Application of specifically regulated membrane protein in inhibition of development of kidney cancer cells

Abstract

The invention relates to the technical field of biology, and discloses an application of a specifically regulated membrane protein in inhibition of development of kidney cancer cells, and the membrane protein is a membrane protein SLC6A3; the amino acid sequence of the membrane protein SLC6A3 is as shown in SEQ ID NO. 1. The development of a new molecular marker membrane of the kidney cancer cells is realized, the SLC6A3 liposome is used as a delivery target of a delivery system and is used for delivering an inhibitor or siRNA of HIF2A, the growth of the kidney cancer cells can be inhibited, the toxicity is low, the side effect is reduced, and the SLC6A3 liposome can be widely applied to the inhibition of HIF2alpha-positive kidney cancer cells and VHL wild type kidney cancer cells and has a good application prospect. A key strategy is provided for developing a novel medicine for treating renal cell carcinoma.

Description

Technical Field

[0001] The invention relates to the field of biotechnology, and in particular to the application of a specifically regulated membrane protein in inhibiting the development of renal cancer cells. Background Art

[0002] The VHL gene is a tumor suppressor gene that plays a critical role in the pathogenesis of clear cell renal cell carcinoma (ccRCC), the most common subtype of renal clear cell carcinoma. In most ccRCC cases, the VHL gene is inactivated by mutation, promoter hypermethylation or other mechanisms, leading to the stabilization and accumulation of HIF protein. HIF2α (hypoxia-inducible factor 2α) is a transcription factor that has been identified as a key driver of the pathogenesis of ccRCC. In the absence of functional VHL protein, HIF2α aggregates and translocates to the cell nucleus. HIF2α can promote key signaling pathways for cancer progression, such as angiogenesis, cell proliferation and metabolic reprogramming, leading to the development of clear cells with VHL gene mutations.

[0003] The HIF-2α inhibitor currently approved for the treatment of renal clear cell carcinoma is Belzutifan (trade name WELIREG). In the LITESPARK-005 Phase III trial, Belzutifan was superior to everolimus in terms of progression-free survival and overall response rate in patients with severe advanced RCC. Belzutifan has made significant progress in the treatment of renal cancer, but there are still some shortcomings that need to be improved:

[0004] Although Belzutifan improves progression-free survival (PFS), the advantage of improved overall survival (OS) is not yet obvious. In the LITESPARK-005 study, the median overall survival of patients using belzutifan was only slightly higher than that of the everolimus group, and the difference was not statistically significant; and common side effects of Belzutifan include anemia, fatigue, and hypoxemia, among which the incidence of anemia is high, with about 84.2% of patients experiencing varying degrees of anemia. These side effects may require additional treatment, such as the use of erythropoietin or oxygen therapy, and sometimes even lead to dose adjustments or discontinuation.

[0005] At the same time, the reason why Belzutifan affects oxygenation status is closely related to its inhibitory mechanism of HIF-2α. HIF-2α is a transcription factor that is upregulated under hypoxic conditions and plays a key role in tumor cells adapting to hypoxic environments, promoting angiogenesis and cell proliferation. Under normal circumstances, when oxygen levels are low, HIF-2α can increase cells' tolerance to hypoxia and increase the oxygen supply capacity of tissues by inducing pro-angiogenic genes (such as VEGF). However, belzutifan prevents these reactions by inhibiting HIF-2α, so patients may experience hypoxemia in some cases. Especially in patients who are already at risk of hypoxia or those with respiratory diseases, belzutifan may aggravate hypoxia and affect the oxygen supply balance throughout the body.

[0006] This is because HIF2α plays an important physiological role in the body, so extensive inhibition of HIF2α may lead to mechanistic side effects, such as anemia, fatigue, hypertension, and other cardiovascular or metabolic disorders. At the same time, HIF2α-targeted drugs may be more effective for clear cell RCC patients carrying VHL gene mutations, and their efficacy may be limited in patients with other RCC subtypes (such as VHL wild-type). Summary of the invention

[0007] The purpose of the present invention is to provide an application of a specifically regulated membrane protein in inhibiting the development of renal cancer cells, so as to solve the technical problems in the prior art that the inhibition of HIF2α has high side effects and the HIF2α targeted drug has limited inhibitory effect on other RCC subtypes of renal cancer cells.

