Limiting tumor seeding as a therapeutic approach for metastatic disease

Abstract

Here we propose that therapeutic targeting of circulating tumor cells (CTCs), which are widely understood to be the seeds of metastasis, would represent an effective strategy towards limiting numerical expansion of secondary lesions and containing overall tumor burden in cancer patients. However, the molecular mediators of tumor seeding have not been well characterized. This is in part due to the limited number of pre-clinical in vivo approaches that appropriately interrogate the mechanisms by which cancer cells home to arresting organs. It is critical that we continue to investigate the mediators of tumor seeding as it is evident that the ability of CTCs to colonize in distant sites is what drives disease progression even after the primary tumor has been ablated by local modalities. In addition to slowing disease progression, containing metastatic spread by impeding tumor cell seeding may also provide a clinical benefit by increasing the duration of the residence of CTCs in systemic circulation thereby increasing their exposure to pharmacological agents commonly used in the treatment of patients such as chemotherapy and immunotherapies. In this review we will examine the current state of knowledge about the mechanisms of tumor cells seeding as well as explore how targeting this stage of metastatic spreading may provide therapeutic benefit to patients with advanced disease.

Keywords: CX3CR1; Chemokines; Integrins; Metastasis; Seeding; Selectins.

Fig. 1

Cancer cells mobilized from disseminated tumors reseed additional lesions. Mice harboring disseminated tumors generated by 4 T-1 murine breast cancer cells and reproducing early metastatic disease were treated with the CXCR4 antagonist AMD-3100, which dislodged cancer cells from the existing tumors and doubled the number of additional lesions as compared to control animals, (Control 11 mice/group, Treated 7 mice/group; ***P = .0002, One-way Anova with Dunnett’s post-test). Reproduced with permission from (Qian et al., 2018).

Fig. 2

Impeding reseeding by targeting CX3CR1 prolongs the time CTCs spend in circulation and promotes cell death. (A) The chemical structure of JMS-17–2, a novel, potent and selective small-molecule antagonist of CX3CR1. (B) CTCs were enumerated at different time points following administration of AMD-3100 alone or combined with the improved CX3CR1 antagonist FX-68 to mice harboring disseminated tumors. The area under the curve was measured as 485 for AMD-3100 alone and 852 for AMD3100 + FX-68, which equates to a 75% increase induced by the CX3CR1 antagonist (shaded area). The red-dotted box indicates the numerical range of CTCs detected at steady-state, i.e in the absence of any treatment (3 mice/ group; *P = .04 paired Student’s t-test; **P = .02 One-way Anova with Dunnett’s post-test. (C) The combination of FX-68 with AMD-3100 fully blunted the increase in additional lesions caused by the administration of AMD-3100 alone (refer also to Fig. 1) (Control 11 mice, Treated 7 mice/group; ***P = .0002, One-way Anova with Dunnett’s post-test). (D) CTCs collected upon treatment with AMD-3100 alone or AMD-3100 + FX-68 were collected at 6 h and 12 h (refer also to B) and levels of Bax and Bcl2 transcripts were measured by qRT-PCR as indication of the extent of apoptotic cells for each time-point and treatment. Bax expression was found to be dramatically increased at 6 h when the reseeding of CTCs was impaired by FX-68; at 12 h the apoptotic fractions were comparable between CTCs mobilized by AMD-3100 in the presence or absence of FX-68. All results were normalized to Bax and Bcl2 expression measured in CTCs collected at steady state (red dotted line) **P = .01 One-way Anova with Dunnett’s post-test. Reproduced with permission from (Shen et al., 2016) and (Qian et al., 2018).

Fig. 3

Obstructing the reseeding of CTCs improves drug exposure in blood. Retaining CTCs in the blood by administering FX-68 increased the exposure to Doxorubicin, as measured by the percentage of cells showing red fluorescence emitted by the drug. Yellow arrows show two cancer cells that did not incorporate Doxorubicin (3 mice/group; *P = .03, One-way Anova with Dunnett’s post-test. Reproduced with permission from (Qian et al., 2018).

Fig. 4

Rationale for therapeutic targeting of tumor seeding to prevent of metastasis. (A) CTCs utilize the adhesive molecular interactions between CX3CR1 and the cell-anchored form of Fractalkine to facilitate extravasation through the endothelium. After successful extravasation, newly seeded cancer cells migrate in response to the chemoattractant gradient established by the soluble Fractalkine released from cells of the surrounding stroma. Given a conducive microenvironment, the DTCs can proliferate and form metastases in the secondary organ. (B) Administering a CX3CR1 antagonist blocks the initial CX3CR1-fractalkine interaction thereby preventing extravasation. Blocking of tumor cell seeding has a dual benefit. Failed extravasation will cause CTCs to be retained in circulation, which consequentially increases their exposure to chemotherapeutic agents and improves clinical outcome. In addition, CTCs with chemoresistant phenotypes will have extended retention in circulation and eventually undergo programed cell death due to their prolonged detachment from the extracellular matrix (anoikis). Schematic created with BioRender graphics web application.