One of the leading causes that complicate the treatment of some malignancies, including breast cancer, is tumor heterogeneity. In addition to inter-heterogeneity and intra-heterogeneity of tumors that reflect the differences between cancer cell characteristics, heterogeneity in the tumor microenvironment plays a critical role in tumor progression and could be considered an overlooked and a proper target for the effective selection of therapeutic approaches. Due to the difficulty of completely capturing tumor heterogeneity in conventional detection methods, Tumor-on-Chip (TOC) devices with culturing patient-derived spheroids could be an appropriate alternative. In this research, human-derived spheroids from breast cancer individuals were cultured for 6 days in microfluidic devices. To compare TOC data with conventional detection methods, immunohistochemistry (IHC) and ITRAQ data were employed, and various protein expressions were validated using the transcriptomic databases. The behavior of the spheroids in the collagen matrix and the cell viability were monitored over 6 days of culture. IHC and immunocytochemistry (ICC) results revealed that inter and intra-heterogeneity of tumor spheroids are associated with HER2/ER expression. HER2 expression levels revealed a more important biomarker associated with invasion in the 3D culturing of spheroids. The expression levels of CD163 (as a marker for Ma2 macrophages) and CD44 (a marker for cancer stem cells (CSCs)) were also evaluated. Interestingly, the levels of M2a macrophages and CSCs were higher in triple-negative specimens and samples that showed higher migration and invasion. Cell density and extracellular matrix (ECM) stiffness were also important factors affecting the migration and invasion of the spheroids through the matrix. Among these, rigid ECM revealed a more crucial role than cell density. To sum up, these research findings demonstrated that human-derived spheroids from breast cancer specimens in microfluidic devices provide a dynamic condition for predicting tumor heterogeneity in patients, which can help move the field forward for better and more accurate therapeutic strategies.

Introduction  CD44, a multistructural and multifunctional transmembrane glycoprotein, is a promising cancer stem cell (CSC) marker that regulates the properties of CSCs, including self-renewal, tumor initiation, and metastasis, and confers resistance to chemotherapy and radiotherapy. The aim of the present study was to evaluate the gene and protein expression of CD44 and explore its prognostic value in head and neck squamous cell carcinoma (HNSCC). Methodology  The present observational study employs computational tools for analysis. The Cancer Genome Atlas Head-Neck Squamous Cell Carcinoma dataset (520 primary HNSCC and 44 normal tissues) from the University of Alabama at Birmingham Cancer platform was used to study the association of CD44 mRNA transcript levels with various clinicopathological characteristics of HNSCC including age, gender, tumor grade, tumor stage, human papillomavirus (HPV) status, p53 mutation status, and overall survival. The CD44 protein expression in HNSCC and normal tissues was ascertained using the National Cancer Institute\'s Clinical Proteomic Tumor Analysis Consortium Head-and-Neck cancer dataset (108 primary HNSCC and 71 normal tissues). Results  CD44 mRNA transcript and protein expression levels were significantly higher in HNSCC tissues than in normal tissues, and high CD44 expression was correlated with poor survival. CD44 was upregulated in Stage 1 and Grade 2 HNSCC compared with other stages and grades. Overexpression of CD44 was observed in HPV-negative and TP53-positive mutant status in HNSCC. Conclusion  The pleiotropic roles of CD44 in tumorigenesis urge the need to explore its differential expression in HNSCC. The study concludes that CD44 can be a potential diagnostic and prognostic biomarker for HNSCC and offer new molecular targets for CD44-targeted therapy for cancer management.

Background: LGR5 is one of the most important stem cell markers for colorectal cancer (CRC), as it potentiates Wnt/Β-catenin signaling. The well-characterized deregulation of Wnt/Β-catenin signaling that occurs during adenoma/carcinoma sequence in CRC renders LGR5 a hopeful therapeutic target. We assessed the immunohistochemical expression of LGR5 and Β-catenin in normal colonic and tumorous lesions with a clinicopathological correlation. Methods: Tissue blocks and clinical data of 50 selected cases were included: 8 from normal mucosa, 12 cases of adenoma, and 30 cases of CRC, where sections were cut and re-examined and the immunohistochemical technique was conducted using anti-LGR5 and anti-Β-catenin to measure the staining density. Results: There was no expression of LGR5 in normal mucosa compared to samples of adenoma and CRC samples. The association analysis showed that CRC specimens were more likely to have strong LGR5 and Β-catenin expressions than the other two groups (p = 0.048 and p < 0.001, respectively). Specimens with high-grade dysplastic adenoma were more likely to express moderate-to-strong expression of LGR5 and Β-catenin (p = 0.013 and p = 0.036, respectively). In contrast, there were no statistically significant associations between LGR5 and Β-catenin expression with grade and stage. Conclusion: These results suggest and support the possible role of LGR5 as a potential marker of cancer stem cells in sporadic colorectal carcinogenesis in addition to a prognostic value for LGR5 and Β-catenin in adenomatous lesions according to immunohistochemical expression density. A potential therapeutic role of LGR5 in CRC is suggested for future studies based on its role in pathogenesis.