[0008] In order to solve the above technical problems, the present invention specifically provides the following technical solutions:

[0009] The present invention provides an application of a specifically regulated membrane protein in inhibiting the development of renal cancer cells, wherein the membrane protein is the membrane protein SLC6A3;

[0010] The amino acid sequence of the membrane protein SLC6A3 is shown in SEQ ID NO.1.

[0011] As a preferred embodiment of the present invention, the development of VHL wild-type renal cancer cells can be inhibited by delivering HIF2α inhibitors or siRNA through liposomes that target and inhibit the membrane protein SLC6A3.

[0012] As a preferred embodiment of the present invention, the development of HIF2α-positive renal cancer cells can be inhibited by delivering HIF2α inhibitors or siRNA via liposomes targeting the membrane protein SLC6A3.

[0013] As a preferred embodiment of the present invention, it also includes a reagent that inhibits the expression of the membrane protein SLC6A3.

[0014] As a preferred embodiment of the present invention, the reagent is selected from siRNA, and the nucleotide sequence of the siRNA is shown as SEQ ID NO.2 and SEQ ID NO.3, and SEQ ID NO.2 and SEQ ID NO.3 are complementary sequences of double-stranded RNA.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] The present invention provides a new molecular marker membrane protein SLC6A3 for renal cancer cells, inhibits the specific expression of membrane protein SLC6A3 on the surface of renal cancer cells, takes SLC6A3 liposomes as the delivery target of the delivery system, and is used to deliver HIF2A inhibitors or siRNA, which can inhibit the growth of renal cancer cells, thereby inhibiting the development of HIF2α-positive renal cancer cells and VHL wild-type renal cancer cells, and has low toxicity and reduced side effects, and can be widely used in the inhibition of HIF2α-positive renal cancer cells and VHL wild-type renal cancer cells, and provides a siRNA for inhibiting SLC6A3. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the implementation methods of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for the implementation methods or the description of the prior art. Obviously, the drawings in the following description are only exemplary, and for ordinary technicians in this field, other implementation drawings can be derived from the provided drawings without creative work.

[0018] Figure 1 Provides the transcriptional expression level diagram of SLC6A3 and EPAS1 (HIF2α) shown in Example 1 for the present invention;

[0019] Figure 2 The present invention provides a protein immunoblot image showing that the expression of SLC6A3 protein is regulated by EPAS1 (HIF2α) as shown in Example 2;

[0020] Figure 3 Provided is a line graph of the growth level of the cell line using siRNA to inhibit SLC6A3 in the 786O cell line as shown in Example 2 of the present invention;

[0021] Figure 4 The present invention provides a line graph of the growth level of the cell line using siRNA to inhibit SLC6A3 in the A498 cell line as shown in Example 2;

[0022] Figure 5The present invention provides a schematic diagram of the extent of the fourth day of the growth level of the 786O cell line using siRNA to inhibit SLC6A3 in the cell line as shown in Example 2;

[0023] Figure 6 The present invention provides a schematic diagram of the extent of the fourth day of the growth level of the A498 cell line using siRNA to inhibit SLC6A3 in the cell line as shown in Example 2;

[0024] Figure 7 The present invention provides a line graph of the growth level of the A498 cell line by the HIF2α siRNA liposome delivery system targeting SLC6A3 as shown in Example 3;

[0025] Figure 8 The present invention provides a line graph of the growth level of 786-O cell line by the HIF2α siRNA liposome delivery system targeting SLC6A3 as shown in Example 3;

[0026] Fig. 9 A schematic diagram of the extent to which the HIF2α siRNA liposome delivery system targeting SLC6A3 shown in Example 3 inhibits the growth of the 786-O cell line on the 14th day is provided for the present invention;

[0027] Fig.10 The present invention provides a schematic diagram of the growth level of organoids derived from RCC patients after being treated with the HIF2α siRNA liposome delivery system targeting SLC6A3 as shown in Example 3;

[0028] Fig.11 The present invention provides the HIF2α siRNA liposome delivery system targeting SLC6A3 as shown in Example 3 to inhibit the growth of mouse 786-O xenograft tumors, and a line graph of the volume change of xenograft tumors;

[0029] Fig.12 A line graph showing the growth level of the cell line in which SLC6A3 is inhibited in the ACHN cell line using siRNA as shown in Example 4 is provided for the present invention. DETAILED DESCRIPTION

[0030] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0031] like Figure 1As shown, the present invention provides an application of a specifically regulated membrane protein in inhibiting the development of renal cancer cells. The membrane protein is the membrane protein SLC6A3, and its amino acid sequence is shown in SEQ ID NO.1.

