ORCID Profile
0000-0002-0141-5975
Current Organisation
oNKo-Innate
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Publisher: Cold Spring Harbor Laboratory
Date: 29-11-2020
DOI: 10.1101/2020.11.29.398867
Abstract: The mechanisms by which cells respond and adapt to oxidative stress are largely unknown but are key to developing a rationale for cancer therapies that target antioxidant pathways. APR-246 is a mutant-p53 targeted therapeutic currently under clinical investigation in myeloid dysplastic syndrome (MDS) and acute myeloid leukemia 1 . Whilst the mechanism of action of APR-246 is thought to be reactivation of wild-type p53 activity through covalent modification of cysteine residues in the core domain of mutant-p53 protein 2,3 , here we report that the anti-neoplastic capacity of APR-246 lies predominantly in the conjugation of free cysteine. Genome-wide CRISPR perturbation screening, metabolite profiling and proteomics in response to APR-246 treatment in mutant-p53 cancer cells highlighted the role of GSH and mitochondrial metabolism in determining APR-246 efficacy. APR-246 sensitivity was increased through loss of key enzymes in mitochondrial one-carbon metabolism, SHMT2 and MTHFD1L , due to diminished glycine supply for de novo GSH synthesis. Critically, we show that APR-246 induces iron-dependent, apoptotic machinery-independent cell death, ferroptosis. Whole-cell proteomics analyses indicated an upregulation of proteins involved in iron-sulfur cluster biogenesis (eg. FDX1). GSH, acetyl-CoA and NADH levels were also depleted in APR-246 treated cells. Importantly, we found that APR-246 inhibits iron-sulfur cluster biogenesis in the mitochondria of cancer cells through cysteine conjugation. This work not only details novel determinants of APR-246 activity in cancer cells, but also provides a clinical roadmap for targeting antioxidant pathways in tumours - beyond targeting mutant-p53 tumours.
Publisher: Frontiers Media SA
Date: 17-07-2020
Publisher: Springer New York
Date: 2018
DOI: 10.1007/978-1-4939-7568-6_17
Abstract: This chapter details a compendium of protocols that collectively enable the reader to perform a pooled shRNA and/or CRISPR screen-with methods to identify and validate positive controls and subsequent hits establish a viral titer in the cell line of choice create and screen libraries, sequence strategies, and bioinformatics resources to analyze outcomes. Collectively, this provides an overarching resource from the start to finish of a screening project, making this technology possible in all laboratories.
Publisher: Elsevier
Date: 2021
Publisher: Publiverse Online S.R.L
Date: 2010
Publisher: Springer Science and Business Media LLC
Date: 27-03-2015
DOI: 10.1038/NCOMMS7548
Abstract: Basal-like breast cancer (BLBC) is a heterogeneous disease with poor prognosis however, its cellular origins and aetiology are poorly understood. In this study, we show that inhibitor of differentiation 4 (ID4) is a key regulator of mammary stem cell self-renewal and marks a subset of BLBC with a putative mammary basal cell of origin. Using an ID4GFP knock-in reporter mouse and single-cell transcriptomics, we show that ID4 marks a stem cell-enriched subset of the mammary basal cell population. ID4 maintains the mammary stem cell pool by suppressing key factors required for luminal differentiation. Furthermore, ID4 is specifically expressed by a subset of human BLBC that possess a very poor prognosis and a transcriptional signature similar to a mammary stem cell. These studies identify ID4 as a mammary stem cell regulator, deconvolute the heterogeneity of BLBC and link a subset of mammary stem cells to the aetiology of BLBC.
