Application of K46 lactic acid of AARS2 mediated ULK1 as target spot in preparation of anti-kidney cancer product

Abstract

The invention provides application of K46 lactic acid of AARS2 mediated ULK1 as a target spot in preparation of an anti-kidney cancer product, so that the K46 lactic acid of the AARS2 mediated ULK1 as the target spot can be used for preparing the anti-kidney cancer product. The use comprises partial or complete knockout of AARS2 and / or ULK1 genes; the AARS2 protein activity is inhibited; the K46 gene of the ULK1 is subjected to specific mutation; the invention relates to a cell penetrating peptide aiming at a ULK1 K46 site and other methods. And inhibiting the expression of AARS2 and / or controlling the K46 lactylation level of ULK1 to play a role in resisting kidney cancer.

Description

Technical Field

[0001] This application belongs to the field of biomedical technology, and in particular relates to the application of AARS2-mediated K46 lactation of ULK1 as a target in the preparation of anti-renal cancer products. Background Technology

[0002] Renal cell carcinoma (RCC) is a common malignant tumor of the urinary system, with clear cell renal cell carcinoma (ccRCC) accounting for the largest proportion. Nephrectomy, as the standard treatment, presents challenges due to recurrence and postoperative metastasis. Despite advances in targeted therapy and immunotherapy, tumor metastasis, recurrence, and drug resistance remain major challenges in clinical treatment. Therefore, identifying new and effective therapeutic targets is crucial.

[0003] Autophagy plays a major role in promoting cancer development in renal cell carcinoma by providing metabolic support to tumor cells, promoting metastasis, and enhancing treatment resistance, thereby driving disease progression. Therefore, regulating autophagy may become a new direction for the treatment of renal cell carcinoma.

[0004] Protein lactation is an emerging post-translational modification that links lactate metabolism with gene expression and cellular function regulation. Lactic acidification plays a crucial role in the metabolic reprogramming of renal cell carcinoma cells: it regulates gene expression, promoting cell proliferation and survival; it modulates the expression of vascular endothelial growth factor (VEGF) and its receptor, promoting tumor angiogenesis and providing nutritional support for cell growth and metastasis; and it regulates gene expression related to extracellular matrix degradation and cell migration, promoting cell invasion and metastasis.

[0005] Therefore, regulating lactation modification levels or developing targeted inhibitors may be one approach to treating renal cell carcinoma. Summary of the Invention

[0006] This application provides an application of AARS2-mediated K46 lactation of ULK1 as a target in the preparation of anti-renal cancer products, in order to solve the problems existing in related technologies. The technical solution is as follows: In a first aspect, embodiments of this application provide an application of AARS2-mediated K46 lactation of ULK1 as a target in the preparation of anti-renal cancer products. In one implementation, the anti-renal carcinoma effect is exerted by inhibiting the expression of AARS2 and / or controlling the K46 lactation level of ULK1.

[0007] Secondly, embodiments of this application provide the application of an AARS2 inhibitor or a K46 lactation inhibitor of ULK1 in the preparation of an anti-renal cancer product.

[0008] In one embodiment, the AARS2 inhibitor includes any one of the following: (1) a reagent for partially or completely knocking out the AARS2 gene; (2) a reagent for inhibiting the activity of the AARS2 protein.

[0009] In one embodiment, the K46 lactation inhibitor of ULK1 includes any one of the following: (1) a reagent for partially or completely knocking out the ULK1 gene; (2) a reagent for specifically mutating the K46 gene of ULK1; and (3) a cell-penetrating peptide targeting the K46 site of ULK1.

[0010] In one embodiment, the amino acid sequence of the cell-penetrating peptide is as shown in SEQ ID No. 1: Ac-HLYVSPWGGDLEVAVKCINKKN-NH2.

[0011] Thirdly, embodiments of this application provide an anti-renal cancer drug that can inhibit the expression of AARS2 and / or control the K46 lactation level of ULK1.

[0012] In one embodiment, it comprises a cell-penetrating peptide as shown in SEQ ID No. 1.

