CCR7, and PD-1, was used to screen for populations sensitive to immunotherapy. By calculating four indicators, namely the ratios of CD4+Tem / CD4T and CD8+Tem / CD4T, and PD1+ / CD4+Tem and PD1+ / CD8+Tem, a combined T cell score was obtained as a prediction index for immune efficacy.

[0014] Furthermore, by comparing the changes in peripheral blood biomarkers before and after immunotherapy in the two validation set cohorts, it was found that the proportions of CD69+Tem cells and CXCR5+Tfh cells only increased in the treatment response group after treatment, while there was no significant change in the resistant group. It is suggested that the immune efficacy can be dynamically monitored by detecting the proportions of CD69+Tem and CXCR5+Tfh cells in peripheral blood, that is, through a combination of 7 indicators, namely CD3, CD4, CD8, CD45RA, CCR7, CD69, and CXCR5.

[0015] Compared with the prior art, the beneficial effects of the present invention are:

[0016] In the present invention, by detecting the levels of CD3, CD4, CD8, CD45RA, CCR7, and PD-1 molecules in the peripheral blood of nasopharyngeal carcinoma patients before immunotherapy, the efficacy of immunotherapy is predicted, and patients with potential benefits are screened for immunotherapy, which is worthy of large-scale clinical promotion and application.

[0017] In the present invention, by detecting the levels of CD3, CD4, CD8, CD45RA, CCR7, CD69, and CXCR5 molecules in the peripheral blood of nasopharyngeal carcinoma patients after immunotherapy, the dynamic monitoring of the efficacy of immunotherapy is carried out, and the subsequent treatment plan is guided.

[0018] The above summary is only for the purpose of the specification and is not intended to be limiting in any way. In addition to the illustrative aspects, embodiments, and features described above, further aspects, embodiments, and features of the present invention will be readily apparent by reference to the drawings and the following detailed description. Brief Description of the Drawings

[0019] Figure 1 It is a flow cytometry gating method diagram of the present invention;

[0020] Figure 2 It is a classification, content analysis of the main cell populations in the blood specimen of the present invention, and an inter-group comparison diagram of different efficacy groups;

[0021] Figure 3 It is a diagram showing the inter-group comparison of the main T cell subsets in different efficacy groups, the expression changes of activation molecules and immune checkpoint molecules expressed by effector CD4+ and CD8+ T cells before and after treatment in Validation Set 1 of the present invention;

[0022] Figure 4 It is a survival analysis diagram of the correlation analysis between the four indexes of CD4+Tem / CD4T and CD8+Tem / CD4T ratios, PD1+ / CD4+Tem, and PD1+ / CD8+Tem and plasma EBV DNA, and the prediction of the progression after nasopharyngeal carcinoma treatment in the present invention;

[0023] Figure 5 It is a diagram showing the changes in the levels of PD-1 and CD69 molecules expressed by CD4+Tem and CD8+Tem before and after treatment and the inter-group comparison in different efficacy groups in Validation Set 2 of the present invention;

[0024] Figure 6 It is a diagram showing the inter-group comparison of each Th cell subset and Treg cell before and after treatment in different efficacy groups, and the trend diagram of Tfh after treatment in the CR group in Validation Set 2 of the present invention;

[0025] Figure 7 It is an effect diagram of pretreatment and post-treatment monitoring of the present invention. Detailed Description of the Invention

[0026] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0027] Example 1

[0028] The present invention provides a technical solution:

[0029] Using the blood specimens of the patients enrolled in the prospective clinical trial (Jupiter-02 study) for research (training set). The experimental group patients in the Jupiter-02 study received 6 cycles of GP chemotherapy combined with immunotherapy, and then were followed up. According to whether the patients had tumor progression during the follow-up after treatment, the patients were divided into a treatment response group (R) and a treatment resistance group (NR) for comparative analysis.

[0030] As shown in the attached Figure 1 figure, through spectral flow cytometry detection, 8 major cell populations in peripheral blood were identified. As shown in the attached Figure 2 figure, including T cells, B cells, monocytes, dendritic cells, etc. Subsequently, the main cell components of the two groups were compared, and it was found that the treatment response group had more CD8+ T cells before treatment.

