Minimally invasive thermal therapy is a successful alternative treatment to surgery in solid tumors with high complete ablation rates, however, tumor recurrence remains a concern. Central memory CD8+ T cells (TCM) play important roles in protection from chronic infection and cancer. Here we find, by single-cell RNA analysis of human breast cancer samples, that although the memory phenotype of peripheral CD8+ T cells increases slightly after microwave ablation (MWA), the metabolism of peripheral CD8+ T cells remains unfavorable for memory phenotype. In mouse models, glycolysis inhibition by 2-deoxy-D-glucose (2DG) in combination with MWA results in long-term anti-tumor effect via enhancing differentiation of tumor-specific CD44hiCD62L+CD8+ TCM cells. Enhancement of CD8+ TCM cell differentiation determined by Stat-1, is dependent on the tumor-draining lymph nodes (TDLN) but takes place in peripheral blood, with metabolic remodeling of CD8+ T cells lasting the entire course of the the combination therapy. Importantly, in-vitro glycolysis inhibition in peripheral CD8+ T cells of patients with breast or liver tumors having been treated with MWA thrice leads to their differentiation into CD8+ TCM cells. Our work thus offers a potential strategy to avoid tumor recurrence following MWA therapy and lays down the proof-of-principle for future clinical trials.

Background Cancer associated fibroblasts (CAFs) can remodel tumor microenvironment by secreting exosomes. This study aimed to investigate the role of exosomes derived from cancer-associated fibroblasts in colorectal cancer (CRC) progression. Methods Circular RNA (circRNA) array was used to identify differentially expressed circRNAs in exosomes from normal fibroblasts (NFs) and CAFs, and confirmed one differentially expressed circRNA circ_0067557 by real-time PCR. The effect of circ_0067557 on proliferation, metastasis, chemoresistance and apoptosis was verified by wound heal, tranwell, CCK8, sphere-forming and flow cytometry assay. Results Circ_0067557 expression in exosomes from CAFs was higher than those from NFs. CAF-derived exosomes promoted the proliferation, migration, invasion and chemoresistance of CRC cells while suppressed apoptosis. Silencing of circ_0067557 inhibited malignant phenotypes of CRC cells by targeting Lin28A and Lin28B. Moreover, CAF-derived exosomes enhanced the growth of CRC xenograft tumors. Conclusion Circ_0067557/Lin28A and Lin28B signal axis may be a potential therapy target for CRC.

Abstract Introduction Locoregional recurrent breast cancers have a poor prognosis. Little is known about the prognostic impact of immune microenvironment, and tertiary lymphoid structures (TLSs) in particular have not been reported. Thus, we aimed to characterize the immune microenvironment in locoregional recurrent breast tumors and to investigate its relationship with prognosis. Methods We retrospectively included 112 patients with locoregional recurrent breast cancer, and hematoxylin–eosin staining and immunohistochemical staining (CD3, CD4, CD8, CD19, CD38, and CD68) were performed on locoregional recurrent tumor samples. The association of immune cells and TLSs with progression‐free survival (PFS) were analyzed by survival analysis. Results We found more immune cells in the peritumor than stroma. After grouping according to estrogen receptor (ER) status, a low level of peritumoral CD3+ cells in ER+ subgroup (p = 0.015) and a low level of stromal CD68+ cells in ER− subgroup (p = 0.047) were both associated with longer PFS. TLSs were present in 68% of recurrent tumors, and CD68+ cells within TLSs were significantly associated with PFS as an independent prognostic factor (p = 0.035). TLSs and immune cells (CD3, CD38, and CD68) within TLSs were associated with longer PFS in ER− recurrent tumors (p = 0.044, p = 0.012, p = 0.050, p < 0.001, respectively), whereas CD38+ cells within TLSs were associated with shorter PFS in ER+ recurrent tumors (p = 0.037). Conclusion Our study proposes potential predictors for the clinical prognosis of patients with locoregional recurrent breast cancer, emphasizing the prognostic value of immune cells within TLSs, especially CD68+ cells.

