bims-tucedo Biomed News
on Tumor cell dormancy
Issue of 2021‒01‒31
thirty-two papers selected by
Isabel Puig Borreil
Vall d’Hebron Institute of Oncology

  1. Trends Cancer. 2021 Jan 25. pii: S2405-8033(20)30341-1. [Epub ahead of print]
      Despite gains in knowledge of the intrinsic signals governing cancer progression, effective clinical management of cancer remains a challenge. Drug resistance and relapse, pose the greatest barriers to cancer care, and are often driven by the co-option of stem cell programs by subpopulations of aggressive cancer cells. Here, we focus on the role of the microenvironment in the acquisition and/or maintenance of stem cell states in cancer in the context of resistance and metastasis. We further discuss the role of cancer stem cells in immune evasion through the course of metastasis, dormancy, and relapse. Understanding the niche in which cancer stem cells live and the signals that sustain them may lead to new strategies that target them by disrupting microenvironmental support.
    Keywords:  Cancer; Metastasis; Stem cell; Therapy resistance; Tumor microenvironment
  2. Cancer Cell. 2021 Jan 27. pii: S1535-6108(21)00053-2. [Epub ahead of print]
      Neutrophils can facilitate the metastatic spread of cancer; however, how neutrophils are activated at metastatic sites remains poorly understood. In this issue, Xiao et al. demonstrate that the protease cathepsin C, secreted by breast cancer cells, triggers neutrophils to form neutrophil extracellular traps in the metastatic niche, thereby promoting lung metastasis.
  3. Cancer Res. 2021 Jan 28. pii: canres.1810.2020. [Epub ahead of print]
      Epidermal growth factor receptor (EGFR) is frequently amplified, mutated, and overexpressed in malignant gliomas. Yet the EGFR-targeted therapies have thus far produced only marginal clinical responses, and the underlying mechanism remains poorly understood. Using an inducible oncogenic EGFR-driven glioma mouse model system, our current study reveals that a small population of glioma cells can evade therapy-initiated apoptosis and potentiate relapse development by adopting a mesenchymal-like phenotypic state that no longer depends on oncogenic EGFR signaling. Transcriptome analyses of proximal and distal treatment responses identified TGFβ/YAP/Slug signaling cascade activation as a major regulatory mechanism that promotes therapy-induced glioma mesenchymal lineage transdifferentiation. Following anti-EGFR treatment, TGFβ secreted from stressed glioma cells acted to promote YAP nuclear translocation which stimulated upregulation of the pro-mesenchymal transcriptional factor Slug and subsequent glioma lineage transdifferentiation towards a stable therapy-refractory state. Blockade of this adaptive response through suppression of TGFβ-mediated YAP activation significantly delayed anti-EGFR relapse and prolonged animal survival. Together, our findings shed new insight into EGFR-targeted therapy resistance and suggest that combinatorial therapies of targeting both EGFR and mechanisms underlying glioma lineage transdifferentiation could ultimately lead to deeper and more durable responses.
  4. Oncogene. 2020 Dec 15.
      Prostate cancer is one of the leading causes of mortality in men. The major cause of death in prostate cancer patients can be attributed to metastatic spread of disease or tumor recurrence after initial treatment. Prostate tumors are known to remain undetected or dormant for a long period of time before they progress locoregionally or at distant sites as overt tumors. However, the molecular mechanism of dormancy is yet poorly understood. In this study, we performed a differential gene expression analysis and identified a gene, Regucalcin (RGN), which promotes dormancy of prostate cancer. We found that cancer patients expressing higher level of RGN showed significantly longer recurrence-free and overall- survival. Using a doxycycline-inducible RGN expression system, we showed that ectopic expression of RGN in prostate tumor cells induced dormancy in vivo, while following suppression of RGN triggered recurrence of tumor growth. On the other hand, silencing RGN in LNCap cells promoted its outgrowth in the tibia of mice. Importantly, RGN promoted multiple known hallmarks of tumor dormancy including activation of p38 MAPK, decrease in Erk signaling and inhibition of FOXM1 expression. Furthermore, we found that RGN significantly suppressed angiogenesis by increasing secretory miR-23c level in the exosomes. Intriguingly, FOXM1 was found to negatively regulate miR-23c expression in prostate cancer. In addition, we identified 11 RGN downstream target genes that independently predicted longer recurrence-free survival in patients. We found that expression of these genes was regulated by FOXM1 and/or p38 MAPK. These findings suggest a critical role of RGN in prostate cancer dormancy, and the utility of RGN signaling and exosomal miR-23c as biomarkers for predicting recurrence.
