Lung cancer is the leading cause of cancer-related mortality worldwide due to its early asymptomatic and late metastasis. While cancer stem cells (CSCs) may play a vital role in oncogenesis and development of lung cancer, mechanisms underlying CSCs self‐renewal remain less clear. In the present study, we constructed a clinically relevant CSCs enrichment recognition model and evaluated the potential functions of phosphatidylinositol 3-kinase (PI3K)/AKT pathway (PI3K/AKT) and mitogen-activated protein kinases/extracellular signal-regulated kinase (MAPK/ERK) pathways in lung cancer via bioinformatic analysis, providing the basis for in depth mechanistic inquisition. Experimentally, we confirmed that PI3K/AKT pathway predominantly promotes proliferation through anti-apoptosis in lung adenocarcinoma cells, while MAPK/ERK pathway has an overwhelming superiority in regulating the proliferation in lung CSCs. Further, utilizing stemness score model, LLC-Symmetric Division (LLC-SD) cells and mouse orthotopic lung transplantation model, we elucidated an intricate cross-talk between the oncogenic pathway and the stem cell reprograming pathway that impact stem cell characteristics as well as cancer biology features of lung CSCs both in vitro and in vivo. In summary, our findings uncovered a new insight that PI3K/AKT and MAPK/ERK pathways as oncogenic signaling pathway and/or stem cell signaling pathway act distinctively and synergistically to regulate lung CSCs self-renewal.

Mitophagy, a form of autophagy, plays a role in cancer development, progression and recurrence. Cancer stem cells (CSCs) also play a key role in these processes, although it not known whether mitophagy can regulate the stemness of CSCs. Here, we employed the A549‐SD human non‐small cell lung adenocarcinoma CSC model that we have developed and characterized to investigate the effect of mitophagy on the stemness of CSCs. We observed a positive relationship between mitophagic activity and the stemness of lung CSCs. At the mechanistic level, our results suggest that augmentation of mitophagy in lung CSCs can be induced by FIS1 through mitochondrial fission. In addition, we assessed the clinical relevance of FIS1 in lung adenocarcinoma using The Cancer Genome Atlas database. An elevation in FIS1, when observed together with other prognostic markers for lung cancer progression, was found to correlate with shorter overall survival.

The mean survival of metastatic lung adenocarcinoma is less than 1 year, highlighting the urgent need to understand the mechanisms underlying its high mortality rate. The role of Extracellular vesicles (EVs) in facilitating the interactions between cancer cells and the metastatic microenvironment has garnered increasing attention. Previous studies on the role of EVs in metastasis have been primarily focused on cancer cell-derived EVs in modulating the functions of stromal cells. However, whether cancer stem cells (CSCs) can alter the metastatic properties of non-CSC cells, and whether EV crosstalk can mediate such interaction, have not been demonstrated prior to this report. In the present study, we integrated multi-omics sequencing and public database analysis with experimental validation to demonstrate, for the first time, the exosomal Mir100hg, derived from CSCs, could enhance the metastatic potential of non-CSCs both in vitro and in vivo. Mechanistically, HNRNPF and HNRNPA2B1 directly binds to Mir100hg, facilitating its trafficking via exosomes to non-CSCs. In non-CSCs, Mir100hg upregulates ALDOA expression, subsequently leading to elevated lactate production. Consequently, the increased lactate levels enhance H3K14 lactylation by 2.5-fold and promote the transcription of 169 metastasis-related genes. This cascade of events ultimately results in enhanced ALDOA-driven glycolysis and histone lactylation-mediated metastatic potential of non-CSC lung cancer cells. We have delineated a complex regulatory network utilized by CSCs to transfer their high metastatic activity to non-CSCs through exosomal Mir100hg, providing new mechanistic insights into the communication between these two heterogeneous tumor cell populations. These mechanistic insights provide novel therapeutic targets for metastatic lung cancer, including HNRNPF/HNRNPA2B1-mediated Mir100hg trafficking and the histone lactylation pathway, advancing our understanding of CSC-mediated metastasis while suggesting promising strategies for clinical intervention.

Despite the unprecedented advancement of cancer treatment, the prognosis for patients with metastatic stage of cancer remains poor. The challenge that underlines this clinical dilemma is the complexity of metastasis. The conventional experiment-driven discovery approaches (the "wet lab") yield overly simplified one-to-one mechanistic relationships that are inept of elucidating the complexity of metastasis. Metastasis research also suffers from the knowledge and skill deficiency of the individual investigators. The importance of the present study is the demonstration that the "dry-lab-driven discovery and wet-lab validation" approach can improve the efficiency of studying complex biological behaviors, and can yield more reliable, objective and comprehensive mechanistic findings that are have clinical significance. Specifically, we applied this approach to study the mechanisms that underline the involvement of exosomal miRNAs in transferring the metastatic capability between heterogenous melanoma cancer cells. We show that the highly metastatic melanoma tumor cells (POL) can transfer their metastatic competency to the low-metastatic melanoma tumor cells (OL) by exosomal miR-211-5p. The oncogenic activity of miR-211-5p is mediated by the target gene guanine nucleotide-binding protein subunit alpha-15 (GNA15) through modifying the immune function of the tumor microenvironment extrinsically; as well as through inhibiting pyroptosis and augmenting glycolysis within OL cells intrinsically. In addition, we show that exosomal sorting of miR-211-5p is like selective and is subjected to regulation by a transcriptional feedback loop between miR-211-5p and zinc finger FYVE-type containing 26 (ZFYVE26). Furthermore, the "8-genes pyroptosis Risk model" derived from LASSO regression analysis was verified as an independent prognostic factor for melanoma.

Background: Exosomes are a new class of molecular entities in the metastatic microenvironment, which can mediate bidirectional communication between cells. While exosomes-mediated interactions between tumor cells and other cell populations in the tumor microenvironment have attracted most attention, little is known about the significance of exosomes in mediating the interaction between non-stemness cancer cells and cancer stem cells during cancer progression. Methods: The structure, sequence and downstream target miRNAs of lncRNA Mir100hg were predicted by online web resources. The bioinformatics prediction results were validated with experimental verification: exosome tracing, electron microscopy, Luciferase assay, metabolomics sequencing and mouse tail vein model of pulmonary metastasis. A complex regulatory network of "cancer stem cells-exosomal lncRNA-non-stem cancer cells" was constructed. Results: This study demonstrates firstly that lncRNA Mir100hg is upregulated in lung cancer stem cell LLC-SD (Lung cancer stem cells) and can be delivered to non-stemness cancer cells LLC (Lewis lung cancer cells) via exosomes. In LLC, Mir100hg targets miR-15a-5p and miR-31-5p which leads to the increase of the global glycolytic activity of lung cancer cells and consequently, the enhancement of their metastatic capability. Conclusion: We delineated a complex regulatory network that utilized by cancer stem cells to transfer their high metastatic activity to the low-metastatic non-stemness cancer cells through exosomal Mir100hg, thereby providing new mechanistic insights into the communication between two heterogeneous tumor cells.