[0032] SEQ ID NO.1:

[0033] MSKSKCSVGLMSSVVAPAKEPNAVGPKEVELILVKEQNGVQLTSSTLTNPRQSPVEAQDRETWGKKIDFLLSVIGFAVDLANVWRFPYLCYKNGGGAFLVPYLLFMVIAGMPLFYMELALGQFNREGAAGVWKICPILKGVGFTVILISLYVGFF YNVIIAWALHYLFSSFTTELPWIHCNNSWNSPNCSDAHPGDSSGDSSGLNDTFGTTPAAEYFERGVLHLHQSHGIDDLGPPRWQLTACLVLVIVLLYFSLWKGVKTSGKVVWITATMPYVVLTALLLRGVTLPGAIDGIRAYLSVDFYRLCEASV WIDAATQVCFSLGVGFGVLIAFSSYNKFTNNCYRDAIVTTSINSLTSFSSGFVVFSFLGYMAQKHSVPIGDVAKDGPGLIFIIYPEAIATLPLSSAWAVVFFIMLLTLGIDSAMGGMESVITGLIDEFQLLHRHRELFTLFIVLATFLLSLFCVT NGGIYVFTLLDHFAAGTSILFGVLIEAIGVAWFYGVGQFSDDIQQMTGQRPSLYWRLCWKLVSPCFLLFVVVVSIVTFRPPHYGAYIFPDWANALGWVIATSSMAMVPIYAAYKFCSLPGSFREKLAYAIAPEKDRELVDRGEVRQFTLRHWLKV

[0034] The present invention also discloses a method for delivering a HIF2α inhibitor or siRNA by liposomes that target and inhibit the membrane protein SLC6A3, thereby inhibiting the development of VHL wild-type renal cancer cells.

[0035] The present invention also discloses a method for delivering a HIF2α inhibitor or siRNA by liposomes targeting the membrane protein SLC6A3, which can inhibit the development of HIF2α-positive renal cancer cells.

[0036] Furthermore, it also includes a reagent for inhibiting the expression of the membrane protein SLC6A3, wherein the reagent contains a HIF2α inhibitor or siRNA.

[0037] The nucleotide sequences of the siRNA are shown in SEQ ID NO.2 and SEQ ID NO.3, and SEQ ID NO.2 and SEQ ID NO.3 are complementary sequences of double-stranded RNA.

[0038] SEQ ID NO.2:

[0039] GCUACAACAAGUUCACCAA / dT / / dT /

[0040] SEQ ID NO.3:

[0041] UUGGUGAACUUGUUGUAGC / dT / / dT /

[0042] The name of SEQ ID NO.2 is SLC6A3-1137-S, and the name of SEQ ID NO.3 is SLC6A3-1137-a.

[0043] The application of membrane protein SLC6A3 is described below by way of examples:

[0044] Embodiment 1:

[0045] 1. Screening of SLC6A3

[0046] A list of membrane protein encoding genes was compiled through literature reports, and a total of 4346 membrane proteins and related molecules were compiled. Then, the mRNA expression of membrane protein encoding genes with significant differences in expression between EPAS1 (HIF2α encoding gene) and EPAS1 (HIF2α encoding gene) were screened in the tumor tissue gene expression matrix of the TCGA-KIRC cohort (Pearson correlation coefficient>0.2 or <-0.2, p<0.05). In the obtained gene set, genes with baseline expression baseMean>500 were screened, and genes with low baseline expression levels in renal cancer / high baseline expression levels in other normal organs / low specificity were manually excluded through the GEPIA database, and arranged in descending order according to the Log2FC value. The first gene obtained was CD70, which has been widely used as an immunotherapy marker for renal cancer and is considered to be a key marker gene for renal cancer. The second gene is SLC6A3.