Publisher: Cold Spring Harbor Laboratory
Date: 15-04-2020
Abstract: Alternative polyadenylation (APA) determines stability, localization and translation potential of the majority of mRNA in eukaryotic cells. The heterodimeric mammalian cleavage factor II (CF II m ) is required for pre-mRNA 3′ end cleavage and is composed of the RNA kinase hClp1 and the termination factor hPcf11 the latter protein binds to RNA and the RNA polymerase II carboxy-terminal domain. Here, we used siRNA mediated knockdown and poly(A) targeted RNA sequencing to analyze the role of CF II m in gene expression and APA in estrogen receptor positive MCF7 breast cancer cells. Identified gene ontology terms link CF II m function to regulation of growth factor activity, protein heterodimerization and the cell cycle. An overlapping requirement for hClp1 and hPcf11 suggested that CF II m protein complex was involved in the selection of proximal poly(A) sites. In addition to APA shifts within 3′ untranslated regions (3′-UTRs), we observed shifts from promoter proximal regions to the 3′-UTR facilitating synthesis of full-length mRNAs. Moreover, we show that several truncated mRNAs that resulted from APA within introns in MCF7 cells cosedimented with ribosomal components in an EDTA sensitive manner suggesting that those are translated into protein. We propose that CF II m contributes to the regulation of mRNA function in breast cancer.
Publisher: Elsevier BV
Date: 03-2022
Publisher: American Society of Hematology
Date: 11-04-2013
DOI: 10.1182/BLOOD-2011-11-394882
Abstract: EGFL7 promotes angiogenesis via its interaction with integrin αvβ3. EGFL7 is involved in physiological and pathological angiogenesis.
Publisher: Elsevier BV
Date: 08-2021
DOI: 10.1016/J.JMII.2020.07.010
Abstract: Human papilloma viruses (HPV) are the main culprit in cervical and oropharyngeal cancers. HPV positive (+) cancers are regarded as 'oncogene addicted', displaying an absolute requirement for the continued expression of the oncogenes for their viability owing their survival, and thus making these genes salient targets for developing specific therapeutic agents. There is a strong association between HPV and oropharyngeal squamous cell carcinomas (OPSCC), a subset of head and neck cancers (HNCs). Alarmingly, HPV-associated OPSCC are on the rise globally, and the number of cases of HPV + OPSCCs surpasses that of cervical cancer in the USA. Here, we show that major HPV oncogenes, E6 and E7, are essential for the survival of HPV positive (+) OPSCCs, making these oncogenes salient targets for HPV-driven OPSCCs. HPV E7 is known to interact with STING, a component of the viral DNA-sensing cGAS-STING machinery which activates a pro-typical anti-viral type I interferon (IFN) response. Our recent work showed that E7 from HPV type 16 is responsible for the blockade of cGAS-STING responses in HPV + OPSCC cells. In this study, we show that CRISPR/Cas9-mediated loss of E7 from HPV + OPSCC cells, SCC2 and SCC104, restored cGAS-STING responses. Future work could involve HPV oncogene targeting leading to HPV + OPSCC tumour regression and that the combined use of STING agonists would induce favourable tumour clearance by activating appropriate anti-tumour responses.
Publisher: Impact Journals, LLC
Date: 16-11-2016
Publisher: Springer Science and Business Media LLC
Date: 07-06-2009
DOI: 10.1038/NCB1896
Abstract: Epidermal growth factor-like domain 7 (EGFL7) is a secreted factor implicated in cellular responses such as cell migration and blood vessel formation however the molecular mechanisms underlying the effects of EGFL7 are largely unknown. Here we have identified transmembrane receptors of the Notch family as EGFL7-binding molecules. Secreted EGFL7 binds to a region in Notch involved in ligand-mediated receptor activation, thus acting as an antagonist of Notch signalling. Expression of EGFL7 in neural stem cells (NSCs) in vitro decreased Notch-specific signalling and consequently, reduced proliferation and self-renewal of NSCs. Such altered Notch signalling caused a shift in the differentiation pattern of cultured NSCs towards an excess of neurons and oligodendrocytes. We identified neurons as a source of EGFL7 in the brain, suggesting that brain-derived EGFL7 acts as an endogenous antagonist of Notch signalling that regulates proliferation and differentiation of subventricular zone-derived adult NSCs.
