Cancer Stem Cell-like Cells Research Articles

Identification of the Neurokinin-1 Receptor as Targetable Stratification Factor for Drug Repurposing in Pancreatic Cancer.

Simple SummaryPancreatic ductal adenocarcinoma is the most common form of pancreatic cancer. It is known for low life expectancies after diagnosis and very limited treatment options. The identification of new therapeutic molecular targets is urgent as it might allow faster development of new treatment strategies. Targeting the neurokinin-1 receptor with small molecules has previously shown anti-tumoral effects in a large variety of cancers. Here, we found specific types of pancreatic cells to express the neurokinin-1 receptor while at the same time showing positive treatment response represented by cell growth reduction in number and size when treated with aprepitant. Our results suggest the neurokinin-1 receptor as a promising targetable structure and therefore interesting in the concept of personalized medicine.The SP/NK1R-complex plays an important role in tumor proliferation. Targeting of the neurokinin-1 receptor in previous studies with its antagonist aprepitant (AP) resulted in anti-tumoral effects in colorectal cancer and hepatoblastoma. However, there is still a lack of knowledge regarding its effects on pancreatic cancer. Therefore, we treated human pancreatic ductal adenocarcinoma (PDAC) cell lines (Capan-1, DanG, HuP-T3, Panc-1, and MIA PaCa-2) and their cancer stem cell-like cells (CSCs) with AP and analyzed functional effects by MTT-, colony, and sphere formation assays, respectively; moreover, we monitored downstream mechanisms by flow cytometry. NK1R inhibition resulted in dose-dependent growth reduction in both CSCs and non-CSCs without induction of apoptosis in most PDAC cell lines. More importantly, we identified striking AP dependent cell cycle arrest in all parental cells. Furthermore, gene expression and the importance of key genes in PDAC tumorigenesis were analyzed combining RT-qPCR in eight PDAC cell lines with publicly available datasets (TCGA, GEO, CCLE). Surprisingly, we found a better overall survival in patients with high NK1R levels, while at the same time, NK1R was significantly decreased in PDAC tissue compared to normal tissue. Interestingly, there is currently no differentiation between the isoforms of NK1R (truncated and full; NK1R-tr and -fl) in any of the indicated public transcriptomic records, although many publications already emphasize on important regulatory differences between the two isoforms of NK1R in many cancer entities. In conclusion, analysis of splice variants might potentially lead to a stratification of PDAC patients for NK1R-directed therapies. Furthermore, we presume PDAC patients with high expressions of NK1R-tr might benefit from treatment with AP to improve chemoresistance. Therefore, analysis of splice variants might potentially lead to a stratification of PDAC patients for NK1R-directed therapies.

CRISPR/Cas9-mediated knockout of PIM3 suppresses tumorigenesis and cancer cell stemness in human hepatoblastoma cells.

Hepatoblastoma remains one of the most difficult childhood tumors to treat and is alarmingly understudied. We previously demonstrated that Proviral Insertion site in Maloney murine leukemia virus (PIM) kinases, specifically PIM3, are overexpressed in human hepatoblastoma cells and function to promote tumorigenesis. We aimed to use CRISPR/Cas9 gene editing with dual gRNAs to introduce large inactivating deletions in the PIM3 gene and achieve stable PIM3 knockout in the human hepatoblastoma cell line, HuH6. PIM3 knockout of hepatoblastoma cells led to significantly decreased proliferation, viability, and motility, inhibited cell-cycle progression, decreased tumor growth in a xenograft murine model, and increased animal survival. Analysis of RNA sequencing data revealed that PIM3 knockout downregulated expression of pro-migratory and pro-invasive genes and upregulated expression of genes involved in apoptosis and differentiation. Furthermore, PIM3 knockout decreased hepatoblastoma cancer cell stemness as evidenced by decreased tumorsphere formation, decreased mRNA abundance of stemness markers, and decreased cell surface expression of CD133, a marker of hepatoblastoma stem cell-like cancer cells. Reintroduction of PIM3 into PIM3 knockout cells rescued the malignant phenotype. Successful CRISPR/Cas9 knockout of PIM3 kinase in human hepatoblastoma cells confirmed the role of PIM3 in promoting hepatoblastoma tumorigenesis and cancer cell stemness.