UNASSIGNED: Anaplastic thyroid cancer (ATC) is one of the most aggressive malignancies in humans that accounts for a considerable rate of cancer-associated mortality. Since conventional therapies are lacking sufficient efficacy, new treatment approaches are required. This goal could be achieved through a better understanding of the molecular pathogenesis of ATC. Thyroid tumorigenesis is initiated by a subpopulation of cells known as cancer stem cells (CSCs) with specific markers such as CD133 that confers to processes such as self-renewal and metastasis. Besides, some long non-coding RNAs (lncRNAs) promote tumorigenesis by mediating the aforementioned processes.
UNASSIGNED: Here, we designed an exploratory study to investigate the role of lncRNAs ROR and MALAT1 and their related genes in CSC stemness. Using magnetic-activated cell sorting (MACS), the CD133- and CD133+ subpopulations were separated in SW1736 and C643 ATC cell lines. Next, the expression profiles of the CD133 marker, MALAT1, and its associated genes (CCND1, NESTIN, MYBL2, MCL1, IQGAP1), as well as ROR and its related genes (POU5F1, SOX2, NANOG), were explored by qRT-PCR.
UNASSIGNED: We found significant up-regulation of ROR, POU5F1, SOX2, NANOG, CD133, MALAT1, IQGAP1, and MCL1 in CD133+ SW1736 cells compared to CD133- cells. As for CD133+ C643 cells, CCND1, IQGAP1, POU5F1, SOX2, NANOG, and NESTIN were significantly up-regulated compared to CD133- cells.
UNASSIGNED: This study suggests that these lncRNAs in CD133-positive SW1736 and C643 cells might regulate stemness behaviors in ATC.

MicroRNA-140 (miR-140) acts as a tumor suppressor and plays a vital role in cell biological functions such as cell proliferation, apoptosis, and DNA repair. The expression of this miRNA has been shown to be considerably decreased in cancer tissues and cell lines compared with normal adjacent tissues. Consequently, aberrant expression of some miR-140 target genes can lead to the initiation and progression of various human cancers, such as breast cancer, gastrointestinal cancers, lung cancer, and prostate cancer. The dysregulation of the miR-140 network also affects cell proliferation, invasion, metastasis, and apoptosis of cancer cells by affecting various signaling pathways. Besides, up-regulation of miR-140 could enhance the efficacy of chemotherapeutic agents in different cancer. We aimed to cover most aspects of miR-140 function in cancer development and address its importance in different stages of cancer progression.

The human body is endowed with an extraordinary ability to maintain different oxygen levels in various tissues and organs. The maintenance of physiological levels of oxygen is known as physoxia. The development of hypoxic conditions plays an important role in the biology of several pathologies, including cancer. In vitro studies using normal and neoplastic cells require that culture conditions be carried out under appropriate oxygen levels, either physoxic or hypoxic conditions. Such requirements are difficult to widely implement in laboratory practice, mainly due to the high costs of specialized equipment. In this work, we present and characterize a cost-effective method to culture cells under a range of oxygen levels using deoxidizing pouches. Our results show that physoxic and hypoxic levels using deoxidizing absorbers can be achieved either by implementing a gradual change in oxygen levels or by a regimen of acute depletion of oxygen. This approach triggers the activation of an epithelial-mesenchymal transition in cancer cells while stimulating the expression of HIF-1α. Culturing cancer cells with deoxidizing agent pouches revealed PI3K oncogenic pathway exacerbations compared to tumor cells growing under atmospheric levels of oxygen. Similar to the PI3K signaling disturbance, we also observed augmented oxidative stress and superoxide levels and increased cell cycle arrest. Most interestingly, the culture of cancer cells under hypoxia resulted in the accumulation of cancer stem cells in a time-dependent manner. Overall, we present an attractive, cost-effective method of culturing cells under appropriate physoxic or hypoxic conditions that is easily implementable in any wet laboratory equipped with cell culture tools.

