Tumor Cell Types Research Articles

OS10.5.A PERIVASCULAR CONTAINED T CELLS IN IDH-MUTANT ASTROCYTOMA CO-LOCALIZE WITH GEMISTOCYTIC TUMOR CELLS OF AN ASTROCYTE-STATE AND MACROPHAGES WITH AN IMMUNE-STIMULATORY PHENOTYPE

Abstract BACKGROUND Isocitrate dehydrogenase mutant (IDHmt) astrocytoma is considered a T cell excluded tumor, yet little is known regarding the underlying cues that may drive T cell exclusion. To aid better understanding of this mechanism in IDHmt astrocytoma, we systematically characterized the immune microenvironment of this tumor type. METHODS We performed and analyzed multiplex immunofluorescence stainings for lymphocytes, myeloid cells and IDH1-R132H of 1293 regions from 75 IDHmt astrocytomas. We consequently performed bulk, single nucleus and spatial RNA sequencing of samples from the same cohort. Integrative analysis of data from these platforms allowed for the identification of unique tumor and immune cell subsets that are crucial in the spatial organization of the immune microenvironment and T cell containment in IDHmt astrocytoma. RESULTS IDHmt astrocytoma showed three spatially distinct distribution patterns of T cells: regions with perivascular accumulation of T cells, regions with T cells in the tumor stroma and T cell absent regions. Perivascular accumulation of T cells, known as T-cell cuffs, was the most abundant pattern in our data. Spatially, there was a significant increase of tumor cells around T cell cuffs that showed a gemistocytic phenotype, a tumor cell type that is histologically characterized by a large eosinophilic cytoplasm and ectopically localized nucleus. Gemistocyte-high tumors (> 20% of all tumor cells) demonstrated enhanced expression of genes related to immune cell migration and activation that were reliably expressed by tumor associated macrophages and T cells. Notably, gemistocytic tumor cells were identified as a distinct sub-cluster of the ‘astrocyte-like’ transcriptomic cell state with a phenotype that is linked to astrogliosis and expressed CD44, TNC and TGFB2. Moreover, gemistocytic tumor cells were spatially associated with a CD83+ subpopulation of tumor-associated macrophages that resemble reactive microglia in astrogliosis and show increased expression of the chemo-attractants CCL3, CCL4 and CCL4L2. CONCLUSION Using different modalities for spatial and single cell profiling of RNA and protein, we uncovered a previously unknown relationship between the immune system and tumor cells in IDHmt astrocytoma. We found that gemistocytic tumor cells showing an ‘astrocyte-like’ state co-occur with perivascular contained T cells and immune-stimulatory macrophages in IDHmt astrocytomas. The transcriptional profiles of these tumor cell subsets and macrophages point towards a CNS specific defense mechanism of glial cells associated with T cell exclusion.

Abl kinases can function as suppressors of tumor progression and metastasis.

Abl family kinases function as proto-oncogenes in various leukemias, and pro-tumor functions have been discovered for Abl kinases in many solid tumors as well. However, a growing body of evidence indicates that Abl kinases can function to suppress tumor cell proliferation and motility and tumor growth in vivo in some settings. To investigate the role of Abl kinases in tumor progression, we used RNAi to generate Abl-deficient cells in a model of androgen receptor-indifferent, metastatic prostate cancer. The effect of Abl kinase depletion on tumor progression and metastasis was studied in an in vivo orthotopic model, and tumor cell motility, 3D growth, and signaling was studied in vitro. Reduced Abl family kinase expression resulted in a highly aggressive, metastatic phenotype in vivo that was associated with AKT pathway activation, increased growth on 3D collagen matrix, and enhanced cell motility in vitro. Inhibiting AKT pathway signaling abolished the increased 3D growth of Abl-deficient cells, while treatment with the Abl kinase inhibitor, imatinib, promoted 3D growth of multiple additional tumor cell types. Moreover, Abl kinase inhibition also promoted soft-agar colony formation by pre-malignant fibroblasts. Collectively, our data reveal that Abl family kinases can function to suppress malignant cell phenotypes in vitro, and tumor progression and metastasis in vivo.