PIM kinases mediate resistance to cisplatin chemotherapy in hepatoblastoma

Despite increasing incidence, treatment for hepatoblastoma has not changed significantly over the past 20 years. Chemotherapeutic strategies continue to rely on cisplatin, as it remains the most active single agent against hepatoblastoma. However, chemoresistance remains a significant challenge with 54–80% of patients developing resistance to chemotherapy after 4–5 cycles of treatment. Stem cell-like cancer cells (SCLCCs) are a subset of cells thought to play a role in chemoresistance and disease recurrence. We have previously demonstrated that Proviral Integration site for Moloney murine leukemia virus (PIM) kinases, specifically PIM3, play a role in hepatoblastoma cell proliferation and tumor growth and maintain the SCLCC phenotype. Here, we describe the development of a cisplatin-resistant hepatoblastoma xenograft model of the human HuH6 cell line and a patient-derived xenograft, COA67. We provide evidence that these cisplatin-resistant cells are enriched for SCLCCs and express PIM3 at higher levels than cisplatin-naïve cells. We demonstrate that PIM inhibition with AZD1208 sensitizes cisplatin-resistant hepatoblastoma cells to cisplatin, enhances cisplatin-mediated apoptosis, and decreases the SCLCC phenotype seen with cisplatin resistance. Together, these findings indicate that PIM inhibition may be a promising adjunct in the treatment of hepatoblastoma to effectively target SCLCCs and potentially decrease chemoresistance and subsequent disease relapse.

Characterization of stem cell-like property in cancer cells based on single-cell impedance measurement in a microfluidic platform

Investigation of stem cell-like property in cancer cells is important for the development of new therapeutic drugs targeting at malignant tumors. Currently, the standard approach for identifying cancer stem cell-like cells relies on the recognition of stem cell surface markers. However, the reliability remains controversial among biologists. In the current work, a dielectrophoretic and impedimetric hybrid microfluidic platform was developed for capturing single cells and characterizing their stem cell-like property. Single cells were captured in 20 μm trapping wells by dielectrophoretic force and their impedance spectra were measured by an impedance analyzer. The result showed that different cancer cell lines could be differentiated by impedance magnitude ranging between 2 and 20 kHz. Moreover, cancer cells and cancer stem cell-like cells could be categorized by a 2-dimensional graph of the impedance magnitudes at 2 and 20 kHz. The stem cell-like property in cancer cells was verified by stem cell surface markers and single-cell derived colony assay. Comparing with bio-chemical approach, i.e., surface markers, bio-physical approach, i.e., cell impedance, is a label-free technique to identify cancer stem cell-like cells.

Novel second-generation rexinoid induces growth arrest and reduces cancer cell stemness in human neuroblastoma patient-derived xenografts

IntroductionThe poor therapeutic efficacy seen with current treatments for neuroblastoma may be attributed to stem cell-like cancer cells (SCLCCs), a subpopulation of cancer cells associated with poor prognosis and disease recurrence. Retinoic acid (RA) is a differentiating agent used as maintenance therapy for high-risk neuroblastoma but nearly half of children treated with RA relapse. We hypothesized that 6-Methyl-UAB30 (6-Me), a second-generation rexinoid recently developed with a favorable toxicity profile compared to RA, would reduce cancer cell stemness in human neuroblastoma patient-derived xenografts (PDXs). MethodsCells from three neuroblastoma PDXs were treated with 6-Me and proliferation, viability, motility, and cell-cycle progression were assessed. CD133 expression, sphere formation, and mRNA abundance of stemness and differentiation markers were evaluated using flow cytometry, in vitro extreme limiting dilution analysis, and real-time PCR, respectively. ResultsTreatment with 6-Me decreased proliferation, viability, and motility, and induced cell-cycle arrest and differentiation in all three neuroblastoma PDXs. In addition, 6-Me treatment led to decreased CD133 expression, decreased sphere-forming ability, and decreased mRNA abundance of Oct4, Nanog, and Sox2, indicating decreased cancer cell stemness. Conclusions6-Me decreased oncogenicity and reduced cancer cell stemness of neuroblastoma PDXs, warranting further exploration of 6-Me as potential novel therapy for neuroblastoma.

