It is hypothesized that tumor-initiating cells (TICs) with stem cell-like properties constitute a sustaining force to drive tumor growth and renew fully established malignancy. However, the identification of such a population in non-small cell lung carcinoma (NSCLC) has been hindered by the lacking of reliable surface markers, and very few of the currently available surface markers are of functional significance. Here, we demonstrate that a subpopulation of TICs could be specifically defined by the voltage-gated calcium channel α2δ1 subunit from non-small cell lung carcinoma (NSCLC) cell lines and clinical specimens. The α2δ1⁺ NSCLC TICs are refractory to conventional chemotherapy, and own stem cell-like properties such as self-renewal, and the ability to generate heterogeneous tumors in NOD/SCID mice. Moreover, α2δ1⁺ NSCLC cells are more enriched for TICs than CD133⁺, or CD166⁺ cells. Interestingly, α2δ1 is functionally sufficient and indispensable to promote TIC properties by mediating Ca²⁺ influx into cells, which subsequently activate Calcineurin/NFATc2 signaling that directly activates the expression of NOTCH3, ABCG2. Importantly, a specific antibody against α2δ1 has remarkably therapeutic effects on NSCLC xenografts by eradicating TICs. Hence, targeting α2δ1 to prevent calcium influx provides a novel strategy for targeted therapy against TICs of NSCLC.

Analysis of T cell receptor (TCR) repertoires may contribute to better understanding of the response to immunotherapy. By deep sequencing of the TCR β chain complementarity-determining regions in the paired biopsies and peripheral blood specimens of 31 patients with non–small cell lung cancer (NSCLC) treated with anti–programmed death 1 (PD-1) or PD-ligand 1 (PD-L1) therapy, we developed a previously unidentified index, the TCR-based immunotherapy response index (TIR index), that estimated the degree of overlap of the TCR repertoire between tumor-infiltrating lymphocytes and circulating PD-1⁺CD8⁺T cells (shared TCR clones). This index correlated with response and survival outcomes of anti–PD-(L)1 treatment. All the TCR sequences of neoantigen-stimulated T cells were included in the shared TCR clones, indicating that TCR clones involved in TIR index estimation represented tumor-specific T cells. Therefore, the TIR index is a feasible approach for assessing tumor-specific TCR and stratifying patients with NSCLC for anti–PD-(L)1 therapy.

Accumulating evidence has shown that the traits of tumor-initiating cells (TICs) are controlled by the microenvironment niches (MENs), but the composition and remodeling mechanisms of the MENs of TICs are poorly defined. Here, we report that the voltage-gated calcium channel α2δ1 subunit-positive TICs of hepatocellular carcinoma (HCC) specifically secret lysyl oxidase (LOX), which leads to the cross-linking of collagen, forming a stiff extracellular matrix (ECM) that is sufficient to drive the formation of TICs with a stiff mechanical trait and is subsequently required for the maintenance the properties of HCC TICs. Furthermore, the cross-linked collagen results in the upregulation of integrin α7 (ITGA7), increased phosphorylation of FAK and extracellular signal-regulated kinase 1/2 (ERK1/2). Inhibition of ITGA7 abolishes all the effects of cross-linked collagen mediated by LOX. Hence, the α2δ1⁺ HCC TICs initiate ECM remodeling by secreting LOX to create a stiff MEN of TIC with cross-linked collagen, which drives the acquisition and subsequent maintenance of the properties of HCC TICs through ITGA7-FAK-ERK1/2 signaling pathway.