MiRNA/epithelial-mesenchymal axis (EMT) axis as a key player in cancer progression and metastasis: A focus on gastric and bladder cancers

The metastasis a major hallmark of tumors that its significant is not only related to the basic research, but clinical investigations have revealed that majority of cancer deaths are due to the metastasis. The metastasis of tumor cells is significantly increased due to EMT mechanism and therefore, inhibition of EMT can reduce biological behaviors of tumor cells and improve the survival rate of patients. One of the gaps related to cancer metastasis is lack of specific focus on the EMT regulation in certain types of tumor cells. The gastric and bladder cancers are considered as two main reasons of death among patients in clinical level. Herein, the role of EMT in regulation of their progression is evaluated with a focus on the function of miRNAs. The inhibition/induction of EMT in these cancers and their ability in modulation of EMT-related factors including ZEB1/2 proteins, TGF-β, Snail and cadherin proteins are discussed. Moreover, lncRNAs and circRNAs in crosstalk of miRNA/EMT regulation in these tumors are discussed and final impact on cancer metastasis and response of tumor cells to the chemotherapy is evaluated. Moreover, the impact of miRNAs transferred by exosomes in regulation of EMT in these cancers are discussed.

Synthesis and in Vitro Cytotoxicity Evaluation of Jadomycins.

Jadomycins, which are benzo[b]phenanthridine-type alkaloids isolated from Streptomyces venezuelae ISP5230, exhibit cytotoxic activity against multidrug-resistant breast cancer cells. We have previously achieved the total synthesis of jadomycins using the direct arylation of juglone as a key step. In this study, we achieved the total synthesis of jadomycin T and jadomycin aglycons using L-threonine and 1-amino-2-propanol as nitrogen sources. Additionally, we evaluated the cytotoxic activity of eight compounds, including glycosides, jadomycin T, and their corresponding aglycons, in eight types of tumor cells. The evaluated jadomycins tended to exhibit stronger cytotoxic activity as aglycons than as glycosides. Although the presence of a 1,3-oxazolidine ring derived from an amino acid was not essential, the presence of the 1,3-oxazolidine ring showed strong activity when the ring had a carboxyl group. Furthermore, compared to the non-natural isomer at a different position on the phenolic hydroxyl group, the naturally occurring phenanthroviridin aglycon exhibited stronger cytotoxic activity. In addition, this study suggests that jadomycins may become lead compounds for the treatment of brain tumors; however, further studies on their ability to penetrate the blood-brain barrier are required.

Diphtheria toxin-derived, anti-PD-1 immunotoxin, a potent and practical tool to selectively deplete PD-1+ cells.

Programmed death-1 (PD-1), an immune checkpoint receptor, is expressed on activated lymphocytes, macrophages, and some types of tumor cells. While PD-1+ cells have been implicated in outcomes of cancer immunity, autoimmunity, and chronic infections, the exact roles of these cells in various physiological and pathological processes remain elusive. Molecules that target and deplete PD-1+ cells would be instrumental in defining the roles unambiguously. Previously, an immunotoxin has been generated for the depletion of PD-1+ cells though its usage is impeded by its low production yield. Thus, a more practical molecular tool is desired to deplete PD-1+ cells and to examine functions of these cells. We designed and generated a novel anti-PD1 diphtheria immunotoxin, termed PD-1 DIT, targeting PD-1+ cells. PD-1 DIT is comprised of two single chain variable fragments (scFv) derived from an anti-PD-1 antibody, coupled with the catalytic and translocation domains of the diphtheria toxin. PD-1 DIT was produced using a yeast expression system that has been engineered to efficiently produce protein toxins. The yield of PD-1 DIT reached 1-2 mg/L culture, which is 10 times higher than the previously reported immunotoxin. Flow cytometry and confocal microscopy analyses confirmed that PD-1 DIT specifically binds to and enters PD-1+ cells. The binding avidities between PD-1 DIT and two PD-1+ cell lines are approximately 25 nM. Moreover, PD-1 DIT demonstrated potent cytotoxicity toward PD-1+ cells, with a half maximal effective concentration(EC50 ) value of 1 nM. In vivo experiments further showed that PD-1 DIT effectively depleted PD-1+ cells and enabled mice inoculated with PD-1+ tumor cells to survive throughout the study. Our findings using PD-1 DIT revealed the critical role of pancreatic PD-1+ T cells in the development of type-1 diabetes (T1D). Additionally, we observed that PD-1 DIT treatment ameliorated relapsing-remitting experimental autoimmune encephalomyelitis (RR-EAE), a mouse model of relapsing-remitting multiple sclerosis (RR-MS). Lastly, we did not observe significant hepatotoxicity in mice treated with PD-1 DIT, which had been reported for other immunotoxins derived from the diphtheria toxin. With its remarkable selective and potent cytotoxicity toward PD-1+ cells, coupled with its high production yield, PD-1 DIT emerges as a powerful biotechnological tool for elucidating the physiological roles of PD-1+ cells. Furthermore, the potential of PD-1 DIT to be developed into a novel therapeutic agent becomes evident.

