An Integrin Alpha 6-Targeted Radiotracer with Improved Receptor Binding Affinity and Tumor Uptake

In this study, we reported a ^99mTc-labeled integrin α₆-targeted peptide as the molecular imaging probe for tumor imaging by single-photon emission computed tomography (SPECT). We found that replacing Cys-Cys cyclized RWY peptide (sequence: cCRWYDENAC) with lactam-bridged cyclic cKiE peptide (sequence: cKRWYDENAisoE) did not sacrifice the integrin α₆-binding affinity and specificity of cKiE radiotracer. To further improve the radiotracer's tumor targeting capability, the dimerized cKiE peptide (termed cKiE2) was designed, and the corresponding radiotracer ^99mTc-cKiE2 was evaluated for tumor uptake and in vivo pharmacokinetics properties in tumor models. We found that cKiE2 showed higher binding affinity to integrin α₆ than did monomeric RWY or cKiE peptide. The biodistribution results showed that the tumor uptake of ^99mTc-cKiE2 was twice higher than that of ^99mTc-RWY (3.20 ± 0.12 vs 1.26 ± 0.06 %ID/g, P < 0.001) at 0.5 h postinjection. The tumor to nontargeting tissue ratios were also enhanced in most normal organs. Specificity of ^99mTc-cKiE2 for integrin α₆ was demonstrated by competitive blocking of tumor uptake with excess cold peptide (3.20 ± 0.24 to 1.38 ± 0.23 %ID/g, P < 0.001). The integrin α₆-positive tumors were clearly visualized by ^99mTc-cKiE2/SPECT with low background except with a relatively high kidney uptake. The tumor uptake of ^99mTc-cKiE2 correlates well with the tumor integrin α₆ expression levels in a linear fashion (R² = 0.9623). We also compared ^99mTc-cKiE2 with an integrin α_vβ₃-targeted radiotracer ^99mTc-3PRGD₂ in the orthotopic hepatocellular carcinoma tumor models. We found that the orthotopic tumor was clearly visualized with ^99mTc-cKiE2. ^99mTc-3PRGD₂ imaging did not show tumor contours in situ as clearly as ^99mTc-cKiE2. The tumor-to-liver ratios of ^99mTc-cKiE2 and ^99mTc-3PRGD₂ were 2.20 ± 0.17 and 0.85 ± 0.20. In conclusion, ^99mTc-cKiE2 is an improved SPECT radiotracer for imaging integrin α₆-positive tumors and has great potential for further clinical application.