Common Chemical Inductors of Replication Stress:  Focus on Cell‐Based Studies

Vesela, Eva; Chroma, Katarina; Turi, Zsofia; Mistrík, Martin

doi:10.3390/biom7010019

Cited by 93 publications

(85 citation statements)

References 324 publications

(363 reference statements)

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“…To further assess whether TIAR prevents mitotic entry in the presence of unreplicated DNA, we induced mild replication stress by treatment with a low dose (0.4 μM) of aphidicolin (APH), an inhibitor of DNA polymerases. At this concentration, cells are affected in S‐phase specifically, and replication is perturbed only partially . Quantification of p‐H3‐positive cells showed that the G2/M checkpoint was strongly activated after 8 h in control cells, whereas TIAR kd cells activated the checkpoint only partially and entered mitosis prematurely (Fig A).…”

Section: Resultsmentioning

confidence: 94%

TIAR marks nuclear G2/M transition granules and restricts CDK 1 activity under replication stress

et al. 2018

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The G2/M checkpoint coordinates DNA replication with mitosis and thereby prevents chromosome segregation in the presence of unreplicated or damaged DNA. Here, we show that the RNA‐binding protein TIAR is essential for the G2/M checkpoint and that TIAR accumulates in nuclear foci in late G2 and prophase in cells suffering from replication stress. These foci, which we named G2/M transition granules (GMGs), occur at low levels in normally cycling cells and are strongly induced by replication stress. In addition to replication stress response proteins, GMGs contain factors involved in RNA metabolism as well as CDK1. Depletion of TIAR accelerates mitotic entry and leads to chromosomal instability in response to replication stress, in a manner that can be alleviated by the concomitant depletion of Cdc25B or inhibition of CDK1. Since TIAR retains CDK1 in GMGs and attenuates CDK1 activity, we propose that the assembly of GMGs may represent a so far unrecognized mechanism that contributes to the activation of the G2/M checkpoint in mammalian cells.

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Section: Resultsmentioning

confidence: 94%

TIAR marks nuclear G2/M transition granules and restricts CDK 1 activity under replication stress

et al. 2018

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“…In recent studies, we have shown that 7a treatment further increases S phase arrest of the cell cycle over hydroxyurea (HU) treatment [13], which is a pure inducer of replication stressdependent S phase arrest [60,61]. In the present study, HCT116 cells were treated with 25 mM of 5-FU for 24 h, then with 20 mM of 7a or 7b for additional 8 h (Fig.…”

Section: Compound 7b Abrogates S Phase Arrest Of Hct116 Cellsmentioning

confidence: 68%

“…It is well-established that ATR-mediated Chk1 phosphorylation at S317P and S345P plays a critical role in DNA replication, DNA repair, S-G 2 /M phase checkpoint control [62], and has been the target for cancer therapy [63], It has been also shown that the inhibition of Chk1(S317P) and Chk1(S296P) abrogates Sphase progression [64,65]. Since 7b abrogates S phase arrest, we determined the phosphorylation level of Chk1 after treatment with 7b either alone or in combination with HU, a pure inducer of S phase arrest [60,61]. The experimental protocol is described in Fig.…”

Section: Compound 7b Blocks Hu-induced Phosphorylation Of Chk1 In Hctmentioning

confidence: 99%

ASR352, A potent anticancer agent: Synthesis, preliminary SAR, and biological activities against colorectal cancer bulk, 5-fluorouracil/oxaliplatin resistant and stem cells

Narayan

Ramisetti

Jaiswal

et al. 2019

European Journal of Medicinal Chemistry

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a b s t r a c tDespite new agent development and short-term benefits in patients with colorectal cancer (CRC), metastatic CRC cure rates have not improved due to high rates of 5-fluorouracil (5-FU)/leucovorin/ oxaliplatin (FOLFOX)-resistance and a clinical therapeutic plateau. At the same time, this treatment regime leads to significant toxicity, cost, and patient inconvenience. Drug-resistance is linked to CRC stem cells, which are associated with the epidermal-to-mesenchymal transition (EMT) pathway. Thus, to optimally treat CRC, a therapy that can target the cell survival and EMT pathways in both CRC bulk and stem cell populations is critical. We recently identified a novel small molecule NSC30049 (7a) that is effective alone, and in combination potentiates 5-FU-mediated growth inhibition of CRC bulk, FOLFOXresistant, and CRC stem cells both in vitro and in vivo models. In the present study, we report the synthesis and anti-CRC evaluation of several stable and effective 7a analogs. ASR352 (7b) was identified as one of the equipotent 7a analogs that inhibited the growth of CRC bulk cells, sensitized FOLFOX-resistant cells, and reduced the sphere formation capacity of CRC stem cells. It appears that the complex mechanism of cytotoxicity for 7b includes abrogation of 5-FU-induced the S phase, reduction of the phosphorylation of Chk1 at S317P, S345P and S296P, increased gH2AX staining, activation of caspase 3/PARP1 cleavage, and enhancement of Bax/Bcl2 ratio. Further 7b-mediated reduced phosphorylation of Chk1 was an indirect effect, since it did not inhibit Chk1 activity in an in vitro kinase assay. Our findings suggest that 7b as a single agent, or in combination with 5-FU can be developed as a therapeutic agent in CRC bulk, FOLFOX-resistant, and CRC stem cell populations for unmanageable metastatic CRC conditions.

