Increased intratumoral fluorothymidine uptake levels following multikinase inhibitor sorafenib treatment in a human renal cell carcinoma xenograft model

Abstract

An early identification of the tumor response to sorafenib treatment is indispensable for selecting optimal personalized treatment strategies. However, at present, no reliable predictors are clinically available. ¹⁸F-fluorothymidine (¹⁸F-FLT) positron emission tomography (PET) is used to assess tumor proliferation, since the FLT uptake level reflects thymidine kinase-1 (TK-1) activity. Thus, the present study determined whether FLT was able to evaluate the early tumor response to sorafenib treatment in a human renal cell carcinoma (RCC; A498) xenograft in comparison with the tumor proliferation marker, Ki-67. Mice bearing A498 tumors were assigned to the control and sorafenib-treated groups and the tumor volume was measured every day. [Methyl-3H(N)]-3'-fluoro-3'-deoxythymidine (³H-FLT) was injected 2 h prior to the sacrifice of the mice on days three and seven following the treatment. ³H-FLT autoradiography (ARG) and Ki-67 immunohistochemistry (IHC) were performed using adjacent tumor sections. In the visual assessment, the intratumoral ³H-FLT uptake level diffusely increased following the treatment, while no significant changes were observed in Ki-67 IHC. The intratumoral ³H-FLT uptake levels significantly increased by 2.7- and 2.6-fold on days three and seven following the treatment, while the tumor volume and Ki-67 index did not significantly change. Thus, an increased FLT uptake level was demonstrated following the treatment, which may indicate the suppression of thymidylate synthase (TS) and the compensatory upregulation of TK-1 activity by sorafenib.

Keywords: Ki-67 labeling index; anti-angiogenic therapy; fluorothymidine; positron emission tomography; tumor proliferation.

Figure 1

Experimental procedures of the present study. RCC, renal cell carcinoma; Control, control group; Sorafenib, sorafenib-treated group; ³H-FLT, [methyl-3H(N)]-3′-fluoro-3′-deoxythymidine; ARG, autoradiography; IHC; immunohistochemistry; HE, hematoxylin and eosin.

Figure 2

Changes in tumor volume following treatment with the vehicle or sorafenib. Dotted arrow, treatment period. Control, control group; Sorafenib, sorafenib-treated group..

Figure 3

Representative images of ³H-FLT ARG, immunohistochemical stainings of Ki-67 and HE stainings on days three and seven following treatment with the vehicle or sorafenib. The dotted line represents the tumor outline. The solid line represents the muscle outline. ³H-FLT, [methyl-3H(N)]-3′-fluoro-3-′deoxythymidine; ARG, autoradiography; Control, control group; Sorafenib, sorafenib-treated group; HE, hematoxylin and eosin; IHC, immunohistochemistry.

Figure 4

Quantitative analysis of intratumoral (A) ³H-FLT distribution and (B) Ki-67 labeling index on days three and seven following treatment with vehicle or sorafenib. ³H-FLT, [methyl-3H(N)]-3′-fluoro-3-′deoxythymidine; ID, injected dose; Control, control group; Sorafenib, sorafenib-treated group. ^*P<0.01.

Figure 5

Schematic pathways of the thymidine supply for DNA synthesis. dUMP, deoxyuridine monophosphate; TMP, thymidine monophosphate; TDP, thymidine diphosphate; TTP, thymidine triphosphate; TS, thymidate synthase; FLT, fluorothymidine; TK-1, thymidine kinase-1.