  5. Cancer Discov. 2021 Jan 26. pii: candisc.1375.2020. [Epub ahead of print]
      Acute myeloid leukemia (AML) patients frequently relapse after chemotherapy, yet the mechanism by which AML reemerges is not fully understood. Herein, we show that primary AML cells enter a senescence-like phenotype following chemotherapy in vitro and in vivo. This is accompanied by induction of senescence/inflammatory and embryonic diapause transcriptional programs, with downregulation of MYC and leukemia stem cell genes. Single-cell RNA-seq suggested depletion of leukemia stem cells in vitro and in vivo, and enrichment for subpopulations with distinct senescence-like cells. This senescence effect was transient and conferred superior colony forming and engraftment potential. Entry into this senescence-like phenotype was dependent on ATR, and persistence of AML cells was severely impaired by ATR inhibitors. Altogether, we propose that AML relapse is facilitated by a senescence-like resilience phenotype that occurs regardless of their stem cell status. Upon recovery, these post-senescence AML cells give rise to relapsed AMLs with increased stem cell potential.
  6. Biomolecules. 2021 Jan 20. pii: E130. [Epub ahead of print]11(2):
      Progression from localized to metastatic disease requires cancer cells spreading to distant organs through the bloodstream. Only a small proportion of these circulating tumor cells (CTCs) survives dissemination due to anoikis, shear forces and elimination by the immune system. However, all metastases originate from CTCs capable of surviving and extravasating into distant tissue to re-initiate a tumor. Metastasis initiation is not always immediate as disseminated tumor cells (DTCs) may enter a non-dividing state of cell dormancy. Cancer dormancy is a reversible condition that can be maintained for many years without being clinically detectable. Subsequently, late disease relapses are thought to be due to cancer cells ultimately escaping from dormant state. Cancer dormancy is usually associated with minimal residual disease (MRD), where DTCs persist after intended curative therapy. Thus, MRD is commonly regarded as an indicator of poor prognosis in all cancers. In this review, we examine the current understanding of MRD and immunity during cancer progression to metastasis and discuss clinical perspectives for oncology.
    Keywords:  CTC; DTC; MRD; dormancy; immunity; metastasis; therapy
  7. Proc Natl Acad Sci U S A. 2021 Feb 02. pii: e2020478118. [Epub ahead of print]118(5):
      Beclin 1, an autophagy and haploinsufficient tumor-suppressor protein, is frequently monoallelically deleted in breast and ovarian cancers. However, the precise mechanisms by which Beclin 1 inhibits tumor growth remain largely unknown. To address this question, we performed a genome-wide CRISPR/Cas9 screen in MCF7 breast cancer cells to identify genes whose loss of function reverse Beclin 1-dependent inhibition of cellular proliferation. Small guide RNAs targeting CDH1 and CTNNA1, tumor-suppressor genes that encode cadherin/catenin complex members E-cadherin and alpha-catenin, respectively, were highly enriched in the screen. CRISPR/Cas9-mediated knockout of CDH1 or CTNNA1 reversed Beclin 1-dependent suppression of breast cancer cell proliferation and anchorage-independent growth. Moreover, deletion of CDH1 or CTNNA1 inhibited the tumor-suppressor effects of Beclin 1 in breast cancer xenografts. Enforced Beclin 1 expression in MCF7 cells and tumor xenografts increased cell surface localization of E-cadherin and decreased expression of mesenchymal markers and beta-catenin/Wnt target genes. Furthermore, CRISPR/Cas9-mediated knockout of BECN1 and the autophagy class III phosphatidylinositol kinase complex 2 (PI3KC3-C2) gene, UVRAG, but not PI3KC3-C1-specific ATG14 or other autophagy genes ATG13, ATG5, or ATG7, resulted in decreased E-cadherin plasma membrane and increased cytoplasmic E-cadherin localization. Taken together, these data reveal previously unrecognized cooperation between Beclin 1 and E-cadherin-mediated tumor suppression in breast cancer cells.
    Keywords:  Beclin 1; E-cadherin; breast cancer
  8. Cancer Res. 2021 Jan 26. pii: canres.CAN-20-2434-A.2020. [Epub ahead of print]
      In the bone marrow (BM), breast cancer (BC) cells (BCC) can survive in dormancy for decades as cancer stem cells (CSC), resurging as tertiary metastasis. The endosteal region where BCCs exist as CSCs poses a challenge to target them, mostly due to the co-existence of endogenous hematopoietic stem cells. This study addresses the early period of dormancy when BCCs enter BM at the perivascular region to begin the transition into CSCs, which we propose as the final step in dormancy. A two-step process comprises the Wnt-β-catenin pathway mediating BCC dedifferentiation into CSCs at the BM perivascular niche. At this site, BCCs responded to two types of mesenchymal stem cell (MSC)-released extracellular vesicles (EV) that may include exosomes. Early released EVs began the transition into cycling quiescence, DNA repair, and reorganization into distinct BCC subsets. After contact with BC, the content of EVs changed (primed) to complete dedifferentiation into a more homogeneous population with CSC properties. BCC progenitors (Oct4alo), which are distant from CSCs in a hierarchical stratification, were sensitive to MSC EVs. Despite of CSC function, Oct4alo BCCs expressed multipotent pathways similar to CSCs. Oct4alo BCCs dedifferentiated and co-localized with MSCs (murine and human BM) in vivo. Overall, these findings elucidate a mechanism of early dormancy at the BM perivascular region and provide evidence of epigenome reorganization as a potential new therapy for BC.