[0013] In one embodiment, the dosage form of the drug includes oral liquid, injection, tablet, pill, dispersant, capsule or granule.

[0014] The advantages or beneficial effects of the above technical solutions include at least the following: This application reveals a crucial role of AARS2-mediated K46 lactation modification of ULK1 in the initiation and progression of autophagy in renal cell carcinoma. This modification directly enhances ULK1's phosphorylation ability on its downstream substrate ATG14, thereby activating autophagy and ultimately promoting the proliferation, migration, invasion, and metastasis of renal cell carcinoma cells. Therefore, AARS2 and ULK1 lactation modification may be potential targets for renal cell carcinoma therapy.

[0015] The above overview is for illustrative purposes only 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 this application will become readily apparent from the accompanying drawings and the following detailed description. Attached Figure Description

[0016] In the accompanying drawings, unless otherwise specified, the same reference numerals throughout the various drawings denote the same or similar parts or elements. These drawings are not necessarily drawn to scale. It should be understood that these drawings depict only some embodiments disclosed in this application and should not be construed as limiting the scope of this application.

[0017] Figure 1The study analyzed the correlation between serum lactate levels, autophagy levels in renal cell carcinoma, and patient prognosis. A represents a heatmap analysis based on the TCGA-KIRC cohort; B divides TCGA-KIRC data into high-lactate and low-lactate groups and compares the differences in autophagy levels between the two groups; C represents a survival analysis based on serum lactate levels. Figure 2 The study aimed to regulate the effects of lactate metabolism on autophagy in renal cell carcinoma cells. A represents the intracellular lactation level in renal cell carcinoma lines Caki-1 and 786O after exogenous addition of sodium lactate and L-lactic acid. B represents the expression level of the autophagy marker P62 protein and the LC3B-II / LC3B-I ratio under high lactate conditions. CD represents the intracellular lactation level, P62 protein expression level, and LC3B-II / LC3B-I ratio after inhibiting endogenous lactate synthesis using the LDH inhibitor sodium oxalate. Figure 3 The number of LC3B fluorescent spots and their co-localization with lysosomes in cells of the high lactate and low lactate groups; Figure 4 To observe the ultrastructural changes of autophagy precursors, mature autophagosomes, and autolysosomes under high lactate microenvironment using transmission electron microscopy. Figure 5 This study aims to identify lactation-modified substrate proteins during autophagy in renal cell carcinoma cells. Image A shows the results of co-immunoprecipitation (CoIP) of renal cell carcinoma cell lines using Pan Kla antibody, combined with Coomassie brilliant blue staining. Image B shows the mass spectra of lactation-modified proteins from the co-immunoprecipitation experiment. Figure 6 This study investigates the regulation of ULK1 lactation and its relationship with autophagy. A shows the use of co-immunoprecipitation to verify ULK1 lactation; B and C show the addition of sodium lactate, L-lactic acid, and sodium oxalate to cells, respectively, and the detection of changes in ULK1 lactation levels; F shows the detection of changes in ULK1 lactation levels by knocking down the LDHA gene; and G shows the detection of dynamic changes in ULK1 lactation levels using an EBSS-induced cell starvation environment. Figure 7 The study investigated the regulatory effects of ULK1 and LDHA on autophagy levels in renal cell carcinoma cells. A showed the effect of ULK1 or LDHA knockdown on autophagy levels in renal cell carcinoma cells, detected by immunofluorescence assay. B showed the morphological and quantitative changes of autophagosomes in renal cell carcinoma cells after ULK1 or LDHA knockdown, observed using transmission electron microscopy. Figure 8To investigate the regulatory mechanism of AARS2-mediated ULK1 lactation modification; where A represents the effect of HA-AARS2 overexpression on ULK1 lactation modification level detected by CoIP experiment; BC represents the verification of the specific binding of ULK1 and AARS2 under physiological conditions by CoIP experiment; and D represents the analysis of changes in ULK1 lactation