[0047] 2. Verification of the relationship between SLC6A3 and renal cancer

[0048] In the RCC patient cohort of TCGA-KIRC, the transcriptional expression levels of SLC6A3 and EPAS1 were detected, and the results are shown in Figure 1 As shown, Figure 1The results showed that SLC6A3 was only significantly expressed in renal cancer tissues and was significantly correlated with the expression of EPAS1.

[0049] Delivery systems play a vital role in targeted tumor therapy. Through targeted delivery, drugs can be concentrated at the tumor site, reducing damage to healthy tissues, improving efficacy, and reducing systemic toxicity. When selecting proteins as targets for cancer-specific delivery systems, the following requirements must be met: ① expressed on the cell surface; ② abnormally expressed on tumor cells.

[0050] Example 1 shows that SLC6A3 is specifically expressed on the surface of renal cancer cells and can be used as a molecular marker of renal cancer cells in the detection of renal cancer. At the same time, the membrane protein SLC6A3 can be used as a therapeutic target for renal cancer in the treatment of renal cancer.

[0051] Embodiment 2:

[0052] The siRNA related to the present invention is synthesized by Sangon Biotechnology (Shanghai) Co., Ltd. and is used to inhibit the transcription of the corresponding gene.

[0053] (1) Observe that the cells are growing well and the confluency is about 50-60%, then start transfecting the cells.

[0054] (2) Aspirate the culture medium in the culture plate, wash the cells with PBS, and add 1 ml of fresh culture medium without fetal bovine serum.

[0055] (3) Add 3 μl siRNA and 3 μl lipo2000 into 100 ml opti-MEM medium respectively and let stand for 5 min.

[0056] (4) Mix siRNA with the diluted Lipo 2000 solution and let stand for another 15-20 minutes.

[0057] (5) Add the mixed solution to the cells in a medium that has been replaced with a medium that does not contain fetal bovine serum and culture in an incubator for 6-8 hours.

[0058] (6) Observe cell morphology. Replace the culture medium with fresh culture medium containing fetal bovine serum. Continue culturing. RNA can be extracted after 24 hours, and protein can be extracted or cell function experiments can be performed after 48 hours.

[0059] After using interfering RNA to inhibit HIF2α in 786-O renal cancer cells, specific results were seen Figure 2 ,from Figure 2 It can be seen that the expression level of SLC6A3 decreased.

[0060] Furthermore, siRNA was used to inhibit SLC6A3 in 786O and A498 cell lines. Figures 3 to 6,As can be seen from the figure, after silencing the expression of SLC6A3, the growth of the cell line was inhibited.

[0061] In vitro growth assay (CCK-8 assay)

[0062] (1) 48 hours after cell transfection, if the cells are in good condition, the experiment can be started.

[0063] (2) After counting the cells, adjust the cell concentration to 1000 cells per 100 μl. Add 100 μl of cell suspension to each well of a 96-well plate, which equals 1000 cells. Each experimental group is measured uniformly for four days, so four batches of replicate samples are required, with 6 replicates in each batch. Place the 96-well culture plate in an incubator for culture.

[0064] (3) After 24h, 48h, 72h, and 96h, measure the samples respectively.

[0065] Aspirate the culture medium in the wells. Add 110 μl of pre-prepared culture medium containing CCK-8 reagent to each well (CCK-8: culture medium = 10:100). Continue to incubate in the incubator for 3 hours.

[0066] (4) Set the wavelength of the microplate reader to 450 nm and measure the absorbance of each well after incubation. Keep away from light.

[0067] (5) After 96 hours, plot the four-day data into a growth curve. This experiment needs to be repeated three times independently.

[0068] Embodiment 3:

[0069] In vitro cell lines and renal cancer organoids, the present system, HIF2α inhibitor or HIF2α siRNA targeting SLC6A3 (hereinafter referred to as HIF2αi), was used for treatment. The results are shown in Figures 7 to 10 ,As can be seen from the figure, compared with PBS, using this system to target SLC6A3 and inhibit HIF2α can significantly inhibit the progression of RCC.

[0070] 1. In vitro growth experiment

[0071] (1) After counting the cells, adjust the cell concentration to 1000 cells per 100 μl. Add 100 μl of cell suspension to each well of a 96-well plate, which equals 1000 cells. Each experimental group is measured uniformly for four days, so four batches of replicate samples are required, with 6 replicates in each batch. Place the 96-well culture plate in an incubator for culture.