Regulation of the stem‑like properties of estrogen receptor‑positive breast cancer cells through NR2E3/NR2C2 signaling.

Cancer stem cells (CSCs) are major drivers of metastasis, drug resistance and recurrence in numerous cancers. However, critical factors that can modulate CSC stemness have not been clearly identified. Nuclear receptor subfamily 2 group E member 3 (nr2e3) expression has been previously reported to be positively associated with drug sensitivity and favorable clinical outcomes in patients with estrogen receptor (ER)+ breast cancer. This suggests that nr2e3 expression may be inversely associated with CSC stemness in this type of tumor cells. The present study aimed to investigate the regulatory roles of NR2E3 in the stem-like properties of ER+ breast cancer cells and to identify the underlying mechanisms. Bioinformatics analysis was performed using the data derived from the Cancer Genome Atlas database. Nr2e3-specific shRNA and nuclear receptor subfamily 2 group C member 2 (nr2c2) overexpressed plasmids were constructed to silence and enhance the expression of nr2e3 and nr2c2, respectively. Transwell and wound healing experiments were conducted to evaluate the migration and invasion ability of MCF7 cells, while colony formation tests were used to evaluate the clonality. Flow cytometry was used to detect the percentage of CD44+CD24-/low cells. Reverse transcription-quantitative PCR and western blotting were performed to detect expression at the mRNA and protein levels. The results showed that compared with normal breast tissues and MCF10A cells, the expression of nr2e3 was increased in ER+ breast tumor tissues and cell lines. Nr2e3 silencing promoted the migration, invasion and colony-forming ability of the ER+ MCF7 cells. It also increased the expression of epithelial-mesenchymal transition markers and stem cell-related transcription factors, in addition to the percentage of CD44+CD24-/low cells. The expression of nr2e3 and nr2c2 was found to be positively correlated. Nr2e3 knockdown decreased the mRNA and protein expression levels of nr2c2, whereas nr2c2 overexpression reversed the elevated CD44+CD24-/low cell ratio and the increased migratory activity caused by nr2e3 silencing. The results of the present study suggest that NR2E3 may serve an important role in modulating the stem-like properties of ER+ breast cancer cells, where NR2E3/NR2C2 signaling may be a therapeutic target in ER+ breast cancer.

Preclinical evaluation and pilot clinical study of [68Ga]Ga-THP-APN09, a novel PD-L1 targeted nanobody radiotracer for rapid one-step radiolabeling and PET imaging.