  9. Cancer Res. 2021 Jan 29. pii: canres.2743.2020. [Epub ahead of print]
      Inflammatory breast cancer (IBC) is a highly metastatic breast carcinoma with high frequency of estrogen receptor α (ER)α-negativity. Here we explored the role of the second ER subtype, ERβ, and report expression in IBC tumors and its correlation with reduced metastasis. Ablation of ERβ in IBC cells promoted cell migration and activated gene networks that control actin reorganization, including G-protein coupled receptors and downstream effectors that activate Rho GTPases. Analysis of preclinical mouse models of IBC revealed decreased metastasis of IBC tumors when ERβ was expressed or activated by chemical agonists. Our findings support a tumor-suppressive role of ERβ by demonstrating the ability of the receptor to inhibit dissemination of IBC cells and prevent metastasis. Based on these findings, we propose ERβ as a potentially novel biomarker and therapeutic target that can inhibit IBC metastasis and reduce its associated mortality.
  10. Cancer Cell. 2021 Jan 09. pii: S1535-6108(20)30671-1. [Epub ahead of print]
      Immunotherapy induces durable clinical responses in a fraction of patients with cancer. However, therapeutic resistance poses a major challenge to current immunotherapies. Here, we identify that expression of tumor stanniocalcin 1 (STC1) correlates with immunotherapy efficacy and is negatively associated with patient survival across diverse cancer types. Gain- and loss-of-function experiments demonstrate that tumor STC1 supports tumor progression and enables tumor resistance to checkpoint blockade in murine tumor models. Mechanistically, tumor STC1 interacts with calreticulin (CRT), an "eat-me" signal, and minimizes CRT membrane exposure, thereby abrogating membrane CRT-directed phagocytosis by antigen-presenting cells (APCs), including macrophages and dendritic cells. Consequently, this impairs APC capacity of antigen presentation and T cell activation. Thus, tumor STC1 inhibits APC phagocytosis and contributes to tumor immune evasion and immunotherapy resistance. We suggest that STC1 is a previously unappreciated phagocytosis checkpoint and targeting STC1 and its interaction with CRT may sensitize to cancer immunotherapy.
    Keywords:  PD-1; T cell immunity; calreticulin; checkpoint; dendritic cell; eat-me signal; macrophages; phagocytosis; stanniocalcin 1; tumor
  11. Theranostics. 2021 ;11(5): 2460-2474
      Rationale: Inflammatory stimuli from the tumor microenvironment play important roles in cancer progression. However, the mechanism of promotion of cancer metastasis by inflammation in gastric cancer (GC) is poorly understood. Methods: The roles of NEK9 were validated via loss-of-function and gain-of-function experiments in vitro and in an animal model of metastasis. Cytoskeletal reorganization-associated molecules were detected by GST pull-down. The regulation of ARHGEF2 by NEK9 was investigated by phosphoproteomics analysis, immunoprecipitation (IP) and in vitro kinase assay. The transcriptional regulation of miR-520f-3p was studied using luciferase reporter and chromatin immunoprecipitation (ChIP). The expression of these proteins in GC tissues was examined by immunohistochemistry. Results: NEK9 directly regulates cell motility and RhoA activation in GC. The phosphorylation of ARHGEF2 by NEK9 is the key step of this process. NEK9 is a direct target of miR-520f-3p, which is transcriptionally suppressed by IL-6-mediated activation of STAT3. A decrease in miR-520f-3p leads to the amplification of IL-6/STAT3 by targeting GP130. A simultaneous elevation of the levels of NEK9, GP130 and p-STAT3 was confirmed in the lymph nodes and distant metastases. An increase in NEK9, GP130 and STAT3 is associated with reduced overall survival of GC patients. Conclusion: This study demonstrates that activation of STAT3 by IL-6 transcriptionally suppresses miR-520f-3p and diminishes the inhibitory effects of miR-520f-3p on NEK9 and GP130. An increase in GP130 enhances this signaling, and NEK9 directly influences cell motility and RhoA activation by targeting the phosphorylation of ARHGEF2. Targeting the IL-6-STAT3-NEK9 pathway may be a new strategy for GC treatment.
    Keywords:  Gastric cancer; Inflammation; Metastasis; NEK9; Phosphorylation
  12. Theranostics. 2021 ;11(5): 2108-2122
      Rationale: Protein kinases are critical therapeutic targets for curing hepatocellular carcinoma (HCC). As a serine/threonine kinase, the potential roles of serine/threonine kinase 39 (STK39) in HCC remain to be explored. Methods: The expression of STK39 was examined by RT-qPCR, western blotting and immunohistochemistry. Cell proliferation and apoptosis were detected by CCK8 and TUNEL kit. Cell migration and invasion assays were performed using a transwell system with or without Matrigel. RNA-seq, mass spectrometry and luciferase reporter assays were used to identify STK39 binding proteins. Results: Here, we firstly report that STK39 was highly overexpressed in clinical HCC tissues compared with adjacent tissues, high expression of STK39 was induced by transcription factor SP1 and correlated with poor patient survival. Gain and loss of function assays revealed that overexpression of STK39 promoted HCC cell proliferation, migration and invasion. In contrast, the depletion of STK39 attenuated the growth and metastasis of HCC cells. Moreover, knockdown of STK39 induced the HCC cell cycle arrested in the G2/M phase and promoted apoptosis. In mechanistic studies, RNA-seq revealed that STK39 positively regulated the ERK signaling pathway. Mass spectrometry identified that STK39 bound to PLK1 and STK39 promoted HCC progression and activated ERK signaling pathway dependent on PLK1. Conclusions: Thus, our study uncovers a novel role of STK39/PLK1/ERK signaling axis in the progress of HCC and suggests STK39 as an indicator for prognosis and a potential drug target of HCC.