Publisher: Springer New York
Date: 2015
Publisher: Cold Spring Harbor Laboratory
Date: 05-02-2021
DOI: 10.1101/2021.02.04.429849
Abstract: Patients with colorectal cancer (CRC) frequently develop liver metastases during the course of their disease. A substantial proportion of them receive neoadjuvant FOLFOX (5-Fluorouracil, Oxaliplatin, Leucovorin) prior to surgery in an attempt to enable successful surgical removal of their metastases and to reduce the risk of recurrence. Yet, the majority of patients progress during treatment or recur following surgery, and molecular mechanisms that contribute to FOLFOX resistance remain poorly understood. Here, using a combination of phenotypic, transcriptomic and genomic analyses of both tumor s les derived from patients with metastatic CRC and matching patient-derived tumor organoids (PDTOs), we characterize a novel FOLFOX resistance mechanism and identify inhibitors that target this mechanism to resensitize metastatic organoids to FOLFOX. Resistant PDTOs, identified after in vitro exposure to FOLFOX, exhibited elevated expression of E2F pathway, S phase, G 2 /M and spindle assembly checkpoints (SAC) genes. Similar molecular features were detected in CRLM from patients with progressive disease while under neoadjuvant FOLFOX treatment, highlighting the relevance of this finding. FOLFOX resistant PDTOs displayed inactivating mutations of TP53 and exhibited transcriptional features of P53 pathway downregulation. We found that they accumulated in early S-phase and underwent significant DNA damage during FOLFOX exposure, thereafter arresting in G 2 /M while they repaired their DNA after FOLFOX withdrawal. In parallel, results of a large kinase inhibitor screen indicated that drugs targeting regulators of the DNA damage response, G 2 M checkpoint and SAC had cytotoxic effects on PDTOs generated from patients whose disease progressed during treatment with FOLFOX. Corroborating this finding, CHK1 and WEE1 inhibitors were found to synergize with FOLFOX and sensitize previously resistant PDTOs. Additionally, targeting the SAC master regulator MPS1 using empesertib after exposure to FOLFOX, when cells accumulate in G 2 M, was also very effective to kill FOLFOX-resistant PDTOs. Our results indicate that targeted and timely inhibition of specific cell cycle checkpoints shows great potential to improve response rates to FOLFOX in patients with metastatic CRC, for whom therapeutic alternatives remain extremely limited.
Publisher: Oxford University Press (OUP)
Date: 16-11-2017
DOI: 10.1093/NAR/GKX1072
Publisher: Springer Science and Business Media LLC
Date: 13-06-2015
Publisher: Springer Science and Business Media LLC
Date: 23-09-2013
DOI: 10.1038/ONC.2013.368
Abstract: The HER2 (ERBB2) and MYC genes are commonly lified in breast cancer, yet little is known about their molecular and clinical interaction. Using a novel chimeric mammary transgenic approach and in vitro models, we demonstrate markedly increased self-renewal and tumour-propagating capability of cells transformed with Her2 and c-Myc. Coexpression of both oncoproteins in cultured cells led to the activation of a c-Myc transcriptional signature and acquisition of a self-renewing phenotype independent of an epithelial-mesenchymal transition programme or regulation of conventional cancer stem cell markers. Instead, Her2 and c-Myc cooperated to induce the expression of lipoprotein lipase, which was required for proliferation and self-renewal in vitro. HER2 and MYC were frequently co lified in breast cancer, associated with aggressive clinical behaviour and poor outcome. Lastly, we show that in HER2(+) breast cancer patients receiving adjuvant chemotherapy (but not targeted anti-Her2 therapy), MYC lification is associated with a poor outcome. These findings demonstrate the importance of molecular and cellular context in oncogenic transformation and acquisition of a malignant stem-like phenotype and have diagnostic and therapeutic consequences for the clinical management of HER2(+) breast cancer.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 16-09-2022
Abstract: The mechanism of action of eprenetapopt (APR-246, PRIMA-1 MET ) as an anticancer agent remains unresolved, although the clinical development of eprenetapopt focuses on its reported mechanism of action as a mutant-p53 reactivator. Using unbiased approaches, this study demonstrates that eprenetapopt depletes cellular antioxidant glutathione levels by increasing its turnover, triggering a nonapoptotic, iron-dependent form of cell death known as ferroptosis. Deficiency in genes responsible for supplying cancer cells with the substrates for de novo glutathione synthesis ( SLC7A11 , SHMT2 , and MTHFD1L ), as well as the enzymes required to synthesize glutathione ( GCLC and GCLM ), augments the activity of eprenetapopt. Eprenetapopt also inhibits iron-sulfur cluster biogenesis by limiting the cysteine desulfurase activity of NFS1, which potentiates ferroptosis and may restrict cellular proliferation. The combination of eprenetapopt with dietary serine and glycine restriction synergizes to inhibit esophageal xenograft tumor growth. These findings reframe the canonical view of eprenetapopt from a mutant-p53 reactivator to a ferroptosis inducer.
