Circulating Tumour Cells (CTCs) in NSCLC: From Prognosis to Therapy Design

Abstract

Designing optimal (neo)adjuvant therapy is a crucial aspect of the treatment of non-small-cell lung carcinoma (NSCLC). Standard methods of chemotherapy, radiotherapy, and immunotherapy represent effective strategies for treatment. However, in some cases with high metastatic activity and high levels of circulating tumour cells (CTCs), the efficacy of standard treatment methods is insufficient and results in treatment failure and reduced patient survival. CTCs are seen not only as an isolated phenomenon but also a key inherent part of the formation of metastasis and a key factor in cancer death. This review discusses the impact of NSCLC therapy strategies based on a meta-analysis of clinical studies. In addition, possible therapeutic strategies for repression when standard methods fail, such as the administration of low-toxicity natural anticancer agents targeting these phenomena (curcumin and flavonoids), are also discussed. These strategies are presented in the context of key mechanisms of tumour biology with a strong influence on CTC spread and metastasis (mechanisms related to tumour-associated and -infiltrating cells, epithelial-mesenchymal transition, and migration of cancer cells).

Keywords: CTCs; NSCLCs; curcumin; flavonoids; metastasis suppression.

Figure 1

Structure of curcuminoids and flavonoids tested for suppression of NSCLC metastasis.

Figure 2

Simplified model of curcumin and flavonoids effects on the NSCLC microenvironment [83,119,148,158,159,162,168,170,174,175,186,187,188,189,190,191,192,193,194,195,196]. A necessary part of igenesis and CTC spreading is the interaction of NSCLC cells with tumour-associated cells. NSCLC cells recruit monocytes via IL-17 into tumour tissue. Signalling factors (e.g., IL-6, IL-8, TNF-α, and PGE2) produced in the tumour microenvironment stimulate monocyte differentiation into TAM2 (which support NSCLC cell proliferation, EMT, chemoresistance and immune resistance, and EM disruption). MDSCs recruited via IL-8 and TGF-β1 repress the cytotoxic effects of T cells against NSCLC cells and induce their differentiation into Treg cells that are responsible for the suppression of the host immune response. VEGF-recruited TECs affect EMT, chemoresistance, and the proliferation of NSCLC cells, recruit MDSCs and induce angiogenesis. IL-8-activated CAFs decrease the anticancer immune response (with the support of TAM2 and MDSCs) and repress T cells. CAFs stimulate EMT and induce NSCLC cell proliferation, migration, and drug resistance. Tumour-associated cells help sustain the tumour microenvironment and aggressive metastatic phenotype. TAM2, CAFs, and TECs are co-inducers of EMT and thereby CTC spreading, chemoresistance, and immune resistance. MDSCs and Treg cells repress the host immune response and thereby support CTC survival in the blood. Nevertheless, the anticancer effect of curcuminoids and flavonoids is not dependent on targeting of NSCLC cells, as they repress other important parts of the complex tumour ecosystem. The application of such agents is associated with stimulation of the immune system (higher levels of TAM1 and T cells; lower levels of Th1 cells, TAM2, and Treg cells; and lower recruitment of monocytes and MDSCs). Curcuminoids and flavonoids also decrease the levels of CAFs and TECs and repress their interaction with NSCLC cells. In addition, the application of curcuminoids and flavonoids leads to a decrease in the proliferation, survival, chemoresistance, immunoresistance, and migration ability of NSCLC cells. CAF, cancer-associated fibroblast; EM, extracellular matrix; EMT, epithelial–mesenchymal transition; EGF, endothelial growth factor; FasL, Fas ligand; IL-1β, interleukin 1β; IL-6, interleukin 6; IL-8, interleukin 8; PGE2, prostaglandin E2; NOS, nitric oxide species; ROS, reactive oxygen species; TAM1, tumour-associated macrophage M1; TAM2, tumour-associated macrophage M2; TEC, tumour endothelial cell; TGF-β1, transforming growth factor beta 1; Th1, T helper 1; Treg, regulatory T; TNF-α, tumour necrosis factor alpha; VEGF, vascular endothelial growth factor. Green arrow = induction/activation of factor/phenomenon/cell; red arrow = repression/inhibition of factor/phenomenon/cell; blue arrow = differentiation of immune cells; ↑ = curcumin/flavonoids activation/induction; ↓ = curcumin/flavonoids repression/inhibition.