modification level in renal cancer cells under starvation-induced conditions by endogenous CoIP experiment. Figure 9 To elucidate the interaction mechanism between ULK1 and AARS2; A shows the co-localization of ULK1 and AARS2 analyzed using immunofluorescence co-localization assays; B shows the design schematics of the ULK1 functional domain plasmids Flag-ULK1KD, Flag-ULK1STD, and Flag-ULK1CTD; C and D show the binding affinity of different functional domains of ULK1 to AARS2 detected by CoIP assays. Figure 10 The results of AARS2-catalyzed lactation modification experiments are presented in vitro. Figure 11 Results of changes in ULK1 lactation modification levels to show the effects of knocking down ULK1 lactotransferase in renal cell carcinoma cells; Figure 12 IP experiments were conducted to detect the modification level in the Flag-ULK1 point mutant (K46R / K132R); Figure 13 A schematic diagram of the K46 site of ULK1; Figure 14 This describes the regulatory mechanism of ATG14 phosphorylation by ULK1 lactation modification; where C represents the co-transfection of AARS2 and ULK1 in 293T cells. WT or ULK1 K46R Co-transfection results of ATG14 phosphorylation modification; DF represents ULK1 under autophagy-induced conditions in renal cell carcinoma cells as shown in Western blot experiments. WT and ULK1 K46R Changes in ATG14 phosphorylation modification in cells, and the results of knocking down AARS2 to reduce ATG14 phosphorylation modification; Figure 15 The effect of ULK1 lactation modification on autophagy levels in renal cell carcinoma cells; where A represents the effect of immunofluorescence assay on ULK1. WT and ULK1 K46R Expression and localization of autophagy-related markers in renal cell carcinoma cells; B shows ULK1 observed by transmission electron microscopy. WT and ULK1 K46R Morphological characteristics of autophagosomes and autophagy-related structures in renal cell carcinoma cells; Figure 16The results show the effect of CPP-K46 treatment on the lactation modification level of ULK1 in renal cell carcinoma cells. Figure 17 The results show the regulation of ULK1 lactation modification level in renal cell carcinoma cells by treatment with the control peptide CPP-K46R. Figure 18 This study investigated the effects of CPP-K46 on ULK1 lactation modification and autophagy levels under autophagy-inducing conditions. AB represents the effect of CPP-K46 on ULK1 lactation modification levels under autophagy-inducing conditions; C shows the expression and distribution of autophagy-related markers in renal cell carcinoma cells after CPP-K46 treatment using immunofluorescence assays; and D shows the formation of autophagosomes in renal cell carcinoma cells after CPP-K46 treatment, observed using transmission electron microscopy. Figure 19 The regulation of renal cell carcinoma proliferation, migration, and invasion by ULK1 lactation modification; where AB represents ULK1. WT ULK1 KO and ULK1 K46R Results of proliferation, migration, and invasion of renal cell carcinoma cells; CD represents the effects of CPP-K46 and CPP-K46R treatment on the proliferation, migration, and invasion of renal cell carcinoma cells; Figure 20 The regulation of the in vivo invasive ability of renal cell carcinoma by ULK1 lactation modification; where A represents the ULK1 tail vein metastasis experiment. WT ULK1 KO and ULK1 K46R B represents the in vivo invasive ability of renal cell carcinoma cells; B represents the in vivo invasion of renal cell carcinoma cells after treatment with CPP-K46 and CPP-K46R. Figure 21 This is the mechanism by which AARS2-mediated ULK1 lactation modification regulates autophagy. Detailed Implementation

[0018] In the following description, only certain exemplary embodiments are briefly described. As those skilled in the art will recognize, the described embodiments can be modified in various ways without departing from the spirit or scope of this application. Therefore, the drawings and description are considered to be exemplary in nature and not restrictive.

[0019] Example 1 Based on the TCGA database, a heatmap analysis of serum lactate and autophagy levels in the KIRC cohort was performed. The results are as follows: Figure 1 As shown in Figure A; further, the TCGA-KIRC data were divided into a high-lactate group and a low-lactate group for comparative analysis, and the results are as follows. Figure 1 As shown in Figure B; and the patient's survival status was analyzed, with the results as follows. Figure 1 As shown in C.