[0072] (2) After the cells adhered to the wall, they were treated with PBS or this system according to the grouping;

[0073] (3) After 24h, 48h, 72h, and 96h, measure the samples respectively.

[0074] Aspirate the culture medium in the wells. Add 110 μl of pre-prepared culture medium containing CCK-8 reagent to each well (CCK-8: culture medium = 10:100). Continue to incubate in the incubator for 3 hours.

[0075] (4) Set the wavelength of the microplate reader to 450 nm and measure the absorbance of each well after incubation. Keep away from light.

[0076] (5) After 96 hours, plot the four-day data into a growth curve. This experiment needs to be repeated three times independently.

[0077] 2. In vitro growth experiment (clone formation experiment)

[0078] (1) After counting the cells, adjust the cell concentration to 1000 cells per 100 μl. Add 100 μl of cell suspension to each well of a 6-well plate, add 2 ml of fresh complete culture medium, and place the 6-well plate in an incubator for culture.

[0079] (2) After the cells adhered to the wall, they were treated with PBS or this system according to the grouping.

[0080] (3) Measurement after 14 days: Take out the 6-well culture plate, aspirate the culture medium in the wells to be measured, and rinse with PBS three times.

[0081] (4) Add 1 ml of 4% paraformaldehyde to each well for fixation for 15 min and rinse three times with PBS.

[0082] (5) Stain with crystal violet solution for 30 min (room temperature), then take photos and count the number of colonies.

[0083] 3. Organoid extraction, culture and passaging

[0084] 1. Organoid extraction

[0085] (1) After obtaining the surgical specimen from the RCC patient, process it within 2 hours

[0086] (2) Rinse the specimen in DPBS and chop it as finely as possible on a glass slide with a scalpel. Transfer the tissue fragments to a 15 ml centrifuge tube, add 5 ml of DPBS and centrifuge at 300 x g for 5 min. Discard the supernatant and add 1-2 ml of preheated digestion solution to mix well.

[0087] (3) Incubate at 37°C for 15-30 min. Monitor the digestion status under a microscope and pipette every 10 min.

[0088] (4) Add 5-10 ml of cold DMEM / F12 to terminate digestion. If there is still visible undigested tissue, filter it using a 40 μm cell strainer.

[0089] (5) Centrifuge at 300 g for 5 min, discard the supernatant, add 40 μl of Matrigel / diluted Matrigel for every 500-1000 cells, resuspend and inoculate.

[0090] 2. Organoid Passaging

[0091] 1. Aspirate the culture medium in the well (avoid touching the matrix gel)

[0092] 2. Add 1 mL of Gentle Cell Dissociation Reagent (GCDR) to the top of the matrix gel and incubate at room temperature for 1 minute

[0093] 3. Wet the 1ml pipette tip with GCDR in the well, blow and beat repeatedly 20 times to destroy the matrix gel, aspirate the suspension into a 15ml centrifuge tube, and then rinse the well wall with 1mL GCDR and aspirate into the centrifuge tube.

[0094] 4. Incubate at room temperature on a shaker (20 rpm) for 10 minutes

[0095] Centrifuge at 290 g for 5 minutes at 5.4°C and discard the supernatant and matrix gel.

[0096] 6. Moisten 1 ml pipette tip with DMEM / F12, resuspend organoids in 10 ml pre-cooled DMEM / F12, centrifuge at 200 g for 5 minutes at 4°C, and discard DMEM / F12 as much as possible.

[0097] 7. Add 50ul complete culture medium and 40ul matrix gel to each tube, and plant organoids in each well with 40ul of culture medium in the same way as the original culture.

[0098] (III) After the organoids were cultured for 3 generations, they were treated with PBS or this system according to the group, and the changes in organoid volume were detected using a microscope;

[0099] 4. Construction and measurement of subcutaneous tumor model

[0100] (1) Digest, centrifuge, resuspend and count the 786-O cell line. Adjust the cell count to 1x10 6 / 50ul. Transfer the resuspended cells to an EP tube and store on ice.

[0101] 3.4. Construction and measurement of subcutaneous tumor model

[0102] (1) Digest, centrifuge, resuspend and count the established stable low-dose 786-O cell line. Adjust the concentration to 1x106 / 50ul. Transfer the resuspended cells to an EP tube and store on ice.

