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Results for “Cancer Biology”

Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2018

Cell Cycle, Energy Metabolism and DNA Repair Pathways in Cancer Cells are Suppressed by Compound Kushen Injection.

In this report we examine candidate pathways perturbed by Compound Kushen Injection (CKI) a Traditional Chinese Medicine (TCM) that we have previously shown to alter the gene expression patterns of multiple pathways and induce apoptosis in cancer cells. We have measured protein levels in HEPG2 and MDA-MB-231 cells for genes in the cell cycle pathway, DNA repair pathway and DNA double strand breaks (DSBs) previously shown to have altered expression by CKI. We have also examined energy metabolism by measuring [ADT]/[ATP] ratio (cell energy charge), lactate production and glucose consumption. Our results demonstrate that CKI can suppress protein levels for cell cycle regulatory proteins and DNA repair while increasing the level of DSBs. We also show that energy metabolism is reduced based on reduced glucose consumption and reduced cellular energy charge. Our results validate these pathways as important targets for CKI. We also examined the effect of the major alkaloid component of CKI, oxymatrine and determined that it had no effect on DSBs, a small effect on the cell cycle and increased the cell energy charge. Our results indicate that CKI likely acts through the effect of multiple compounds on multiple targets where the observed phenotype is the integration of these effects and synergistic interactions.

cancer biology↗

bioRxiv2018

WDR5 regulates epithelial-to-mesenchymal transition in breast cancer cells via TGFB

Even if the mortality rate in breast cancer (BC) has recently decreased, development of metastases and drug resistance are still challenges to successful systemic treatment. The epithelial-to-mesenchymal transition (EMT), as well as epigenetic dynamic modifications, plays a pivotal role in invasion, metastasis, and drug resistance. Here, we report that WDR5, the core subunit of histone H3 K4 methyltransferase complexes, is crucial in coordinating EMT and regulating epigenetic changes that drive metastasis. We show that silencing of WDR5 in BC up-regulates an epithelial signature in triple negative and luminal B like patients by transcriptional repression of mesenchymal genes and reduction of the metastatic properties of these cells. Moreover, we demonstrate that this regulation is mediated by inhibition of the TGF{beta} signaling both at the transcriptional and post-translational level, suggesting an active role of WDR5 in guiding tumor plasticity upon oncogenic insults, regardless of the pathological BC subtypes.\n\nWe therefore suggest that WDR5 inhibition could be a successful pharmacologic approach to inhibit EMT and sensitize breast cancer cells to chemotherapy.

cancer biology↗

bioRxiv2018

Multiparametric Signature of Glioblastoma Differentiation Revealed by Imaging of Cellular Epigenetic Landscapes

The resistance of Glioblastoma (GBM) to conventional cytotoxic drugs has prompted novel therapeutic strategies, including differentiating tumor propagating cells (TPCs) into less tumorigenic cells using small molecule inducers of TPC differentiation. However, high-throughput screening for such molecules is hampered by the lack of robust markers of GBM differentiation. To obtain a signature of differentiated TPCs, we developed \"Microscopic Imaging of Epigenetic Landscapes\" (MIEL), which captures patterns of nuclear staining for epigenetic marks to derive feature-fingerprints of individual cells. We confirmed MIELs ability to accurately distinguish multiple cell fates and identified a multiparametric epigenetic signature of differentiated TPCs. Critically, we validated epigenetic imaging-based signature using global gene expression thus providing the proof of principle for the MIELs ability to select and prioritize small molecules, which induce TPC differentiation.

cancer biology↗

bioRxiv2018

Notch3 Promotes Prostate Cancer-Induced Bone Osteoblastic Metastasis in an MMP-3-Dependent Manner