[0020] Figure 1 The results of the A-analysis showed that as serum lactate levels increased, the autophagy level in patients with renal cell carcinoma also showed a significant upward trend. Figure 1 The results of the B-cell analysis showed that the autophagy level in the high-lactate group was significantly higher than that in the low-lactate group. Furthermore, Figure 1 Mid-C survival analysis showed that patients in the low lactate group had significantly longer survival times than those in the high lactate group, further suggesting a close correlation between serum lactate levels and patient prognosis. In conclusion, these data indicate a significant positive correlation between serum lactate levels and autophagy levels in renal cell carcinoma patients, and that serum lactate levels may serve as a potential biomarker for predicting patient prognosis.

[0021] Example 2 (1) Renal cell carcinoma cells Caki-1 and 786O (both from the American Type Culture Collection (ATCC)) were inoculated into culture containers containing complete medium (DMEM basal medium supplemented with 10% fetal bovine serum (FBS) and 1% penicillin and streptomycin) and cultured in a constant temperature incubator at 37°C and 5% CO2. Sodium lactate and L-lactic acid were added exogenously to simulate the high lactate microenvironment of tumors. Culture was continued to obtain the high lactate group. (2) In the reverse experiment, the LDH inhibitor sodium oxalate was added and cultured for a longer period to inhibit endogenous lactic acid synthesis and obtain a low lactic acid group.

[0022] Intracellular lactation modification levels, P62 protein expression (an autophagy marker), and the LC3B-II / LC3B-I ratio were measured in high-lactate and low-lactate groups. The results for intracellular lactation modification levels in the high-lactate group are as follows: Figure 2 As shown in Figure A; the expression levels of the autophagy marker P62 protein and the LC3B-II / LC3B-I ratio are shown in Figure A. Figure 2 As shown in Figure B. The results of intracellular lactation modification levels in the low-lactate group are as follows: Figure 2 As shown in Figure C; the expression levels of the autophagy marker P62 protein and the LC3B-II / LC3B-I ratio are shown in Figure C. Figure 2 As shown in D; Immunofluorescence assays were performed to detect the number of LC3B fluorescent spots and their co-localization with lysosomes in cells of the high-lactate and low-lactate groups; results are as follows. Figure 3 As shown; Transmission electron microscopy was used to observe the ultrastructural changes of autophagy precursors, mature autophagosomes, and autolysosomes in a high-lactate microenvironment; the results are as follows: Figure 4 As shown.

[0023] In the renal cell carcinoma lines Caki-1 and 786O, the exogenous addition of sodium lactate and L-lactic acid to simulate the high-lactate microenvironment of tumors significantly increased intracellular lactation levels. Correspondingly, autophagy markers exhibited typical activation characteristics: the expression level of the autophagy substrate P62 protein decreased significantly, while the LC3B-II / LC3B-I ratio increased significantly. In the reverse experiment, inhibition of endogenous lactate synthesis by the LDH inhibitor sodium oxalate significantly reduced intracellular lactation levels, and P62 protein accumulation and a decrease in the LC3B-II / LC3B-I ratio were observed, suggesting a bidirectional regulatory relationship between autophagy activity and lactate metabolism.

[0024] Immunofluorescence assays showed a significant increase in the number of LC3B fluorescent spots and the proportion of lysosome colocalization in the high-lactate group, while the low-lactate group exhibited inhibited autophagosome-lysosome fusion. Transmission electron microscopy further captured typical changes in the dynamic process of autophagy: the number of autophagy precursor structures, mature autophagosomes, and autolysosomes was significantly increased in the high-lactate microenvironment, fully demonstrating the activation state of the entire autophagic flux from initiation to degradation.

[0025] This indicates that there is a clear positive regulatory relationship between the level of lactation modification and autophagy activity in renal cell carcinoma cells, and that this process can be reversibly regulated by intervening in lactate metabolism.