    Keywords:  ERK; HCC; PLK1; STK39; proliferation
  13. Cancer Res. 2021 Jan 28.
      Triple-negative breast cancers (TNBC) are resistant to standard-of-care chemotherapy and lack known targetable driver gene alterations. Identification of novel drivers could aid the discovery of new treatment strategies for this hard-to-treat patient population, yet studies using high-throughput and accurate models to define the functions of driver genes in TNBC to date have been limited. Here, we employed unbiased functional genomics screening of the 200 most frequently mutated genes in breast cancer, using spheroid cultures to model in vivo-like conditions, and identified the histone acetyltransferase CREBBP as a novel tumor suppressor in TNBC. CREBBP protein expression in patient tumor samples was absent in 8% of TNBCs and at a high frequency in other tumors, including squamous lung cancer, where CREBBP-inactivating mutations are common. In TNBC, CREBBP alterations were associated with higher genomic heterogeneity and poorer patient survival and resulted in upregulation and dependency on a FOXM1 proliferative program. Targeting FOXM1-driven proliferation indirectly with clinical CDK4/6 inhibitors (CDK4/6i) selectively impaired growth in spheroids, cell line xenografts, and patient-derived models from multiple tumor types with CREBBP mutations or loss of protein expression. In conclusion, we have identified CREBBP as a novel driver in aggressive TNBC and identified an associated genetic vulnerability in tumor cells with alterations in CREBBP and provide a preclinical rationale for assessing CREBBP alterations as a biomarker of CDK4/6i response in a new patient population. SIGNIFICANCE: This study demonstrates that CREBBP genomic alterations drive aggressive TNBC, lung cancer, and lymphomas and may be selectively treated with clinical CDK4/6 inhibitors.
  14. Mol Cancer Res. 2021 Jan 26. pii: molcanres.0532.2020. [Epub ahead of print]
      Several master transcription factors (TFs) can activate the epithelial-to-mesenchymal transition (EMT). However, their individual and combinatorial contributions to EMT in breast cancer are not defined. We show that overexpression of EMT-TFs individually in epithelial cells upregulated endogenous SNAI2, ZEB1/2, TCF4 and TWIST1/2 as a result of positive feedback mediated in part by suppression of their negative regulator microRNAs miR200s/203/205. We identified TCF4 as a potential new target of miR200s. Expression of ZEB1/2 strongly correlated with the mesenchymal phenotype in breast cancer cells, with the CD24-/CD44+ stemness profile, and with lower expression of core epithelial genes in human breast tumors. Knockdown of EMT-TFs identified the key role of ZEB1 and its functional cooperation with other EMT-TFs in the maintenance of the mesenchymal state. Inducible ZEB1+2 knockdown in xenograft models inhibited pulmonary metastasis, emphasizing their critical role in dissemination from primary site and in extravasation. However, ZEB1+2 depletion one week post intravenous injection did not inhibit lung colonization, suggesting that ZEB1/2 and EMT are not essential for macrometastatic outgrowth. These results provide strong evidence that EMT is orchestrated by coordinated expression of several EMT-TFs and establish ZEB1 as a key master regulator of EMT and metastasis in breast cancer. Implications: The EMT program is orchestrated by coordinated expression of multiple EMT transcription factors, while ZEB1 integrates the EMT master regulatory network and plays the major role in promoting EMT and metastasis.
  15. Cancer Cell. 2021 Jan 27. pii: S1535-6108(21)00054-4. [Epub ahead of print]
      Pathologists use histological features to classify tumors and assign site of origin for metastasis. How and why tumors organize the way they do and recreate their histological organization during metastasis is unknown. Here, I discuss the concept of "histostasis" conferring tumors a histological memory and hypothesize its implications for metastasis.