Radiosensitization of breast cancer cells using AS1411 aptamer-conjugated gold nanoparticles

BackgroundGold nanoparticles (GNPs) have been used to sensitize cancer cells and enhance the absorbed dose delivered to such cells. Active targeting can provide specific effect and higher uptake of the GNPs in the tumor cells, while having small effect on healthy cells. The aim of this study was to assess the possible radiosensitiazation effect of GNPs conjugated with AS1411 aptamer (AS1411/GNPs) on cancer cells treated with 4 MeV electron beams.Materials and methodsCytotoxicity studies of the GNPs and AS1411/GNPs were carried out with MTT and MTS assay in different cancer cell lines of MCF-7, MDA-MB-231 and mammospheres of MCF-7 cells. Atomic absorption spectroscopy confirmed the cellular uptake of the gold particles. Radiosensitizing effect of the GNPs and AS1411/GNPs on the cancer cells was assessed by clonogenic assay.ResultAS1411 aptamer increased the Au uptake in MCF-7 and MDA-MB-231 cells. Clonogenic survival data revealed that AS1411/GNPs at 12.5 mg/L could result in radiosensitization of the breast cancer cells and lead to a sensitizer enhancement ratio of 1.35 and 1.66 and 1.91 for MCf-7, MDA-MB-231 and mammosphere cells.ConclusionGold nanoparticles delivery to the cancer cells was enhanced by AS1411 aptamer and led to enhanced radiation induced cancer cells death. The combination of our clonogenic assay and Au cell uptake results suggested that AS1411 aptamer has enhanced the radiation-induced cell death by increasing Au uptake. This enhanced sensitization contributed to cancer stem cell-like cells to 4 MeV electron beams. This is particularly important for future preclinical testing to open a new insight for the treatment of cancers.

A `one-two punch' therapy strategy to target chemoresistance in estrogen receptor positive breast cancer

Cancer cell phenotypes evolve during a tumor's treatment. In some cases, tumor cells acquire cancer stem cell-like (CSL) traits such as resistance to chemotherapy and diminished differentiation; therefore, targeting these cells may be therapeutically beneficial. In this study we show that in progressive estrogen receptor positive (ER+) metastatic breast cancer tumors, resistant subclones that emerge following chemotherapy have increased CSL abundance. Further, in vitro organoid growth of ER+ patient cancer cells also shows that chemotherapy treatment leads to increased abundance of ALDH+/CD44+ CSL cells. Chemotherapy induced CSL abundance is blocked by treatment with a pan-HDAC inhibitor, belinostat. Belinostat treatment diminished both mammosphere formation and size following chemotherapy, indicating a decrease in progenitor CSL traits. HDAC inhibitors specific to class IIa (HDAC4, HDAC5) and IIb (HDAC6) were shown to primarily reverse the chemo-resistant CSL state. Single-cell RNA sequencing analysis with patient samples showed that HDAC targets and MYC signaling were promoted by chemotherapy and inhibited upon HDAC inhibitor treatment. In summary, HDAC inhibition can block chemotherapy-induced drug resistant phenotypes with ‘one-two punch’ strategy in refractory breast cancer cells.

Teratoma Growth Retardation by HDACi Treatment of the Tumor Embryonal Source.

Simple SummaryTesticular germ cell tumors are the most common neoplasms in young male populations, with a rising incidence. Among them, teratomas may often be very aggressive and resistant to therapy. Our aim was to investigate the impact of two potential anti-tumor epigenetic drugs (Valproate and Trichostatin A) in a mammalian model of teratoma development from an early trilaminar mouse embryo. Both drugs applied to the embryonic tissue had a significant negative impact on the teratoma growth in a three-dimensional in vitro culture. However, Trichostatin A did not diminish some potentially dangerous features of teratomas in contrast to Valproate. This research is an original contribution to the basic knowledge of the origin and development of teratomas. Such knowledge is necessary for envisioning therapeutic strategies against human testicular tumors.Among testicular germ cell tumors, teratomas may often be very aggressive and therapy-resistant. Our aim was to investigate the impact of histone deacetylase inhibitors (HDACi) on the in vitro growth of experimental mouse teratoma by treating their embryonic source, the embryo-proper, composed only of the three germ layers. The growth of teratomas was measured for seven days, and histopathological analysis, IHC/morphometry quantification, gene enrichment analysis, and qPCR analysis on a selected panel of pluripotency and early differentiation genes followed. For the first time, within teratomas, we histopathologically assessed the undifferentiated component containing cancer stem cell-like cells (CSCLCs) and differentiated components containing numerous lymphocytes. Mitotic indices were higher than apoptotic indices in both components. Both HDACi treatments of the embryos-proper significantly reduced teratoma growth, although this could be related neither to apoptosis nor proliferation. Trichostatin A increased the amount of CSCLCs, and upregulated the mRNA expression of pluripotency/stemness genes as well as differentiation genes, e.g., T and Eomes. Valproate decreased the amount of CSCLCs, and downregulated the expressions of pluripotency/stemness and differentiation genes. In conclusion, both HDACi treatments diminished the inherent tumorigenic growth potential of the tumor embryonal source, although Trichostatin A did not diminish the potentially dangerous expression of cancer-related genes and the amount of CSCLC.

