Abstract Background: Cervical adenocarcinoma exhibits poor responsiveness to radiotherapy and inferior survival compared with squamous cell carcinoma. The molecular mechanisms underlying its intrinsic radiation resistance remain largely unknown. Methods: Integrative analysis of TCGA and our RNA-seq cohort identified CYP4A22-AS1 as one of the most upregulated lncRNAs in cervical adenocarcinoma. qRT-PCR confirmed higher CYP4A22-AS1 expression in tumors with short disease-free survival (DFS). RNA pulldown, mass spectrometry, and RIP assays defined YBX1 as a direct CYP4A22-AS1-binding partner. Immunofluorescence and ChIP-qPCR assays examined YBX1 nuclear localization and promoter occupancy of PGK1, respectively. Functional effects of CYP4A22-AS1, YBX1, and PGK1 silencing were assessed by CCK-8, colony, EdU, and TUNEL assays in HeLa and C33A cells, and validated in xenograft models. Results: CYP4A22-AS1 was markedly overexpressed in cervical adenocarcinoma relative to normal cervix. Clinically, high CYP4A22-AS1 and PGK1 levels correlated with shorter DFS. Mechanistically, CYP4A22-AS1 binds YBX1 and enhances its nuclear translocation, thereby promoting YBX1 recruitment to the PGK1 promoter and activating glycolytic metabolism. Knockdown of CYP4A22-AS1 or PGK1 suppressed proliferation and markedly increased radiosensitivity both in vitro and in vivo. Conclusions: This study identifies CYP4A22-AS1 as a novel oncogenic lncRNA that drives radiation resistance through YBX1-mediated PGK1 transactivation. Clinically, CYP4A22-AS1 overexpression predicts poor outcome, while its inhibition restores radiation sensitivity. Targeting the CYP4A22-AS1-YBX1-PGK1 signaling axis offers a promising therapeutic strategy to overcome radioresistance in cervical adenocarcinoma. Citation Format: Mingyi Zhou, Chunlai Li, Cristina Ivan, Simone Anfossi, Linda Fabris, Melanie Winkle, Recep Bayraktar, Meng Chen, Lan Pang, Masayoshi Shimizu, Francois Claret, George Calin, . CYP4A22-AS1-YBX1 axis drives radiation resistance in cervical adenocarcinoma via PGK1 transactivation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1366.

