N‐terminal sequences are important sites for post‐translational modifications that alter protein localization, activity, and stability. Dipeptidyl peptidase 9 (DPP9) is a serine aminopeptidase with ...the rare ability to cleave off N‐terminal dipeptides with imino acid proline in the second position. Here, we identify the tumor‐suppressor BRCA2 as a DPP9 substrate and show this interaction to be induced by DNA damage. We present crystallographic structures documenting intracrystalline enzymatic activity of DPP9, with the N‐terminal Met1‐Pro2 of a BRCA21‐40 peptide captured in its active site. Intriguingly, DPP9‐depleted cells are hypersensitive to genotoxic agents and are impaired in the repair of DNA double‐strand breaks by homologous recombination. Mechanistically, DPP9 targets BRCA2 for degradation and promotes the formation of RAD51 foci, the downstream function of BRCA2. N‐terminal truncation mutants of BRCA2 that mimic a DPP9 product phenocopy reduced BRCA2 stability and rescue RAD51 foci formation in DPP9‐deficient cells. Taken together, we present DPP9 as a regulator of BRCA2 stability and propose that by fine‐tuning the cellular concentrations of BRCA2, DPP9 alters the BRCA2 interactome, providing a possible explanation for DPP9's role in cancer.
Synopsis
Dipeptidyl peptidase 9 triggers BRCA2 degradation and promotes repair of DNA double strand breaks by homologous recombination.
The amino dipeptidase DPP9 cleaves BRCA2 and removes the N‐terminal dipeptide Met‐Pro.
In response to DNA damage, DPP9 targets BRCA2 for proteasomal degradation.
DPP9 limits BRCA2‐PALB2 interactions and promotes RAD51 foci formation.
Depletion of DPP9 leads to hypersensitivity to genotoxic stress, accumulation of γH2AX, and defects in repair of DNA double strand breaks by homologous recombination.
Dipeptidyl peptidase 9 triggers BRCA2 degradation and promotes repair of DNA double strand breaks by homologous recombination.
The standard treatment of locally advanced esophageal cancer comprises multimodal treatment concepts including preoperative chemoradiotherapy (CRT) followed by radical surgical resection. However, ...despite intensified treatment approaches, 5-year survival rates are still low. Therefore, new strategies are required to overcome treatment resistance, and to improve patients’ outcome. In this study, we investigated the impact of Wnt/β-catenin signaling on CRT resistance in esophageal cancer cells. Experiments were conducted in adenocarcinoma and squamous cell carcinoma cell lines with varying expression levels of Wnt proteins and Wnt/β-catenin signaling activities. To investigate the effect of Wnt/β-catenin signaling on CRT responsiveness, we genetically or pharmacologically inhibited Wnt/β-catenin signaling. Our experiments revealed that inhibition of Wnt/β-catenin signaling sensitizes cell lines with robust pathway activity to CRT. In conclusion, Wnt/β-catenin activity may guide precision therapies in esophageal carcinoma patients.
Abstract Background and purpose Preoperative chemoradiotherapy (CRT) represents the standard treatment for locally advanced rectal cancer. Tumor response and progression vary considerably. MicroRNAs ...represent master regulators of gene expression, and may therefore contribute to this diversity. Material and methods Genome-wide microRNA (miRNA) profiling was performed for 12 colorectal cancer (CRC) cell lines and an individual in vitro signature of chemoradiosensitivity was established. Functional relevance of selected miRNAs was established by transfecting miRNA-mimics into SW480 and SW837 cells. The prognostic value of selected miRNAs was assessed in 128 pretherapeutic patient biopsies. Results Thirty-six miRNAs were identified to significantly correlate with sensitivity to CRT ( Q < 0.05) including miR-320a and other miRNAs involved in the MAPK-, TGF- and Wnt-pathway. Transfection of selected miRNAs (let-7g, miR-132, miR-224, miR-320a) each induced a shift of sensitivity. High expression of let-7g was associated with a good prognosis in rectal cancer patients ( P = 0.03). Conclusions This is the first report of a miRNA expression signature for in vitro chemoradiosensitivity of CRC cell lines. Many of the identified miRNAs have not been linked to the response to CRT and may represent potential molecular targets to sensitize resistant cancers. If further validated, let7g expression may serve as predictive biomarker.