Publisher: American Society of Hematology
Date: 30-03-2022
DOI: 10.1182/BLOODADVANCES.2021004571
Abstract: Current strategies to target RNA splicing mutant myeloid cancers proposes targeting the remaining splicing apparatus. This approach has only been modestly sensitizing and is also toxic to non-mutant-bearing wild-type cells. To explore potentially exploitable genetic interactions with spliceosome mutations, we combined data mining and functional screening for synthetic lethal interactions with an Srsf2P95H/+ mutation. Analysis of missplicing events in a series of both human and murine SRSF2P95H mutant s les across multiple myeloid diseases (acute myeloid leukemia, myelodysplastic syndromes, chronic myelomonocytic leukemia) was performed to identify conserved missplicing events. From this analysis, we identified that the cell-cycle and DNA repair pathways were overrepresented within the conserved misspliced transcript sets. In parallel, to functionally define pathways essential for survival and proliferation of Srsf2P95H/+ cells, we performed a genome-wide Clustered regularly interspaced short palindromic repeat loss-of-function screen using Hoxb8 immortalized R26-CreERki/+Srsf2P95H/+ and R26-CreERki/+Srsf2+/+ cell lines. We assessed loss of single guide RNA representation at 3 timepoints: immediately after Srsf2P95H/+ activation, and at 1 week and 2 weeks after Srsf2P95H/+ mutation. Pathway analysis demonstrated that the cell-cycle and DNA damage response pathways were among the top synthetic lethal pathways with Srsf2P95H/+ mutation. Based on the loss of guide RNAs targeting Cdk6, we identified that palbociclib, a CDK6 inhibitor, showed preferential sensitivity in Srsf2P95H/+ cell lines and in primary nonimmortalized lin−cKIT+Sca-1+ cells compared with wild-type controls. Our data strongly suggest that the cell-cycle and DNA damage response pathways are required for Srsf2P95H/+ cell survival, and that palbociclib could be an alternative therapeutic option for targeting SRSF2 mutant cancers.
Publisher: Springer Science and Business Media LLC
Date: 18-05-2018
DOI: 10.1038/S41598-018-26050-Y
Abstract: Docetaxel and cabazitaxel are taxane chemotherapy treatments for metastatic castration-resistant prostate cancer (CRPC). However, therapeutic resistance remains a major issue. MicroRNAs are short non-coding RNAs that can silence multiple genes, regulating several signalling pathways simultaneously. Therefore, synthetic microRNAs may have therapeutic potential in CRPC by regulating genes involved in taxane response and minimise compensatory mechanisms that cause taxane resistance. To identify microRNAs that can improve the efficacy of taxanes in CRPC, we performed a genome-wide screen of 1280 microRNAs in the CRPC cell lines PC3 and DU145 in combination with docetaxel or cabazitaxel treatment. Mimics of miR-217 and miR-181b-5p enhanced apoptosis significantly in PC3 cells in the presence of these taxanes. These mimics downregulated at least a thousand different transcripts, which were enriched for genes with cell proliferation and focal adhesion functions. In idual knockdown of a selection of 46 genes representing these transcripts resulted in toxic or taxane sensitisation effects, indicating that these genes may be mediating the effects of the microRNA mimics. A range of these genes are expressed in CRPC metastases, suggesting that these microRNA mimics may be functional in CRPC. With further development, these microRNA mimics may have therapeutic potential to improve taxane response in CRPC patients.