Bromodomain and Extra-Terminal (BET) motif inhibitor therapy, alone or in combination with JAK inhibition (JAKi), is emerging as a promising and potentially disease modifying therapy for the treatment of myeloproliferative neoplasms (MPNs), specifically myelofibrosis (MF). While these agents promote spleen volume and bone marrow (BM) fibrosis reduction, their impact on symptom burden is less pronounced, perhaps due to ineffective pro-inflammatory cytokine modulation. Alternative epigenetic modifying therapies that further quell NFkB-mediated inflammation might enhance clinical outcomes in MF. EP300 is a histone acetyltransferase increasingly implicated in MF and other myeloid diseases, and recent studies suggest p300 inhibition, like BRD4 inhibition, can suppress NFkB-mediated inflammation and fibrosis progression in vivo. The expression of many genes is synergistically controlled by BRD4 and p300, leading to the development of EP31670 (aka NEO2734), a dual BRD4 (BET) + p300 inhibitor. EP31670 has shown pre-clinical activity across multiple leukemia cell lines and in myeloid leukemia xenograft models, and a phase-I clinical trial of EP31670 is on-going for treatment-refractory hematologic malignancies, including MF (NCT05488548). Given encouraging pre-clinical activity of combined BRD4/p300 inhibition and the emerging role of both BRD4 and p300 in fibrosis, we sought to investigate efficacy of EP31670 in MF animal models. The hMPLW515L adoptive transfer model was chosen for analysis. Lethally-irradiated BalbC mice were transplanted with lineage-negative BM cells retrovirally transfected with hMPLW515L-IRES-GFP plasmid and monitored for the development of MPN. Approximately 3 weeks post-transplant, mice were bled and cohorted into respective vehicle (VEH), EP31670 (3.5mg/kg BID), ruxolitinib (RUX; 60mg/kg BID), or combination (Combo) therapy arms. At the conclusion of the 3-week trial, mice were sacrificed and BM/spleen harvested for flow cytometric and histopathologic analysis. Serum cytokine profiling was performed using the Millipore 32-plex cytokine array kit on a Luminex platform. hMPLW515L mice treated with EP31670 demonstrated significant reductions in total white blood cell (K/uL: VEH 219, RUX 77, EP31670 36, Combo 15, p<0.005) and platelet counts (K/uL: VEH 2097, RUX 3029, EP31670 1286, Combo 799, p<0.05) compared to VEH and RUX treated arms, with distinct combinatorial activity observed in mice treated with EP31670/RUX together. Spleen volume reductions were observed across all treatment arms but reached statistical significance with combination therapy primarily (mg: VEH 472, RUX 377, EP31670 372, Combo 175, p<0.05). By flow analysis, significant reductions in total peripheral blood mutant cell fraction by GFP% were observed with EP31670, both alone and in combination with RUX (GFP: VEH 85%, RUX 84%, EP31670 71%, Combo 65%, P<0.005)—an effect not observed with RUX alone—consistent with selective loss of mutant cells in this compartment. Total BM Mac1+Gr1+ mature myeloid cell fractions were also significantly reduced with RUX/EP31670 combination (VEH 75%, RUX 76%, EP31670 68%, Combo 56%, p<0.05); however, this reduction did not extend to more primitive myeloid progenitor fractions. Improvements in BM reticulin fibrosis, along with reductions in total megakaryocyte number, were also observed with EP31670—an effect again further enhanced with combined RUX/EP31670. Finally, serum cytokine analysis revealed significant reductions in multiple inflammatory cytokines of EPI31670 +/- RUX treated mice not observed with RUX alone, including NFkB-mediated cytokines LIF, IL13, IP-10, MIG, and RANTES suggesting enhanced anti-inflammatory effects with EP31670, particularly in combination with JAKi. In line with early results from ongoing human clinical trials, a low dose of EP31670 demonstrates significant pre-clinical efficacy in MF models—with reduction of pro-inflammatory cytokines, spleen volume, hematologic parameters, and BM fibrosis—both alone and in combination with JAKi. Notably, the efficacious dose of EP31670 in these mouse studies, in combination with JAKi, was lower than what is usually required for EP31670 efficacy when applied as monotherapy in other mouse tumor models. Dual BET+p300 inhibition suppresses pro-inflammatory cytokine production in vivo and represents a potential alternative to other emerging epigenetic modifying therapies for MF treatment.

Abstract High phylogenetic conservation of genomic loci is a potential marker of essential biological functionality. Ultraconserved elements (UCEs) are DNA sequences entirely conserved through millions of years of evolution, but their functions are not well understood. Several long non-coding RNAs (lncRNAs) are transcribed from UCEs and play active roles in cancer development. Using a combination of in vitro and in vivo experiments with two animal models and 4 patient cohorts, we discovered several remarkable aspects of ultraconservation. Using a multi-step genetically defined carcinogenesis model, we identified one UCE, uc.206, and its associated overexpressed lncRNA TRUC-16 as a regulator of cell proliferation in chronic lymphocytic leukemia (CLL). High levels of TRUC-16 inhibit p16INK4A translation, causing increased proliferation through an E2F1-mediated mechanism in CLL patients, as well as in zebrafish and murine models. uc.206/TRUC-16 function is extremely conserved throughout evolution. We found cell cycle-related effects of TRUC-16 in both mice and zebrafish, in which we observed increased proliferation of blood cells after overexpression of the human TRUC-16. High levels of CDK4/6 consequent to TRUC-16 overexpression and p16 decrease can be specifically targeted in both CLL and Richter’s Transformation (RT), a rare but deadly consequence of CLL. Inhibition of CDK6 is a therapeutic option never tested in CLL, but in use for another type of B cell malignancy, such as the mantle cell lymphoma. Our results highlight the multifaceted contribution of a UCE to human tumorigenesis and open new possibilities for the use of long-noncoding RNAs in cancer therapies. Citation Format: Linda Fabris, Erik Knutsen, Maria Teresa S. Bertilaccio, Steliana Calin, Recep Bayraktar, George T. Eisenhoffer, Leonard Girnita, George A. Calin. Regulation of p16-E2F1-CDK4/6 signaling by TRUC-16 ultraconserved long non-coding RNA. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3799.