Prostate cancer metastases primarily localize in the bone where they induce a unique osteoblastic response. Elevated Notch activity is associated with high-grade disease and metastasis. To address how Notch affects prostate cancer bone lesions, we manipulated Notch expression in mouse tibia xenografts and monitored tumor growth, lesion phenotype, and the bone microenvironment. Prostate cancer cell lines that induce mixed osteoblastic lesions in bone expressed 5-6 times more Notch3, than tumor cells that produce osteolytic lesions. Expression of active Notch3 (NICD3) in osteolytic tumors reduced osteolytic lesion area and enhanced osteoblastogenesis, while loss of Notch3 in osteoblastic tumors enhanced osteolytic lesion area and decreased osteoblastogensis. This was accompanied by a respective decrease and increase in the number of active osteoclasts and osteoblasts at the tumor-bone interface, without any effect on tumor proliferation. Conditioned medium from NICD3-expressing cells enhanced osteoblast differentiation and proliferation in vitro, while simultaneously inhibiting osteoclastogenesis. MMP-3 was specifically elevated and secreted by NICD3-expressing tumors, and inhibition of MMP-3 rescued the NICD3-induced osteoblastic phenotypes. Clinical osteoblastic bone metastasis samples had higher levels of Notch3 and MMP-3 compared to patient matched visceral metastases or osteolytic metastasis samples. We identified a Notch3-MMP-3 axis in human prostate cancer bone metastases that contributes to osteoblastic lesion formation by blocking osteoclast differentiation, while also contributing to osteoblastogenesis. These studies define a new role for Notch3 in manipulating the tumor microenvironment in bone metastases.

cancer biology↗

bioRxiv2018

Androgen Receptor-Induced Integrin α6β1 and Adhesion to Laminin Promotes Survival andDrug Resistance in Castration-Resistant Prostate Cancer through BNIP3

Although castration-resistant prostate cancers no longer respond to anti-androgen therapies, the androgen receptor (AR) is still required to promote tumor survival. However, the signaling pathways downstream of AR that promote this survival are not well known. We recently identified an AR-dependent survival pathway whereby AR induction of integrin 6{beta}1 and adhesion to laminin activates NF-kB/RelA signaling and Bcl-xL. This pathway acts in parallel with the PI3K/Akt pathway in Pten-null tumor cells such that combined inhibition of both PI3K and integrin 6{beta}1 is required to kill tumor cells adherent to laminin. However, PTEN-null castration-resistant tumors were not effectively inhibited by this combination. We discovered that BNIP3, a hypoxia-induced BH3-only, pro-mitophagic Bcl2 family member, is induced by androgen in castration-resistant cells through integrin 6{beta}1 signaling to HIF1. Furthermore, castration-resistant cells adherent to laminin were much more efficient at inducing autophagy in response to androgen. Androgen blocked the ability of the PI3K inhibitor PX-866 to kill castration-resistant tumors, but this was reversed by loss of BNIP3. Although BNIP3 was dispensable for androgen-induced autophagy, its mitophagy function was required for BNIP3 to promote resistance to PI3K inhibition. Thus, adhesion to laminin triggers signaling through AR/6{beta}1/HIF1 in castration-resistant cells to drive the expression of BNIP3 and cooperates with AR/6{beta}1-mediated autophagy, both of which contribute to PI3K resistance through induction of mitophagy.

cancer biology↗

bioRxiv2018

The immune checkpoint kick start: Optimization of neoadjuvant combination therapy using game theory

An upcoming clinical trial at the Moffitt Cancer Center for women with stage 2/3 ER+breast cancer combines an aromatase inhibitor and a PD-L1 checkpoint inhibitor, and aims to lower a preoperative endocrine prognostic index (PEPI) that correlates with relapse-free survival. PEPI is fundamentally a static index, measured at the end of neoadjuvant therapy before surgery. We develop a mathematical model of the essential components of the PEPI score in order to identify successful combination therapy regimens that minimize both tumor burden and metastatic potential, based on time-dependent trade-offs in the system. We consider two molecular traits, CCR7 and PD-L1 which correlate with treatment response and increased metastatic risk. We use a matrix game model with the four phenotypic strategies to examine the frequency-dependent interactions of cancer cells. This game was embedded into an ecological model of tumor population growth dynamics. The resulting model predicts both evolutionary and ecological dynamics that track with changes in the PEPI score. We consider various treatment regimens based on combinations of the two therapies with drug holidays. By considering the trade off between tumor burden and metastatic potential, the optimal therapy plan was found to be a 1 month kick start of the immune checkpoint inhibitor followed by five months of continuous combination therapy. Relative to a protocol with both therapeutics given together from the start, this delayed regimen results in transient sub-optimal tumor regression while maintaining a phenotypic constitution that is more amenable to fast tumor regression for the final five months of therapy. The mathematical model provides a useful abstraction of clinical intuition, enabling hypothesis generation and testing of clinical assumptions.

