The PIM Kinase Inhibitor K00135 Sensitizes Primary Acute Lymphoblastic Leukaemia (ALL) Cells to DNA-Damaging Agents

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3238-3238
Author(s):  
David Da Costa ◽  
Victoria J Weston ◽  
Stefan Knapp ◽  
A. Malcolm R Taylor ◽  
Pamela R. Kearns ◽  
...  
Keyword(s):  
Dna Damage ◽  
Myeloid Leukaemia ◽  
Conflicts Of Interest ◽  
Constitutive Expression ◽  
Treatment Resistance ◽  
Tumour Cells ◽  
Leukaemia Virus ◽  
Pim Kinases ◽  
The Impact

Abstract Abstract 3238 Relapsed ALL is the most common cause of death in children, with only 30% being cured despite intensified treatment and stem cell transplantation. We previously showed that a subset of paediatric ALL tumours have a defect in the apoptotic response to DNA damage in vitro which correlates with a poor clinical response in vivo. Proviral integration site for Moloney murine leukaemia virus (PIM) proteins are a family of serine/threonine kinases which mediate prosurvival signals in response to cytokines, growth factors, hypoxia and DNA damage. PIM kinases have been implicated in several haematological malignancies such as chronic myeloid leukaemia (CML), chronic lymphocytic leukaemia (CLL), acute myeloid leukaemia (AML), and T-ALL. However, the role of PIM kinases and their contribution to treatment resistance in a large cohort of primary paediatric ALL has not been comprehensively addressed. To address the frequency of PIM overexpression in ALL we used quantitative real-time PCR to quantify the levels of PIM-1 and PIM-2 in an unselected cohort of 86 primary ALL tumours compared to the PIM-expressing lymphoid cell line K562. We observed high PIM-1 levels in only 2/86 (2.3%) ALLs, whereas PIM-2 was highly expressed in 20/86 (23%) primary tumours. PIM-2 overexpression was detected in tumours with a range of chromosome translocations or hyperdiploidy. In representative samples, the level of PIM-2 protein expression correlated with abundance of mRNA transcript. We then investigated the role of PIM overexpression on apoptosis using the highly specific PIM inhibitor, imidazo[1,2-b]pyridazine K00135, to sensitize tumour cells with high PIM expression to ionising radiation (IR)-induced apoptosis. Pretreatment with K00135 led to sensitization of representative primary tumour cells with high PIM-2 expression to 5Grays IR at an half maximal lethal concentration (LC50) of 10μM, and killing was mediated by caspase-dependant apoptosis. Furthermore, consistent with the role of PIM-2 in the regulation of c-Myc protein stability (Zhang Y, Wang Z, Li X, Magnuson NS. 2008; Oncogene; 27;4890-4819), K00135-induced killing coincided with the downregulation of c-Myc protein. We are currently addressing the impact of PIM inhibition in an ALL xenograft model. Constitutive expression of PIM-2 may be an underlying mechanism of treatment resistance in a subset of paediatric ALL. Pharmacological inhibition of PIM-2 may prove to be a useful therapeutic option for poorly responding ALL in children in the future. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Potential role of oxidative DNA damage in the impact of PNPLA3 variant (rs 738409 C>G) in hepatocellular carcinoma risk

Hepatology ◽  
2014 ◽  
Vol 60 (3) ◽  
pp. 1110-1111 ◽  
Author(s):  
Emeric Limagne ◽  
Vanessa Cottet ◽  
Alexia Karen Cotte ◽  
Samia Hamza ◽  
Patrick Hillon ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Potential Role ◽  
The Impact ◽  
Carcinoma Risk

scholarly journals Evolving Role of Genomics in Genitourinary Neoplasms

2019 ◽  
Vol 48 (1) ◽  
pp. 68
Author(s):  
Michael E. Devitt ◽  
Robert Dreicer
Keyword(s):  
Dna Damage ◽  
Genomic Data ◽  
Treatment Decisions ◽  
Response To Therapy ◽  
Genomic Alterations ◽  
Dna Damage And Repair ◽  
Genitourinary Cancer ◽  
Genitourinary Cancers ◽  
The Impact

