Targeting metastasis-initiating cancer stem cells in gastric cancer with leukaemia inhibitory factor

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Abstract​

Gastric cancer’s (GC) bad prognosis is usually associated with metastatic spread. Invasive cancer stem cells (CSC) are considered to be the seed of GC metastasis and not all CSCs are able to initiate metastasis. Targeting these aggressive metastasis-initiating CSC (MIC) is thus vital. Leukaemia inhibitory factor (LIF) is hereby used to target Hippo pathway oncogenic members, found to be induced in GC and associated with CSC features. LIF-treated GC cell lines, patient-derived xenograft (PDX) cells and/or CSC tumourspheres underwent transcriptomics, laser microdissection-associated proteomics, 2D and 3D invasion assays and in vivo xenograft in mice blood circulation. LIFR expression was analysed on tissue microarrays from GC patients and in silico from public databases. LIF-treated cells, especially CSC, presented decreased epithelial to mesenchymal transition (EMT) phenotype and invasion capacity in vitro, and lower metastasis initiation ability in vivo. These effects involved both the Hippo and Jak/Stat pathways. Finally, GC’s high LIFR expression was associated with better clinical outcomes in patients. LIF treatment could thus represent a targeted anti-CSC strategy to fight against metastatic GC, and LIFR detection in primary tumours could constitute a potential new prognosis marker in this disease.

 

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Introduction​

GC is a deadly disease accounting for ~768,793 deaths worldwide in 2020, making it the 4th leading cause of cancer-related death [1]. GC therapy comprises surgery with additional adjuvant or neo-adjuvant chemo- and radio-therapies and the number of relapses remains high. Surgical resection which remains the only curative therapeutic option is possible in case of early tumours but GC is commonly a late-diagnosed disease with 80% of patients detected at metastatic state and with a 5-year survival rate of <5% [2, 3]. It is thus crucial to find appropriate diagnostic tools for earlier detection of GC as well as strategies for better-targeted therapy of disseminated cancers.
The most aggressive component of heterogeneous gastric tumours is the rare population of cancer stem cells (CSC) which carry specific self-renewal and chemo-/radio-resistant properties [4,5,6,7], allowing them to initiate tumours and cause recurrence. In the metastatic process, part of these CSC corresponding to metastasis-initiating CSC (MIC), is able to acquire invasive characteristics, evade the primary tumour, disseminate, and colonise distant organs to initiate metastases [8, 9]. We have recently identified CD44v3+ cells as MIC in gastric tumours, associated with bad prognosis in GC [8] and which could constitute an interesting therapeutical target for GC metastatic disease. CD44v3+ cells are a subpopulation of gastric CD44+ CSC and our previous work has demonstrated an enrichment of the Hippo pathway oncogenic members Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) in GC and especially in gastric CSC [10,11,12]. YAP and TAZ are well-described for their role in invasion and metastasis and their targeting could affect GC MIC.
We have demonstrated that activating the Hippo kinase core with leukaemia inhibitory factor (LIF) decreases tumorigenic properties of gastric CSC [4]. Nevertheless, LIF/LIF receptor (LIFR) signalling was never tested in the gastric metastatic context despite its promising anti-metastatic effects in hepatocellular carcinoma [13], clear cell renal carcinoma [14] and breast cancer [15].
LIF–LIF receptor (LIFR) canonical pathway is mediated by JAK/STAT activation, causing STAT3 phosphorylation, dimerisation, and nuclear translocation, resulting in the transcription of target genes involved in cell survival and tumorigenesis [16]. Other cell signalisation processes have been identified downstream of LIF-LIFR, which could explain its pleiotropy [16,17,18,19,20,21,22,23]. Nonetheless, LIF-LIFR-dependent anti-metastatic properties in breast cancer implicated the Hippo pathway activation [15].
The aim of this study was to decipher LIF’s potential as anti-metastatic therapy in GC and the underlying signalling mechanisms. We have shown that LIF decreased invasive properties of GC cell lines and patient-derived xenograft (PDX) cells and more especially gastric CSC in vitro through activation of Hippo kinases and repression of YAP/TAZ oncogenic signalling. LIF’s anti-invasive function involves the repression of CD44v3 expression and of the epithelial-to-mesenchymal transition (EMT) programme in CSC, important in the GC metastatic process. LIF-treated cells were also less keen to form metastasis compared to non-treated cells when injected into mice’ blood circuits. Finally, LIFR’s high expression in gastric tumours seems to be protective since it is correlated to better prognosis of GC patients despite them having CD44v3+ MIC-rich or mesenchymal ZEB1+ tumours.

 

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Surgical resection which remains the only curative therapeutic option is possible in case of early tumours but GC is commonly a late-diagnosed disease with 80% of patients detected at metastatic state and with a 5-year survival rate of <5% [2, 3]. It is thus crucial to find appropriate diagnostic tools for earlier detection of GC as well as strategies for better-targeted therapy of disseminated cancers.

Now you know why gastric cancer is deadly because surgical resection “remains the only curative therapeutic option” for gastric cancer in 2024 and this can only be done in case of early tumors provided the cancer has not metastasized.