Upregulation of Peroxiredoxin-2 in Well-Differentiated Pancreatic Neuroendocrine Tumors and Its Utility as a Biomarker for Predicting the Response to Everolimus.

Pancreatic neuroendocrine neoplasms (pNENs) account for 2–3% of pancreatic malignancies. Peroxiredoxins (Prdxs), which are major cellular antioxidants, are involved in multiple oncogenic signaling pathways. We investigated the role of peroxiredoxin-2 in QGP-1 human pNEN cell line and patient-derived pNEN tissue. To validate the cancer stem cell-like cell characteristics of QGP-1 cells in spheroid culture, in vitro analyses and xenografting were performed. Furthermore, immunohistochemical staining was conducted to verify the overexpression of Prdx2 in pNEN tissue. Prdx2 expression was high at the mRNA and protein levels in QGP-1 cells. Prdx2 was also overexpressed in patient-derived pNEN tissue. Silencing of Prdx2 using siRNA induced overexpression and phosphorylation of ERK and AKT in QGP-1. Cell proliferation was increased by treating QGP-1 cells with siPrdx2, and the IC50 of everolimus increased suggesting resistance to everolimus. Interestingly, QGP-1 spheroid cells, which exhibited cancer stem cell-like features, exhibited lower expression of Prdx2 and mTOR. The results suggest that Prdx2 expression level and its activity may be a potential predictive biomarker for therapeutic response or resistance to everolimus in pNEN.

CD44+ cells determine fenofibrate-induced microevolution of drug-resistance in prostate cancer cell populations.

Combinations of metabolic blockers (including fenofibrate) with chemotherapeutic drugs interfere with the drug‐resistance of prostate cancer cells. However, their effect on cancer stem cells‐dependent microevolution of prostate cancer malignancy remains unaddressed. Here, we hypothesize that the combined docetaxel/fenofibrate treatment prompts the selective expansion of cancer stem cells that affects the microevolution of their progenies. Accordingly, we adapted a combined in vitro/in vivo approach to identify biological and therapeutic consequences of this process. Minute subpopulations of docetaxel‐resistant CD133high and/or CD44high cancer stem cell‐like (SCL) cells were found in prostate cancer DU145 and PC3 cell populations. When pretreated with docetaxel, they readily differentiated into docetaxel‐resistant CD44negative “bulk” cells, thus accounting for the microevolution of drug‐resistant cell lineages. Combined docetaxel/fenofibrate treatment induced the generation of poly(morpho)nuclear giant cells and drug‐resistant CD44high SCL cells. However, the CD44negative offspring of docetaxel‐ and docetaxel/fenofibrate‐treated SCLs remained relatively sensitive to the combined treatment, while retaining enhanced resistance to docetaxel. Long‐term propagation of drug‐resistant SCL‐derived lineages in the absence of docetaxel/fenofibrate resulted in their reverse microevolution toward the drug‐sensitivity and invasive phenotype. Consequently, prostate tumors were able to recover from the combined docetaxel/fenofibrate stress after the initial arrest of their expansion in vivo. In conclusion, we have confirmed the potential of fenofibrate for the metronomic treatment of drug‐resistant prostate tumors. However, docetaxel/fenofibrate‐induced selective expansion of hyper‐resistant CD44high SCL prostate cells and their “bulk” progenies prompts the microevolution of prostate tumor drug‐resistance. This process can limit the implementation of metabolic chemotherapy in prostate cancer treatment.