<p>The aim of this article is to review the current role of genomic testing in the risk, prognosis, and treatment of genitourinary malignancies. The authors selected guidelines, publications, and abstracts relevant to the current and emerging role of genomics in genitourinary cancers. The risk of developing genitourinary cancer can be stratified based on genomic data. Prostate cancer has the strongest degree of heritability, with <em>BRCA1/2 </em>and <em>HOXB13 </em>mutations playing a role in familial disease. Genomic data is on the verge of informing treatment decisions across genitourinary cancers. mCRPC has diverse genomic alterations that represent potential therapeutic targets, including alterations in the AR pathway, DNA damage and repair pathways, cell cycle pathways, PI3K pathway, and Wnt signaling. Genomic alterations in clear cell renal cell carcinoma can inform prognosis and mutations in mTOR pathways predict response to mTOR inhibitors. Urothelial carcinoma can be classified into different subtypes based on gene expression profiling, which provides prognostic information and predicts response to chemotherapy and immunotherapy. Specific mutations have been identified that predict response to therapy including <em>ERCC2 </em>mutations and cisplatin, DNA damage and repair mutations and checkpoint inhibitors, and <em>FGFR3 </em>mutations and FGFR tyrosine kinase inhibitors such as erdafitinib.</p><p><strong>Conclusion. </strong>Genitourinary malignancies have not felt the impact of genomic data as greatly as other cancer types. The majority of benefit lies in identifying patients at high risk of genitourinary cancer. Fortunately, breakthroughs are on the horizon that will result in a greater incorporation of genomic information into treatment decisions for patients with genitourinary cancer.</p>

Impaired Hydroxylation of 5-Methylcytosine In TET2 mutated Patients with Myeloid Malignancies

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1-1
Author(s):  
Anna Jankowska ◽  
Myunggon Ko ◽  
Yun Huang (equal contribution) ◽  
Utz J. Pape ◽  
Hadrian Szpurka ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Genomic Dna ◽  
Conflicts Of Interest ◽  
Methylation Pattern ◽  
Myeloid Malignancies ◽  
Cpg Sites ◽  
Low Levels ◽  
Methylation Patterns ◽  
The Impact

Abstract Abstract 1 TET2 mutations are frequently found across broad spectrum of myeloid malignancies but how these mutations contribute to diseases is still unknown. Preliminary results from our laboratory have suggested that TET2 converts 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) and consequently, the levels of 5-hmC may be lower in genomes of mutant bone marrow cells. To facilitate study of TET2 function we developed a blot assay to detect 5-hmC in genomic DNA with a specific antiserum to 5-hmC. In a second improved assay with increased sensitivity and precision, we treated genomic DNA with bisulfite in order to convert 5-hmC to cytosine 5-methylenesulfonate (CMS) and measured 5-hmC levels indirectly using a specific anti-CMS serum. Based on the results of this technique we demonstrate here for the first time that indeed TET2 mutations in predicted catalytic residues and other positions compromised TET2 function. We studied 102 