Metastasis is the major cause of cancer-related death, and lymph node is the most common site of metastasis in breast cancer. However, the alterations that happen in tumor-draining lymph nodes (TDLNs) to form a premetastatic microenvironment are largely unknown. Here, we first report the dynamic changes in size and immune status of TDLNs before metastasis in breast cancer. With the progression of tumor, the TDLN is first enlarged and immune-activated at early stage that contains specific antitumor immunity against metastasis. The TDLN is then contracted and immunosuppressed at late stage before finally getting metastasized. Mechanistically, B and follicular helper T (Tfh) cells parallelly expand and contract to determine the size of TDLN. The activation status and specific antitumor immunity of CD8 + T cells in the TDLN are determined by interleukin-21 (IL-21) produced by Tfh cells, thus showing parallel changes. The turn from activated enlargement to suppressed contraction is due to the spontaneous contraction of germinal centers mediated by follicular regulatory T cells. On the basis of the B-Tfh-IL-21-CD8 + T cell axis, we prove that targeting the axis could activate TDLNs to resist metastasis. Together, our findings identify the dynamic alterations and regulatory mechanisms of premetastatic TDLNs of breast cancer and provide new strategies to inhibit lymph node metastasis.

The occurrence and progression of tumors can be established through a complex interplay among tumor cells undergoing epithelial-mesenchymal transition (EMT), invasive factors and immune cells. In this study, we employed single-cell RNA sequencing (scRNA-seq) and spatially resolved transcriptomics (ST) to evaluate the pseudotime trajectory and spatial interactive relationship between EMT-invasive malignant tumors and immune cells in primary colorectal cancer (CRC) tissues at different stages (stage I/II and stage III with tumor deposit). Our research characterized the spatiotemporal relationship among different invasive tumor programs by constructing pseudotime endpoint-EMT-invasion tumor programs (EMTPs) located at the edge of ST, utilizing evolution trajectory analysis integrated with EMT-invasion genes. Strikingly, the invasive and expansive process of tumors undergoes remarkable spatial reprogramming of regulatory and immunosuppressive cells, such as myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), regulatory T cells (Treg), and exhausted T cells (Tex). These EMTP-adjacent cell are linked to EMT-related invasion genes, especially the C-X-C motif ligand 1 (CXCL1) and CXCL8 genes that are important for CRC prognosis. Interestingly, the EMTPs in stage I mainly produce an inflammatory margin invasive niche, while the EMTPs in stage III tissues likely produce a hypoxic pre-invasive niche. Our data demonstrate the crucial role of regulatory and immunosuppressive cells in tumor formation and progression of CRC. This study provides a framework to delineate the spatiotemporal invasive niche in CRC samples.

Background: Tumor cell epithelial-mesenchymal transition (EMT) is crucial for cancer progression. Molecular profiling of single cells has enhanced our understanding of the molecular basis of EMT but losing the crucial spatial information of invasion process in tumor microenvironment.Methods: This study utilized single cell RNA sequencing and spatially resolved transcriptomic (SRT) to assess rectum carcinoma tissues (stage I vs. stage III with tumor deposit). To evaluate the grow direction of malignant programs, we proposed up-regulated and down-regulated invasion scores by associating with pseudo-time. Cell-cell interaction and relocation the tumor programs with EMT-status in SRT are also performed.Findings: The research characterizes the spatio-temporal relationship for different stage of tumor programs by constructing an EMT-time-dependent invasive signature. The research also revealed the spatial and physiologic relationship between advanced tumor programs with highly EMT status. Adjacent immune cells in TME are also have important interaction with the EMT tumor program, including cytotoxic CD8+ T cells, neutrophils, and terminally exhausted T cells, with interaction of MIF- (S100A4/SREBF1/ERRFI1), TGFB1-JAG1, and EGFR-ADAM17.Interpretation: The current study provides a framework for studying the underline mechanisms of tumor cell invasion across EMT in complex tumor progression.Funding: This study was funded by the National Natural Science Foundation of China (U20A20376); the Free Exploration Foundation of Science, 45 Technology, and Innovation Commission of Shenzhen Municipality (No. JCYJ2018307151238174); and the Science and Technology Planning Project of Guangdong Province (No. 2017A020215014).Declaration of Interest: We declare there are no any competing financial interests in relation to this work.Ethical Approval: This study was approved by the Ethics Committee of Harbin Medical University Cancer Hospital. Both study patients provided written informed consent before tissue collection for this study.