Background: Long non-coding RNA (lncRNA) was identified as a novel diagnostic biomarker in gastric cancer (GC). However, the functions of lncRNAs in immuno-microenvironments have not been comprehensively explored. In this study, we explored a critical lncRNA, LOC339059, that can predict the clinical prognosis in GC related to the modulation of PD-L1 and determined its influence upon macrophage polarization via the IL-6/STAT3 pathway. Methods: To date, accumulating evidence has demonstrated that the dysregulation of LOC339059 plays an important role in the pathological processes of GC. It acts as a tumor suppressor, regulating GC cell proliferation, migration, invasion, tumorigenesis, and metastasis. A flow cytometry assay showed that the loss of LOC339059 enhanced PDL1 expression and M2 macrophage polarization. RNA sequencing, RNA pull-down, RNA immunoprecipitation, Chip-PCR, and a luciferase reporter assay revealed the pivotal role of signaling alternation between LOC339059 and c-Myc. Results: A lower level of LOC339059 RNA was found in primary GC tissues compared to adjacent tissues, and such a lower level is associated with a poorer survival period (2.5 years) after surgery in patient cohorts. Moreover, we determined important immunological molecular biomarkers. We found that LOC339059 expression was correlated with PD-L1, CTLA4, CD206, and CD204, but not with TIM3, FOXP3, CD3, C33, CD64, or CD80, in a total of 146 GC RNA samples. The gain of LOC339059 in SGC7901 and AGS inhibited biological characteristics of malignancy, such as proliferation, migration, invasion, tumorigenesis, and metastasis. Furthermore, our data gathered following the co-culture of THP-1 and U937 with genomic GC cells indicate that LOC339059 led to a reduction in the macrophage cell ratio, in terms of CD68⁺/CD206⁺, to 1/6, whereas the selective knockdown of LOC339059 promoted the abovementioned malignant cell phenotypes, suggesting that it has a tumor-suppressing role in GC. RNA-Seq analyses showed that the gain of LOC339059 repressed the expression of the interleukin family, especially IL-6/STAT3 signaling. The rescue of IL-6 in LOC339059-overexpressing cells reverted the inhibitory effects of the gain of LOC339059 on malignant cell phenotypes. Our experiments verified that the interaction between LOC339059 and c-Myc resulted in less c-Myc binding to the IL-6 promoter, leading to the inactivation of IL-6 transcription. Conclusions: Our results establish that LOC339059 acts as a tumor suppressor in GC by competitively inhibiting c-Myc, resulting in diminished IL-6/STAT3-signaling-mediated PDL1 expression and macrophage M2 polarization.

Purpose: Cancer stem cells (CSCs) are a subpopulation of cancer cells with self-renewal property and responsible for tumor malignancy, progression and drug resistance. Researches on CSC-specific markers in gastric cancer remain limited. Our current study explored the expression of voltage-dependent calcium channel α2δ1 subunit and the potential of using α2δ1 as a CSC marker in gastric cancer. We also compared the specificity of α2δ1 and CD44 in identifying gastric cancer stem cells (GCSCs). Materials and methods: Expression of α2δ1 was analyzed in gastric cancer cell lines, patient-derived xenograft (PDX) models and clinical samples of malignant ascites of gastric cancer patients. α2δ1^+, gastric cancer cells were isolated from gastric cancer cell lines. CSC properties of α2δ1^+, gastric cancer cells were then verified by subsequent tests both in vitro and in vivo. Results: The expression level of α2δ1 was found to differ drastically among gastric cancer cell lines, PDX models and clinical samples of malignant ascites. α2δ1^+, gastric cancer cells sorted from HGC-27 and SGC-7901 cell lines demonstrated significant self-renewal properties, including tumorigenic capacity, sphere-formation capacity and asymmetric differentiation potential. Knockdown of α2δ1 in α2δ1^+, HGC-27 significantly inhibited CSC properties and rendered HGC-27 more sensitive to chemotherapy. Flow cytometry showed that α2δ1^+, gastric cancer cells accounted for a small fraction of CD44^+, gastric cancer cells. Isolated CD44^+,α2δ1^+, HGC-27 cells displayed more significant tumorigenic capacity and sphere-forming capacity compared with their CD44^+,α2δ1^−, counterparts. Conclusion: α2δ1^+, gastric cancer cells possessed CSC properties. α2δ1 could be a proper marker in identifying GCSCs with superior specificity than CD44. The combination of α2δ1 and CD44 could be used to identify GCSCs with improved accuracy. Knockdown of α2δ1 combined with chemotherapy displayed higher therapeutic efficacy on gastric cancer cells, suggesting that α2δ1 could be a potential target for anticancer treatment.

Lung adenocarcinoma (LUAD) is a leading cause of cancer-related mortality, with heme metabolism playing a critical role in tumor progression and treatment resistance. This study investigates the clinical implications of heme metabolism in LUAD, focusing on its link to ferroptosis and drug sensitivity. Using multi-omics data from TCGA-LUAD, GEO databases, and a single-cell RNA-seq cohort, we identified two molecular subtypes based on heme metabolism-related genes. We further developed a prognostic panel, termed the heme metabolism risk score (HMRS), using LASSO and multivariate Cox regression analyses. The HMRS panel effectively stratified patients into high- and low-risk groups, with high-risk patients showing enhanced tumor proliferation, suppressed ferroptosis, and resistance to chemotherapy. Single-cell analysis revealed elevated heme metabolism risk in epithelial cells correlated with tumor progression. Drug sensitivity predictions were validated in platinum-based chemotherapy cohorts, confirming HMRS as a robust prognostic tool. ABCC2 was identified as a key regulator of ferroptosis and cisplatin resistance, with in vitro experiments demonstrating that ABCC2 knockdown enhanced cisplatin-induced ferroptosis. These findings highlight HMRS as a critical tool for patient stratification and ABCC2 as a promising therapeutic target to overcome cisplatin resistance.