patients with various myeloid malignancies (4/28 MDS, 14%, 26/48 MDS/MPN, 54% and 1/4 MPN, 2% and primary 2/11 AML 18% and 3/11 sAML, 27% TET2 mutants, respectively) and compared to wt cases or controls (N=17). Mutations were found throughout the entire coding region and were mostly inactivating (33/45 TET2 mutations). The levels of 5-hmC in genomic DNA from TET2 mutants were significantly decreased in comparison to wt cases and controls (p=4.5e-08 and p=1.8e-09, respectively). Particularly low levels of 5-hmC were found in patients with homozygous (UPD)/hemizygous (deletion) TET2 mutations and those with biallelic mutations. Surprisingly, 18% of all TET2 WT patients also showed low levels of genomic 5-hmC (despite normal TET2 mRNA expression), suggesting that these patients may carry not yet identified variants/lesions in TET2 or other partner proteins involved in TET2-mediated catalysis. To further investigate the impact of TET2 mutations associated with myeloid malignancies we also introduced 9 different missense mutations corresponding to those found in patients into murine Tet2 cells; severe loss of enzymatic activity was observed in 7/9 cases as measured by greatly diminished 5-hmC levels. To study the role of Tet2 in normal hematopoiesis we depleted Tet2 in C57BL/6 mice by retrovirus-mediated transduction of shRNA against Tet2. Tet2 depletion is associated with skewing of hematopoietic differentiation towards the monocyte/macrophage lineage. To further investigate the function of TET2 we transduced the myeloid THP-1 cell line with lentiviral vector containing TET2 cDNA (TET2+) or an empty vector. This manipulation allowed us to select clones showing 19-fold increase in TET2 mRNA expression without significantly alterations of proliferation kinetics. Using this model we studied the impact of TET2 overexpression on resultant methylation pattern of CpG sites. We have applied Illumina Infinium HumanMethylation27 arrays (27,5K CpG sites/14.4K genes). Overexpression of TET2 resulted in a distinct promoter methylation patterns with 169 altered CpG sites with difference of averaged β>0.5 (considered significant as compared to control). Among these differentially methylated loci, 27 promoters were significantly hypomethylated while 42 were hypermethylated as compared to control cells. Change in methylation pattern observed through overexpression of TET2 in vitro prompted us to analyze methylation patterns in patients with and without TET2 mutations or those with decreased 5-hmC levels. Using methylation arrays a total of 62 cases were analyzed. When patients were grouped based on the levels of 5hmC, an associated methylation signature can be clearly discerned with 2512 differentially methylated loci and distinct skewing towards hypomethylation (2510 sites; e.g., TMEM102, ABCC11) vs. hypermethylation (2 sites, AIM2 and SP140), consistent with the observation made in the TET2+ cells line. In sum, our results provide strong evidence for TET2 as the first mutated gene in myeloid malignancies that is involved in conversion of 5-mC to 5-hmC in DNA, indicating the novel role of TET2 in a substantial component of epigenetic deregulation in myeloid malignancies. Disclosures: No relevant conflicts of interest to declare.