[0026] Example 3 After EBSS-induced autophagy in renal cell carcinoma cells, co-immunoprecipitation (CoIP) experiments were performed on the renal cell carcinoma cell lines using Pan Kla antibody, followed by Coomassie brilliant blue staining analysis; the experimental results are as follows. Figure 5 As shown in Figure A, the protein with the specific band was identified by mass spectrometry analysis, and the mass spectrum is shown in Figure A. Figure 5 As shown in B.

[0027] A specific band exists in the region with a molecular weight of approximately 150 kDa. Mass spectrometry analysis identified several potential candidate proteins for lactation modification in this band, including ULK1, LRPPRC, ACLY, COPA, and DHX9. Among them, ULK1 was identified with a significantly higher frequency than other proteins, suggesting that it may serve as a key target for lactation modification during autophagy.

[0028] The lactation modification of ULK1 protein was verified by further co-immunoprecipitation (CoIP) of protein extracts from renal cell carcinoma cells using either IgG control antibody or ULK1 antibody. Figure 6 As shown in Figure A.

[0029] Sodium lactate, L-lactic acid, and sodium oxalate were added to cells to alter intracellular lactation levels, and significant changes in the lactation level of ULK1 cells were observed. The results are as follows: Figure 6 As shown in the BE diagram. Furthermore, by reducing LDHA expression levels through gene knockdown experiments, we found that while intracellular lactate levels decreased, the lactation modification level of ULK1 also significantly decreased, as shown in the diagram. Figure 6 As shown in Figure F. Given that ULK1 is a key regulatory protein for autophagy initiation, a cellular starvation environment induced by EBSS was used to observe the dynamic changes in ULK1 lactation modification. The results are shown in Figure F. Figure 6 The results shown in Figure G indicate that the lactation modification level of ULK1 increases significantly with increasing starvation levels. These experiments systematically reveal the regulatory role of ULK1 lactation modification in cellular metabolism and autophagy.

[0030] Further verification using immunofluorescence and transmission electron microscopy experiments showed that knocking down ULK1 or LDHA significantly inhibited autophagy levels in renal cell carcinomas. Figure 7 As shown in A and B, these results indicate that ULK1 and LDHA play crucial roles in regulating autophagy, potentially serving as targets and therapeutic agents against renal cell carcinoma.

[0031] Example 4 To identify the key catalytic enzymes regulating ULK1 lactation modification, candidate acyltransferases were screened using the CoIP experimental system.

[0032] like Figure 8 As shown in Figure A, plasmids that may modify ULK1 lactation-modifying transferases and ULK1 tag plasmids, such as HA-AARS1 and HA-AARS2, were first constructed in vitro. By co-transfecting 293T cells with the Flag-ULK1 plasmid and the modifying enzyme plasmid, it was found that the lactation modification level of ULK1 was significantly enhanced in the HA-AARS2 overexpression group. Furthermore, interaction verification experiments confirmed that ULK1 and AARS2 specifically bind under physiological conditions. Figure 8 As shown in Figures B and C, AARS2 is the direct catalytic enzyme for ULK1 lactation. Further endogenous CoIP experiments revealed that starvation-induced metabolic stress significantly upregulated the lactation modification level of ULK1 in renal cell carcinoma cells, such as... Figure 8 As shown in D, this indicates that the modification event is dynamically regulated by pathophysiological stimuli. The binding of the two was confirmed using an immunofluorescence co-localization assay, such as... Figure 9As shown in Figure A. To resolve the functional domains of ULK1, Flag-tagged ULK1 functional domain plasmids were designed and constructed, including Flag-ULK1KD, Flag-ULK1STD, and Flag-ULK1CTD, as follows. Figure 9 As shown in Figure B, CoIP experiments clarified that the STD domain of ULK1 is the key region mediating its direct binding to the AARS2 protein, as... Figure 9 As shown in C and D.

[0033] The hydroxy-carboxylic acid structure of L-lactic acid is highly homologous to the amino-carboxylic acid skeleton of alanine. This structural feature suggests that AARS2 may catalyze the transfer of L-lactic acid to protein substrates through a similar substrate recognition mechanism. In vitro lactation modification reactions using recombinant protein systems revealed that AARS2 can directly catalyze the acyl transfer of Lactyl-CoA, such as... Figure 10 As shown, this is the first time that it has been confirmed to have lactation-modifying enzyme activity.