Bone Marrow-Derived Myofibroblasts Promote Gastric Cancer Metastasis by Activating TGF-β1 and IL-6/STAT3 Signalling Loop.

BackgroundMurine bone marrow-derived myofibroblasts (BMFs) have previously been shown to promote gastric cancer growth. However, whether BMFs promote gastric cancer cell metastasis remains largely unknown.MethodsWound healing assay, Transwell invasion and migration assay and 3D organotypic co-culture systems were conducted to study the effects of BMFs on invasion and migration of gastric cancer cells and the invasion and migration ability of gastric cancer stem cell-like cells (CSC-LCs) induced by BMFs. We employed two animal model to study the role of BMFs on the in vivo metastasis of gastric cancer cells and the metastatic ability of gastric BMF-induced CSC-LCs. A human gastric cancer tissue microarray and TCGA gastric cancer database were analysed to study the relationship between the expression of IL-6 and TGF-β1 and clinicopathological characteristics and survival in gastric cancer.ResultsWe found that BMFs promoted the in vitro migration and invasion of gastric cancer cells. BMFs promoted liver, lung, subcutaneous, and splenic metastases of MKN28 cells in the spleen injection liver metastasis model and co-injection of caudal vein (IOCV) mouse model. BMFs reprogrammed non-gastric cancer stem cell (CSC) to CSC-LCs and enhanced CSC-LC migration and metastasis. BMF-derived IL-6 and gastric cancer cell-secreted TGF-β1 mediated the interaction between BMFs and gastric cancer cells, promoting tumour metastasis. BMFs enhanced the expressions of STAT3 and p-STAT3 in co-cultured gastric cancer cells. A combination of Napabucasin and Galunisertib exhibited the strongest inhibition of cell migration compared to when administered alone. Gastric cancer tissue array and TCGA database indicated that the overexpression of IL-6 and TGF-β1 was associated with gastric cancer metastasis.ConclusionOur results demonstrated that BMFs promote gastric cancer metastasis through the activation of the TGF-β1 and IL-6/STAT3 signalling pathways. Targeting the inhibition of these interactions may be a potent therapeutic strategy for addressing gastric cancer metastasis.

9-cis-UAB30, a novel rexinoid agonist, decreases tumorigenicity and cancer cell stemness of human neuroblastoma patient-derived xenografts

Retinoic acid (RA) therapy has been utilized as maintenance therapy for high-risk neuroblastoma, but over half of patients treated with RA relapse. Neuroblastoma stem cell-like cancer cells (SCLCCs) are a subpopulation of cells characterized by the expression of the cell surface marker CD133 and are hypothesized to contribute to drug resistance and disease relapse. A novel rexinoid compound, 9-cis-UAB30 (UAB30), was developed having the same anti-tumor effects as RA but a more favorable toxicity profile. In the current study, we investigated the efficacy of UAB30 in neuroblastoma patient-derived xenografts (PDX). Two PDXs, COA3 and COA6, were utilized and alterations in the malignant phenotype were assessed following treatment with RA or UAB30. UAB30 significantly decreased proliferation, viability, and motility of both PDXs. UAB30 induced cell-cycle arrest as demonstrated by the significant increase in percentage of cells in G1 (COA6: 33.7 ± 0.7 vs. 43.3 ± 0.7%, control vs. UAB30) and decrease in percentage of cells in S phase (COA6: 44.7 ± 1.2 vs. 38.6 ± 1%, control vs. UAB30). UAB30 led to differentiation of PDX cells, as evidenced by the increase in neurite outgrowth and mRNA abundance of differentiation markers. CD133 expression was decreased by 40% in COA6 cells after UAB30. The ability to form tumorspheres and mRNA abundance of known stemness markers were also significantly decreased following treatment with UAB30, further indicating decreased cancer cell stemness. These results provide evidence that UAB30 decreased tumorigenicity and cancer cell stemness in neuroblastoma PDXs, warranting further exploration as therapy for high-risk neuroblastoma.