Mesodermal Development From Reprogrammed Fanconi Anemia Cells Is Affected by ALDH2 Enzymatic Activity

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 648-648
Author(s):  
Naoya Suzuki ◽  
Asuka Hira ◽  
Akira Niwa ◽  
Megumu Saito ◽  
Keitaro Matsuo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Dna Damage ◽  
Developmental Stage ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
A Genome ◽  
The Impact ◽  
Mesodermal Development

Abstract Abstract 648 Introduction Fanconi anemia (FA) is a genome instability disorder with clinical characteristics including progressive bone marrow failure (BMF), developmental abnormalities, and increased occurrence of leukemia and cancer. To date 15 genes have been implicated in FA, and their products form a common DNA repair network often referred to as “FA pathway”. Following DNA damage or replication stress, the FA pathway is activated, leading to the monoubiquitination of FANCD2 and FANCI proteins (the ID complex). The monoubiquitinated ID complex is loaded on damaged chromatin with subnuclear foci formation, and mediates homologous recombination. Since cells derived from FA patients are hypersensitive to treatments that induce DNA interstrand cross-links (ICLs), the FA pathway has been considered to function in ICL repair. However, it still remains unclear what type of endogenous DNA damage is repaired through the FA pathway and is the cause of phenotypes in FA patients. Recent studies have suggested that cells deficient in the FA pathway are also sensitive to formaldehyde and acetaldehyde. Aldehydes may create DNA adducts including ICLs or protein DNA crosslinking. These results raise a possibility that the FA pathway prevents BMF by mitigating genotoxicity due to endogenous aldehydes. It has been known that ALDH2 deficiency resulting from Glu487Lys substitution (A allele) is prevalent in East Asian populations. While the Glu487 form (G allele) is proficient in aldehyde catabolism, even the GA heterozygote displayed strongly reduced catalysis because ALDH2 is a tetrameric enzyme and the variant form can suppress the activity in a dominant negative manner. Therefore some Japanese FA patients are expected to be deficient in ALDH2, providing an opportunity to test role of ALDH2 and aldehyde metabolism in human FA patients. Results and discussion In FA fetus, p53/p21 axis has already activated in fetal liver (Ceccaldi, Cell stem cell, 2012), indicating the possibility that hematopoietic defects in FA patients originates from an earlier developmental stage. Since human hematopoietic system originates from embryonic mesoderm, we set out to estimate the role of ALDH2 and FANCA pathway during early embryogenesis. For this, we reprogrammed somatic cells from a patient with ALDH2 GA genotype and observed their in vitro mesodermal differentiation. We first introduced reprogramming factors into fibroblasts by episomal vectors, and obtained colonies which are morphologically compatible with human induced pluripotent stem cells (iPSCs). These iPSC-like cells (designated as FA-iPLCs) showed close similarity to conventional ES/iPSCs regarding marker gene expressions and differentiation ability into three germ layers. We obtained gene-complemented FA-iPLCs (designated as cFA-iPLCs) for control study. To evaluate the impact of ALDH2 activity on iPSC- or iPLC-derived mesodermal differentiation, we next adapted the previously reported serum-free monolayer culture system. Both FA- and cFA-iPLCs showed similar differentiation manners with conventional embryonic stem cells and iPSCs, and percentages of KDR+ mesodermal progenitors including KDR+CD34+ common hemoangiogenic progenitors were comparable. Notably, ALDH2 agonist Alda1 did increase only FA-iPLC-derived mesodermal progenitors but not cFA-iPLCs. These data supported the hypothesis that mesodermal development towards hematopoietic cells in human can be affected by ALDH2 activity in the absence of FA pathway. To confirm the hypothesis, next we set out to assess whether the variation in ALDH2 affects symptoms in Japanese FA patients. Strikingly, we found that progression of BMF was strongly accelerated in heterozygous carrier of the variant A allele compared to homozygous GG patients. Furthermore we looked at occurrence of leukemia and/or myelodysplasia and the somatic developments. Interestingly, these were not significantly difference between patients with each variation of ALDH2, indicating the possibility that aldehydes affect only in early hematopoietic development, not other mesodermal tissues. Overall, our results from FA-iPLCs and clinical study indicate that the variation in ALDH2 affects the occurrence of bone marrow failure in FA patients, and that hematopoietic defect in FA patients is caused by aldehydes in early mesodermal developmental stage. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Vitamin D Status in Children with Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3973-3973 ◽  
Author(s):  
Natalja Jackmann ◽  
Arja H Harila-Saari ◽  
Outi Mäkitie ◽  
Jan Gustafsson ◽  
Dzeneta Nezirevich Dernroth ◽  
...  
Keyword(s):  
Vitamin D ◽  
Overall Survival ◽  
Myeloid Leukaemia ◽  
Vitamin D Deficiency ◽  
Conflicts Of Interest ◽  
Significant Negative Correlation ◽  
Vitamin D Status ◽  
Cross Sectional ◽  
Pediatric Cancer Patients ◽  
The Impact