cancer biology↗

bioRxiv2018

High throughput proteomics identifies 484 high-accuracy plasma protein biomarker signatures for ovarian cancer

Ovarian cancer is usually detected at a late stage with the 5-year survival at only 30-40%. Additional means for early detection and improved diagnosis are acutely needed. To search for novel biomarkers, we compared circulating plasma levels of 981 proteins in patients with ovarian cancer and benign tumours, using the proximity extension assay. A novel combinatorial strategy was developed for identification of multivariate biomarker signatures, resulting in 484 mutually exclusive models out of which 448 did not contain the present biomarker MUCIN-16. The top-ranking model consisted of 14 proteins and had a AUC=0.95, PPV=1.0, sensitivity=0.99 and specificity=1.0 for detection of stage III-IV ovarian cancer in the discovery data, and an AUC=0.89, PPV=0.93, sensitivity=0.89 and specificity=0.95 in the replication data. The novel plasma protein signature could be used to improve the diagnosis of women with adnexal ovarian mass or in screening to identify women that should be referred to specialized examination.

cancer biology↗

bioRxiv2018

Childhood cerebellar tumors mirror conserved fetal transcriptional programs

The study of the origin and development of cerebellar tumours has been hampered by the complexity and heterogeneity of cerebellar cells that change over the course of development. We used single-cell transcriptomics to study >60,000 cells from the developing murine cerebellum, and show that different molecular subgroups of childhood cerebellar tumors mirror the transcription of cells from distinct, temporally restricted cerebellar lineages. Sonic Hedgehog medulloblastoma transcriptionally mirrors the granule cell hierarchy as expected, whereas Group 3 medulloblastoma resemble Nestin+ve stem cells, Group 4 medulloblastomas resemble unipolar brush cells, and PFA/PFB ependymoma and cerebellar pilocytic astrocytoma resemble the prenatal gliogenic progenitor cells. Furthermore, single-cell transcriptomics of human childhood cerebellar tumors demonstrates that many bulk tumors contain a mixed population of cells with divergent differentiation. Our data highlight cerebellar tumors as a disorder of early brain development, and provide a proximate explanation for the peak incidence of cerebellar tumors in early childhood.

cancer biology↗

bioRxiv2018

Targeting integrin alpha5 receptor in pancreatic stellate cells to diminish tumor-promoting effects in pancreatic cancer

Pancreatic stellate cells (PSCs) are the main precursors of cancer-associated fibroblasts (CAFs) in pancreatic ductal adenocarcinoma (PDAC), known to induce cancer aggressiveness. Integrin alpha5 (ITGA5), a fibronectin receptor, was found to be overexpressed by CAFs in stroma and linked to poor overall survival (log-rank p=0.022, n=137) of patients with PDAC. In vitro, knockdown of ITGA5 in human PSCs (hPSCs) inhibited their adhesion, migration, and proliferation and also inhibited TGF-{beta}-mediated differentiation. In vivo, co-injection of PANC-1 tumor cells and hPSCs (sh-ITGA5) developed tumors with reduced fibrosis and slower growth rate compared to those composed of PANC-1 and hPSC (sh-Ctrl). Furthermore, we developed a ITGA5-antagonizing peptidomimetic (AV3) which inhibited TGF{beta}-mediated hPSC differentiation by blocking ITGA5/FAK pathway. In vivo, treatment with AV3 intraperitoneally attenuated tumor fibrosis and thereby enhanced the efficacy of gemcitabine in patient-derived xenografts in mice. Altogether, this study reports the therapeutic importance of ITGA5 in PDAC and provides novel therapeutic peptidomimetic to enhance the effect of chemotherapy.