[0031] Subsequently, the T cells were further divided, and CD4+ and CD8+ T cells were divided into naive T cells (Tn), central memory T cells (Tcm), effector T cells (Tem), and terminal effector T cells (Temra). As shown in the attached Figure 3 figure, the proportions of CD4+Tem and CD8+Tem cells in the response group before treatment were significantly higher than those in the resistance group, and the CD4+Tem and CD8+Tem cells in the response group highly expressed PD-1. After receiving immunotherapy, the expression level of the PD-1 molecule decreased significantly, indicating that CD4+Tem and CD8+Tem cells are the action targets of immunotherapy.

[0032] The correlation between the proportions of CD4+Tem and CD8+Tem before treatment and the plasma EBV DNA level before treatment was analyzed. As shown in the attached Figure 3 figure, the proportions of CD4+Tem and CD8+Tem were significantly negatively correlated with EBV DNA. The higher the disease burden of nasopharyngeal carcinoma, the lower the proportion of effector T cells in peripheral blood.

[0033] Example 2

[0034] As shown in the attached Figure 4As shown, survival analysis was performed on patients according to the proportions of CD4+ Tem and CD8+ Tem cells in the blood. The progression-free survival of patients with high proportions of peripheral blood CD4+ Tem and CD8+ Tem was significantly prolonged after immunotherapy, suggesting that the levels of CD4+ Tem and CD8+ Tem can be important indicators for predicting the survival prognosis of patients. Four indicators, namely the ratios of CD4+ Tem / CD4T and CD8+ Tem / CD4T, PD1+ / CD4+ Tem, and PD1+ / CD8+ Tem, were modeled to obtain the T cell immune score (integrated score). The higher the score, the more likely the patient is to benefit from immunotherapy, and the progression-free survival is significantly longer than that of patients with a low score.

[0035] Example 3

[0036] The blood specimens after immunotherapy were detected to dynamically evaluate the phenotypic or functional changes of peripheral blood T cells during immunotherapy. As shown in the appendix Figure 5 As shown, the levels of CD69 molecules expressed by CD4+ Tem and CD8+ Tem were significantly increased in patients with treatment response (CR) after immunotherapy, while there was no obvious change in drug-resistant patients (NCR). At the same time, as shown in the appendix Figure 6 As shown, the number of CXCR5+ Tfh cells generated after treatment increased in patients with treatment response, while the proportion of Tfh cells did not change in drug-resistant patients.

[0037] In summary, the efficacy of immunotherapy can be inferred by detecting the levels of CD69 molecules expressed by CD4+ Tem and CD8+ Tem and the proportion of CXCR5+ Tfh after immunotherapy, as shown in the appendix Figure 7 As shown.

[0038] The parts not involved in the present invention are the same as or can be implemented by the prior art. Although the embodiments of the present invention have been shown and described, those of ordinary skill in the art can understand that various changes, modifications, substitutions, and variations can be made to these embodiments without departing from the principles and spirit of the present invention. The scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A peripheral blood marker combination for predicting the immune efficacy and dynamic monitoring of nasopharyngeal carcinoma, characterized in that: include: The levels of CD3, CD4, CD8, CD45RA, CCR7, and PD-1 molecules in peripheral blood before immunotherapy predict the efficacy of immunotherapy; After immunotherapy, the levels of CD3, CD4, CD8, CD45RA, CCR7, CD69, and CXCR5 molecules in peripheral blood were used to dynamically monitor the efficacy of immunotherapy.

2. The peripheral blood marker combination for predicting the immunotherapy efficacy and dynamic monitoring of nasopharyngeal carcinoma according to claim 1, characterized in that: The samples for the marker combination were collected from blood specimens of patients enrolled in the prospective clinical trial Jupiter-02 study and the prospective clinical trial NCT04833257.

3. A model for predicting the effect of immunotherapy on patients with nasopharyngeal carcinoma, characterized in that: Modeling was performed based on four indicators: CD4+Tem / CD4T and CD8+Tem / CD4T ratios, PD1+ / CD4+Tem and PD1+ / CD8+Tem.

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