[0034] Genetic intervention experiments revealed that conditional knockdown of AARS2 in renal cell carcinoma cells significantly reduced the lactation modification level of ULK1, such as... Figure 11 As shown.

[0035] Example 5 Based on the DeepKla algorithm, the lactation modification sites of the ULK1 protein were systematically predicted, and candidate sites such as K46 and K132 were screened out, as shown in Table 1.

[0036] Table 1

[0037] Further functional validation was performed using K46R and K132R site-specific mutant plasmids. IP experiments showed that the lactation modification level of the ULK1K46R mutant was significantly reduced, such as... Figure 12 As shown, the K132R mutation did not significantly affect the modification level, ultimately confirming K46 as the key site for ULK1 lactation modification; sequence conservation analysis is shown in Table 2; schematic diagram is shown below. Figure 13 As shown.

[0038] Table 2

[0039] Example 6 Further exploration was conducted in 293T cells, where the HA-AARS2 plasmid and ULK1 were co-transfected. WT or ULK1 K46R AARS2 and ULK1 were found WTCo-transfection significantly enhanced the phosphorylation modification of ATG14, while co-transfection with ULK1... K46R Co-transfection does not have this effect, such as Figure 14 C. Subsequently, endogenous validation experiments were conducted in renal cell carcinoma cells to construct a ULK1 gene knockout (ULK1) model. KO ), wild type ULK1 (ULK1) WT ) and K46 site mutation (ULK1) K46R ) cell model; discovered ULK1 WT Under autophagy-induced conditions, the phosphorylation level of ATG14 was significantly increased, while knockdown of AARS2 significantly decreased the phosphorylation level of ATG14. ULK1 K46R Cells cannot induce phosphorylation of ATG14, such as Figure 14 These results indicate that ULK1 lactation regulates ATG14 phosphorylation via an AARS2-dependent mechanism, thereby affecting the activation of downstream autophagy signaling pathways. Therefore, the K46 mutation and AARS2 knockdown can inhibit ATG14 phosphorylation.

[0040] Immunofluorescence assay was performed (results as follows) Figure 15 A) and transmission electron microscopy techniques (results as follows) Figure 15 B) A systematic analysis of autophagy levels in renal cell carcinoma cells was performed. Experimental results showed that, compared to ULK1... WT Compared to the group, ULK1 K46R The autophagy level in the group of renal cell carcinomas was significantly reduced, a finding that further confirms the key role of ULK1 lactation modification in autophagy regulation at the subcellular structural level.

[0041] Example 7 A cell-penetrating peptide (CPP-K46) targeting the ULK1 K46 site was designed and synthesized. The amino acid sequence is shown in SEQ ID No. 1: Ac-HLYVSPWGGDLEVAV K CINKKN-NH2. And the control peptide (CPP-K46R), the amino acid sequence of which is shown in SEQ ID No. 2: Ac-HLYVSPWGGDLEVAV R CINKKN-NH2 (synthesized by Anhui Guoping Pharmaceutical Co., Ltd.). Cell-penetrating peptide interference experiments were conducted; the results showed that: CPP-K46 can significantly inhibit the lactation modification level of ULK1 in renal cell carcinoma cells (e.g., ...). Figure 16 As shown), while the control peptide CPP-K46R did not show a similar regulatory effect (as shown). Figure 17 As shown). Under autophagy-induced conditions, CPP-K46 can still effectively inhibit the lactation modification of ULK1 (results are shown in Figure 1). Figure 18 (As shown in A and B).

[0042] In addition, through immunofluorescence experiments (results as follows) Figure 18 (as shown in C) and transmission electron microscopy (results are shown in...). Figure 18 As shown in Figure D, CPP-K46 significantly reduced autophagy levels in renal cell carcinomas. These results further support the role of CPP-K46 in regulating ULK1 lactation modification and its mediated autophagy. WT The cell proliferation and migration / invasion capabilities of the group were significantly higher than those of ULK1. KO Group, and ULK1 K46R These biological behaviors of the group are compared with ULK1 WT The group showed a significant decrease (e.g.) Figure 19 (As shown in A and B).