cancer biology↗

bioRxiv2018

Comparison of the effects of metformin on MDA-MB-231 breast cancer cells in a monolayer culture and in tumour spheroids as a function of nutrient concentrations

Metabolic pathways of cancer cells depend on the concentrations of nutrients in their micro-environment. However, they can vary also between monolayer cultures of cancer cells and tumour spheroids. Here we examined whether the absence of glucose, pyruvate and glutamine increases the sensitivity of MDA-MB-231 cells to metabolic drug metformin using two in vitro cell models (monolayer culture and tumour spheroids). To evaluate the effects of nutrient depletion in more detail, we tested the effects of metformin in commonly used media (DMEM, MEM and RPMI-1640) that differ mainly in the concentrations of amino acids. We used MTS, Hoechst and propidium iodide assay to determine cell number, viability and survival, respectively. We evaluated the effects of metformin on the size of tumour spheroids and determined cell survival by calcein and propidium iodide staining. Finally, we observed the effects of metformin in nutrient depleted conditions on the phosphorylation of AMP-activated protein kinase using Western blotting. Our main finding is that the effects of metformin on MDA-MB-231 cells depend on in vitro cell model used (monolayer culture vs. tumour spheroids). While metformin did not have any major effect on proliferation of MDA-MB-231 cells grown in complete cell culture media in a monolayer culture, it disintegrated tumour spheroids in MEM and RPMI-1640 medium. The effects of metformin on tumour spheroids were most pronounced in MEM, which is deficient of several non-essential amino acids. Glutamine depletion had no effect on the sensitivity of MDA-MB-231 cells to metformin in all tested conditions, whereas pyruvate depletion sensitized MDA-MB-231 cells to metformin in a monolayer culture only in MEM. Taken together, our results show that media formulation as well as in vitro cell model (monolayer culture vs. tumour spheroids) must be considered, when we evaluate the effects of metformin on MDA-MB-231 cells as a function of nutrient availability.

cancer biology↗

bioRxiv2018

Regulatory network controlling tumor-promoting inflammation in human cancers

Using an inducible, inflammatory model of breast cellular transformation, we describe the transcriptional regulatory network mediated by STAT3, NF-{kappa}B, and AP-1 factors on a genomic scale. These regulators form transcriptional complexes that directly regulate the expression of hundreds of genes in oncogenic pathways via a positive feedback loop. This inflammatory feedback loop, which functions to various extents in many types of cancer cells and patient tumors, is the basis for an \"inflammation\" index that defines cancer types by functional criteria. We identify a network of non-inflammatory genes whose expression is well correlated with the cancer inflammatory index. Conversely, the inflammation index is negatively correlated with expression of genes involved in DNA metabolism, and transformation is associated with genome instability. Inflammatory tumors are preferentially associated with infiltrating immune cells that might be recruited to the site of the tumor via inflammatory molecules produced by the cancer cells.

cancer biology↗

bioRxiv2018

Biased signaling downstream of epidermal growth factor receptor regulates proliferative versus apoptotic response to ligand

