Copy number alterations in metastatic and early breast tumours: prognostic and acquired biomarkers of resistance to CDK4/6 inhibitors.

BACKGROUND: Copy number alterations (CNA) are acquired during the evolution of cancers from their early stage to metastatic stage. This study aims at analysing the clinical value of the identified metastasis-associated CNAs both in metastatic breast cancers (mBCs) and early breast cancers (eBCs). METHODS: Single-nucleotide polymorphism (SNP)-array was performed on 926 biopsies from mBC patients, enrolled in SAFIR02-BREAST prospective trial. CNA profiles of eBCs from The Cancer Genome Atlas Breast Invasive Carcinoma (n = 770), Molecular Taxonomy of Breast Cancer International Consortium (n = 1620) and PACS04 trial (n = 243) cohorts were used as references for comparing mBCs and eBCs CNA profiles. Overall survival was the considered survival endpoint. RESULTS: Among the twenty-one genes frequently altered in ER + /HER2- mBCs: focal amplification of TERT was associated with poor outcome in the ER + /HER2- mBC population. Among the ER + /HER2- mBCs patients for whom CDK4/6 inhibitors information before biopsies collection was available: we identified seven genes on post-treatment biopsies, including the cyclin-dependent kinase 4 (CDK4), which was amplified in 9.8% of the ER + /HER2- mBCs pretreated population, as compared to 1.5% in the ER + /HER2- mBCs unpretreated population (P = 2.82E-04) as well as the 3 eBC populations. CDK4 amplification was associated with poor outcome in the ER + /HER2- eBCs. CONCLUSIONS: This study provides insights into the biology of mBCs and identifies clinically useful genomic features for future improvement of breast cancer patient management.

Metastatic renal cell carcinoma with occult primary: a multicenter prospective cohort.

Metastatic carcinoma of presumed renal origin (rCUP) has recently emerged as a new entity within the heterogeneous entity of Cancers of Unknown Primary (CUP) but their biological features and optimal therapeutic management remain unknown. We report the molecular characteristics and clinical outcome of a series of 25 rCUP prospectively identified within the French National Multidisciplinary Tumor Board for CUP. This cohort strongly suggests that rCUP share similarities with common RCC subtypes and benefit from renal-tailored systemic treatment. This study highlights the importance of integrating clinical and molecular data for optimal diagnosis and management of CUP.

Escape from X inactivation is directly modulated by levels of Xist non-coding RNA.

In placental females, one copy of the two X chromosomes is largely silenced during a narrow developmental time window, in a process mediated by the non-coding RNA Xist1. Here, we demonstrate that Xist can initiate X-chromosome inactivation (XCI) well beyond early embryogenesis. By modifying its endogenous level, we show that Xist has the capacity to actively silence genes that escape XCI both in neuronal progenitor cells (NPCs) and in vivo, in mouse embryos. We also show that Xist plays a direct role in eliminating TAD-like structures associated with clusters of escapee genes on the inactive X chromosome, and that this is dependent on Xist's XCI initiation partner, SPEN2. We further demonstrate that Xist's function in suppressing gene expression of escapees and topological domain formation is reversible for up to seven days post-induction, but that sustained Xist up-regulation leads to progressively irreversible silencing and CpG island DNA methylation of facultative escapees. Thus, the distinctive transcriptional and regulatory topologies of the silenced X chromosome is actively, directly - and reversibly - controlled by Xist RNA throughout life.

VC-resist glioblastoma cell state: vessel co-option as a key driver of chemoradiation resistance.