Programmed cell death protein-1/ligand-1 (PD-1/L1) blockade has been a breakthrough in the treatment of patients with non-small cell lung cancer (NSCLC), but there is still a lack of effective methods to screen patients. Here we report a novel 68Ga-labeled nanobody [68Ga]Ga-THP-APN09 for PET imaging of PD-L1 status in mouse models and a first-in-human study in NSCLC patients. [68Ga]Ga-THP-APN09 was prepared by site-specific radiolabeling, with no further purification. Cell uptake assays were completed in the human lung adenocarcinoma cell line A549, NSCLC cell line H1975 and human PD-L1 gene-transfected A549 cells (A549PD-L1). The imaging to image PD-L1 status and biodistribution were investigated in tumor-bearing mice of these three tumor cell types. The first-in-human clinical translational trial was registered as NCT05156515. The safety, radiation dosimetry, biodistribution, and correlations of tracer uptake with immunohistochemical staining and major pathologic response (MPR) were evaluated in NSCLC patients who underwent adjuvant immunotherapy combined with chemotherapy. Radiosynthesis of [68Ga]Ga-THP-APN09 was achieved at room temperature and a pH of 6.0-6.5 in 10min with a high radiochemical yield (> 99%) and 13.9-27.8GBq/μmol molar activity. The results of the cell uptake study reflected variable levels of surface PD-L1 expression observed by flow cytometry in the order A549PD-L1 > H1975 > A549. In small-animal PET/CT imaging, H1975 and A549PD-L1 tumors were clearly visualized in an 8.3:1 and 2.2:1 ratios over PD-L1-negative A549 tumors. Ex vivo biodistribution studies showed that tumor uptake was consistent with the PET results, with the highest A549PD-L1 being taken up the most (8.20 ± 0.87%ID/g), followed by H1975 (3.69 ± 0.50%ID/g) and A549 (0.90 ± 0.16%ID/g). Nine resectable NSCLC patients were enrolled in the clinical study. Uptake of [68Ga]Ga-THP-APN09 was mainly observed in the kidneys and spleen, followed by low uptake in bone marrow. The radiation dose is within a reliable range. Tumor uptake was positively correlated with PD-L1 expression TPS (rs = 0.8763, P = 0.019). Tumor uptake of [68Ga]Ga-THP-APN09 (SUVmax) in MPR patients was higher than that in non-MPR patients (median SUVmax 2.73 vs. 2.10, P = 0.036, determined with Mann-Whitney U-test). [68Ga]Ga-THP-APN09 has the potential to be transformed into a kit-based radiotracer for rapid, simple, one-step, room temperature radiolabeling. The tracer can detect PD-L1 expression levels in tumors, and it may make it possibility to predict the response of PD-1 immunotherapy combined with chemotherapy. Confirmation in a large number of cases is needed. Clinical Trial (NCT05156515). Registered 12 December 2021.

Fortuitously detected primary ovarian carcinoid tumor: A case report.

Carcinoid tumors, derived from the cells of the disseminated neuroendocrine system, are rare, slow-growing neuroendocrine neoplasms that display a relatively indolent disease course. The majority of carcinoids are found within the gastrointestinal tract and bronchopulmonary system. Primary ovarian carcinoids are rare and account for merely 1% of all carcinoid tumors. We describe our experience of a rare case of primary ovarian carcinoid, presenting as chronic constipation, with no other carcinoid symptoms such as flushing, diarrhea, and wheezing. A 51-year-old postmenopausal woman with chronic constipation visited the clinic for routine check-up of her preexisting uterine fibroids. She had undergone hemorrhoidectomy 3 years ago. Physical examination revealed a soft abdomen without direct or rebound tenderness. Transvaginal ultrasonography revealed two subserosal fibroids, which had increased in size compared with previous ultrasonographic findings. A 3 cm hyperechoic mass was also detected in the right ovary. Her blood and urine tests were unremarkable, with no ascites in the pelvic cavity. She had a normal CA-125 level of 5.5 units/mL. The patient underwent a robot-assisted hysterectomy and right salpingo-oophorectomy because of enlarging fibroids and the right ovarian mass. Subsequently, based on the pathological and immunohistochemical findings, she was diagnosed with a primary ovarian carcinoid. The mass consisted of the insular and trabecular types of tumor cells. It was positive for pan-cytokeratin and synaptophysin, and the Ki-67 proliferation index was less than 1%. A follow-up positron emission tomography-computed tomography revealed no distant metastasis. Six months postoperatively, the patient was doing well without any signs of recurrence. Primary ovarian carcinoids without teratoma components are rare. It is crucial to make an accurate diagnosis based on the immunohistochemical staining results. Diagnosis in the early stages of the disease are associated with a favorable prognosis, but regular follow-up is mandatory.

Single-cell profiling reveals the trajectory of FOLR2-expressing tumor-associated macrophages to regulatory T cells in the progression of lung adenocarcinoma