Exosomal lncRNA DOCK9-AS2 derived from cancer stem cell-like cells activated Wnt/\u03b2-catenin pathway to aggravate stemness, proliferation, migration, and invasion in papillary thyroid carcinoma

Exosomal long non-coding RNAs (lncRNAs) are crucial factors that mediate the extracellular communication in tumor microenvironment. DOCK9 antisense RNA2 (DOCK9-AS2) is an exosomal lncRNA which has not been investigated in papillary thyroid carcinoma (PTC). Based on the result of differentially expressed lncRNAs in PTC via bioinformatics databases, we discovered that DOCK9-AS2 was upregulated in PTC, and presented elevation in plasma exosomes of PTC patients. Functionally, DOCK9-AS2 knockdown reduced proliferation, migration, invasion, epithelial-to-mesenchymal (EMT) and stemness in PTC cells. PTC-CSCs transmitted exosomal DOCK9-AS2 to improve stemness of PTC cells. Mechanistically, DOCK9-AS2 interacted with SP1 to induce catenin beta 1 (CTNNB1) transcription and sponged microRNA-1972 (miR-1972) to upregulate CTNNB1, thereby activating Wnt/β-catenin pathway in PTC cells. In conclusion, PTC-CSCs-derived exosomal lncRNA DOCK9-AS2 activated Wnt/β-catenin pathway to aggravate PTC progression, indicating that DOCK9-AS2 was a potential target for therapies in PTC.

Glycolysis, via NADH-dependent dimerisation of CtBPs, regulates hypoxia-induced expression of CAIX and stem-like breast cancer cell survival.

Adaptive responses to hypoxia are mediated by the hypoxia-inducible factor (HIF) family of transcription factors. These responses include the upregulation of glycolysis to maintain ATP production. This also generates acidic metabolites, which require HIF-induced carbonic anhydrase IX (CAIX) for their neutralisation. C-terminal binding proteins (CtBPs) are coregulators of gene transcription and couple glycolysis with gene transcription due to their regulation by the glycolytic coenzyme NADH. Here, we find that experimental manipulation of glycolysis and CtBP function in breast cancer cells through multiple complementary approaches supports a hypothesis whereby the expression of known HIF-inducible genes, and CAIX in particular, adapts to available glucose in the microenvironment through a mechanism involving CtBPs. This novel pathway promotes the survival of stem cell-like cancer (SCLC) cells in hypoxia.

Abstract B053: Expression of alternative mRNA splicing variant MBD2_v2 promotes triple-negative breast cancer tumor initiation and is associated with body mass index

Abstract According to epidemiologic research, obesity is a risk factor for triple-negative breast cancer (TNBC). The underlying molecular biology remains unknown. We reasoned that obesity-induced chronic inflammation, reactive oxygen species (ROS) being central, serves as the general link to TNBC. We are the first to report that expression of the epigenetic reader methyl-CpG-binding domain protein 2 mRNA variant MBD2_v2 in TNBC cell cultures depends on ROS and is necessary to maintain and promote expansion of cancer stem cell-like cells (CSCs). The relevance of CSCs is that they are a subpopulation of cancer cells recognized as the source of malignant tumor initiation, and they give rise to drug resistance and metastatic recurrence. We also previously reported evidence that MBD2_v2 expression underlies TNBC resistance to EGFR inhibitor drugs. Now, having used a diet-induced obesity (DIO) mouse model that mimics human obesity, we report that MBD2_v2 and serine/arginine-rich mRNA splicing factor 2 (SRSF2) levels were increased in tumors that formed more frequently in DIO mice relative to lean controls. To more directly test if increased MBD2_v2 drives increased tumor initiation capacity, we stably modified MBD2_v2 or SRSF2 expression in TNBC cells prior to inoculation. MBD2_v2 overexpression increased tumor initiation while SRSF2 knockdown, resulting in decreased MBD2_v2 expression, attenuated tumor formation. In addition, our analysis of TNBC patient tumors revealed a significant positive association for MBD2_v2 expression and body mass index (BMI). African American (AA) women are 1.7 times more often obese relative to European American women, and a TNBC driver mechanism fueled by obesity-coupled inflammation could underlie the higher incidence of TNBC among AA women. Citation Format: Emily A. Teslow, Cristina Mitrea, Bin Bao, Ramzi M. Mohammad, Lisa A. Polin, Greg Dyson, Kristen S. Purrington, Aliccia Bollig-Fischer. Expression of alternative mRNA splicing variant MBD2_v2 promotes triple-negative breast cancer tumor initiation and is associated with body mass index [abstract]. In: Proceedings of the Eleventh AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2018 Nov 2-5; New Orleans, LA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl):Abstract nr B053.

