Bruton’s tyrosine kinase (BTK) is a non-receptor intracellular kinase playing a key role in the proliferation and survival of normal and malignant B-lymphocytes. Its targeting by Ibrutinib, the first specific inhibitor, represented a turning point for the therapy of certain types of B-cell leukemias/lymphomas and several more BTK inhibitors are today in the clinic or advanced clinical trials. BTK expression was successively found to occur also outside of the hematopoietic compartment. In fact, we identified p65BTK, a novel 65 kDa isoform lacking an N-term stretch of 86 amino acids (compared to the 77 kDa protein expressed in B cells) as highly expressed in colon cancer patients. We demonstrated that p65BTK is a powerful oncogene acting downstream of the RAS/MAPK pathway and necessary for RAS-mediated transformation. Notably, the kinase domain is conserved and therefore inhibited by the available BTK-targeting drugs (Ibrutinib, Spebrutinib, etc.) which we used to demonstrate that p65BTK is an actionable target in drug-resistant colorectal carcinomas. We found p65BTK expressed also in >50% non-small cell lung cancers (NSCLC) and demonstrated that it is an actionable target in KRAS-mutated/EGFR-wild type drug-resistant NSCLC models (for which no targeted therapy is available). We also reported a significant correlation between p65BTK expression and low-grade tumors and overall survival of patients with grade III gliomas and showed that its targeting induced a significant decrease in the viability of in glioma stem cells. Finally, in ovarian cancer patients, p65BTK expression levels correlate with early relapse and shorter progression-free survival, both indicators of resistance to therapy. Remarkably, Ibrutinib is more effective than standard of care (SOC) therapeutics in in vitro and ex vivo settings. On the whole, our preclinical data indicate that, depending on the tumor type, BTK inhibitors used alone can induce cytotoxicity (gliomas), be more effective than SOC chemotherapy (ovarian cancer) or can kill drug-resistant tumor cells when used in combination with SOC chemotherapy (colon cancer and NSCLC) or targeted therapy (NSCLC and ovarian cancer), thus suggesting that p65BTK may be an actionable target in different solid tumors. In addition, our data also give the proof-of-concept for starting clinical trials using BTK inhibitors, alone or in combination, to improve the therapeutic options for solid tumors treatment.
The transgenic technologies represent potent biotechnological tools that allow the generation of genetically modified animals useful for basic research and for biomedical, veterinary, and agricultural applications. Among transgenic techniques, we describe here the sperm-mediated gene transfer methods that is gene transfer based on the spontaneous ability of sperm cells to bind and internalize exogenous DNA and to carry it to oocyte during fertilization, producing genetically modified animals with high efficiency.
Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with a malignant prognosis. GBM is characterized by high cellular heterogeneity and its progression relies on the interaction with the central nervous system, which ensures the immune-escape and tumor promotion. This interplay induces metabolic, (epi)-genetic and molecular rewiring in both domains. In the present study, we aim to characterize the time-related changes in the GBM landscape, using a syngeneic mouse model of primary GBM. GL261 glioma cells were injected in the right striatum of immuno-competent C57Bl/6 mice and animals were sacrificed after 7, 14, and 21 days (7D, 14D, 21D). The tumor development was assessed through 3D tomographic imaging and brains were processed for immunohistochemistry, immunofluorescence, and western blotting. A human transcriptomic database was inquired to support the translational value of the experimental data. Our results showed the dynamic of the tumor progression, being established as a bulk at 14D and surrounded by a dense scar of reactive astrocytes. The GBM growth was paralleled by the impairment in the microglial/macrophagic recruitment and antigen-presenting functions, while the invasive phase was characterized by changes in the extracellular matrix, as shown by the analysis of tenascin C and metalloproteinase-9. The present study emphasizes the role of the molecular changes in the microenvironment during the GBM progression, fostering the development of novel multi-targeted, time-dependent therapies in an experimental model similar to the human disease.
It is known that Bruton's tyrosine kinase (BTK) is essential for B-cell proliferation/differentiation and it is generally believed that its expression and function are limited to bone marrow-derived cells. Here, we report the identification and characterization of p65BTK, a novel oncogenic isoform expressed in colon carcinomas. Standard procedures were used for cloning the full length p65BTK-encoding mRNA and raise anti-p65BTK specific polyclonal antibodies. Quantitative PCR, western blot, RNA immunoprecipitation, silencing experiments and fluorescence assay upon transfection with bi-cistronic vectors were used to demonstrate post-transcriptional regulation of p65BTK expression. Immunohystochemistry was employed to study p65BTK expression in colon cancer patients specimens. Soft agar assay and foci assay were carried out to assess p65BTK oncogenic properties. Cell growth, cell viability and colony assays were performed to study the effects of p65BTK inhibition (by a specific kinase inhibitor) on colon cancer cells biology. We found that p65BTK differs from the already known 77 kDa isoform for the lack of the N-terminal PH domain and is translated - through an IRES-dependent mechanism -from a transcript containing an alternative first exon in the 5'UTR. Moreover, p65BTK mRNA translation requires phosho-hnRNPK binding to its cognate sites (located in the alternative first exon) and is post-transcriptionally regulated, via hnRNPK, by the MAPK pathway. We demonstrate that p65BTK is endowed with strong transforming activity that depends on active ERK1/2 and its inhibition abolishes RAS transforming activity. Accordingly, p65BTK overexpression in colon cancer tissues correlates with ERK1/2 activation. Finally we show that p65BTK inhibition affects growth and survival of colon cancer cells. In conclusion, our data reveal that BTK, via p65BTK expression, is a novel and powerful oncogene acting downstream of the RAS/MAPK pathway and suggest that its targeting may be a promising therapeutic approach. Citation Format: Emanuela Grassilli, Fabio Pisano, Annamaria Cialdella, Sara Bonomo, Carola Missaglia, Maria Grazia Cerrito, Laura Masiero, Leonarda Ianzano, Robert Narloch, Filomena D'Amato, Barbara Noli, Gian Luca Ferri, Biagio E. Leone, Giorgio Stanta, Serena Bonin, Kristian Helin, Roberto Giovannoni, Marialuisa Lavitrano. A novel oncogenic BTK isoform is overexpressed in colon cancers and required for RAS-mediated transformation. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B121.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.