Inhibition of EGFR signaling by small molecule kinase inhibitors and monocloncal antibodies has proven effective in the treatment of multiple cancers. In contrast, metastatic breast cancers (BC) derived from EGFR-expressing mammary tumors are inherently resistant to EGFR-targeted therapies. Mechanisms that contribute to this inherent resistance remain poorly defined. Here we show that in contrast to primary tumors, ligand-mediated activation of EGFR in metastatic BC is dominated by STAT1 signaling. This change in downstream signaling leads to apoptosis and growth inhibition in response to EGF in metastatic BC cells. Mechanistically, these changes in downstream signaling result from an increase in the internalized pool of EGFR in metastatic cells, increasing physical access to the nuclear pool of STAT1. Along these lines, an EGFR mutant that is defective in endocytosis is unable to elicit STAT1 phosphorylation and apoptosis. Additionally, inhibition of endosomal signaling using an EGFR inhibitor linked to a nuclear localization signal specifically prevents EGF-induced STAT1 phosphorylation and cell death, without affecting EGFR:ERK1/2 signaling. Pharmacologic blockade of ERK1/2 signaling through the use of the allosteric MEK1/2 inhibitor, trametinib, dramatically biases downstream EGFR signaling toward a STAT1 dominated event, resulting in enhanced EGF-induced apoptosis in metastatic BC cells. Importantly, combined administration of trametinib and EGF also facilitated an apoptotic switch in EGFR-transformed primary tumor cells, but not normal mammary epithelial cells. These studies reveal a fundamental distinction for EGFR function in metastatic BC. Furthermore, the data demonstrate that pharmacological biasing of EGFR signaling toward STAT1 activation is capable of revealing the apoptotic function of this critical pathway.

cancer biology↗

bioRxiv2018

Cancer stemness, intratumoral heterogeneity, and immune response across cancers

Regulatory programs that control the function of stem cells are active in cancer and confer properties that promote progression and therapy resistance. However, the impact of a stem cell-like tumor phenotype (\"sternness\") on the immunological properties of cancer has not been systematically explored. Using gene expression-based metrics, we evaluate the association of stemness with immune cell infiltration and genomic, transcriptomic, and clinical parameters across 21 solid cancers. We find pervasive negative associations between cancer stemness and anticancer immunity. This occurs despite high stemness cancers exhibiting increased mutation load, cancer-testis antigen expression, and intratumoral heterogeneity. Stemness was also strongly associated with cell-intrinsic suppression of endogenous retroviral expression and type I interferon signaling and increased expression of several therapeutically accessible signaling pathways. Thus, stemness is not only a fundamental process in cancer progression but may represent a unifying mechanism linking antigenicity, intratumoral heterogeneity, and immune suppression across cancers.

cancer biology↗

bioRxiv2018

Pan-cancer systematic identification of lncRNAs associated with cancer prognosis

The \"dark matter\" of the genome harbors several non-coding RNA species including IncRNAs, which have been implicated in neoplasias but remain understudied. RNA-seq has provided deep insights into the nature of lncRNAs in cancer but current RNA-seq data are rarely accompanied by longitudinal patient survival information. In contrast, a plethora of microarray studies have collected these clinical metadata that can be leveraged to identify novel associations between gene expression and clinical phenotypes. In this study, we developed an analysis framework that computationally integrates RNA-seq and microarray data to systematically screen 9,463 lncRNAs for association with mortality risk across 20 cancer types. In total, we identified a comprehensive list of associations between lncRNAs and patient survival and demonstrate that these prognostic lncRNAs are under selective pressure and may be functional. Our results provide valuable insights that facilitate further exploration of lncRNAs and their potential as cancer biomarkers and drug targets.

cancer biology↗

bioRxiv2018

Genome-wide CNV study and functional evaluation identified CTDSPL as tumour suppressor gene for cervical cancer

We have investigated copy number variations (CNVs) in relation to cervical cancer by analyzing 731,422 single-nucleotide polymorphisms (SNPs) in 1,034 cervical cancer cases and 3,948 controls, followed by replication in 1,396 cases and 1,057 controls. We found that a 6367bp deletion in intron 1 of the CTD small phosphatase like gene (CTDSPL) was associated with 2.54-fold increased risk of cervical cancer (odds ratio =2.54, 95% confidence interval =2.08-3.12, P=2.0x10-19). This CNV is one of the strongest genetic risk variants identified so far for cervical cancer. The deletion removes the binding sites of zinc finger protein 263, binding protein 2 and interferon regulatory factor 1, and hence downregulates the transcription of CTDSPL. HeLa cells expressing CTDSPL showed a significant decrease in colony-forming ability. Compared with control groups, mice injected with HeLa cells expressing CTDSPL exhibited a significant reduction in tumour volume. Furthermore, CTDSPL-depleted immortalized End1/E6E7 could form tumours in NOD-SCID mice.