Glioblastoma (GBM) is a highly lethal type of cancer. GBM recurrence following chemoradiation is typically attributed to the regrowth of invasive and resistant cells. Therefore, there is a pressing need to gain a deeper understanding of the mechanisms underlying GBM resistance to chemoradiation and its ability to infiltrate. Using a combination of transcriptomic, proteomic, and phosphoproteomic analyses, longitudinal imaging, organotypic cultures, functional assays, animal studies, and clinical data analyses, we demonstrate that chemoradiation and brain vasculature induce cell transition to a functional state named VC-Resist (vessel co-opting and resistant cell state). This cell state is midway along the transcriptomic axis between proneural and mesenchymal GBM cells and is closer to the AC/MES1-like state. VC-Resist GBM cells are highly vessel co-opting, allowing significant infiltration into the surrounding brain tissue and homing to the perivascular niche, which in turn induces even more VC-Resist transition. The molecular and functional characteristics of this FGFR1-YAP1-dependent GBM cell state, including resistance to DNA damage, enrichment in the G2M phase, and induction of senescence/stemness pathways, contribute to its enhanced resistance to chemoradiation. These findings demonstrate how vessel co-option, perivascular niche, and GBM cell plasticity jointly drive resistance to therapy during GBM recurrence.

Tumor mutational burden assessment and standardized bioinformatics approach using custom NGS panels in clinical routine.

BACKGROUND: High tumor mutational burden (TMB) was reported to predict the efficacy of immune checkpoint inhibitors (ICIs). Pembrolizumab, an anti-PD-1, received FDA-approval for the treatment of unresectable/metastatic tumors with high TMB as determined by the FoundationOne®CDx test. It remains to be determined how TMB can also be calculated using other tests. RESULTS: FFPE/frozen tumor samples from various origins were sequenced in the frame of the Institut Curie (IC) Molecular Tumor Board using an in-house next-generation sequencing (NGS) panel. A TMB calculation method was developed at IC (IC algorithm) and compared to the FoundationOne® (FO) algorithm. Using IC algorithm, an optimal 10% variant allele frequency (VAF) cut-off was established for TMB evaluation on FFPE samples, compared to 5% on frozen samples. The median TMB score for MSS/POLE WT tumors was 8.8 mut/Mb versus 45 mut/Mb for MSI/POLE-mutated tumors. When focusing on MSS/POLE WT tumor samples, the highest median TMB scores were observed in lymphoma, lung, endometrial, and cervical cancers. After biological manual curation of these cases, 21% of them could be reclassified as MSI/POLE tumors and considered as "true TMB high." Higher TMB values were obtained using FO algorithm on FFPE samples compared to IC algorithm (40 mut/Mb [10-3927] versus 8.2 mut/Mb [2.5-897], p < 0.001). CONCLUSIONS: We herein propose a TMB calculation method and a bioinformatics tool that is customizable to different NGS panels and sample types. We were not able to retrieve TMB values from FO algorithm using our own algorithm and NGS panel.

p4EBP1 staining predicts outcome in ER-positive endocrine-resistant metastatic breast cancer patients treated with everolimus and exemestane.

BACKGROUND: To identify patients most likely to respond to everolimus, a mammalian target of rapamycin (mTOR) inhibitor, a prospective biomarker study was conducted in hormone receptor-positive endocrine-resistant metastatic breast cancer patients treated with exemestane-everolimus therapy. METHODS: Metastatic tumor biopsies were processed for immunohistochemical staining (p4EBP1, PTEN, pAKT, LKB1, and pS6K). ESR1, PIK3CA and AKT1 gene mutations were detected by NGS. The primary endpoint was the association between the p4EBP1 expression and clinical benefit rate (CBR) at 6 months of everolimus plus exemestane treatment. RESULTS: Of 150 patients included, 107 were evaluable for the primary endpoint. p4EBP1 staining above the median (Allred score ≥6) was associated with a higher CBR at 6 months (62% versus 40% in high-p4EBP1 versus low-p4EBP1, χ2 test, p = 0.026) and a longer progression-free survival (PFS) (median PFS of 9.2 versus 5.8 months in high-p4EBP1 versus low-p4EBP1; p = 0.02). When tested with other biomarkers, only p4EBP1 remained a significant predictive marker of PFS in multivariate analysis (hazard ratio, 0.591; p = 0.01). CONCLUSIONS: This study identified a subset of patients with hormone receptor-positive endocrine-resistant metastatic breast cancer and poor outcome who would derive less benefit from everolimus and exemestane. p4EBP1 may be a useful predictive biomarker in routine clinical practice. CLINICAL TRIAL REGISTRATION: NCT02444390.