A considerable percentage of rectal cancers are resistant to standard preoperative chemoradiotherapy. Because patients with a priori-resistant tumors do not benefit from multimodal treatment, ...understanding and overcoming this resistance remains of utmost clinical importance. We recently reported overexpression of the Wnt transcription factor TCF4, also known as TCF7L2, in rectal cancers that were resistant to 5-fluorouracil-based chemoradiotherapy. Because Wnt signaling has not been associated with treatment response, we aimed to investigate whether TCF4 mediates chemoradioresistance. RNA interference-mediated silencing of TCF4 was employed in three colorectal cancer (CRC) cell lines, and sensitivity to (chemo-) radiotherapy was assessed using a standard colony formation assay. Silencing of TCF4 caused a significant sensitization of CRC cells to clinically relevant doses of X-rays. This effect was restricted to tumor cells with high T cell factor (TCF) reporter activity, presumably in a β-catenin-independent manner. Radiosensitization was the consequence of (i) a transcriptional deregulation of Wnt/TCF4 target genes, (ii) a silencing-induced G2/M phase arrest, (iii) an impaired ability to adequately halt cell cycle progression after radiation and (iv) a compromised DNA double strand break repair as assessed by γH2AX staining. Taken together, our results indicate a novel mechanism through which the Wnt transcription factor TCF4 mediates chemoradioresistance. Moreover, they suggest that TCF4 is a promising molecular target to sensitize resistant tumor cells to (chemo-) radiotherapy.
Abrogation of growth factor-dependent signaling represents an effective therapeutic strategy for patients with colorectal cancer (CRC). Here we evaluated the effectiveness of targeting the epidermal ...growth factor (EGF) receptors HER-2 and HER-3 in the three cell lines LS513, LS1034 and SW837.
Treatment with HER-2-specific antibodies trastuzumab and pertuzumab resulted in a mild reduction of cellular viability. In contrast, the antibody-drug conjugate T-DM1 mediated a strong and dose-dependent decrease of viability and Akt phosphorylation.
The most striking effects were observed with the dual tyrosine kinase inhibitor lapatinib, and the Pan-ErbB inhibitor afatinib. Selectively, the effect of EGF receptor inhibition was augmented by a combination with 5-fluorouracil and oxaliplatin. Finally, high expression of HER-3 was detected in 121 of 172 locally advanced rectal cancers (70.3%). In conclusion, inhibition of EGF receptors effectively blocks downstream signaling and significantly impairs viability of CRC cells. However, the effectiveness of receptor inhibition highly depends on the inhibitors' mode of action, as targeting HER-2 alone is not sufficient.
Since HER-2 and HER-3 are expressed in a relevant number of patients, targeting both receptors may represent a promising therapeutic strategy for CRC.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Chemoradiotherapy (CRT) represents a standard treatment for many human cancers, frequently combined with radical surgical resection. However, a considerable percentage of primary cancers are at least ...partially resistant to CRT, which represents a substantial clinical problem, because it exposes cancer patients to the potential side effects of both irradiation and chemotherapy. It is therefore exceedingly important to determine the molecular characteristics underlying CRT-resistance and to identify novel molecular targets that can be manipulated to re-sensitize resistant tumors to CRT. In this review, we highlight much of the recent evidence suggesting that the signal transducer and activator of transcription 3 (STAT3) plays a prominent role in mediating CRT-resistance, and we outline why inhibition of STAT3 holds great promise for future multimodal treatment concepts in oncology.
The debate is ongoing regarding the potential role of preoperative chemoradiotherapy (CRT) for patients with pancreatic ductal adenocarcinoma (PDAC), and whether it should be reserved for borderline ...resectable or unresectable tumors. However, treatment response is heterogeneous, implicating the need to unveil and overcome the underlying mechanisms of resistance. Activation of the transcription factor STAT3 was recently linked to CRT resistance in other gastrointestinal cancers such as rectal and esophageal cancers, but its role in PDAC needs to be clarified. Protein expression and phosphorylation of STAT3 was determined in PDAC cell lines and connected to transcriptional activity measured by dual-luciferase reporter gene assays. Inhibition of STAT3 signaling was achieved by RNAi or the small-molecule inhibitor napabucasin. We observed a positive correlation between STAT3 signaling activity and CRT resistance. Importantly, genetical and pharmacological perturbation of the IL-6/STAT3 pathway resulted in CRT sensitization specifically in those cell lines, in which STAT3 activity was augmented by IL-6. In conclusion, our data underscore the general importance of IL-6/STAT3 signaling for CRT resistance and suggest that pathway inhibition may represents a putative treatment strategy in order to increase the fraction of patients with PDAC who are candidates for surgical approaches.
