Selective killing of circulating tumor cells prevents metastasis and extends survival
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Distant metastasis is initiated by circulating tumor cells (CTCs), which are considered to be a determining factor for the degree of metastasis and the survival of cancer patients. Although CTC-based diagnostic approaches are being rapidly developed, limited studies have proven the benefits of CTC elimination, with most studies providing only hypothetical inference because of the technical difficulty in examining the effects of CTC elimination in vivo. We modified photodynamic therapy to specifically eliminate green fluorescent protein (GFP)-expressing CTCs and evaluated the therapeutic efficacy of CTC elimination. When circulating blood is illuminated with a blue laser (λ = 473 nm), the combination of GFP and photosensitizers induces a selective elimination of GFP-expressing CTCs, with limited effect on normal cells. In GFP-expressing cancer cell-infused or transplanted mice models, the treatment suppressed distant metastasis and extended the survival of the tumor-bearing mice. Taken together, CTCs are a core seed to be metastasized into secondary organs and elimination of CTCs may improve the survival of cancer patients.
KeywordsCirculating tumor cells Green fluorescent protein Metastasis Photodynamic therapy Photosensitizers
Circulating tumor cells
Green fluorescent protein
Circulating tumor cells (CTCs) present in the vascular system are tumor cells that will metastasize from primary or disseminated tumors . Rapid advancements in detection and isolation techniques have led to the remarkable discoveries on the role of CTCs and their association with cancer prognosis [2, 3, 4, 5, 6, 7]. Since an increased number of CTCs are associated with poor prognosis, CTC-targeted therapies may provide a promising new approach which could improve cancer prognosis [8, 9]. However, the unpredictable nature and dynamics of CTCs and the lack of adequate treatment modalities hamper the selective targeting of CTCs.
CTC-targeting PDT was also performed in mice with GFP+ metastatic 4T1 cells transplanted into their flanks (Fig. 2c). No changes in primary tumor size (Additional file 3) were observed between treated (irradiated) mice and untreated mice, implying limited effects on GFP− normal cells; however, the numbers of CTCs observed in the fluorescent images were significantly decreased in the treated mice compared to those the untreated mice (Fig. 2d and Additional file 4). In the treated group, the number of lung metastatic nodules in the treated mice was significantly lower compared to that in the untreated group (Fig. 2e). Mice receiving treatment for 1 week showed survival gain compared with untreated mice (P = 0.0325) (Fig. 2f). However, the difference was more significant in the mice treated for 2 weeks (P = 0.0026). There was no hematologic difference between the untreated group and the 2 weeks treatment group (Additional file 5). Materials and methods are described in Additional file 6.
To prove the benefits of CTC elimination, we developed an energy transfer-based PDT that targets GFP-expressing CTCs. Using this technique, we attempted to eliminate CTCs and optimize conditions to specifically target CTCs, with minimum damage to normal cells. To our knowledge, this is the first experimental study to demonstrate that the direct killing of CTCs extends survival in vivo. The present study highlights the concept of energy distinction between normal and cancer cells by using a new factor, i.e., cancer cell-specific fluorescence.
Although this is a preliminary study using the externally fluorescence-labeled cancer cells and the injected mouse models, thus, this strategy is not suitable for in vivo targeting therapeutics of CTC; we reveal that clearance of CTC is associated with the reduction of metastasis and extension of survival. In addition, this experiment directly suggests CTCs are a core seed to be metastasized into secondary organs. Advancements in the field of molecular diagnostics have made it possible to use combinations of fluorescence proteins and photosensitizers or molecular-targeted photosensitizers in diverse biological fields, including cancer stem cell-targeted therapy.
This work was supported by the grants from Kyung Hee University (KHU-20170844 for JW Choi) and the National R&D Program for Cancer Control, Ministry of Health and Welfare, Republic of Korea (HA17C0039 for YR Kim and CW Jeong).
Availability of data and materials
All data generated or analyzed during this study are included in this published article and its supplementary information files.
YRK and JWC designed the study. YRK and JKY performed in vitro and in vivo experiments. YRK and JWC analyzed the data. JWC created figures for the results. YRK, CWJ, and JWC wrote the manuscript with inputs from all the authors and reviewed the manuscript. All authors read and approved the final manuscript.
Ethics approval and consent to participate
All animal experiments were approved by the Institutional Animal Care and Use Committee at Kyung Hee University (KHSIRB 18-014) and were performed in compliance with the institutional guidelines.
Consent for publication
The authors declare that they have no competing interests.
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