Abstract Exosomes derived from cancer-associated fibroblasts (CAFs) may be crucial in colorectal cancer (CRC) progression, although the underlying function and mechanism of CAF-derived exosomes in CRC are still unclear. We used a circular RNA (circRNA) array to identify differentially expressed circRNAs in exosomes from normal fibroblasts (NFs) and CAFs. One differentially expressed circRNA was circ_0067557, whose expression was verified by RT-qPCR. The target genes of circ_0067557 (Lin28A and Lin28B) were identified by RNA-pull down, RNA-IP, and RNA-EMSA. The biological behavior of CRC cells and CRC xenograft tumors were examined by multiple assays. Circ_0067557 expression in exosomes from CAFs was higher than in exosomes from NFs. Functionally, CAF-derived exosomes could induce proliferation, migration, invasion, chemoresistance, EMT, and the cancer stem cell (CSC) features of CRC cells while suppressing apoptosis. In addition, we confirmed that circ_0067557 could bind to its target genes (Lin28A and Lin28B). Silencing of circ_0067557 prevented the malignant phenotype of the CRC cell by targeting Lin28A and Lin28B. Moreover, CAF-derived exosomes enhanced the growth of CRC xenograft tumors, suppressed apoptosis, and upregulated Lin28A and Lin28B through circ_0067557. Circ_0067557/Lin28A and Lin28B signal axis may be a potential therapy target for CRC patients.

Ephrin Type-A Receptor 3 (EphA3) and Ephrin Type-B Receptor 6 (EphB6) belong to the ephrin receptor group consisting of the largest subset of receptor tyrosine kinases (RTKs) and are essential for neurogenesis and embryogenesis.The current study aimed to evaluate their functional roles in transforming colorectal epithelial cells and dissect the underlying molecular mechanisms.We observed altered EphA3 and EphB6 expression in tumor tissues as compared to normal tissues in a tissue microarray study.Enforced EphB6 expression promoted IMCE cell proliferation, migration, and invasion in vitro and tumor formation in nude mice, with a stronger oncogenic activity than EphA3.Pathway analysis of differentially expressed genes from a gene microarray study provided important insight into potential mechanisms through which EphB6 may regulate the malignant transformation of colorectal epithelial cells.This study represents the first demonstration of EphB6 in enhancing colorectal epithelial cell transformation, suggesting its stipulative role in the early stage of colorectal tumorigenesis.Our findings primarily uncover novel biomarkers and therapeutic targets of colorectal cancer.

Postmenopausal osteoporosis (PMOP) is a common metabolic bone disease in postmenopausal women in the Worldwide, and seriously affects the quality of life of middle-aged and elderly women. Therefore, there is an urgent need to discover a highly effective drug for PMOP treatment. In this study, ultra-high performance liquid tandem quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS) was used to analyze the urine metabolic profiling and potential biomarkers, the relevant metabolic network of PMOP rats, and further to evaluate the intervention effect of Eleutheroside E (EE) against PMOP. Using multivariate statistical analysis combined with UPLC-Q/TOF-MS, a total of 27 biomarkers were identified, which related with 16 metabolic pathways, mainly involving steroidogenesis, beta oxidation of very long chain fatty acids, glutathione metabolism, carnitine synthesis, estrone metabolism, oxidation of branched chain fatty acids, etc. After treatment of EE, these biomarkers were markedly regulated, mainly involving steroid hormone biosynthesis, arachidonic acid metabolism, primary bile acid biosynthesis, indicating that EE had the therapeutic effect on PMOP. This study identified the potential urine metabolic markers and related metabolic pathways of the PMOP, explained the metabolic effect and pharmacological mechanisms of EE against PMOP, and provided a basis for the pharmacological study of EE.