The causes and consequences of abnormal biogenesis of extracellular vesicles (EVs) are not yet well understood in malignancies, including in breast cancers (BCs). Given the hormonal signaling dependence of estrogen receptor-positive (ER+) BC, we hypothesized that 17β-estradiol (estrogen) might influence EV production and microRNA (miRNA) loading. We report that physiological doses of 17β-estradiol promote EV secretion specifically from ER+ BC cells via inhibition of miR-149-5p, hindering its regulatory activity on SP1, a transcription factor that regulates the EV biogenesis factor nSMase2. Additionally, miR-149-5p downregulation promotes hnRNPA1 expression, responsible for the loading of let-7's miRNAs into EVs. In multiple patient cohorts, we observed increased levels of let-7a-5p and let-7d-5p in EVs derived from the blood of premenopausal ER+ BC patients, and elevated EV levels in patients with high BMI, both conditions associated with higher levels of 17β-estradiol. In brief, we identified a unique estrogen-driven mechanism by which ER+ BC cells eliminate tumor suppressor miRNAs in EVs, with effects on modulating tumor-associated macrophages in the microenvironment.

The ultraconserved regions (UCRs) are the genomic segments manifesting perfect conservation between the orthologous genomes of humans, rats, and mice that get transcribed into mono-exonic long non-coding RNAs (lncRNAs) known as transcribed ultraconserved regions (T-UCRs). Increasing evidence demonstrates the importance of T-UCRs in human cancers, however, their involvement in the pathogenesis of Chronic Lymphocytic Leukemia (CLL) is poorly understood. Our lab has recently identified a lncRNA transcribed from a UCR and named it TRUC-16 (Translational Regulatory UltraConserved gene affecting p16), which is overexpressed in CLL and correlates with treatment-free survival. We have developed a transgenic C57BL6 mouse model to overexpress TRUC-16 in B cells. We monitored the number and development of B cell subsets over time in peripheral blood and lymphoid organs (lymph nodes, spleen, and bone marrow), and performed immunophenotyping to assess clonality and proportion of the cells considered to be the origin for CLL (i.e. B1 cells and IgM memory B cells that are CD5+). We followed mice survival (n=10 for each gender) and performed necropsy with histopathologic and immunophenotypic analysis. If blood sampling suggests developing a CLL-like state, we will monitor for evolution to Richter Syndrome by flow cytometry analysis and biochemistry markers. We have validated the TRUC-16 overexpression specific to the B-cell compartment in the mice cohorts and have observed lymphoma in several organs. Interestingly, TRUC-16 overexpressing mice showed significantly higher splenic weights compared to controls. The mice were found to develop adulthood lymphoma that we are now analyzing molecularly and phenotypically. We are planning to further characterize the subtype of lymphoma by utilizing specific diagnostic markers through immunohistochemistry and flow cytometry. This is the first-ever transgenic mice modeling overexpression of UCR in the context of cancer. Validation of the functional correlation between the TRUC-16 phenotype and CLL patients’ clinical data will provide a valuable resource to pursue a multitude of preclinical studies to find targets that might help better understand the disease pathophysiology. Citation Format: Swati Mohapatra, Erik Knutsen, Mihai Iurascu Gagea, Linda Fabris, George Adrian Calin. Deciphering the in vivo roles of a novel long non-coding RNA in chronic lymphocytic leukemia and Richter syndrome. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3795.

Background: Ultraconserved regions or elements (UCEs) are greater than 100-base pairs in length and are perfectly (100%) conserved across large evolutionary distances in the genomes of at least 3 of 5 placental mammals: human, cow, dog, rat, and mouse. Most UCEs are located in non-coding regions of the genome; however, some UCEs overlap with coding exons. In addition, some UCEs have been found to be transcriptionally active as long non-coding RNAs and are involved in a variety of cellular processes such as cellular proliferation. Although a few UCE sequence variations were reported to be associated with human diseases, the molecular functions of somatic mutations in UCEs remain largely unexplained in human cancers. In this study, we characterized the distribution of somatic UCE mutations throughout a spectrum of cancers and investigated the biological functions of cancer associated UCEs. Methods: We examined somatic UCE mutations in 2,449 cases of 22 cancer types using the PCAWG and ICGC platforms. To validate these mutation patterns, UCE sequencing was performed on the Illumina NovaSeq 6000 platform for MD Anderson patient cohorts. A custom AsCpf1 guide library was built to identify potential regulatory UCE functions in colorectal cancer models, and 3 guides were designed to target 2,247 UCEs. The proliferation ratio of knockout UCEs was analyzed. We then generated stable mutated UCE clones using the CRISPR AsCpf1 technology in colorectal cancer cells. We conducted RNA sequencing (RNA-seq) of mutated UCE cells. The expression of target proteins and genes were analyzed by qRT-PCR and western blotting. To address the effects of the mutated UCE_11311 tumor growth, DLD1 cells were injected in athymic nude mice. Results: We analyzed the WGS data on 2,449 cases of 22 cancer types and identified 24,039 somatic mutations in 10,090 (73.46%) UCEs. These were mostly located in non-coding DNAs, mainly in introns.Based on the RNA-seq data, one of the strongest impacts of mutated UCEs was on ARID1B. We confirmed that specific mutated UCE decreased ARID1B mRNA and protein levels. Our in vivo results demonstrated that UCE mutations enhanced the tumorigenicity of DLD1 xenograft tumors. As a summary, UCE_11311 has a transcriptional enhancer activity on the ARID1B gene and UCE_11311 mutations are actively participating in the tumorigenesis at least in part through regulating ARID1B expression. Discussion: We identified hundreds of unexplored tumorigenic UCEs that need to be further characterized functionally and clinically, and proved one of them to be a transcriptional enhancer of the tumor suppressor ARID1B. Collectively, these data support the concept that certain somatic UCE mutations are frequent and functional in cancer evolution, acting as driver mutations that can be used as new therapeutic targets; other somatic UCE mutations are cancer specific or patient specific, are markers of aggressiveness, and can be used to personalized therapy. Citation Format: Recep Bayraktar, Yitao Tang, Mihnea P. Dragomir, Linda Fabris, Giulio F. Draetta, Han Liang, George A. Calin. The mutational landscape of ultraconserved elements in human cancers. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5750.