Disruptor of telomeric silencing 1-like (DOT1L) is a non-SET domain containing methyltransferase known to catalyze mono-, di-, and tri-methylation of histone 3 on lysine 79 (H3K79me). DOT1L-mediated ...H3K79me has been implicated in chromatin-associated functions including gene transcription, heterochromatin formation, and DNA repair. Recent studies have uncovered a role for DOT1L in the initiation and progression of leukemia and other solid tumors. The development and availability of small molecule inhibitors of DOT1L may provide new and unique therapeutic options for certain types or subgroups of cancer.
In this study, we examined the role of DOT1L in DNA double-strand break (DSB) response and repair by depleting DOT1L using siRNA or inhibiting its methyltransferase activity using small molecule inhibitors in colorectal cancer cells. Cells were treated with different agents to induce DNA damage in DOT1L-depleted or -inhibited cells and analyzed for DNA repair efficiency and survival. Further, rectal cancer patient samples were analyzed for H3K79me3 levels in order to determine whether it may serve as a potential marker for personalized therapy.
Our results indicate that DOT1L is required for a proper DNA damage response following DNA double-strand breaks by regulating the phosphorylation of the variant histone H2AX (γH2AX) and repair via homologous recombination (HR). Importantly, we show that small molecule inhibitors of DOT1L combined with chemotherapeutic agents that are used to treat colorectal cancers show additive effects. Furthermore, examination of H3K79me3 levels in rectal cancer patients demonstrates that lower levels correlate with a poorer prognosis.
In this study, we conclude that DOT1L plays an important role in an early DNA damage response and repair of DNA double-strand breaks via the HR pathway. Moreover, DOT1L inhibition leads to increased sensitivity to chemotherapeutic agents and PARP inhibition, which further highlights its potential clinical utility. Our results further suggest that H3K79me3 can be useful as a predictive and or prognostic marker for rectal cancer patients.
Purpose: Voltage-gated Kv potassium channels, like ether a go-go (EAG) channels, have been recognized for their oncogenic potential
in breast cancer and other malignant tumors.
Experimental Design: ...We examined the molecular and functional expression of Kv channels in human colonic cancers and colon of mice treated with
the chemical carcinogens dimethylhydrazine and N -methyl- N -nitrosourea. The data were compared with results from control mice and animals with chemically induced DSS colitis.
Results: Electrogenic salt transport by amiloride-sensitive Na + channels and cyclic AMP–activated cystic fibrosis transmembrane conductance regulator Cl − channels were attenuated during tumor development and colitis, whereas Ca 2+ -dependent transport remained unchanged. Kv channels, in particular Eag-1, were enhanced during carcinogenesis. Multiplex
reverse transcription-PCR showed increased mRNA expression for Kv1.3, Kv1.5, Kv3.1, and members of the Eag channel family,
after dimethylhydrazine and N -methyl- N -nitrosourea treatment. Eag-1 protein was detected in the malignant mouse colon and human colonic cancers. Genomic amplification
of Eag-1 was found in 3.4% of all human colorectal adenocarcinoma and was an independent marker of adverse prognosis.
Conclusions: The study predicts an oncogenic role of Kv and Eag channels for the development of colonic cancer. These channels may represent
an important target for a novel pharmacotherapy of colonic cancer.
Resistance of tumor cells to chemoradiotherapy represents a fundamental problem in clinical oncology. The underlying mechanisms are actively debated. Here we show that blocking inflammatory cytokine ...receptor signaling via STAT3 re-sensitized treatment-refractory cancer cells and abolished tumor growth in a xenograft mouse model when applied together with chemoradiotherapy. STAT3 executed treatment resistance by triggering the expression of RBPJ, the key transcriptional regulator of the NOTCH pathway. The mandatory RBPJ interaction partner, NOTCH intracellular domain, was provided by tumor cell-intrinsic expression of NOTCH ligands that caused tonic NOTCH proteolysis. In fact, NOTCH inhibition phenocopied the effect of blocking STAT3 signaling. Moreover, genetic profiling of rectal cancer patients revealed the importance of the STAT3/NOTCH axis as NOTCH expression correlated with clinical outcome. Our data uncovered an unprecedented signal alliance between inflammation and cellular development that orchestrated resistance to chemoradiotherapy. Clinically, our findings allow for biomarker-driven patient stratification and offer novel treatment options.
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