  16. Theranostics. 2021 ;11(5): 2297-2317
      Rationale: Considerable evidence suggests that breast cancer metastasis and recurrence occur due to emergence of cancer stem cells (CSCs). In our previous study, we designed a high-throughput siRNA screening platform that identifies inflammation genes involved in the regulation of cancer cell stemness. We reported that CCL16 protein decreases OCT4 expression and reduces the ALDH+ subpopulation. However, the mechanism by which CCL16 maintains stem cell-like properties remains unclear. Methods: Tissue microarrays were used to evaluate CCL16 expression. Cancer stemness assays were performed in CCL16 knockdown and overexpressing cells in vitro and in a xenograft model in vivo. Human phosphokinase array, immunofluorescence and chromatin immunoprecipitation assays were performed to explore the underlying mechanism. Results: We report that CCL16 was overexpressed in breast tumors and significantly correlated with clinical progression. We found that silencing CCL16 in MDA-MB-231 and BT549 cells diminished CSC properties including ALDH+ subpopulation, side population, chemo-resistance, and sphere formation. Furthermore, mice bearing CCL16-silenced MDA-MB-231 xenografts had lower tumorigenic frequency and developed smaller tumors. Exploration of the underlying mechanism found that CCL16 selects CCR2 to activate p-AKT/GSK3β signaling and facilitate β-catenin nuclear translocation. Further, CCL16 binds to the OCT4 promoter and promotes OCT4 expression. In addition, shRNAs targeting CCR2 and XAV939 targeting β-catenin abolished CCL16-mediated cancer stemness. Upstream, IL10 mediates STAT3 activation, which binds to the CCL16 promoter and enhances its expression. The STAT3-targeted inhibitor Stattic suppressed CCL16 expression in vitro and restrained tumor progression in vivo. Conclusions: We identified a potential CSC regulator and suggest a novel mechanism for how CCL16 governs cancer cell stemness. We propose that CCL16 could be an effective target for breast cancer therapy.
    Keywords:  CCL16; CCR2; CSCs; OCT4; breast cancer; β-catenin
  17. Cancer Res. 2021 Jan 25. pii: canres.2808.2020. [Epub ahead of print]
      Small cell lung cancer (SCLC) is a pulmonary neuroendocrine cancer with very poor prognosis and limited effective therapeutic options. Most patients are diagnosed at advanced stages, and the exact reason for the aggressive and metastatic phenotype of SCLC is completely unknown. Despite a high tumor mutational burden, responses to immune checkpoint blockade are minimal in SCLC patients. This may reflect defects in immune surveillance. Here we illustrate that evading NK surveillance contributes to SCLC aggressiveness and metastasis, primarily through loss of NK cell recognition of these tumors by reduction of NK-activating ligands (NKG2DL). SCLC primary tumors expressed very low level of NKG2DL mRNA and SCLC lines express little to no surface NKG2DL at the protein level. ChIP-Seq showed NKG2DL loci in SCLC are inaccessible compared to NSCLC, with few H3K27Ac signals. Restoring NKG2DL in preclinical models suppressed tumor growth and metastasis in an NK cell-dependent manner. Likewise, HDAC inhibitor treatment induced NKG2DL expression and led to tumor suppression by inducing infiltration and activation of NK and T cells. Among all the common tumor types, SCLC and neuroblastoma were the lowest NKG2DL-expressing tumors, highlighting a lineage dependency of this phenotype. In conclusion, these data show that epigenetic silencing of NKG2DL results in a lack of stimulatory signals to engage and activate NK cells, highlighting the underlying immune avoidance of SCLC and neuroblastoma.
  18. Proc Natl Acad Sci U S A. 2021 Feb 02. pii: e2017421118. [Epub ahead of print]118(5):
      Multiplex assays, involving the simultaneous use of multiple circulating tumor DNA (ctDNA) markers, can improve the performance of liquid biopsies so that they are highly predictive of cancer recurrence. We have developed a single-tube methylation-specific quantitative PCR assay (mqMSP) that uses 10 different methylation markers and is capable of quantitative analysis of plasma samples with as little as 0.05% tumor DNA. In a cohort of 179 plasma samples from colorectal cancer (CRC) patients, adenoma patients, and healthy controls, the sensitivity and specificity of the mqMSP assay were 84.9% and 83.3%, respectively. In a head-to-head comparative study, the mqMSP assay also performed better for detecting early-stage (stage I and II) and premalignant polyps than a published SEPT9 assay. In an independent longitudinal cohort of 182 plasma samples (preoperative, postoperative, and follow-up) from 82 CRC patients, the mqMSP assay detected ctDNA in 73 (89.0%) of the preoperative plasma samples. Postoperative detection of ctDNA (within 2 wk of surgery) identified 11 of the 20 recurrence patients and was associated with poorer recurrence-free survival (hazard ratio, 4.20; P = 0.0005). With subsequent longitudinal monitoring, 14 patients (70%) had detectable ctDNA before recurrence, with a median lead time of 8.0 mo earlier than seen with radiologic imaging. The mqMSP assay is cost-effective and easily implementable for routine clinical monitoring of CRC recurrence, which can lead to better patient management after surgery.
    Keywords:  DNA methylation; circulating tumor DNA; colorectal cancer; liquid biopsy; recurrence
  19. Clin Cancer Res. 2021 Jan 25. pii: clincanres.4391.2020. [Epub ahead of print]
      PURPOSE: Abiraterone acetate (AA), an inhibitor of CYP17A1, is an FDA-approved drug for advanced prostate cancer. However, not all patients respond to AA, and AA resistance ultimately develops in patients who initially respond. We aimed to identify AA resistance mechanisms in prostate cancer cells.EXPERIMENTAL DESIGN: We established several AA-resistant cell lines and performed a comprehensive study on mechanisms involved in AA resistance development. RNA-seq and phospho-kinase array screenings were performed to discover that the cAMP response element-CRE binding protein 1 (CREB1) was a critical molecule in AA resistance development.