Extensive efforts recently witnessed the complexity of cancer biology; however, molecular medicine still lacks the ability to elucidate hidden mechanisms for the maintenance of specific subclasses of rare tumors characterized by the silent onset and a poor prognosis (e.g., ovarian cancer, pancreatic cancer, and glioblastoma). Recent mutational fingerprints of human cancers highlighted genomic alteration occurring on epigenetic modulators. In this scenario, the epigenome dependency of cancer orchestrates a broad range of cellular processes critical for tumorigenesis and tumor progression, possibly mediating escaping mechanisms leading to drug resistance. Indeed, in this review, we discuss the pivotal role of chromatin remodeling in shaping the tumor architecture and modulating tumor fitness in a microenvironment-dependent context. We will also present recent advances in the epigenome targeting, posing a particular emphasis on how this knowledge could be translated into a feasible therapeutic approach to individualize clinical settings and improve patient outcomes.

BACKGROUND & AIMS: Chromosomal instability (CIN) is a carcinogenesis event that promotes metastasis and resistance to therapy by unclear mechanisms. Expression of the colon cancer-associated transcript 2 gene (CCAT2), which encodes a long noncoding RNA (lncRNA), associates with CIN, but little is known about how CCAT2 lncRNA regulates this cancer enabling characteristic. METHODS: We performed cytogenetic analysis of colorectal cancer (CRC) cell lines (HCT116, KM12C/SM, and HT29) overexpressing CCAT2 and colon organoids from C57BL/6N mice with the CCAT2 transgene and without (controls). CRC cells were also analyzed by immunofluorescence microscopy, g-H2AX, and senescence assays. CCAT2 transgene and control mice were given azoxymethane and dextran sulfate sodium to induce colon tumors. We performed gene expression array and mass spectrometry to detect downstream targets of CCAT2 lncRNA. We characterized interactions between CCAT2 with downstream proteins using MS2 pull-down, RNA immunoprecipitation, and selective 20-hydroxyl acylation analyzed by primer extension analyses. Downstream proteins were overexpressed in CRC cells and analyzed for CIN. Gene expression levels were measured in CRC and non-tumor tissues from 5 cohorts, comprising more than 900 patients. RESULTS: High expression of CCAT2 induced CIN in CRC cell lines and increased resistance to 5-fluorouracil and oxaliplatin. Mice that expressed the CCAT2 transgene developed chromosome abnormalities, and colon organoids derived from crypt cells of these mice had a higher percentage of chromosome abnormalities compared with organoids from control mice. The transgenic mice given azoxymethane and dextran sulfate sodium developed more and larger colon polyps than control mice given these agents. Microarray analysis and mass spectrometry indicated that expression of CCAT2 increased expression of genes involved in ribosome biogenesis and protein synthesis. CCAT2 lncRNA interacted directly with and stabilized BOP1 ribosomal biogenesis factor (BOP1). CCAT2 also increased expression of MYC, which activated expression of BOP1. Overexpression of BOP1 in CRC cell lines resulted in chromosomal missegregation errors, and increased colony formation, and invasiveness, whereas BOP1 knockdown reduced viability. BOP1 promoted CIN by increasing the active form of aurora kinase B, which regulates chromosomal segregation. BOP1 was overexpressed in polyp tissues from CCAT2 transgenic mice compared with healthy tissue. CCAT2 lncRNA and BOP1 mRNA or protein were all increased in micro satellite stable tumors (characterized by CIN), but not in tumors with microsatellite instability compared with nontumor tissues. Increased levels of CCAT2 lncRNA and BOP1 mRNA correlated with each other and with shorter survival times of patients. CONCLUSIONS: We found that overexpression of CCAT2 in colon cells promotes CIN and carcinogenesis by stabilizing and inducing expression of BOP1 an activator of aurora kinase B. Strategies to target this pathway might be developed for treatment of patients with microsatellite stable colorectal tumors.