Abstract Children and adolescents with leukemia are potentially at a high risk of developing vitamin D deficiency, due to limited physical activity and sunlight exposure, poor nutrition, chemotherapy, and its complications. The prevalence of vitamin D inadequacy in European pediatric cancer patients has been reported to be high. It is not known how many patients already have vitamin D deficiency at the time of diagnosis and whether vitamin D status at the time of diagnosis influences clinical outcome. We aimed to investigate vitamin D status in children with leukemia at the time of diagnosis and explore possible factors (age, type of leukemia, gender, year and season of sampling) contributing to a low level of 25-hydroxyvitamin D (25-OHD). Furthermore, we aimed to investigate if vitamin D status at the time of diagnosis influences overall survival. We carried out a cross-sectional study including all 295 children (169 boys, 57.3%) aged <18 years who were diagnosed with leukemia in our institution between 1991 and 2016 and had a stored serum sample available from the time of diagnosis. All samples had been stored at -80C. We analysed serum 25-OHD and PTH with reagents from the same batch in January 2018; 25-OHD levels <25 nmol/L were considered deficient, 25-50 nmol/L insufficient, 50-75 nmol/L sufficient, and ≥75nmol/L optimal. Clinical data (sex, age, diagnosis, date of the diagnosis, overall survival) were collected from the Swedish Childhood Cancer Registry. Altogether 295 children were included: 232 of them had acute lymphoblastic leukaemia (ALL), 52 acute myeloid leukaemia (AML), and 11 other types of leukemia (8 chronic myeloid leukaemia and 3 juvenile myelomonocytic leukaemia). Mean 25-OHD concentration was 60.7 nmol/L (SD 23.3). One third of the children (33.2%) had a subnormal 25-OHD level (6.4% had deficiency and 26.8% insufficiency), 39.7% were sufficient and 27.1% had an optimal level. There was a significant negative correlation between serum 25-OHD and PTH (p<0.001). Season affected serum 25-OHD: it was lowest in the spring (55.2 nmol/L, SD 21.7) and highest in the summer (68.4 nmol/L, SD 19.6). Multiple linear regression with unadjusted and adjusted analyses to explore the impact of age, diagnosis, gender, season, and time of sampling (calendar year) on 25-OHD level indicated that significant predictors of lower 25-OHD level were older age (p<0.001), sampling in the spring (p<0.001), sampling in more recent calendar year (p=0.001) and sampling in the winter (p=0.001). When exploring the impact of 25-OHD on survival, we used Cox proportional hazard regression. In the whole cohort only the diagnosis and the age at diagnosis were significant. However, when the younger patients (≤ 6 year of age) were analysed separately, 25-OHD level <50 nmol/L at the time of diagnosis was associated with inferior overall survival independently of other factors (HR 3.05, p=0.03) as compared with those with 25-OHD ≥50 nmol/L. This patient group included 163 patients with 16 events. Conclusion: Subnormal 25-OHD levels are common in pediatric patients with leukemia already at the time of diagnosis. In younger children with leukemia 25-OHD level <50 nmol/L is associated with inferior survival. Disclosures No relevant conflicts of interest to declare.

scholarly journals TRIM33 Loss in Multiple Myeloma Impairs the DNA Damage Response Resulting in Sensitivity to PARP and ATR Inhibitors

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1570-1570
Author(s):  
Roisin M McAvera ◽  
Jonathan J Morgan ◽  
Ken I Mills ◽  
Lisa J Crawford
Keyword(s):  
Multiple Myeloma ◽  
Dna Damage ◽  
Dna Damage Response ◽  
High Throughput ◽  
Chromosomal Instability ◽  
Conflicts Of Interest ◽  
Parp Cleavage ◽  
Structural Variants ◽  
Damage Response ◽  
The Impact