Interleukin-31 Receptor α Is Required for Basal-Like Breast Cancer Progression.

Purpose: Interleukin-31 receptor α (IL31RA) usually mediates IL-31 induced inflammation and allergic diseases. However, the functional roles of IL-31/IL31RA signaling in basal-like breast cancer (BLBC) progression remain totally unclear.Methods: Tumorsphere formation, transwell, and wound healing assays were used to measure the BLBC progression. We implanted tumor cells in mammary fat pad and tail vein of nude mice to detect the growth and metastasis of BLBC cells. Luciferase and ChIP assays were employed to measure the transcriptional regulation. Western blot and real-time PCR assays as well as bio-informatics analyses were conducted to observe the expression of IL31RA.Results: We found that silencing of IL31RA suppresses the cancer stem cell-like properties, migration and invasion of BLBC cells in vitro as well as tumor growth and metastasis in vivo. Knockdown of IL31RA ameliorates IL-31-mediated pro-oncogenic functions. Overexpression of IL31RA in luminal breast cancer cells enhances the cancer stem cell-like properties and cell motility. Our data further identified IL31RA as a target gene of Twist/BRD4 transcription complex.Conclusion: Overall, these data indicate that IL31RA promotes basal-like breast cancer progression and metastasis, suggesting that targeting of IL-31/IL31RA axis might be beneficial to treatment of BLBC.

All-trans retinoic acid reduces cancer stem cell-like cell-mediated resistance to gefitinib in NSCLC adenocarcinoma cells

BackgroundThe enrichment of cancer stem cell-like cells (CSCs) has been considered to be responsible for tumor progression after an initial response to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (EGFR-TKIs) in patients with non-small cell lung adenocarcinoma (NSCLC/ADC). CSCs with ALDH1A1bright /CD44high expression contribute to the TKIs resistance in NSCLC/ADC cells. All-trans retinoic acid (ATRA) has been shown to be a potential targeted therapy against CSCs due to its ability to inhibit ALDH1A1 activity. We therefore investigated whether ATRA could circumvent the resistance to improve the response to gefitinib in NSCLC/ADC cells.MethodsTreatment of NSCLC/ADC A549 and H1650 cells with gefitinib enriched the gefitinib surviving cells (GSCs). The expression of ALDH1A1 and CD44 and the IC50 values for gefitinib were determined by flow cytometry (FCM) and crystal violet assay in GSCs and ATRA-treated GSCs, respectively. Using DEAB as the positive control, direct inhibitory effect of ATRA on ALDH1A1 activity was determined by ALDEFLUOR assay,ResultsGSCs showed higher expression of ALDH1A1 and CD44 and IC50 values for gefitinib than their respective parental cells, suggesting that gefitinib can lead to propagation of CSC-enriched gefitinib-resistant cells. Treatment with ATRA was found to significantly reduce the increased expression of ALDH1A1 and CD44 and the IC50 values for gefitinib in A549GSC and H1650GSC cells, and ATRA could directly inhibit active ALDH1A1 as compared to DEAB.ConclusionOur findings suggest that combination treatment with ATRA prevents gefitinib-induced enrichment of ALDH1A1bright/CD44high CSCs and enhances gefitinib-induced growth inhibition of NSCLC/ADC cells.

WIP1 promotes cancer stem cell properties by inhibiting p38 MAPK in NSCLC

Cancer stem cells (CSCs) are a small population of stem cell-like cancer cells that can initiate tumors in vivo, and are the major source of cancer initiation, relapse, and drug resistance. We previously reported that the p38 MAPK, through its downstream effectors MK2 and HSP27, suppressed CSC properties by downregulating the expression of transcription factors that mediate stemness in non-small-cell lung cancer (NSCLC) cells, and that despite unaltered total expression of total p38 proteins, the levels of activated p38 were reduced in NSCLC tissues. However, the mechanism underlying the reduced levels of activated p38 in NSCLC is unknown. In this study, we identified WIP1, a p38 phosphatase frequently overexpressed in cancer, as a suppressor of p38 in a pathway that regulates CSC properties in NSCLC. Increased WIP1 expression correlated with reduced levels of activated p38, and with increased levels of a CSC marker in NSCLC tissues. Further investigation revealed that WIP1 promoted stemness-related protein expression and CSC properties by inhibiting p38 activity in NSCLC cells. WIP1 inhibitors are currently under development as anticancer drugs based on their ability to reactivate p53. We found that a WIP1 inhibitor suppressed stemness-related protein expression and CSC properties by activating p38 in NSCLC cells in vitro and in vivo. These studies have identified the WIP1–p38–MK2–HSP27 cascade as a novel signaling pathway that, when altered, promotes CSC properties in NSCLC development, and have defined novel mechanisms underlying the oncogenic activity of WIP1 and the anticancer efficacy of WIP1 inhibitors.