Liquid biopsy—the determination of circulating cells, proteins, DNA or RNA from biofluids through a “less invasive” approach—has emerged as a novel approach in all cancer entities. Circulating non-(protein) coding RNAs including microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and YRNAs can be passively released by tissue or cell damage or actively secreted as cell-free circulating RNAs, bound to lipoproteins or carried by exosomes. In renal cell carcinoma (RCC), a growing body of evidence suggests circulating non-coding RNAs (ncRNAs) such as miRNAs, lncRNAs, and YRNAs as promising and easily accessible blood-based biomarkers for the early diagnosis of RCC as well as for the prediction of prognosis and treatment response. In addition, circulating ncRNAs could also play a role in RCC pathogenesis and progression. This review gives an overview over the current study landscape of circulating ncRNAs and their involvement in RCC pathogenesis as well as their potential utility as future biomarkers in RCC diagnosis and treatment.

The discovery of non-coding RNAs (ncRNAs) and their role in tumor onset and progression has revolutionized the way scientists and clinicians study cancers. This discovery opened new layers of complexity in understanding the fine-tuned regulation of cellular processes leading to cancer. NcRNAs represent a heterogeneous group of transcripts, ranging from a few base pairs to several kilobases, that are able to regulate gene networks and intracellular pathways by interacting with DNA, transcripts or proteins. Deregulation of ncRNAs impinge on several cellular responses and can play a major role in each single hallmark of cancer. This review will focus on the most important short and long non-coding RNAs in chronic lymphocytic leukemia (CLL), highlighting their implications as potential biomarkers and therapeutic targets as they relate to the well-established hallmarks of cancer. The key molecular events in the onset of CLL will be contextualized, taking into account the role of the "dark matter" of the genome.

Myelofibrosis (MF) is a myeloproliferative neoplasm characterized by cytopenia and extramedullary hematopoiesis, resulting in splenomegaly. Multiple pathological mechanisms (e.g., circulating cytokines and genetic alterations, such as JAKV617F mutation) have been implicated in the etiology of MF, but the molecular mechanism causing resistance to JAK2V617F inhibitor therapy remains unknown. Among MF patients who were treated with the JAK inhibitor ruxolitinib, we compared noncoding RNA profiles of ruxolitinib therapy responders versus nonresponders and found miR-543 was significantly upregulated in nonresponders. We validated these findings by reverse transcription-quantitative PCR. in this same cohort, in 2 additional independent MF patient cohorts from the United States and Romania, and in a JAK2V617F mouse model of MF. Both in vitro and in vivo models were used to determine the underlying molecular mechanism of miR-543 in MF. Here, we demonstrate that miR-543 targets the dioxygenases ten-eleven translocation 1 (TET1) and 2 (TET2) in patients and in vitro, causing increased levels of global 5-methylcytosine, while decreasing the acetylation of histone 3, STAT3, and tumor protein p53. Mechanistically, we found that activation of STAT3 by JAKs epigenetically controls miR-543 expression via binding the promoter region of miR-543. Furthermore, miR-543 upregulation promotes the expression of genes related to drug metabolism, including CYP3A4, which is involved in ruxolitinib metabolism. Our findings suggest miR-543 as a potentially novel biomarker for the prognosis of MF patients with a high risk of treatment resistance and as a potentially new target for the development of new treatment options.