An immunosuppressive microenvironment enriched with regulatory CD4+ T lymphocytes (Tregs) facilitates the progression of lung adenocarcinoma (LUAD). This study aims to investigate the cellular mechanism underlying the formation of the immunosuppressive microenvironment in LUAD. LUAD samples (n = 12) and normal lung samples (n = 3) were obtained from patients with different pathological stages of LUAD. Single-cell RNA sequencing was performed to classify cellular components and analyze the transcriptomes, including transcription factors/targets and chemokine ligands/receptors, followed by bioinformatics study such as pseudotime analysis. Myeloid cells and T cells were the most abundant cell types in tumors and normal lung tissues, while tumor-associated macrophage-folate receptor 2 (TAM-FOLR2) and CD4+ nuclear receptor subfamily 4 group A member 3 (NR4A3) exhibited sharp increases in invasive adenocarcinoma (IA). The enrichment of TAM-FOLR2 in IA might result from alveolar resident macrophage-resistin (ARM-RETN) transformation and recruitment of dendritic cells (DCs) and other TAMs, as evidenced by temporal trajectories and differential expression profiles of chemokine ligands/receptors versus those in the early stages of tumors. High expression of CCL17/19/22 was observed in IA as well as in DCs, along with the strong interaction of TAM-FOLR2 with DCs. The results of pseudotime analysis suggested that CD4+NR4A3 might potentially convert to CD4+FOXP3, further supported by the high expression of NR4A3 target genes in CD4+FOXP3 cells. This study provides a single-cell transcriptome atlas from preinvasive to invasive LUAD and reveals a potential ARM-RETN/TAM-FOLR2/DCs/CD4+NR4A3/CD4+FOXP3 trajectory in shaping the immune suppressive microenvironment along the pathogenesis of LUAD.

Raman Spectroscopy Combined with Malaria Protein for Early Capture and Recognition of Broad-Spectrum Circulating Tumor Cells.

Early identification of tumors can significantly reduce the mortality rate. Circulating tumor cells (CTCs) are a type of tumor cell that detaches from the primary tumor and circulates through the bloodstream. Monitoring CTCs may allow the early identification of tumor progression. However, due to their rarity and heterogeneity, the enrichment and identification of CTCs is still challenging. Studies have shown that Raman spectroscopy could distinguish CTCs from metastatic cancer patients. VAR2CSA, a class of malaria proteins, has a strong broad-spectrum binding effect on various tumor cells and is a promising candidate biomarker for cancer detection. Here, recombinant malaria VAR2CSA proteins were synthesized, expressed, and purified. After confirming that various types of tumor cells can be isolated from blood by recombinant malaria VAR2CSA proteins, we further proved that the VAR2CSA combined with Raman spectroscopy could be used efficiently for tumor capture and type recognition using A549 cell lines spiked into the blood. This would allow the early screening and detection of a broad spectrum of CTCs. Finally, we synthesized and purified the malaria protein fusion antibody and confirmed its in vitro tumor-killing activity. Herein, this paper exploits the theoretical basis of a novel strategy to capture, recognize, and kill broad-spectrum types of CTCs from the peripheral blood.

The GLI2/CDH6 axis enhances migration, invasion and mitochondrial fission of stomach adenocarcinoma cells

It has been reported that cadherin 6 (CDH6) upregulation is associated with enhanced epithelial-to-mesenchymal transition (EMT) in several types of solid tumor cells. The current study aimed to explore the effect of CDH6 on the migration and invasion of stomach adenocarcinoma (STAD) cells, the transcription factors involved in CDH6 dysregulation and their effect on mitochondrial fission. Bioinformatics analysis was performed using data extracted from the Genotype-Tissue Expression Project, the Cancer Genome Atlas and Kaplan-Meier plotter. AGS and HGC27 cells were used to establish an in vitro STAD cell model. The results showed that higher CDH6 expression was associated with significantly shorter overall survival in patients with STAD. In addition, CDH6 overexpression promoted wound healing, enhanced the invasion ability of tumor cells and increased mitochondrial fission. Glioma-associated oncogene family zinc finger 2 (GLI2) could bind to the CDH6 promoter and activate its transcription. Fluorescent labeling also showed that GLI2 overexpression promoted mitochondrial fission. However, CDH6 silencing significantly reduced mitochondrial fragmentation. Besides, GLI2 overexpression notably upregulated phosphorylated-focal adhesion kinase and dynamin-related protein 1. However, the above effects were largely abrogated by CDH6 knockdown. In conclusion, the present study suggested that the novel GLI2/CDH6 axis could enhance the migration, invasion and mitochondrial fission of STAD cells.

Tumor cell-intrinsic c-Myb upregulation stimulates antitumor immunity in a murine colorectal cancer model.