    RESULTS: The drug-resistant cell lines are phenotypically stable without drug selections, and exhibit permanent global gene expression changes. The phosphorylated CREB1 (pCREB1) is increased in AA-resistant cell lines and is critical in controlling global gene expression. Upregulation of pCREB1 desensitized prostate cancer cells to AA, while blocking CREB1 phosphorylation re-sensitized AA-resistant cells to AA. AA treatment increases intracellular cAMP levels, induces kinases activity, and leads to the phosphorylation of CREB1, which may subsequently augment the essential role of the CBP/p300 complex in AA-resistant cells, since AA-resistant cells exhibit a relatively higher sensitivity to CBP/p300 inhibitors. Further pharmacokinetic studies demonstrated that AA significantly synergizes with CBP/p300 inhibitors in limiting the growth of prostate cancer cells.
    CONCLUSIONS: Our studies suggest that AA treatment upregulates pCREB1 which enhances CBP/p300 activity, leading to global gene expression alterations, subsequently resulting in drug resistance development. Combining AA with therapies targeting resistance mechanisms may provide a more effective treatment strategy.
  20. Mol Cancer. 2021 Jan 23. 20(1): 20
      Hepatocellular carcinoma (HCC) is one of the most common human malignancies worldwide with very poor prognosis. Resistance to targeted therapeutic drugs such as sorafenib remains one of the major challenges in clinical treatment. In the present study, PARP1 was found to be highly expressed in human embryonic stem cells, but progressively decreased upon specified hepatic differentiation. Reactivation of PARP1 expression was also detected in HCC residual tumors after sorafenib treatment in xenograft mouse model, indicating the potential important roles of PARP1 in stem cell pluripotency and HCC sorafenib treatment resistance. Overexpression of PARP1 was frequently observed in HCC patients, and closely associated with poor clinical outcome. Treatment of Sorafenib induced activation of DNA damage repair signaling, which is highly active and essential for maintenance of stem cell pluripotency in HCC residual tumors. PARP inhibitor Olaparib extensively suppressed the DNA damage repair signaling, and significantly inhibited the global pluripotent transcriptional network. The repression of key pluripotent transcriptional factors and DNA damage repair signaling by Olaparib was mainly through CHD1L-mediated condensation of the chromatin structure at their promotor regions. The global reshaping of the pluripotent transcriptome by Olaparib might reinforce Sorafenib in eliminating HCC residual tumors and enhance therapeutic efficiency.
    Keywords:  Embryonic stem cell; PARP inhibitors; Pluripotent transcriptome; Sorafenib resistance
  21. Nat Commun. 2021 01 25. 12(1): 574
      In contrast to primary colorectal cancer (CRC) little is known about the genomic landscape of metastasized CRC. Here we present whole genome sequencing data of metastases of 429 CRC patients participating in the pan-cancer CPCT-02 study (NCT01855477). Unsupervised clustering using mutational signature patterns highlights three major patient groups characterized by signatures known from primary CRC, signatures associated with received prior treatments, and metastasis-specific signatures. Compared to primary CRC, we identify additional putative (non-coding) driver genes and increased frequencies in driver gene mutations. In addition, we identify specific genes preferentially affected by microsatellite instability. CRC-specific 1kb-10Mb deletions, enriched for common fragile sites, and LINC00672 mutations are associated with response to treatment in general, whereas FBXW7 mutations predict poor response specifically to EGFR-targeted treatment. In conclusion, the genomic landscape of mCRC shows defined changes compared to primary CRC, is affected by prior treatments and contains features with potential clinical relevance.
  22. Nat Commun. 2021 01 27. 12(1): 608
      Haematopoietic stem cells (HSCs) are characterized by their self-renewal potential associated to dormancy. Here we identify the cell surface receptor neogenin-1 as specifically expressed in dormant HSCs. Loss of neogenin-1 initially leads to increased HSC expansion but subsequently to loss of self-renewal and premature exhaustion in vivo. Its ligand netrin-1 induces Egr1 expression and maintains quiescence and function of cultured HSCs in a Neo1 dependent manner. Produced by arteriolar endothelial and periarteriolar stromal cells, conditional netrin-1 deletion in the bone marrow niche reduces HSC numbers, quiescence and self-renewal, while overexpression increases quiescence in vivo. Ageing associated bone marrow remodelling leads to the decline of netrin-1 expression in niches and a compensatory but reversible upregulation of neogenin-1 on HSCs. Our study suggests that niche produced netrin-1 preserves HSC quiescence and self-renewal via neogenin-1 function. Decline of netrin-1 production during ageing leads to the gradual decrease of Neo1 mediated HSC self-renewal.