Properties of Stem Cell-like Cancer Cells in Cultured Head and Neck Squamous Cell Carcinoma

Properties of Stem Cell-like Cancer Cells in Cultured Head and Neck Squamous Cell Carcinoma

MP51-17 SPINK1 IS ASSOCIATED WITH CASTRATION RESISTANCE IN PROSTATE CANCER

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology II (MP51)1 Apr 2020MP51-17 SPINK1 IS ASSOCIATED WITH CASTRATION RESISTANCE IN PROSTATE CANCER Ikenna Madueke, Wen-Yang Hu, Lishi Xie, Donald Vander Griend, Michael Abern, Gail Prins, and Ikenna Madueke* Ikenna MaduekeIkenna Madueke More articles by this author , Wen-Yang HuWen-Yang Hu More articles by this author , Lishi XieLishi Xie More articles by this author , Donald Vander GriendDonald Vander Griend More articles by this author , Michael AbernMichael Abern More articles by this author , Gail PrinsGail Prins More articles by this author , and Ikenna Madueke*Ikenna Madueke* More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000913.017AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: SPINK1 is a secreted serine protease inhibitor that has been implicated in prostate cancer although its contribution has been controversial. Previous RNA-seq data from our lab revealed SPINK1 to be one of the most differentially expressed genes enriched in PCa stem cells compared to non-diseased progenitor population. We thus sought to investigate its role in disease progression in a prostate cancer stem cell-like cell line. METHODS: We analyzed the STand Up to Cancer database of human RNA-seq data sets for differential expression of SPINK1 in metastatic and localized PCa. We utilized ShRNA to create stable knockdown of SPINK1 in HusLC cells, a PCa stem-like cell line. MTT assay was utilized to study its effects on proliferation. Migration was assessed with scratch assay. Stemness was assayed with prostasphere formation and RT-PCR. We then assessed xenograft formation and propagation in the presence or absence of testosterone. RESULTS: RNA-seq data set of annotated human PCa metastasis revealed an increase in SPINK1 mRNA expression compared with localized tumors. SPINK1 mRNA levels are increased in HuSLC prostaspheres compared to HuSLC cells cultured in monolayer supporting a role in maintenance of a progenitor population. Knockdown of SPINK1 (SPINK1-KD) however did not change the sphere-forming capacity by size or number but did decrease stemness genes Nanog, SOX2, and Oct4 suggesting a role in affecting stemness. Knockdown of SPINK1 decreased migration and proliferation of HuSLC in monolayer indicating a decrease in SPINK1 is associated with a less aggressive phenotype. Subcutaneous xenografts developed in non-castrate nude mice and with supplemental exogenous testosterone did not reveal a difference in tumor weight or with serial transplantation. SPINK1-KD xenografts however were more secretory as assessed by PSA suggesting an increased responsiveness to testosterone. Strikingly, when the knock-down and control xenografts were cultured in castrate conditions, SPINK1-KD xenografts were drastically smaller than control HuSLC suggesting that SPINK1 is associated with promotion of castration resistance. CONCLUSIONS: The clinical significance of SPINK1 in PCa remains controversial. However, taken together, our results present supporting evidence to suggest a role for SPINK1 in promoting castration resistance in the progression of PCa disease with implications for targeted treatment. Source of Funding: None © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e770-e770 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Ikenna Madueke More articles by this author Wen-Yang Hu More articles by this author Lishi Xie More articles by this author Donald Vander Griend More articles by this author Michael Abern More articles by this author Gail Prins More articles by this author Ikenna Madueke* More articles by this author Expand All Advertisement PDF downloadLoading ...