Publisher: Springer Science and Business Media LLC
Date: 11-06-2020
DOI: 10.1186/S13058-020-01306-6
Abstract: Basal-like breast cancer (BLBC) is a poorly characterised, heterogeneous disease. Patients are diagnosed with aggressive, high-grade tumours and often relapse with chemotherapy resistance. Detailed understanding of the molecular underpinnings of this disease is essential to the development of personalised therapeutic strategies. Inhibitor of differentiation 4 (ID4) is a helix-loop-helix transcriptional regulator required for mammary gland development. ID4 is overexpressed in a subset of BLBC patients, associating with a stem-like poor prognosis phenotype, and is necessary for the growth of cell line models of BLBC through unknown mechanisms. Here, we have defined unique molecular insights into the function of ID4 in BLBC and the related disease high-grade serous ovarian cancer (HGSOC), by combining RIME proteomic analysis, ChIP-seq mapping of genomic binding sites and RNA-seq. These studies reveal novel interactions with DNA damage response proteins, in particular, mediator of DNA damage checkpoint protein 1 (MDC1). Through MDC1, ID4 interacts with other DNA repair proteins (γH2AX and BRCA1) at fragile chromatin sites. ID4 does not affect transcription at these sites, instead binding to chromatin following DNA damage. Analysis of clinical s les demonstrates that ID4 is lified and overexpressed at a higher frequency in BRCA1 -mutant BLBC compared with sporadic BLBC, providing genetic evidence for an interaction between ID4 and DNA damage repair deficiency. These data link the interactions of ID4 with MDC1 to DNA damage repair in the aetiology of BLBC and HGSOC.
Publisher: Research Square Platform LLC
Date: 13-05-2020
DOI: 10.21203/RS.3.RS-16182/V2
Abstract: Background Basal-like breast cancer (BLBC) is a poorly characterised, heterogeneous disease. Patients are diagnosed with aggressive, high-grade tumours and often relapse with chemotherapy resistance. Detailed understanding of the molecular underpinnings of this disease is essential to the development of personalised therapeutic strategies. Inhibitor of Differentiation 4 (ID4) is a helix-loop-helix transcriptional regulator required for mammary gland development. ID4 is overexpressed in a subset of BLBC patients, associating with a stem-like poor prognosis phenotype, and is necessary for the growth of cell line models of BLBC, through unknown mechanisms. Methods Here, we have defined unique molecular insights into the function of ID4 in BLBC and the related disease high-grade serous ovarian cancer (HGSOC), by combining RIME proteomic analysis, ChIP-seq mapping of genomic binding sites and RNA-seq. Results These studies reveal novel interactions with DNA damage response proteins, in particular, mediator of DNA damage checkpoint protein 1 (MDC1). Through MDC1, ID4 interacts with other DNA repair proteins (γH2AX and BRCA1) at fragile chromatin sites. ID4 does not affect transcription at these sites, instead binding to chromatin following DNA damage. Analysis of clinical s les demonstrates that ID4 is lified and overexpressed at a higher frequency in BRCA1 -mutant BLBC compared with sporadic BLBC, providing genetic evidence for an interaction between ID4 and DNA damage repair deficiency. Conclusions These data link the interactions of ID4 with MDC1 to DNA damage repair in the aetiology of BLBC and HGSOC.
No related grants have been discovered for Iva Nikolic.