The transcription factor c-Myb is overexpressed in many different types of solid tumors, including colorectal cancer. However, its exact role in tumorigenesis is unclear. In this study, we show that tumor-intrinsic c-Myb expression in mouse models of colon cancer and melanoma suppresses tumor growth. While no differences in proliferation, apoptosis, and angiogenesis of tumors were evident tumors with distinct levels of c-Myb expression, we observed changes in intratumoral immune cell infiltrates. MC38 tumors with upregulated c-Myb expression showed increased numbers of CD103+ dendritic cells and eosinophils, but decreased tumor-associated macrophages (TAMs). Concomitantly, an increase in the number of activated cytotoxic CD8+ T cells upon c-Myb upregulation was observed, which correlated with a pro-inflammatory tumor microenvironment and increased numbers of M1 polarized TAMs. Mechanistically, c-Myb upregulation in immunogenic MC38 colon cancer cells resulted in enhanced expression of immunomodulatory genes, including those encoding 2-microglobulin and IFNβ, and decreased expression of the gene encoding the chemokine receptor CCR2. The increased numbers of activated cytotoxic CD8+ T cells contributed to tumor growth attenuation. In poorly immunogenic CT26, LLC, and B16-BL6 tumor cells, c-Myb upregulation did not affect the immunomodulatory gene expression. Despite this, c-Myb upregulation led to reduced B16-BL6 tumor growth but it did not affect tumor growth of CT26 and LLC tumors. Altogether, we postulate that c-Myb functions as a tumor suppressor in a tumor cell-type specific manner and modulates antitumor immunity.

Different Molecular Phenotypes of Progression in BRAF- and RAS-Like Papillary Thyroid Carcinoma.

Papillary thyroid carcinoma (PTC) can be classified into two distinct molecular subtypes, BRAF-like (BL) and RASlike (RL). However, the molecular characteristics of each subtype according to clinicopathological factors have not yet been determined. We aimed to investigate the gene signatures and tumor microenvironment according to clinicopathological factors, and to identify the mechanism of progression in BL-PTCs and RL-PTCs. We analyzed RNA sequencing data and corresponding clinicopathological information of 503 patients with PTC from The Cancer Genome Atlas database. We performed differentially expressed gene (DEG), Gene Ontology, and molecular pathway enrichment analyses according to clinicopathological factors in each molecular subtype. EcoTyper and CIBERSORTx were used to deconvolve the tumor cell types and their surrounding microenvironment. Even for the same clinicopathological factors, overlapping DEGs between the two molecular subtypes were uncommon, indicating that BL-PTCs and RL-PTCs have different progression mechanisms. Genes related to the extracellular matrix were commonly upregulated in BL-PTCs with aggressive clinicopathological factors, such as old age (≥55 years), presence of extrathyroidal extension, lymph node metastasis, advanced tumor-node-metastasis (TNM) stage, and high metastasis-age-completeness of resection- invasion-size (MACIS) scores (≥6). Furthermore, in the deconvolution analysis of tumor microenvironment, cancer-associated fibroblasts were significantly enriched. In contrast, in RL-PTCs, downregulation of immune response and immunoglobulin-related genes was significantly associated with aggressive characteristics, even after adjusting for thyroiditis status. Conclusion: The molecular phenotypes of cancer progression differed between BL-PTC and RL-PTC. In particular, extracellular matrix and cancer-associated fibroblasts, which constitute the tumor microenvironment, would play an important role in the progression of BL-PTC that accounts for the majority of advanced PTCs.

Expression, Intracellular Localization, and Maturation of Cysteine Cathepsins in Renal Embryonic and Cancer Cell Lines

Cysteine cathepsins play an important role in tumor development and metastasis. The expression of these enzymes is often increased in many types of tumor cells. Cysteine cathepsins contribute to carcinogenesis through a number of mechanisms, including proteolysis of extracellular matrix and signaling molecules on the cell surface, as well as degradation of transcription factors and disruption of signaling cascades in the cell nucleus. Distinct oncogenic functions have been reported for several members of the cysteine cathepsin family in various types of cancer, but a comparative study of all eleven cysteine cathepsins in one experimental model is still missing. In this work, we assessed and compared the expression, localization, and maturation of all eleven cysteine cathepsins in embryonic kidney cells HEK293 and kidney cancer cell lines 769-P and A-498. We found that the expression of cathepsins V, B, Z, L, and S was 3- to 9-fold higher in kidney tumor cells than in embryonic cells. We also showed that all cysteine cathepsins were present in varying amounts in the nucleus of both embryonic and tumor cells. Notably, more than half of the cathepsin Z or K and over 88% of cathepsin F were localized in tumor cell nuclei. Moreover, mature forms of cysteine cathepsins were more prevalent in tumor cells than in embryonic cells. These results can be further used to develop novel diagnostic tools and may assist in the investigation of cysteine cathepsins as potential therapeutic targets.