Abstract Introduction Chromosomal instability is a hallmark of Multiple Myeloma (MM), with most patients displaying cytogenetic abnormalities which can arise due to DNA damage response (DDR) defects. TRIM33 is an E3 ligase and transcriptional co-repressor located on chromosome 1p13.2, a region frequently deleted in MM. Previous studies have shown that TRIM33 plays a role in the DDR and can regulate chromosomal stability, but its precise function remains unknown. In this study we investigated the impact of TRIM33 loss in MM on genomic stability and DDR pathways and whether this could be exploited therapeutically. Methods The CoMMpass dataset (IA15 release) was screened to identify patients with copy number (CN) loss of TRIM33 and this was correlated with overall survival (OS) and structural variants. TRIM33 shRNA knockdown models were established in JJN3 and U266 cells. The effect on DDR signalling was determined by western blotting and immunofluorescence. The Selleckchem DNA Damage/Repair Compound Library was screened on the JJN3 model in a high-throughput manner using the CellTox™ Green cytotoxicity assay. Validation of selected compounds was performed using CellTiter® Glo viability assay or clonogenic assays. Combination indices (CI) were calculated using CompuSyn software. Results Data on CN, OS and structural variants were available for 730 newly diagnosed MM patients and of these, 69 (9.5%) were identified to have a CN loss of TRIM33. These patients have poorer OS compared to those without TRIM33 loss (52.3 months vs 72.6 months; p&lt;0.0001). Moreover, they exhibit a significantly higher median number of structural variants (deletions, duplications, inversions, and translocations; 38 vs 26; p&lt;0.0001), indicative of increased chromosomal instability. Our data in MM cell lines has shown that TRIM33 is rapidly recruited to chromatin within 5 minutes of induced DNA damage. TRIM33 knockdown led to an increase in 53BP1 foci formation and endogenous γH2AX (P&lt;0.001) indicating unrepaired DNA double-strand breaks (DSBs) typical of a DDR defect. In response to these DSBs both ATM and ATR kinases were activated as demonstrated by increased pKAP1 Ser824 and pCHK1 Ser345 respectively (p&lt;0.001). Additionally, we observed a reduction in RAD51 (p&lt;0.05) indicative of a potential defect in the DSB repair pathway homologous recombination (HR). To identify therapeutic vulnerabilities relating to TRIM33 loss, we performed a high-throughput screen to assess sensitivity to 160 unique DNA damaging compounds. TRIM33 knockdown cells exhibited increased sensitivity to 27 compounds across a range of drug classes. Additional studies confirmed that compared to control cells, TRIM33 knockdown sensitized cells to the PARP inhibitor Olaparib and ATR inhibitors BAY-1895344 and VE-821. Further investigation with VE-821 demonstrated that whilst treatment induced PARP cleavage and DSBs in both control and knockdown cells within 48 hours, knockdown cells exhibited significantly more pCHK1 Ser345 inhibition (p&lt;0.01). Furthermore, combining VE-821 with bortezomib yielded synergistic effects in TRIM33 knockdown cells across a range of doses (CI range 0.57-0.9) while no synergy was observed in control cells (CI&gt;1 for all combinations). Conclusion We have identified a subset of MM patients with TRIM33 loss who display high-risk disease characterized by chromosomal abnormalities and defective DDR. Alongside this we have identified PARP and ATR inhibitors as therapeutic vulnerabilities in cell line models of TRIM33 loss. Moreover, we demonstrate that ATR inhibition increases the efficacy of bortezomib in TRIM33 knockdown cells. Further investigation into these compounds could lead to novel therapies for patients with TRIM33 loss. Disclosures No relevant conflicts of interest to declare.

scholarly journals Actin dynamics regulation by TTC7A/PI4KIIIα axis limits DNA damage and cell death during leukocyte migration

2021 ◽  
Author(s):  
Tania Gajardo ◽  
Marie Lo ◽  
Mathilde Bernard ◽  
Claire Leveau ◽  
Marie-Therese El-Daher ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Cell Migration ◽  
Actin Cytoskeleton ◽  
Leukocyte Migration ◽  
Actin Dynamics ◽  
Immune Homeostasis ◽  
Cellular Functions ◽  
The Impact

The actin cytoskeleton has a crucial role in the maintenance of the immune homeostasis by controlling various cell processes, including cell migration. Mutations in the TTC7A gene have been described as the cause of a primary immunodeficiency associated to different degrees of gut involvement and alterations in the actin cytoskeleton dynamics. Although several cellular functions have been associated with TTC7A, the role of the protein in the maintenance of the immune homeostasis is still poorly understood. Here we leverage microfabricated devices to investigate the impact of TTC7A deficiency in leukocytes migration at the single cell level. We show that TTC7A-deficient leukocytes exhibit an altered cell migration and reduced capacity to deform through narrow gaps. Mechanistically, TTC7A-deficient phenotype resulted from impaired phosphoinositides signaling, leading to the downregulation of the PI3K/AKT/RHOA regulatory axis and imbalanced actin cytoskeleton dynamic. This resulted in impaired cell motility, accumulation of DNA damage and increased cell death during chemotaxis in dense 3D gels. Our results highlight a novel role of TTC7A as a critical regulator of leukocyte migration. Impairment of this cellular function is likely to contribute to pathophysiology underlying progressive immunodeficiency in patients.

scholarly journals Evidence for hypoxia increasing the tempo of evolution in glioblastoma

2020 ◽  
Vol 123 (10) ◽  
pp. 1562-1569
Author(s):  
David Robert Grimes ◽  
Marnix Jansen ◽  
Robert J. Macauley ◽  
Jacob G. Scott ◽  
David Basanta
Keyword(s):  
Cell Division ◽  
Cancer Progression ◽  
Metastatic Disease ◽  
Evolutionary Dynamics ◽  
Treatment Resistance ◽  
Tumour Cells ◽  
Ecological Niches ◽  
Tumour Hypoxia ◽  
Low Oxygen ◽  
The Impact

