A novel cancer-associated cassette exon in TLN1 alters Talin 1 mechanosensitivity

Talin 1 is the core mechanosensitive adapter protein linking integrins to the cytoskeleton. The TLN1 gene is comprised of 57 exons that encode the 2541 amino acid TLN1 protein. TLN1 was previously considered to be expressed as a single isoform. However, through differential pre-mRNA splicing analysis, we discovered a cancer-enriched, non-annotated 51-nucleotide exon in TLN1 between exons 17 and 18, which we refer to as exon 17b. TLN1 is comprised of an N-terminal FERM domain, linked to 13 force-dependent switch domains, R1-R13. Inclusion of exon 17b results in an in-frame insertion of 17 amino acids immediately after Gln665 in the region between R1-R2 which lowers the force required to open the R1-R2 switches potentially altering downstream mechanotransduction. Biochemical analysis of this isoform revealed enhanced vinculin binding, and cells expressing this variant show altered adhesion dynamics. Finally, we show that the TGF-Beta/SMAD3 signaling pathway regulates this isoform switch. Future studies will need to consider the balance of these two TLN1 isoforms

Current and future diagnostic and treatment strategies for patients with invasive lobular breast cancer.

BACKGROUND: Invasive lobular breast cancer (ILC) is the second most common type of breast cancer after invasive breast cancer of no special type (NST), representing up to 15% of all breast cancers. DESIGN: Latest data on ILC are presented, focusing on diagnosis, molecular make-up according to the European Society for Medical Oncology Scale for Clinical Actionability of molecular Targets (ESCAT) guidelines, treatment in the early and metastatic setting and ILC-focused clinical trials. RESULTS: At the imaging level, magnetic resonance imaging-based and novel positron emission tomography/computed tomography-based techniques can overcome the limitations of currently used imaging techniques for diagnosing ILC. At the pathology level, E-cadherin immunohistochemistry could help improving inter-pathologist agreement. The majority of patients with ILC do not seem to benefit as much from (neo-)adjuvant chemotherapy as patients with NST, although chemotherapy might be required in a subset of high-risk patients. No differences in treatment efficacy are seen for anti-human epidermal growth factor receptor 2 (HER2) therapies in the adjuvant setting and cyclin-dependent kinases 4 and 6 inhibitors in the metastatic setting. The clinical utility of the commercially available prognostic gene expression-based tests is unclear for patients with ILC. Several ESCAT alterations differ in frequency between ILC and NST. Germline BRCA1 and PALB2 alterations are less frequent in patients with ILC, while germline CDH1 (gene coding for E-cadherin) alterations are more frequent in patients with ILC. Somatic HER2 mutations are more frequent in ILC, especially in metastases (15% ILC versus 5% NST). A high tumour mutational burden, relevant for immune checkpoint inhibition, is more frequent in ILC metastases (16%) than in NST metastases (5%). Tumours with somatic inactivating CDH1 mutations may be vulnerable for treatment with ROS1 inhibitors, a concept currently investigated in early and metastatic ILC. CONCLUSION: ILC is a unique malignancy based on its pathological and biological features leading to differences in diagnosis as well as in treatment response, resistance and targets as compared to NST

Selective inhibition of BRCA2-deficient mammary tumor cell growth by AZD2281 and cisplatin

Purpose: To assess efficacy of the novel, selective poly (ADP-ribose) polymerase-1 (PARP-1) inhibitor AZD2281 against newly established BRCA2-deficient mouse mammary tumor cell lines and to determine potential synergy between AZD2281 and cisplatin. Experimental Design: We established and thoroughly characterized a panel of clonal cell lines from independent BRCA2-deficient mouse mammary tumors and BRCA2-proficient control tumors. Subsequently, we assessed sensitivity of these lines to conventional cytotoxic drugs and the novel PARP inhibitor AZD2281. Finally, in vitro combination studies were done to investigate interaction between AZD2281 and cisplatin. Results: Genetic, transcriptional, and functional analyses confirmed the successful isolation of BRCA2-deficient and BRCA2-proficient mouse mammary tumor cell lines. Treatment of these cell lines with 11 different anticancer drugs or with gamma-irradiation showed that AZD2281, a novel and specific PARP inhibitor, caused the strongest differential growth inhibition of BRCA2-deficient versus BRCA2-proficient mammary tumor cells. Finally, drug combination studies showed synergistic cytotoxicity of AZD2281 and cisplatin against BRCA2-deficient cells but not against BRCA2-proficient control cells. Conclusion: We have successfully established the first set of BRCA2-deficient mammary tumor cell lines, which form an important addition to the existing preclinical models for BRCA-mutated breast cancer. The exquisite sensitivity of these cells to the PARP inhibitor AZD2281, alone or in combination with cisplatin, provides strong support for AZD2281 as a novel targeted therapeutic against BRCA-deficient cancers

Loss of E-cadherin leads to Id2-dependent inhibition of cell cycle progression in metastatic lobular breast cancer.

Invasive lobular breast carcinoma (ILC) is characterized by proliferative indolence and long-term latency relapses. This study aimed to identify how disseminating ILC cells control the balance between quiescence and cell cycle re-entry. In the absence of anchorage, ILC cells undergo a sustained cell cycle arrest in G0/G1 while maintaining viability. From the genes that are upregulated in anchorage independent ILC cells, we selected Inhibitor of DNA binding 2 (Id2), a mediator of cell cycle progression. Using loss-of-function experiments, we demonstrate that Id2 is essential for anchorage independent survival (anoikis resistance) in vitro and lung colonization in mice. Importantly, we find that under anchorage independent conditions, E-cadherin loss promotes expression of Id2 in multiple mouse and (organotypic) human models of ILC, an event that is caused by a direct p120-catenin/Kaiso-dependent transcriptional de-repression of the canonical Kaiso binding sequence TCCTGCNA. Conversely, stable inducible restoration of E-cadherin expression in the ILC cell line SUM44PE inhibits Id2 expression and anoikis resistance. We show evidence that Id2 accumulates in the cytosol, where it induces a sustained and CDK4/6-dependent G0/G1 cell cycle arrest through interaction with hypo-phosphorylated Rb. Finally, we find that Id2 is indeed enriched in ILC when compared to other breast cancers, and confirm cytosolic Id2 protein expression in primary ILC samples. In sum, we have linked mutational inactivation of E-cadherin to direct inhibition of cell cycle progression. Our work indicates that loss of E-cadherin and subsequent expression of Id2 drive indolence and dissemination of ILC. As such, E-cadherin and Id2 are promising candidates to stratify low and intermediate grade invasive breast cancers for the use of clinical cell cycle intervention drugs