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Role of IL13RA2 in Sunitinib Resistance in Clear Cell Renal Cell Carcinoma.

Shibasaki N, Yamasaki T, Kanno T, Arakaki R, Sakamoto H, Utsunomiya N, Inoue T, Tsuruyama T, Nakamura E, Ogawa O, Kamba T - PLoS ONE (2015)

Bottom Line: Conversely, shRNA-mediated knockdown of IL13RA2 successfully overcame the sunitinib-resistance in Caki-1 cells.Histopathological analyses revealed that IL13RA2 repressed sunitinib-induced apoptosis without increasing tumor vasculature in vivo.To our knowledge, this is a novel mechanism of developing resistance to sunitinib in a certain population of ccRCC, and these results indicate that IL13RA2 could be one of potential target to overcome sunitinib resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

ABSTRACT
Vascular endothelial growth factor (VEGF) and mammalian target of rapamycin are well-known therapeutic targets for renal cell carcinoma (RCC). Sunitinib is an agent that targets VEGF receptors and is considered to be a standard treatment for metastatic or unresectable clear cell RCC (ccRCC). However, ccRCC eventually develops resistance to sunitinib in most cases, and the mechanisms underlying this resistance are not fully elucidated. In the present study, we established unique primary xenograft models, KURC1 (Kyoto University Renal Cancer 1) and KURC2, from freshly isolated ccRCC specimens. The KURC1 xenograft initially responded to sunitinib treatment, however finally acquired resistance. KURC2 retained sensitivity to sunitinib for over 6 months. Comparing gene expression profiles between the two xenograft models with different sensitivity to sunitinib, we identified interleukin 13 receptor alpha 2 (IL13RA2) as a candidate molecule associated with the acquired sunitinib-resistance in ccRCC. And patients with high IL13RA2 expression in immunohistochemistry in primary ccRCC tumor tends to have sunitinib-resistant metastatic site. Next, we showed that sunitinib-sensitive 786-O cells acquired resistance in vivo when IL13RA2 was overexpressed. Conversely, shRNA-mediated knockdown of IL13RA2 successfully overcame the sunitinib-resistance in Caki-1 cells. Histopathological analyses revealed that IL13RA2 repressed sunitinib-induced apoptosis without increasing tumor vasculature in vivo. To our knowledge, this is a novel mechanism of developing resistance to sunitinib in a certain population of ccRCC, and these results indicate that IL13RA2 could be one of potential target to overcome sunitinib resistance.

No MeSH data available.


Related in: MedlinePlus

Evaluation of apoptosis and microvessel density after sunitinib treatment in xenograft model derived from cell lines.MVD was decreased by sunitinib treatment of each xenograft tumor derived from (A) 786-O or (B) Caki-1 subclones regardless of IL13RA2 expression level. MVD was determined from CD31 staining using Image J software. Statistical analysis was performed using the Students’ t-test (*P < 0.01). Immunoblot analysis of (C) 786-O subclones and (D) Caki-1 subclones. IL13RA2 expression was negatively correlated with the phosphorylation of STAT6. Whole cell extracts were immunoblotted using the indicated antibodies. ssDNA staining of xenograft tumors derived from (E) 786-O subclones and (B) Caki-1 subclones treated with sunitinib or vehicle only. Apoptosis was assessed by calculating the ssDNA positivity rate. Statistical analysis was performed using the Students’ t-test (*P < 0.05, **P < 0.01).
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pone.0130980.g004: Evaluation of apoptosis and microvessel density after sunitinib treatment in xenograft model derived from cell lines.MVD was decreased by sunitinib treatment of each xenograft tumor derived from (A) 786-O or (B) Caki-1 subclones regardless of IL13RA2 expression level. MVD was determined from CD31 staining using Image J software. Statistical analysis was performed using the Students’ t-test (*P < 0.01). Immunoblot analysis of (C) 786-O subclones and (D) Caki-1 subclones. IL13RA2 expression was negatively correlated with the phosphorylation of STAT6. Whole cell extracts were immunoblotted using the indicated antibodies. ssDNA staining of xenograft tumors derived from (E) 786-O subclones and (B) Caki-1 subclones treated with sunitinib or vehicle only. Apoptosis was assessed by calculating the ssDNA positivity rate. Statistical analysis was performed using the Students’ t-test (*P < 0.05, **P < 0.01).

Mentions: Next, we sought to explore the mechanisms of IL13RA2-mediated resistance to sunitinib. Xenograft tumors derived from 786-O and Caki-1 subclone were resected after 21days, 75days of sunitinib treatment, respectively. Since increased MVD was associated with sunitinib resistance [24, 25], we first measured MVD in a series of xenograft tumors from 786-O- or Caki-1-derived cells. As expected, MVD was significantly decreased in sunitinib-sensitive 786-O mock xenografts after the treatment (Fig 4A & S6A Fig). To our surprise, sunitinib significantly suppressed tumor microvasculature in specimens from resistant 786-O IL13RA2 cells (Fig 4A & S6A Fig). Similarly, CD31 immunostaining exhibited significantly decreased MVD in both sunitinib-treated sh-scrambled and shIL13RA2 Caki-1-derived xenografts irrespective of their sensitivity to the drug (Fig 4B & S6B Fig). These results indicated that IL13RA2 did not grossly affect the effect of sunitinib on tumor microvasculature and might play significant roles in the survival of tumor cells under the stressed circumstances of diminished MVD.


