Roles of Tumor Suppressor Signaling on Reprogramming and Stemness Transition in Somatic Cells

link_to_OA_fulltex

Physiological β-catenin signaling controls self-renewal networks and generation of stem-like cells from nasopharyngeal carcinoma

BACKGROUND: A few reports suggested that low levels of Wnt signaling might drive cell reprogramming, but these studies could not establish a clear relationship between Wnt signaling and self-renewal networks. There are ongoing debates as to whether and how the Wnt/β-catenin signaling is involved in the control of pluripotency gene networks. Additionally, whether physiological β-catenin signaling generates stem-like cells through interactions with other pathways is as yet unclear. The nasopharyngeal carcinoma HONE1 cells have low expression of β-catenin and wild-type expression of p53, which provided a possibility to study regulatory mechanism of stemness networks induced by physiological levels of Wnt signaling in these cells. RESULTS: Introduction of increased β-catenin signaling, haploid expression of β-catenin under control by its natural regulators in transferred chromosome 3, resulted in activation of Wnt/β-catenin networks and dedifferentiation in HONE1 hybrid cell lines, but not in esophageal carcinoma SLMT1 hybrid cells that had high levels of endogenous β-catenin expression. HONE1 hybrid cells displayed stem cell-like properties, including enhancement of CD24(+) and CD44(+) populations and generation of spheres that were not observed in parental HONE1 cells. Signaling cascades were detected in HONE1 hybrid cells, including activation of p53- and RB1-mediated tumor suppressor pathways, up-regulation of Nanog-, Oct4-, Sox2-, and Klf4-mediated pluripotency networks, and altered E-cadherin expression in both in vitro and in vivo assays. qPCR array analyses further revealed interactions of physiological Wnt/β-catenin signaling with other pathways such as epithelial-mesenchymal transition, TGF-β, Activin, BMPR, FGFR2, and LIFR- and IL6ST-mediated cell self-renewal networks. Using β-catenin shRNA inhibitory assays, a dominant role for β-catenin in these cellular network activities was observed. The expression of cell surface markers such as CD9, CD24, CD44, CD90, and CD133 in generated spheres was progressively up-regulated compared to HONE1 hybrid cells. Thirty-four up-regulated components of the Wnt pathway were identified in these spheres. CONCLUSIONS: Wnt/β-catenin signaling regulates self-renewal networks and plays a central role in the control of pluripotency genes, tumor suppressive pathways and expression of cancer stem cell markers. This current study provides a novel platform to investigate the interaction of physiological Wnt/β-catenin signaling with stemness transition networks

Tumor suppressive role of chromosomes 11, 13, and 14 in esophageal squamous cell carcinoma studied by functional complementation

Despite the abundant evidence for high allelic loss of chromosome 14q in human cancers (Lee et al., 1997; Chang et al., 1995; Mutirangura et al., 1998; Hu et al., 1999; Hu et al., 2000; Dekken et al., 1999), tumor suppressor genes mapped to this chromosome have yet to be identified due to the complexity of the chromosomal alterations reported. To narrow down the search for candidate genes, we performed monochromosome transfer of chromosome 14 into an esophageal squamous cell carcinoma (ESCC) cell line, SLMT-1 S1. Statistically significant suppression of the tumorigenic potential of microcell hybrids (MCHs), containing the transferred chromosome 14, provides functional evidence that tumor suppressive regions on chromosome 14 are essential for ESCC. Tumor segregants (TSs) emerging in the nude mice during the tumorigenicity assay were analyzed by detailed PCR-microsatellite typing to identify non-randomly eliminated critical regions (CRs). A 680 kb CR mapped to 14q32.13 and a ~2.2 Mb CR mapped to 14q32.33 were delineated. Dual color bacterial artificial chromosome fluorescent in situ hybridization (BAC FISH) analysis of MCHs and TSs verified the selective loss of the 14q32.13 region. In contrast, similar transfers of an intact chromosome 11 into SLMT-1 S1 did not significantly suppress tumor formation. These functional complementation studies showing the correlation of tumorigenic potential with critical regions of chromosome 14 validate the importance of the 14q32 region in tumor suppression in ESCC. The present study also paves the path for further identification of novel tumor suppressor genes (TSGs), which are relevant in the molecular pathogenesis of ESCC. Chromosomal regions with a high rate of loss of heterozygosity (LOH) may implicate candidate TSG(s) that is/are involved in the molecular pathogenesis of ESCC. In ESCC, chromosome 13q LOH ranged from 48% - l00%, as independently observed from different groups (Boynton et al., 1991; Huang et al., 1992; Hu et al., 2000; Li et al., 2001; Li et al., 2003; Hu et al., 2003) and 13q genetic aberrations of 22% - 100% revealed by comparative genomic hybridization (CGH) (Pack et al., 1999; Tada et al., 2000; Wei et al., 2002). However, it is still unclear if the loss of genetic materials is a cause or consequence of ESCC, as this indirect evidence only implicates the presence of candidate TSG(s). Direct functional evidence for tumor suppression after transferring genetic materials is still scanty in ESCC. A tumor suppression effect was observed after the transfer of chromosome 13 into SLMT-1 S1. The tumor suppressive effect observed in MCH13-113 suggested TSGs may be located at 13q34. Three critical regions, CR1 and CR2, at 13q12.3, and CR3 at 13q14.11, were delineated during TS deletion analysis. TSGs important for ESCC may be located on 13q12.3 and 13q14.11. The first functional proof from microcell-mediated chromosome transfer (MMCT) that tumor suppressive regions on chromosomes 13 and 14 are essential for ESCC development is provided in the current study