Clearance of senescent macrophages ameliorates tumorigenesis in KRAS-driven lung cancer.

The accumulation of senescent cells in the tumor microenvironment can drive tumorigenesis in a paracrine manner through the senescence-associated secretory phenotype (SASP). Using a new p16-FDR mouse line, we show that macrophages and endothelial cells are the predominant senescent cell types in murine KRAS-driven lung tumors. Through single cell transcriptomics, we identify a population of tumor-associated macrophages that express a unique array of pro-tumorigenic SASP factors and surface proteins and are also present in normal aged lungs. Genetic or senolytic ablation of senescent cells, or macrophage depletion, result in a significant decrease in tumor burden and increased survival in KRAS-driven lung cancer models. Moreover, we reveal the presence of macrophages with senescent features in human lung pre-malignant lesions, but not in adenocarcinomas. Taken together, our results have uncovered the important role of senescent macrophages in the initiation and progression of lung cancer, highlighting potential therapeutic avenues and cancer preventative strategies.

Profound Diversity of the N-Glycome from Microdissected Regions of Colorectal Cancer, Stroma, and Normal Colon Mucosa

• Striking diversity of N -glycosylation was found between cancerous, stromal and healthy mucosa cells. • Cancer showed increased α2-6 sialylation and monoantennary N -glycans and decreased bisection compared to healthy mucosa. • N -glycans carrying (sialyl-)Lewis A/X were exclusively found in cancer. • Stroma showed higher fucosylation and (α2-3) sialylation and lower oligomannosidics, monoantennaries, Lewis A/X and sulfation compared to cancer. Aberrant glycosylation is considered to be a hallmark of colorectal cancer (CRC), as demonstrated by various studies. While the N -glycosylation of cell lines and serum has been widely examined, the analysis of cancer-associated N -glycans from tissues has been hampered by the heterogeneity of tumors and the complexity of N -glycan structures. To overcome these obstacles, we present a study using laser capture microdissection that makes it possible to largely deconvolute distinct N- glycomic signatures originating from different regions of heterogeneous tissues including cancerous, stromal, and healthy mucosa cells. N -glycan alditols were analyzed by means of porous graphitized carbon liquid chromatography-electrospray ionization tandem mass spectrometry, enabling the differentiation and structural characterization of isomeric species. In total, 116 N- glycans were identified that showed profound differences in expression among cancer, stroma, and normal mucosa. In comparison with healthy mucosa, the cancer cells showed an increase in α2-6 sialylation and monoantennary N -glycans, as well as a decrease in bisected N -glycans. Moreover, specific sialylated and (sialyl-)Lewis A/X antigen-carrying N -glycans were exclusively expressed in cancers. In comparison with cancer, the stroma showed lower levels of oligomannosidic and monoantennary N -glycans, Lewis A/X epitopes, and sulfation, as well as increased expression of (core-)fucosylation and α2-3 sialylation. Our study reveals the distinct N -glycomic profiles of different cell types in CRC tumor and control tissues, proving the necessity of their separate analysis for the discovery of cancer-associated glycans.

Inhibitory Effect of Cell Phones Against Human Breast Cancer and Myeloid Leukemia Cells Proliferation in Culture Media

Objective: There is current news that emerges regarding the relationship between the magnetic effects of cell phones and some types of cancer. In spite of the studies carried out, the level of evidence of this news is low, and also the relationship between the magnetic effects of cell phones and other types of cancer is not certain except for brain cancer. In this study, it is aimed at examining the effects of magnetic field of cell phones on the samples of breast cancer human myeloid leukemia cell growth. 
 Methods: In the study, breast cancer MCF-7 and leukemia K562 cell lines were used as the source of cancer cell. During the six-day cell culture, cancer cells were subjected to the effects of cell phone by using a telephone call program (Automated outbound call software). The system made 6 calls for 1 minute for each call once in 144 minutes from fixed line. The number of cultured cells and proliferation capacities of the two types of tumor cells in the control and experimental groups were assessed. 
 Results: The number of cancer cells, which were subjected to the effects of cell phone as a result of the culture of tumor cells, was found lower when compared with control group (7500000 ± 100000 vs 6625000 ± 225000 for MCF-7; 15412500 ± 112500 vs 13700000 ± 250000 for K562; P < 0.05 for both). In MTT test, it was found out that two types of cell proliferation was inclined to slow down with the effect of cell phone. 
 Conclusion: The results can be translated that cell phone may inhibit neoplastic transformation, and this observation may promote to initiate a new clinical studies both for healthy people and for patients with cancer.