[0043] Cell-penetrating peptide interference experiments revealed that CPP-K46 significantly inhibited the proliferation, migration, and invasion of renal cell carcinoma cells, while the control peptide CPP-K46R failed to show similar regulatory effects (e.g., Figure 19 (As shown in C and D).

[0044] Through tail vein transfer experiments in mice, the results showed that ULK1 WT The invasive ability of cancer cells in group A was significantly higher than that in ULK1. KO Group, and ULK1 K46R The group's attack capability is greater than ULK1 WT The group showed a significant decrease (e.g.) Figure 20 (As shown in A and B). Furthermore, through cell penetration peptide interference experiments, it was found that CPP-K46 significantly inhibited the invasive ability of renal cell carcinoma cells, while the control peptide CPP-K46R did not show a significant regulatory effect (e.g., ...). Figure 20 (As shown in C and D). Combining in vivo and in vitro experimental results, the efficacy of CPP-K46 in treating renal cell carcinoma cells was confirmed.

[0045] In summary, this application validates the crucial role of AARS2-mediated ULK1 lactation in the initiation and progression of autophagy in renal cell carcinoma, elucidating a novel mechanism by which lactation modification regulates autophagy, such as... Figure 21 As shown, targeting AARS2-mediated K46 lactation of ULK1 can enable the preparation of anti-renal cancer products; the anti-renal cancer effect is achieved by inhibiting AARS2 expression and / or controlling the K46 lactation level of ULK1. Methods include partial or complete knockout of the AARS2 and / or ULK1 genes; inhibition of AARS2 protein activity; specific mutation of the ULK1 K46 gene; and cell-penetrating peptides targeting the ULK1 K46 site.

[0046] In the description of this specification, the references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of this application. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of those different embodiments or examples.

[0047] Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this application, "a plurality of" means two or more, unless otherwise explicitly specified.

[0048] The above description is merely a specific embodiment of this application, but the scope of protection of this application is not limited thereto. Any person skilled in the art can easily conceive of various variations or substitutions within the technical scope disclosed in this application, and these should all be included within the scope of protection of this application. Therefore, the scope of protection of this application should be determined by the scope of the claims.

Claims

1. Application of AARS2-mediated K46 lactation of ULK1 as a target in the preparation of anti-renal cancer products.

2. The application according to claim 1, characterized in that, It exerts its anti-renal carcinoma effect by inhibiting AARS2 expression and / or controlling the K46 lactation level of ULK1.

3. Application of AARS2 inhibitors or K46 lactation inhibitors of ULK1 in the preparation of anti-renal cancer products.

4. The application according to claim 3, characterized in that, The AARS2 inhibitor includes any one of the following: (1) a reagent for partially or completely knocking out the AARS2 gene; (2) a reagent for inhibiting the activity of the AARS2 protein.

5. The application according to claim 3, characterized in that, The K46 lactation inhibitor of ULK1 includes any one of the following: (1) a reagent for partially or completely knocking out the ULK1 gene; (2) a reagent for specifically mutating the K46 gene of ULK1; (3) a cell-penetrating peptide targeting the K46 site of ULK1.

6. The application according to claim 3, characterized in that, The amino acid sequence of the cell-penetrating peptide is shown in SEQ ID No. 1: Ac-HLYVSPWGGDLEVAVKCINKKN-NH2.

7. A drug for treating renal cell carcinoma, characterized in that, The drug can inhibit the expression of AARS2 and / or control the K46 lactation level of ULK1.

8. The drug for treating renal cell carcinoma according to claim 7, characterized in that, It contains a cell-penetrating peptide as shown in SEQ ID No.

1.

9. The medicament according to claim 7, characterized in that, The dosage forms of the drug include oral liquid, injection, tablet, pill, dispersant, capsule or granule.

Images