Singapore Programme for Integrated Care for the Elderly (SPICE)—an integrated model of care to enable frail elderly to be cared for in the community

PURPOSE: Alternative models of care for the frail elderly are needed in Singapore in view of the rapidly ageing population, increased longevity and increase in the complexity of elderly care needs. The objective of the programme (SPICE) is to delay and avoid institutional care for the frail elderly by meeting their complex care needs in the community. THEORY: SPICE is modeled after the US-based Programme for All-Inclusive Care of the Elderly (PACE), which delivers comprehensive care for NH eligible individuals. Evidence has shown that the community-based, integrated and comprehensive programme decreases overall hospital re-admission, average length of stay in hospital and visits to the emergency department (ED), decreases caregiver stress and improves overall satisfaction with care arrangements [1, 2]. METHOD: In SPICE, an inter-disciplinary team (IDT) delivers an integrated suite of medical, nursing, rehabilitation, social and personal care services, both in the centre and at the participant’s home, dependent on the needs. A comprehensive assessment, regular re-evaluation and detailed individualised care plan (ICP) is tailored and implemented to enable these frail elderly to avoid institutional care. RESULTS: 37 participants have been recruited for the programme since October 2010. Preliminary 6-month pre-post analysis showed that caregiver stress decreased by 42% and participants perception of their own health increased by 18%. The total number of hospital admissions also decreased by 66.6%, average length of stay in the hospital decreased by 47.8% and the number of visits to the ED decreased by 50% over the 6 months. CONCLUSION: The results suggest that SPICE is effective in enabling the frail elderly to reduce and avoid institutional care and improving their overall satisfaction with care arrangements. Further investigations with matched controls are needed to study if the long-term results

FANCD2 Confers a Malignant Phenotype in Esophageal Squamous Cell Carcinoma by Regulating Cell Cycle Progression

Fanconi anemia patients with germline genetic defects in FANCD2 are highly susceptible to cancers. Esophageal squamous cell carcinoma (ESCC) is a deadly cancer. Little is known about the function of FANCD2 in ESCC. For detailed molecular and mechanistic insights on the functional role of FANCD2 in ESCC, in vivo and in vitro assays and RNA sequencing approaches were used. Utilizing Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) technology, FANCD2 knockout models were established to examine the functional impact in mouse models for tumor growth and metastasis and in vitro assays for cell growth, cell cycle, and cellular localization. Our RNA sequence analyses were integrated with public datasets. FANCD2 confers a malignant phenotype in ESCC. FANCD2 is significantly upregulated in ESCC tumors, as compared to normal counterparts. Depletion of FANCD2 protein expression significantly suppresses the cancer cell proliferation and tumor colony formation and metastasis potential, as well as cell cycle progression, by involving cyclin-CDK and ATR/ATM signaling. FANCD2 translocates from the nucleus to the cytoplasm during cell cycle progression. We provide evidence of a novel role of FANCD2 in ESCC tumor progression and its potential usefulness as a biomarker for ESCC disease management

Depletion of DNA Polymerase Theta Inhibits Tumor Growth and Promotes Genome Instability through the cGAS-STING-ISG Pathway in Esophageal Squamous Cell Carcinoma

Overexpression of the specialized DNA polymerase theta (POLQ) is frequent in breast, colon and lung cancers and has been correlated with unfavorable clinical outcomes. Here, we aimed to determine the importance and functional role of POLQ in esophageal squamous cell carcinoma (ESCC). Integrated analysis of four RNA-seq datasets showed POLQ was predominantly upregulated in ESCC tumors. High expression of POLQ was also observed in a cohort of 25 Hong Kong ESCC patients and negatively correlated with ESCC patient survival. POLQ knockout (KO) ESCC cells were sensitized to multiple genotoxic agents. Both rH2AX foci staining and the comet assay indicated a higher level of genomic instability in POLQ-depleted cells. Double KO of POLQ and FANCD2, known to promote POLQ recruitment at sites of damage, significantly impaired cell proliferation both in vitro and in vivo, as compared to either single POLQ or FANCD2 KOs. A significantly increased number of micronuclei was observed in POLQ and/or FANCD2 KO ESCC cells. Loss of POLQ and/or FANCD2 also resulted in the activation of cGAS and upregulation of interferon-stimulated genes (ISGs). Our results suggest that high abundance of POLQ in ESCC contributes to the malignant phenotype through genome instability and activation of the cGAS pathway