The cancer-risk-associated rs6983267 single nucleotide polymorphism (SNP) and the accompanying long noncoding RNA CCAT2 in the highly amplified 8q24.21 region have been implicated in cancer predisposition, although causality has not been established. Here, using allele-specific CCAT2 transgenic mice, we demonstrate that CCAT2 overexpression leads to spontaneous myeloid malignancies. We further identified that CCAT2 is overexpressed in bone marrow and peripheral blood of myelodysplastic/myeloproliferative neoplasms (MDS/MPN) patients. CCAT2 induces global deregulation of gene expression by down-regulating EZH2 in vitro and in vivo in an allele-specific manner. We also identified a novel non-APOBEC, non-ADAR, RNA editing at the SNP locus in MDS/MPN patients and CCAT2-transgenic mice. The RNA transcribed from the SNP locus in malignant hematopoietic cells have different allelic composition from the corresponding genomic DNA, a phenomenon rarely observed in normal cells. Our findings provide fundamental insights into the functional role of rs6983267 SNP and CCAT2 in myeloid malignancies.

In the originally published version of this Article, the positions of the final two authors in the author list were inadvertently inverted during the production process. This error has now been corrected in both the PDF and HTML versions of the Article.

Transcribed ultraconserved regions (T-UCRs) are genomic regions conserved across large evolutionary distances, which encode for noncoding RNAs that serve as regulators of gene expression. Although T-UCRs have been linked to multiple aspects of mammalian gene regulation, the roles of their extreme evolutionary conservation remain largely unexplained. Growing body of literature is now focusing on T-UCRs as potential cancer biomarkers or as new drug targets. Here we present an overview of the discoveries so far published about the role of T-UCR in cancer and disease.

Clinico-pathological characteristics of the metastatic colorectal carcinoma from the third CRC patient cohort. (XLSX 9 kb)

BACKGROUND:Non-coding RNAs have been drawing increasing attention in recent years as functional data suggest that they play important roles in key cellular processes. N-BLR is a primate-specific long non-coding RNA that modulates the epithelial-to-mesenchymal transition, facilitates cell migration, and increases colorectal cancer invasion.RESULTS:We performed multivariate analyses of data from two independent cohorts of colorectal cancer patients and show that the abundance of N-BLR is associated with tumor stage, invasion potential, and overall patient survival. Through in vitro and in vivo experiments we found that N-BLR facilitates migration primarily via crosstalk with E-cadherin and ZEB1. We showed that this crosstalk is mediated by a pyknon, a short ~20 nucleotide-long DNA motif contained in the N-BLR transcript and is targeted by members of the miR-200 family. In light of these findings, we used a microarray to investigate the expression patterns of other pyknon-containing genomic loci. We found multiple such loci that are differentially transcribed between healthy and diseased tissues in colorectal cancer and chronic lymphocytic leukemia. Moreover, we identified several new loci whose expression correlates with the colorectal cancer patients' overall survival.CONCLUSIONS:The primate-specific N-BLR is a novel molecular contributor to the complex mechanisms that underlie metastasis in colorectal cancer and a potential novel biomarker for this disease. The presence of a functional pyknon within N-BLR and the related finding that many more pyknon-containing genomic loci in the human genome exhibit tissue-specific and disease-specific expression suggests the possibility of an alternative class of biomarkers and therapeutic targets that are primate-specific.

The tumor suppressor protein p27Kip1 plays a pivotal role in the control of cell growth and metastasis formation.Several studies pointed to different roles for p27Kip1 in the control of Ras induced transformation, although no explanation has been provided to elucidate these differences. We recently demonstrated that p27kip1 regulates H-Ras activity via its interaction with stathmin.Here, using in vitro and in vivo models, we show that p27kip1 is an important regulator of Ras induced transformation. In H-RasV12 transformed cells, p27kip1 suppressed cell proliferation and tumor growth via two distinct mechanisms: 1) inhibition of CDK activity and 2) impairment of MT-destabilizing activity of stathmin. Conversely, in K-Ras4BV12 transformed cells, p27kip1 acted mainly in a CDK-dependent but stathmin-independent manner.Using human cancer-derived cell lines and primary breast and sarcoma samples, we confirmed in human models what we observed in mice.Overall, we highlight a pathway, conserved from mouse to human, important in the regulation of H-Ras oncogenic activity that could have therapeutic and diagnostic implication in patients that may benefit from anti-H-Ras therapies.

Our recent study has uncovered an additional mechanism by which the cell cycle inhibitor p27kip1 controls cell proliferation. Through its effect on the activity of the microtubule destabilizing protein Stathmin, p27kip1 modulates full H-Ras activation and, as a consequence, the MAPK signaling cascade. This regulatory mechanism influences the cell cycle in vitro and tissue and/or organ growth in mice, in vivo and, when unbalanced, may lead to uncontrolled proliferation and tumor onset.