Figure 3

Simplified model of curcumin and flavonoids action on NSCLC metastasis [83,190,223,224,229,230,231,236,237,238,239,240,242,243,244,245,246,247,248,249]. 1. NSCLC cells with a mesenchymal phenotype partially induced by TAM2, TECs, and CAFs migrate from tissue into blood vessels. 2. CTCs are transported (via passive transport) to distant metastatic site(s), and TECs can serve as guardians against anoikis apoptosis. 3. Infiltrating NSCLC cells revert to an epithelial phenotype via MErT (which possibly occurs in blood). 4. Dormant cancer cells activated by Wnt signalling start recruiting CAFs and TECs and form micrometastases. 5 Micrometastases stabilised by oxidative stress recruit tumour-associated cells to form macrometastases, at which point metastatic NSCLC can be diagnosed in patients. TECs and CAFs can support the formation of highly aggressive and metastatic CTC clusters. CAF, cancer-associated fibroblast; EMT, epithelial–mesenchymal transition; EGF, endothelial growth factor; IL-1β, interleukin 1β; IL-6, interleukin 6; IL-8, interleukin 8; PGE2, prostaglandin E2; ROS, reactive oxygen species; TAM2, tumour-associated macrophage M2 phenotype; TEC, tumour endothelial cell; TGF-β1, transforming growth factor beta 1; VEGF, vascular endothelial growth factor. Green arrow = induction/activation of factor/phenomenon; red arrow = repression/inhibition of factor/phenomenon; blue arrow = transition between individual steps of metastatic spread; ↓ = curcumin/flavonoids repression/inhibition.

Figure 4

Simplified model of the effects of curcumin on NSCLC cells [128,196,242,243,244,245,248,249,276,279,280,281,282,283,284]. NSCLC is associated with dysregulation of numerous signalling and regulatory pathways. Some signalling factors (e.g., IL-6, IL-8, EGF, TGF-β1, and Wnt) produced by cancer or tumour-associated cells can induce migration and invasion phenotypes of NSCLC cells and thereby support CTC spreading. These pathways are interconnected, and dysregulation of one can induce dysregulation of another and modulate therapeutic targeting. Nevertheless, curcumin targets multiple signalling pathways to repress this phenomenon. Its effect is associated with repression of ERK2, JAK, STAT3, EGFR, PI3K, Akt, SMAD 2/3, and β-catenin. In addition, curcumin induces oxidative stress in the endoplasmic reticulum by inhibiting TrxR1 and mitochondrial-dependent and mitochondrial-independent apoptotic pathways and repressing drug resistance. Reduced levels of signalling factors produced by NSCLC cells, such as IL-6, 8, VEGF, and PGE2, lead to decreased activity of tumour-associated/tumour-infiltrating cells and thereby decreased support of NSCLC cell metastasis, proliferation, and survival in the tumour microenvironment. Curcumin’s effects on the tumour microenvironment also include activation of the immune system and suppression of angiogenesis and chemoresistance and immune resistance. Akt, protein kinase B; Bcl-2, B cell lymphoma 2; CAF, cancer-associated fibroblast; COX-2, cyclooxygenase-2; EGF, endothelial growth factor; EGFR, endothelial growth factor receptor; ERK2, extracellular signal-regulated kinase 2; GEF, guanine nucleotide exchange factor; HIF-1α, hypoxia-inducible factor 1α; JAK, Janus tyrosine kinase; JNK, c-Jun N-terminal kinase; IL-1β, interleukin 1β; IL-6, interleukin 6; IL-6R, interleukin 6 receptor; IL-8, interleukin 8; IL-8R, interleukin 8 receptor; KATP, ATP-sensitive potassium channel; LRP-6, low-density lipoprotein receptor-related protein 6; MAPK, mitogen-activated protein kinase; MDSC, myeloid-derived suppressor cell; mTOR, mammalian target of rapamycin; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; P70S6K, ribosomal protein S6 kinase beta-1; p-eIF2α, phosphorylated eukaryotic translation initiation factor 2 subunit 1; PGE2, prostaglandin E2; PI3K, phosphoinositide 3-kinase; ROS, reactive oxygen species; STAT3, signal transducer and activator of transcription 3; SRC, intracytoplasmic tyrosine kinase; TAM2, tumour-associated macrophage M2 phenotype; TEC, tumour endothelial cell; TGF-β, transforming growth factor beta; TGF-β1R, transforming growth factor beta 1 receptor; TNF-α, tumour necrosis factor alpha; Trx1, thioredoxin reductase 1; VEGF, vascular endothelial growth factor; ΔΨm, mitochondrial membrane potential. Green arrow = induction/activation of factor/phenomenon/cell, dotted (indirect); red arrow = repression/inhibition of factor/phenomenon/tumour-supporting cell; green factor = anticarcinogenic factor; red factor = carcinogenic factor; ↑ = curcumin activation/induction; ↓ = curcumin repression/inhibition.

Figure 5

Curcuminoids and flavonoids in migrastatic and cytostatic (MICY) therapy. Green arrow = induction/activation of CTC; ↑ = curcuminoids/flavonoids activation/induction; ↓ = curcuminoids/flavonoids repression/inhibition.