Role of IL13RA2 in Sunitinib Resistance in Clear Cell Renal Cell Carcinoma.

Shibasaki N, Yamasaki T, Kanno T, Arakaki R, Sakamoto H, Utsunomiya N, Inoue T, Tsuruyama T, Nakamura E, Ogawa O, Kamba T - PLoS ONE (2015)

Evaluation of apoptosis and microvessel density after sunitinib treatment in xenograft model derived from cell lines.MVD was decreased by sunitinib treatment of each xenograft tumor derived from (A) 786-O or (B) Caki-1 subclones regardless of IL13RA2 expression level. MVD was determined from CD31 staining using Image J software. Statistical analysis was performed using the Students’ t-test (*P < 0.01). Immunoblot analysis of (C) 786-O subclones and (D) Caki-1 subclones. IL13RA2 expression was negatively correlated with the phosphorylation of STAT6. Whole cell extracts were immunoblotted using the indicated antibodies. ssDNA staining of xenograft tumors derived from (E) 786-O subclones and (B) Caki-1 subclones treated with sunitinib or vehicle only. Apoptosis was assessed by calculating the ssDNA positivity rate. Statistical analysis was performed using the Students’ t-test (*P < 0.05, **P < 0.01).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4482605&req=5

pone.0130980.g004: Evaluation of apoptosis and microvessel density after sunitinib treatment in xenograft model derived from cell lines.MVD was decreased by sunitinib treatment of each xenograft tumor derived from (A) 786-O or (B) Caki-1 subclones regardless of IL13RA2 expression level. MVD was determined from CD31 staining using Image J software. Statistical analysis was performed using the Students’ t-test (*P < 0.01). Immunoblot analysis of (C) 786-O subclones and (D) Caki-1 subclones. IL13RA2 expression was negatively correlated with the phosphorylation of STAT6. Whole cell extracts were immunoblotted using the indicated antibodies. ssDNA staining of xenograft tumors derived from (E) 786-O subclones and (B) Caki-1 subclones treated with sunitinib or vehicle only. Apoptosis was assessed by calculating the ssDNA positivity rate. Statistical analysis was performed using the Students’ t-test (*P < 0.05, **P < 0.01).
Mentions: Next, we sought to explore the mechanisms of IL13RA2-mediated resistance to sunitinib. Xenograft tumors derived from 786-O and Caki-1 subclone were resected after 21days, 75days of sunitinib treatment, respectively. Since increased MVD was associated with sunitinib resistance [24, 25], we first measured MVD in a series of xenograft tumors from 786-O- or Caki-1-derived cells. As expected, MVD was significantly decreased in sunitinib-sensitive 786-O mock xenografts after the treatment (Fig 4A & S6A Fig). To our surprise, sunitinib significantly suppressed tumor microvasculature in specimens from resistant 786-O IL13RA2 cells (Fig 4A & S6A Fig). Similarly, CD31 immunostaining exhibited significantly decreased MVD in both sunitinib-treated sh-scrambled and shIL13RA2 Caki-1-derived xenografts irrespective of their sensitivity to the drug (Fig 4B & S6B Fig). These results indicated that IL13RA2 did not grossly affect the effect of sunitinib on tumor microvasculature and might play significant roles in the survival of tumor cells under the stressed circumstances of diminished MVD.

Bottom Line: Conversely, shRNA-mediated knockdown of IL13RA2 successfully overcame the sunitinib-resistance in Caki-1 cells.Histopathological analyses revealed that IL13RA2 repressed sunitinib-induced apoptosis without increasing tumor vasculature in vivo.To our knowledge, this is a novel mechanism of developing resistance to sunitinib in a certain population of ccRCC, and these results indicate that IL13RA2 could be one of potential target to overcome sunitinib resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

ABSTRACT
Vascular endothelial growth factor (VEGF) and mammalian target of rapamycin are well-known therapeutic targets for renal cell carcinoma (RCC). Sunitinib is an agent that targets VEGF receptors and is considered to be a standard treatment for metastatic or unresectable clear cell RCC (ccRCC). However, ccRCC eventually develops resistance to sunitinib in most cases, and the mechanisms underlying this resistance are not fully elucidated. In the present study, we established unique primary xenograft models, KURC1 (Kyoto University Renal Cancer 1) and KURC2, from freshly isolated ccRCC specimens. The KURC1 xenograft initially responded to sunitinib treatment, however finally acquired resistance. KURC2 retained sensitivity to sunitinib for over 6 months. Comparing gene expression profiles between the two xenograft models with different sensitivity to sunitinib, we identified interleukin 13 receptor alpha 2 (IL13RA2) as a candidate molecule associated with the acquired sunitinib-resistance in ccRCC. And patients with high IL13RA2 expression in immunohistochemistry in primary ccRCC tumor tends to have sunitinib-resistant metastatic site. Next, we showed that sunitinib-sensitive 786-O cells acquired resistance in vivo when IL13RA2 was overexpressed. Conversely, shRNA-mediated knockdown of IL13RA2 successfully overcame the sunitinib-resistance in Caki-1 cells. Histopathological analyses revealed that IL13RA2 repressed sunitinib-induced apoptosis without increasing tumor vasculature in vivo. To our knowledge, this is a novel mechanism of developing resistance to sunitinib in a certain population of ccRCC, and these results indicate that IL13RA2 could be one of potential target to overcome sunitinib resistance.

No MeSH data available.


Related in: MedlinePlus