  23. Biochim Biophys Acta Rev Cancer. 2021 Jan 23. pii: S0304-419X(21)00013-5. [Epub ahead of print] 188514
      Cancer metastasis is the main reason for the high mortality in patients, contributing to 90% of cancer-related deaths. Biomarkers for early detection and therapeutic monitoring are essential to improve cancer outcomes. Circulating tumour cells (CTCs) arise from solid tumours and are capable of metastatic dissemination via the bloodstream or lymphatic system. Thus, CTCs can potentially be developed as a minimally invasive biomarker for early detection and therapeutic monitoring. Despite its clinical potential, research on CTCs remains limited, and this is likely due to their low numbers, short half-life, and the lack of robust methods for their isolation. There is also a need for molecular characterisation of CTCs to identify tumour-specific features, such as epigenetic signatures of metastasis. This review provides an overview of the epigenetic landscape of CTCs. We discuss the role of epigenetic modifications in CTC dissemination and metastatic tumour formation and progression.
    Keywords:  Cancer; Circulating tumour cell (CTC); Epigenetics; Metastasis; Tumour-specific epigenetic signature
  24. Cancer Res. 2021 Jan 29. pii: canres.2912.2020. [Epub ahead of print]
      Homologous recombination (HR)-deficient cancers are sensitive to inhibitors of Poly-ADP Ribose Polymerase (PARPi), which have shown clinical efficacy in the treatment of high-grade serous cancers (HGSC). However, the majority of patients will relapse, and acquired PARPi resistance is emerging as a pressing clinical problem. Here we generated seven single-cell clones with acquired PARPi resistance derived from a PARPi-sensitive, TP53-/- and BRCA1-/- epithelial cell line generated using CRISPR/Cas9. These clones showed diverse resistance mechanisms, and some clones presented with multiple mechanisms of resistance at the same time. Genomic analysis of the clones revealed unique transcriptional and mutational profiles and increased genomic instability in comparison to a PARPi-sensitive cell line. Clonal evolutionary analyses suggested that acquired PARPi resistance arose via clonal selection from an intrinsically unstable and heterogenous cell population in the sensitive cell line, which contained pre-existing drug tolerant cells. Similarly, clonal and spatial heterogeneity in tumor biopsies from a clinical BRCA1-mutant HGSC patient with acquired PARPi resistance were observed. In an imaging-based drug screening, the clones showed heterogenous responses to targeted therapeutic agents, indicating that not all PARPi-resistant clones can be targeted with just one therapy. Furthermore, PARPi-resistant clones showed mechanism-dependent vulnerabilities to the selected agents, demonstrating that a deeper understanding on the mechanisms of resistance could lead to improved targeting and biomarkers for HGSC with acquired PARPi resistance.
  25. Nat Biotechnol. 2021 Jan 28.
      Although genomic analyses predict many noncanonical open reading frames (ORFs) in the human genome, it is unclear whether they encode biologically active proteins. Here we experimentally interrogated 553 candidates selected from noncanonical ORF datasets. Of these, 57 induced viability defects when knocked out in human cancer cell lines. Following ectopic expression, 257 showed evidence of protein expression and 401 induced gene expression changes. Clustered regularly interspaced short palindromic repeat (CRISPR) tiling and start codon mutagenesis indicated that their biological effects required translation as opposed to RNA-mediated effects. We found that one of these ORFs, G029442-renamed glycine-rich extracellular protein-1 (GREP1)-encodes a secreted protein highly expressed in breast cancer, and its knockout in 263 cancer cell lines showed preferential essentiality in breast cancer-derived lines. The secretome of GREP1-expressing cells has an increased abundance of the oncogenic cytokine GDF15, and GDF15 supplementation mitigated the growth-inhibitory effect of GREP1 knockout. Our experiments suggest that noncanonical ORFs can express biologically active proteins that are potential therapeutic targets.
  26. Theranostics. 2021 ;11(5): 2442-2459
      Cancer growth is usually accompanied by metastasis which kills most cancer patients. Here we aim to study the effect of cisplatin at different doses on breast cancer growth and metastasis. Methods: We used cisplatin to treat breast cancer cells, then detected the migration of cells and the changes of epithelial-mesenchymal transition (EMT) markers by migration assay, Western blot, and immunofluorescent staining. Next, we analyzed the changes of RNA expression of genes by RNA-seq and confirmed the binding of activating transcription factor 3 (ATF3) to cytoskeleton related genes by ChIP-seq. Thereafter, we combined cisplatin and paclitaxel in a neoadjuvant setting to treat xenograft mouse models. Furthermore, we analyzed the association of disease prognosis with cytoskeletal genes and ATF3 by clinical data analysis. Results: When administered at a higher dose (6 mg/kg), cisplatin inhibits both cancer growth and metastasis, yet with strong side effects, whereas a lower dose (2 mg/kg) cisplatin blocks cancer metastasis without obvious killing effects. Cisplatin inhibits cancer metastasis through blocking early steps of EMT. It antagonizes transforming growth factor beta (TGFβ) signaling through suppressing transcription of many genes involved in cytoskeleton reorganization and filopodia formation which occur early in EMT and are responsible for cancer metastasis. Mechanistically, TGFβ and fibronectin-1 (FN1) constitute a positive reciprocal regulation loop that is critical for activating TGFβ/SMAD3 signaling, which is repressed by cisplatin induced expression of ATF3. Furthermore, neoadjuvant administration of cisplatin at 2 mg/kg in conjunction with paclitaxel inhibits cancer growth and blocks metastasis without causing obvious side effects by inhibiting colonization of cancer cells in the target organs. Conclusion: Thus, cisplatin prevents breast cancer metastasis through blocking early EMT, and the combination of cisplatin and paclitaxel represents a promising therapy for killing breast cancer and blocking tumor metastasis.