cancer biology↗

bioRxiv2018

Subjugation of TGFβ Signaling by Human Papilloma Virus in Head and Neck Squamous Cell Carcinoma Shifts DNA Repair from Homologous Recombination to Alternative End-Joining

Purpose: Following cytotoxic therapy, 70% of patients with human papillomavirus (HPV) positive oropharyngeal head and neck squamous cell carcinoma (HNSCC) are alive at 5 years compared to 30% of those with similar HPV-negative cancer, which is thought to be due to dysregulation of DNA repair. Loss of transforming growth factor {beta} (TGF{beta}) signaling is a poorly studied consequence of HPV that could contribute to this phenotype.\n\nExperimental Design: Human HNSCC cell lines (n=9), patient-derived xenografts (n=9), tissue microarray (n=194), TCGA expression data and primary tumor specimens (n=10) were used to define the relationship between TGF{beta} competency, response to DNA damage, and type of DNA repair.\n\nResults: Analysis of HNSCC specimens in situ and in vitro showed that HPV associates with loss of TGF{beta} signaling that increases the response to radiation or cisplatin. TGF{beta} suppressed miR-182 that inhibited both BRCA1, necessary for homologous recombination repair, and FOXO3, which is required for ATM kinase activity. TGF{beta} signaling blockade by either HPV or inhibitors released this control, compromised HRR and increased response to PARP inhibition. Antagonizing miR-182 rescued the homologous recombination deficit in HPV+ cells. Loss of TGF{beta} signaling unexpectedly increased error-prone, alternative end-joining repair.\n\nConclusions: HPV-positive HNSCC cells are unresponsive to TGF{beta}. Abrogated TGF{beta} signaling compromises homologous recombination and shifts reliance on alt-EJ repair that provides a mechanistic basis for sensitivity to PARP inhibitors. The effect of HPV in HNSCC provides critical validation of TGF{beta}s role in DNA repair proficiency and further raises the translational potential of TGF{beta} inhibitors in cancer therapy.

cancer biology↗

bioRxiv2018

Mutational likeliness and entropy help to identify driver mutations and their functional role in cancer

Alterations in cancer genomes originate from mutational processes taking place throughout oncogenesis and cancer progression. We show that likeliness and entropy are two properties of somatic mutations crucial in cancer evolution, as cancer-driver mutations stand out, with respect to both of these properties, as being distinct from the bulk of passenger mutations. Our analysis can identify novel cancer driver genes and differentiate between gain and loss of function mutations.

cancer biology↗

bioRxiv2018

Cancer-associated mRNAs regulated by the Helix-Loop-Helix motif of human EIF3A

Improper regulation of translation initiation, a vital checkpoint of protein synthesis in the cell, has been linked to a number of cancers. Overexpression of protein subunits of eukaryotic translation initiation factor 3 (eIF3) is associated with increased translation of mRNAs involved in cell proliferation. In addition to playing a major role in general translation initiation by serving as a scaffold for the assembly of translation initiation complexes, eIF3 regulates translation of specific cellular mRNAs and viral RNAs. Mutations in the N-terminal Helix-Loop-Helix (HLH) RNA-binding motif of the EIF3A subunit interfere with Hepatitis C Virus Internal Ribosome Entry Site (IRES) mediated translation initiation in vitro. Here we show that the EIF3A HLH motif controls translation of a small set of cellular transcripts enriched in oncogenic mRNAs, including MYC. We demonstrate that the HLH motif of EIF3A acts specifically on the 5-UTR of MYC mRNA and modulates the function of EIF4A1 on select transcripts during translation initiation. In Ramos lymphoma cell lines, which are dependent on MYC overexpression, mutations in the HLH motif greatly reduce MYC expression, impede proliferation and sensitize cells to anti-cancer compounds. These results reveal the potential of the EIF3A HLH motif in eIF3 as a promising chemotherapeutic target.

cancer biology↗