BACKGROUND: This study aims to explore the mechanism of drug resistance in multiple myeloma (MM).
METHODS: In this study, the possible mechanism of chemotherapeutic tolerance was preliminarily explored from two aspects: (I) the changes in cell morphology, cell cycle, cell apoptosis, stem cell markers and the signaling transduction pathway after the irradiation of RPMI-8226 cells; (II) the mechanism of enhancing chemotherapeutic sensitivity through the PI3K/AKT/mTOR signaling pathway.
RESULTS: The results showed that the cell morphology and cell cycle of RPMI-8226 had been significantly changed after receiving a radiation dose of 6 Gy in the logarithmic growth phase, the sensitivity of cells to bortezomib had been decreased, the level of stem cell markers had been upregulated, and the PI3K/AKT/mTOR signaling pathway had been activated. However, blocking the PI3K/AKT/mTOR signaling pathway caused the expression of the stem cell markers of RPMI-8226 cells. In addition, the sensitivity of cells to bortezomib had been increased.
CONCLUSIONS: Blocking the PI3K/AKT/mTOR might decrease the RPMI-8226 cells which survived the radiotherapy and increase the sensitivity of cells to bortezomib.

Cancer stem cells (CSCs) are a relatively rare subpopulation of tumor cell with self-renewal and tumorigenesis capabilities. CSCs are associated with cancer recurrence, progression, and chemoradiotherapy resistance. Establishing a reliable platform for CSC enrichment and study is a prerequisite for understanding the characteristics of CSCs and discovering CSC-related therapeutic strategies. Certain strategies for CSC enrichment have been used in laboratory, particularly fluorescence-activated cell sorting (FACS) and mammosphere culture. However, these methods fail to recapitulate the in vivo chemical and physical conditions in tumors, thus potentially decreasing the malignancy of CSCs in culture and yielding unreliable research results. Accumulating research suggests the promise of a biomaterial-based three-dimensional (3D) strategy for CSC enrichment and study. This strategy has an advantage over conventional methods in simulating the tumor microenvironment, thus providing a more effective and predictive model for CSC laboratory research. In this review, we first briefly discuss the conventional methods for CSC enrichment and study. We then summarize the latest advances and challenges in biomaterial-based 3D CSC platforms. Design strategies for materials, morphology, and chemical and physical cues are highlighted to provide direction for the future construction of platforms for CSC enrichment and study.

Cancer stem cells (CSCs) represent a subset of tumor cells that are responsible for recurrence and metastasis of tumors. These cells are resistant to radiotherapy and chemotherapy. Immunotherapeutic strategies that target CSCs specifically have provided initial results; however, the mechanism of action of these strategies is unclear. The data were requested from The Cancer Genome Atlas and Genotype-Tissue Expression, followed with the survival analysis and weighted gene co-expression network analysis to detect survival and stemness related genes. Patients were divided into three groups based on their immune status by applying single sample GSEA (ssGSEA) with proven dependability by ESTIMATE analysis. The filtered key genes were analyzed using oncomine, GEPIA, HPA, qRT-PCR, and functional analysis. Patients in a group with a higher stemness and a lower immune infiltration showed a worse overall survival probability, stemness and immune infiltration characteristics of breast cancer progressed in a non-linear fashion. Thirteen key genes related to stemness and immunity were identified and the functional analysis indicated their crucial roles in cell proliferation and immune escape strategies. The qRT-PCR results showed that the expression of PIMREG and MTFR2 differed in different stages of patients. Our study revealed a promising potential for CSC-target immunotherapy in the early stage of cancer and a probable value for PIMREG and MTFR2 as biomarkers and targets for immunotherapy.

Standard supportive care during induction therapy for high-risk neuroblastoma (HR-NBL) includes primary prophylactic granulocyte colony-stimulating factor (G-CSF) aimed at limiting duration of neutropenia, reducing infection risk, and minimizing treatment delays. Preclinical models suggest that G-CSF promotes maintenance of neuroblastoma cancer stem cells and may reduce the efficacy of chemotherapy. This study\'s objective was to determine the safety and feasibility of administering induction chemotherapy without routine use of prophylactic G-CSF.
Children with newly diagnosed HR-NBL received six-cycle induction chemotherapy regimen without prophylactic G-CSF in four cycles. G-CSF was administered for stem cell mobilization after cycle 3 and granulocyte-monocyte colony-stimulating factor after cycle 5 prior to surgical resection of primary disease. The primary outcome measure was the incidence of grade 3 or higher infection. We hypothesized that the per patient infection rate would be comparable to our institutional baseline rate of 58% in patients with HR-NBL receiving induction chemotherapy with prophylactic growth factor support. The trial used an A\'Hern single-stage design.
Twelve patients with HR-NBL received 58 cycles of chemotherapy on study. Three patients completed the entire six-cycle regimen with no infections. Nine patients experienced grade 3 infections (bacteremia four, urinary tract infection two, skin/soft tissue infection three). No patients experienced grade 4 infections or required intensive care treatment for infection.
A greater than expected number of serious bacterial infections were observed during administration of induction chemotherapy for HR-NBL without primary prophylactic G-CSF. These results support continued prophylactic administration growth factor during induction chemotherapy.