Aminocyanine Fluorescent Dyes For Optical Imaging Of Tumor Cells

In recent years, although study on development and metastasis mechanism of tumor has achieved much progress, due to a lack of sensitive early diagnosis, highly specific targeting therapy and effective real-time monitoring, malignant tumor is still a serious disease that threats human health. Tumor theranostics is a new strategy for cancer treatment which has been come up with in recent years, its core is the effective combination of tumor treatment and real-time imaging, in order to guide doctors to adjust treatment, improve survival rate and life quality of tumor patients and provide a new approach for the individualized treatment of tumor. Therefore, preparing a diagnostic and therapeutic agent with tumor specific imaging and targeting therapy is very important.In this paper, we designed and synthesized a tumor targeting probe Cy7-RGD based on cyclo (-RGDyK-) and aminocyanine fluorescent dyes, whose structure has not been reported. Cyclo (-RGDyK-) is recognized as an antagonist of cell adhesion molecule integrin avp3, integrin ανβ3 expresses specifically in the surface of several solid tumor cells and in almost all sprouting tumor vasculatures. At present, the research of probe based on cyclo (-RGDyK-) which targets integrin ανβ3 has become a hot research direction of tumor diagnosis and therapy.In this paper, we used the Fmoc solid phase synthesis method, with 2-Chlorotrityl Resin as the carrier, with DIEA, TBTU, HOBT as condensation agents to synthesize the cyclic pentapeptide cyclo(-RGDyK-), and recrystallized it in ethyl ether, the yield was up to 84%. Then, we synthesized an aminocyanine fluorescent dye Cy7-RGD based on our group’s previous research result of aminocyanine with large Stokes shift.Next, we studid its spectral properties and found that its maximum excitation wavelength is 617 nm in PBS (PH=7.4) and maximum emission wavelength is 754 nm, the Stokes shift is up to 137 nm, and it’s in the near-infrared region. These spectral properties show that it is very suitable for studies of fluorescence imaging in vivo.Then, we cultured tumor cells U87MG and MCF-7 with Cy7-RGD, glioma cells U87MG are known to over express integrin ανβ3, but breast cancer cells MCF-7 are known to express almost no integrin ανβ.The result shows that Cy7-RGD can gather in U87MG cells, but has almost no aggregation in MCF-7. And this process is integrin specific, because it can be inhibited by adding the uncoupled cyclo (-RGDyK-). These results indicate that the combination of Cy7-RGD with integrin ανβ3 is highly specific, so Cy7-RGD has the potential to specifically diagnose glioma.At last, MTT experiments showed that Cy7-RGD can specifically inhibit the proliferation of U87MG cells which overexpress integrin ανβ3, and its IC50 value is 8076 nM. However, it has no little effect on normal cells and tumor cells which doesn’t express integrin αvβ3. It indicats that Cy7-RGD has the potential of targeting treat glioma, which is never reported in the literature.