Single-cell transcriptomics and epigenomics unravel the role of monocytes in neuroblastoma bone marrow metastasis

Metastasis is the major cause of cancer-related deaths. Neuroblastoma (NB), a childhood tumor has been molecularly defined at the primary cancer site, however, the bone marrow (BM) as the metastatic niche of NB is poorly characterized. Here we perform single-cell transcriptomic and epigenomic profiling of BM aspirates from 11 subjects spanning three major NB subtypes and compare these to five age-matched and metastasis-free BM, followed by in-depth single cell analyses of tissue diversity and cell-cell interactions, as well as functional validation. We show that cellular plasticity of NB tumor cells is conserved upon metastasis and tumor cell type composition is NB subtype-dependent. NB cells signal to the BM microenvironment, rewiring via macrophage mgration inhibitory factor and midkine signaling specifically monocytes, which exhibit M1 and M2 features, are marked by activation of pro- and anti-inflammatory programs, and express tumor-promoting factors, reminiscent of tumor-associated macrophages. The interactions and pathways characterized in our study provide the basis for therapeutic approaches that target tumor-to-microenvironment interactions.

Effects of the Intake of Deuterium Depleted Water During Cancer Therapy: A Narrative Review

Drinking water has a deuterium (2H) concentration of 150 ppm. The intake of deuteriumdepleted water (DDW) provides a series of effects on the improvement of the state of health, contributing to the prevention of various diseases, as well as the delay in the evolution of certain types of cancer. The objective of this review is to verify the influence of DDW intake on the regression and survival rate of different types of cancer in living beings. The intake of DDW in patients with chronic lymphocytic leukemia, lung, prostate and breast cancer, as well as the antitumor effects of the intake of DDW in living beings, were analyzed. In conclusion, commenting that the intake of DDW has beneficial effects on the increase in the average survival time of cancer patients, favors the inhibition of cancer cell growth, increases the probability of total or partial regression of malignant tumors, as well as the delay in the multiplication of various types of tumor cells (PC-3, MDA, HT-29 and M14) and contributes to the decrease in PSA values in patients with prostate cancer.

Heterogeneity of tumor immune microenvironment in malignant and metastatic change in LUAD is revealed by single-cell RNA sequencing.

Lung adenocarcinoma (LUAD) is the most common type of non-small cell lung cancer and accounts for approximately 40% of all lung cancer cases. Multiple distant metastases are the major cause of mortality in lung cancer. In this study, single-cell sequencing datasets of LUAD were utilized to depict the transcriptome characteristic of LUAD based on the bioinformatic method. Firstly, the transcriptome landscape of heterogeneous cell types in LUAD was analyzed and memory T cells, NK cells, and helper T cells were revealed to be the common immune cells in tumor, normal, and metastasis tissue, respectively. Then, marker genes were calculated and 709 genes were identified to play a vital role in the microenvironment of LUAD. While macrophages were reported to act as one of the cells in LUAD, enrichment analysis of macrophage marker genes revealed the important role of macrophages in the activation of neutrophils. Next, the results of cell-cell communication analysis suggested that pericytes interact with broad immune cells via MDK-NCL pathways in metastasis samples, MIF-(CD74+CXCR4) and MIF-(CD74+CC44) interaction especially occurred between different cell types in tumor and normal samples. Finally, bulk RNA-seq was integrated to validate the prognosis effect of the marker gene and the maker gene of M2 macrophage, CCL20, showed the most related to LUAD prognosis. Besides, ZNF90 (Helper T cells), FKBP4 (memory T, helper T, Cytotoxic T, and B cells), CD79A (B cells), TPI1 (pericyte), and HOPX (epithelial cells, pericytes) were also pivotal in the pathology of LUAD, helping researchers understand the molecular insight of microenvironment in LUAD.