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“…4A and 4B). These treatments seem to act through a common pathway: induction of DDR [45]. Specifically, HU, which inhibits the incorporation of nucleotides by interfering with the enzyme ribonucleotide reductase [46], and APH, which interferes with DNA replication by inhibiting DNA polymerases α, ε and d [47], are both commonly used to induce replication fork stalling that leads to ATR/ATM activation.…”

Section: Discussionmentioning

confidence: 99%

APOBEC3B reporter myeloma cell lines identify DNA damage response pathways leading to APOBEC3B expression

Yamazaki

Shirakawa

Matsumoto

et al. 2019

Preprint

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18 2 19 Text word count: 3635 words 20 Abstract word count: 190 words 21 Number of figures: 4 22 Number of tables: 0 (Supplemental Table: 2) 23 Number of references: 69 3 25 Abstract 26 Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) DNA cytosine 27 deaminase 3B (A3B) is a DNA editing enzyme which induces genomic DNA mutations in 28 multiple myeloma and various other cancers. APOBEC family proteins are highly homologous 29 so it is especially difficult to investigate the biology of A3B alone in cancer cells. To investigate 30 A3B function in myeloma cells easily and comprehensively, we used CRISPR/Cas9 to generate 31 A3B reporter cells that contain 3×FLAG tag and IRES-EGFP sequences integrated at the end of 32 the A3B gene. These reporter cells stably express 3xFLAG tagged A3B and the reporter EGFP 33 and this expression is enhanced under known stimuli, such as PMA. Conversely, shRNA 34 knockdown of A3B decreased EGFP fluorescence and 3xFLAG tagged A3B protein levels. We 35 screened a series of anticancer treatments using these cell lines and identified that most 36 conventional therapies, such as antimetabolites or radiation, exacerbated endogenous A3B 37 expression, but recent molecular targeting drugs, including bortezomib, lenalidomide and 38 elotuzumab, did not. Furthermore, chemical inhibition of ATM, ATR and DNA-PK suppressed 39 the EGFP expression upon treatment with antimetabolites. These results suggest that DNA 40 damage response triggers A3B expression through ATM, ATR and DNA-PK signaling. 42 Introduction43 The apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like DNA cytosine 44 deaminase 3 family (APOBEC3, A3) consists of seven proteins (A3A, A3B, A3C, A3D, A3F, 45 A3G and A3H) that preferentially induce C to U mutations in single strand DNA. A3 proteins 46 were originally identified as factors of the innate immunity due to their mutagenic activity on 47 viral genomes, and have recently joined the growing list of key intrinsic mutagens that play a 48 part in oncogenesis [1]. Evidence for A3 mutagenicity consists in the presence of their 49 mutational signature in cancer genomes [2], in the effects observed when overexpressed in tumor 50 tissues [3, 4], as well as in the correlation of APOBEC signature mutations with poor prognosis 51 [5, 6]. Nevertheless, the precise biology of individual APOBEC3 proteins in cancer cells remains 52 unknown. Due to the high structural homology of APOBEC3 family members, it is particularly 53 difficult to obtain high-affinity-and high-specificity-antibodies against each APOBEC3 protein, 54 which limits our capability to distinguish the precise role of each endogenous APOBEC3 during 55 tumorigenesis. 56Among APOBEC3s, we previously reported that endogenous A3B is overexpressed and 57 seems to be the main source of deamination activity in most of the myeloma cell lines we 58 examined [7]. Notably, high levels of A3B expression in tumor cells were an independent risk 59 factor for overall survival in myeloma patients [7] as well as in...

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Common Chemical Inductors of Replication Stress: Focus on Cell‐Based Studies

Cited by 93 publications

References 324 publications

TIAR marks nuclear G2/M transition granules and restricts CDK 1 activity under replication stress

TIAR marks nuclear G2/M transition granules and restricts CDK 1 activity under replication stress

ASR352, A potent anticancer agent: Synthesis, preliminary SAR, and biological activities against colorectal cancer bulk, 5-fluorouracil/oxaliplatin resistant and stem cells

APOBEC3B reporter myeloma cell lines identify DNA damage response pathways leading to APOBEC3B expression

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