[0103] (2) After wearing sterile gloves, pinch the neck skin of the mouse with the thumb and index finger of the left hand, and then fix the mouse tail to the thenar eminence of the left hand with the ring finger and pinky finger of the left hand. Disinfect the axilla or groin with 75% alcohol three times. Hold the syringe containing tumor cells in the right hand, and insert the needle at a 45-degree angle at the groin or axilla, being careful not to break through the peritoneum, and keep the needle in the subcutaneous position.

[0104] (3) Then, the needle is almost completely inserted into the subcutaneous tissue at a near horizontal position and the tumor cells are injected into the subcutaneous tissue (the amount of tumor cells is about 1x10 6 ), quickly withdraw the needle, press the needle hole lightly with the left index finger for about 1 min, and then put the mouse back into the cage.

[0105] (4) Ten days later, mice were injected intratumorally with PBS or the present system (a delivery system targeting SLC6A3 and inhibiting HIF2α).

[0106] (5) After 10 days, tumor diameter was measured using a vernier caliper.

[0107] Results are shown in Fig.11 , Fig.11 This is a mouse tumor growth curve. It can be seen from the figure that after inhibiting the expression of membrane protein SLC6A3, the growth of RCC xenograft tumors can be significantly inhibited in mouse in vivo experiments, and there is no significant toxicity to normal kidney cells and mouse kidneys and livers.

[0108] Example 4: Validation of VHL wild-type renal cancer cell line ACHN

[0109] In RCC, VHL deletion mutations are often accompanied by abnormal activation of HIF2α, while in patients with VHL wild-type, HIF2α is often in a lower activation state. For this type of tumor, simply targeting HIF2α does not have a good therapeutic effect.

[0110] To this end, we targeted SLC6A3 inhibition to determine the extent of renal cancer cell proliferation in VHL wild-type patients. The specific characterization results are shown in Fig.12 .

[0111] Depend on Fig.12 It can be seen that in VHL wild-type patients, targeting SLC6A3 can also play a role in inhibiting tumors. It can be seen that the expression level of SLC6A3 has a certain positive correlation with HIF2α, but it is not only regulated by the VHL-HIF2α axis, but can also play a certain therapeutic role in patients with other RCC subtypes (such as VHL wild-type).

[0112] Through the application of the specifically regulated membrane protein of this embodiment in inhibiting the development of renal cancer cells, a new molecular marker membrane protein SLC6A3 for renal cancer cells can be discovered, and the specific expression of membrane protein SLC6A3 on the surface of renal cancer cells can be inhibited. At the same time, by targeted inhibition of membrane protein SLC6A3, the growth of renal cancer cells can be inhibited, and the toxicity is low and the side effects are reduced. It can be widely used in the inhibition of HIF2α-positive renal cancer cells and VHL wild-type renal cancer cells, and a siRNA for inhibiting SLC6A3 is provided, which provides a key strategy for the development of new drugs for renal cell carcinoma.

[0113] The above embodiments are only exemplary embodiments of the present application and are not intended to limit the present application. The protection scope of the present application is defined by the claims. Those skilled in the art may make various modifications or equivalent substitutions to the present application within the essence and protection scope of the present application, and such modifications or equivalent substitutions shall also be deemed to fall within the protection scope of the present application.

Claims

1. Use of a specifically regulated membrane protein in inhibiting the development of renal cancer cells, characterized in that: The membrane protein is the membrane protein SLC6A3; The amino acid sequence of the membrane protein SLC6A3 is shown in SEQ ID NO.

1.

2. The use of a specifically regulated membrane protein in inhibiting the development of renal cancer cells according to claim 1, characterized in that: By delivering HIF2α inhibitors or siRNA through liposomes targeting the membrane protein SLC6A3, the development of HIF2α-positive renal cancer cells and VHL wild-type renal cancer cells can be inhibited.

3. The use of a specifically regulated membrane protein in inhibiting the development of renal cancer cells according to claim 2, characterized in that: Also included are agents that inhibit the expression of the membrane protein SLC6A3.

4. The use of a specifically regulated membrane protein in inhibiting the development of renal cancer cells according to claim 3, characterized in that: The reagent is selected from the siRNA, and the nucleotide sequence of the siRNA is shown in SEQ ID NO.2 and SEQ ID NO.3.

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