Abstract Background Tumour hypoxia is associated with metastatic disease, and while there have been many mechanisms proposed for why tumour hypoxia is associated with metastatic disease, it remains unclear whether one precise mechanism is the key reason or several in concert. Somatic evolution drives cancer progression and treatment resistance, fuelled not only by genetic and epigenetic mutation but also by selection from interactions between tumour cells, normal cells and physical micro-environment. Ecological habitats influence evolutionary dynamics, but the impact on tempo of evolution is less clear. Methods We explored this complex dialogue with a combined clinical–theoretical approach by simulating a proliferative hierarchy under heterogeneous oxygen availability with an agent-based model. Predictions were compared against histology samples taken from glioblastoma patients, stained to elucidate areas of necrosis and TP53 expression heterogeneity. Results Results indicate that cell division in hypoxic environments is effectively upregulated, with low-oxygen niches providing avenues for tumour cells to spread. Analysis of human data indicates that cell division is not decreased under hypoxia, consistent with our results. Conclusions Our results suggest that hypoxia could be a crucible that effectively warps evolutionary velocity, making key mutations more likely. Thus, key tumour ecological niches such as hypoxic regions may alter the evolutionary tempo, driving mutations fuelling tumour heterogeneity.

Regulation of PCNA Mono-Ubiqutination by FANCD2 and Rad51 in Response to Hydroxyurea.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2377-2377
Author(s):  
Gary M. Kupfer ◽  
Xiaoyong Chen
Keyword(s):  
Dna Damage ◽  
Ubiquitin Ligase ◽  
Conflicts Of Interest ◽  
Translesion Synthesis ◽  
Deficient Mutant ◽  
Independent Manner ◽  
Knock Down ◽  
Mutant Cells

Abstract Abstract 2377 FANCD2 is a key player in FA pathway. It has been shown that FANCD2 can interact with PCNA and with Rad18, the ubiquitin ligase responsible for PCNA mono-ubiquitination. The mono-ubiquitination of PCNA is very important for its function in translesion synthesis. We found that in response to DNA damage agent hyxdroxyurea (HU) the interaction of FANCD2 with Rad18 or Rad51, and the interaction of Rad51 with Rad18 or PCNA was enhanced. FANCD2 is required for increased interaction between Rad51 and Rad18 indicating that FANCD2, Rad51 and Rad18 form a complex in response to HU. Rad18 was required for PCNA mono-ubiquitination in response to HU. FANCD2 deficient cells failed to enhance the interaction between Rad18 and Rad51. Furthermore, PCNA mono-ubiquitination was impaired in FANCD2 deficient cells in response to HU. FANCD2 mono-ubiquitination deficient mutant partially rescued PCNA mono-ubiquitination. The partial mono-ubiquitination of PCNA in response to HU in FANCA deficient mutant confirmed the role of non-ubiquitinated FANCD2 in PCNA mono-ubiquitination. The normal mono-ubiquitination of PCNA in FANCJ deficient mutant confirmed that the effect of FANCD2 in PCNA mon-ubiquitination is not due to FA pathway deficiency. Rad51 was also involved in regulating PCNA mono-ubiquitination in response to HU. Rad51 siRNA knock down cells showed decreased PCNA mono-ubiquitination in response to HU. The role of Rad51 in regulating PCNA mono-ubiquitination did not require BRCA1, indicating that this function is independent of HR. More importantly FANCD2 deficient cells were hypersensitive to HU, whereas FANCD2 mono-ubiquitination deficient mutant cells, FANCD2 corrected cells, FANCA deficient cells and FANCJ deficient cells were not hypersensitive to HU. Our data indicate that FANCD2 plays an important role in PCNA mono-ubiquitination and translesion synthesis partially in a mono-ubiquitination independent manner. Rad51 also plays an important role in PCNA mono-ubiquitination and translesion synthesis in a HR independent fashion. Disclosures: No relevant conflicts of interest to declare.

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