    Keywords:  TGFβ; cisplatin; metastasis; neoadjuvant therapy; paclitaxel
  27. Cancer Discov. 2021 Jan 28.
      A new study indicates that liver metastases reduce the effectiveness of immunotherapy by triggering apoptosis of tumor-specific T cells. In mice, radiation reversed the loss of T cells and restored the benefits of immunotherapy, providing a potential means of increasing the effectiveness of immunotherapy in patients.
  28. Theranostics. 2021 ;11(5): 2058-2076
      Histone deacetylases (HDACs) are involved in key cellular processes and have been implicated in cancer. As such, compounds that target HDACs or drugs that target epigenetic markers may be potential candidates for cancer therapy. This study was therefore aimed to identify a potential epidrug with low toxicity and high efficiency as anti-tumor agents. Methods: We first screened an epigenetic small molecule inhibitor library to screen for an epidrug for breast cancer. The candidate was identified as PCI-24781 and was characterized for half maximal inhibitory concentration (IC50), for specificity to breast cancer cells, and for effects on carcinogenesis and metastatic properties of breast cancer cell lines in vitro. A series of in silico and in vitro analyses were further performed of PCI-24781 to identify and understand its target. Results: Screening of an epigenetic inhibitor library in MDA-MB-231 cells, a malignant cancer cell line, showed that PCI-24781 is a potential anti-tumor drug specific to breast cancer. Ca2+ related pathways were identified as a potential target of PCI-24781. Further analyses showed that PCI-24781 inhibited Gαq-PLCβ3-mediated calcium signaling by activating the expression of regulator of G-protein signaling 2 (RGS2) to reduce cell proliferation, metastasis, and differentiation, resulting in cell death in breast cancer. In addition, RGS2 depletion reversed anti-tumor effect and inhibition of calcium influx induced by PCI-24781 treatment in breast cancer cells. Conclusions: We have demonstrated that PCI-24781 is an effective anti-tumor therapeutic agent that targets calcium signaling by activating RGS2. This study also provides a novel perspective into the use of HDAC inhibitors for cancer therapy.
    Keywords:  PCI-24781; RGS2; anti-tumor; calcium signaling; epidrug
  29. Nat Mater. 2021 Jan 25.
      The basement membrane (BM) is a special type of extracellular matrix and presents the major barrier cancer cells have to overcome multiple times to form metastases. Here we show that BM stiffness is a major determinant of metastases formation in several tissues and identify netrin-4 (Net4) as a key regulator of BM stiffness. Mechanistically, our biophysical and functional analyses in combination with mathematical simulations show that Net4 softens the mechanical properties of native BMs by opening laminin node complexes, decreasing cancer cell potential to transmigrate this barrier despite creating bigger pores. Our results therefore reveal that BM stiffness is dominant over pore size, and that the mechanical properties of 'normal' BMs determine metastases formation and patient survival independent of cancer-mediated alterations. Thus, identifying individual Net4 protein levels within native BMs in major metastatic organs may have the potential to define patient survival even before tumour formation. The ratio of Net4 to laminin molecules determines BM stiffness, such that the more Net4, the softer the BM, thereby decreasing cancer cell invasion activity.
  30. Nat Rev Mol Cell Biol. 2021 Jan 25.
      The research field of ferroptosis has seen exponential growth over the past few years, since the term was coined in 2012. This unique modality of cell death, driven by iron-dependent phospholipid peroxidation, is regulated by multiple cellular metabolic pathways, including redox homeostasis, iron handling, mitochondrial activity and metabolism of amino acids, lipids and sugars, in addition to various signalling pathways relevant to disease. Numerous organ injuries and degenerative pathologies are driven by ferroptosis. Intriguingly, therapy-resistant cancer cells, particularly those in the mesenchymal state and prone to metastasis, are exquisitely vulnerable to ferroptosis. As such, pharmacological modulation of ferroptosis, via both its induction and its inhibition, holds great potential for the treatment of drug-resistant cancers, ischaemic organ injuries and other degenerative diseases linked to extensive lipid peroxidation. In this Review, we provide a critical analysis of the current molecular mechanisms and regulatory networks of ferroptosis, the potential physiological functions of ferroptosis in tumour suppression and immune surveillance, and its pathological roles, together with a potential for therapeutic targeting. Importantly, as in all rapidly evolving research areas, challenges exist due to misconceptions and inappropriate experimental methods. This Review also aims to address these issues and to provide practical guidelines for enhancing reproducibility and reliability in studies of ferroptosis. Finally, we discuss important concepts and pressing questions that should be the focus of future ferroptosis research.