The incorporation of circulating tumor DNA (ctDNA) into the management of operable breast cancer (BC) has been hampered by the heterogeneous results from different studies. We aimed to assess the prognostic value of ctDNA in patients with operable (non metastatic) BC. A systematic search of databases (PubMed/Medline, Embase, and CENTRAL) and conference proceedings was conducted to identify studies reporting the association of ctDNA detection with disease-free survival (DFS) and overall survival (OS) in patients with stage I-III BC. Log-hazard ratios (HRs) were pooled at each timepoint of ctDNA assessment (baseline, after neoadjuvant therapy, and follow-up). ctDNA assays were classified as primary tumor-informed and non tumor-informed. Of the 3174 records identified, 57 studies including 5779 patients were eligible. In univariate analyses, ctDNA detection was associated with worse DFS at baseline [HR 2.98, 95% confidence interval (CI) 1.92-4.63], after neoadjuvant therapy (HR 7.69, 95% CI 4.83-12.24), and during follow-up (HR 14.04, 95% CI 7.55-26.11). Similarly, ctDNA detection at all timepoints was associated with worse OS (at baseline: HR 2.76, 95% CI 1.60-4.77; after neoadjuvant therapy: HR 2.72, 95% CI 1.44-5.14; and during follow-up: HR 9.19, 95% CI 3.26-25.90). Similar DFS and OS results were observed in multivariate analyses. Pooled HRs were numerically higher when ctDNA was detected at the end of neoadjuvant therapy or during follow-up and for primary tumor-informed assays. ctDNA detection sensitivity and specificity for BC recurrence ranged from 0.31 to 1.0 and 0.7 to 1.0, respectively. The mean lead time from ctDNA detection to overt recurrence was 10.81 months (range 0-58.9 months). ctDNA detection was associated with worse DFS and OS in patients with operable BC, particularly when detected after treatment and using primary tumor-informed assays. ctDNA detection has a high specificity for anticipating BC relapse.

Hormone receptor-positive breast cancer (HR+ BC) accounts for approximately 75% of new BC diagnoses. Despite the undisputable progresses obtained in the treatment of HR+ BC in recent years, primary or acquired resistance to endocrine therapies still represents a clinically relevant issue, and is largely responsible for disease recurrence after curative surgery, as well as for disease progression in the metastatic setting. Among the mechanisms causing primary or acquired resistance to endocrine therapies is the loss of estrogen/progesterone receptor expression, which could make BC cells independent of estrogen stimulation and, consequently, resistant to estrogen deprivation or the pharmacological inhibition of estrogen receptors. This review aims at discussing the molecular mechanisms and the clinical implications of HR loss as a result of the therapies used in the neoadjuvant setting or for the treatment of advanced disease in HR+ BC patients.

Thyroid carcinoma (TC) comprises several histotypes with different aggressiveness, from well (papillary carcinoma, PTC) to less differentiated forms (poorly differentiated and anaplastic thyroid carcinoma, PDTC and ATC, respectively). Previous reports have suggested a functional role for cancer-associated fibroblasts (CAFs) or senescent TC cells in the progression of PTC. In this study, we investigated the presence of CAFs and senescent cells in proprietary human TCs including PTC, PDTC, and ATC. Screening for the driving lesions BRAFV600E and N/H/KRAS mutations, and gene fusions was also performed to correlate results with tumor genotype. In samples with unidentified drivers, transcriptomic profiles were used to establish a BRAF- or RAS-like molecular subtype based on a gene signature derived from The Cancer Genome Atlas. By using immunohistochemistry, we found co-occurrence of stromal CAFs and senescent TC cells at the tumor invasive front, where deposition of collagen (COL1A1) and expression of lysyl oxidase (LOX) enzyme were also detected, in association with features of local invasion. Concurrent high expression of CAFs and of the senescent TC cells markers, COL1A1 and LOX was confirmed in different TC histotypes in proprietary and public gene sets derived from Gene Expression Omnibus (GEO) repository, and especially in BRAF mutated or BRAF-like tumors. In this study, we show that CAFs and senescent TC cells co-occur in various histotypes of BRAF-driven thyroid tumors and localize at the tumor invasive front.

Patients with estrogen receptor–positive breast cancer receive adjuvant endocrine therapies (ET) that delay relapse by targeting clinically undetectable micrometastatic deposits. Yet, up to 50% of patients relapse even decades after surgery through unknown mechanisms likely involving dormancy. To investigate genetic and transcriptional changes underlying tumor awakening, we analyzed late relapse patients and longitudinally profiled a rare cohort treated with long-term neoadjuvant ETs until progression. Next, we developed an in vitro evolutionary study to record the adaptive strategies of individual lineages in unperturbed parallel experiments. Our data demonstrate that ETs induce nongenetic cell state transitions into dormancy in a stochastic subset of cells via epigenetic reprogramming. Single lineages with divergent phenotypes awaken unpredictably in the absence of recurrent genetic alterations. Targeting the dormant epigenome shows promising activity against adapting cancer cells. Overall, this study uncovers the contribution of epigenetic adaptation to the evolution of resistance to ETs. Significance: This study advances the understanding of therapy-induced dormancy with potential clinical implications for breast cancer. Estrogen receptor-positive breast cancer cells adapt to endocrine treatment by entering a dormant state characterized by strong heterochromatinization with no recurrent genetic changes. Targeting the epigenetic rewiring impairs the adaptation of cancer cells to ETs. See related commentary by Llinas-Bertran et al., p. 704. This article is featured in Selected Articles from This Issue, p. 695

The clinical relevance of circulating tumor cell clusters (CTC-clusters) in breast cancer (BC) has been mostly studied using the CellSearch^®, a marker-dependent method detecting only epithelial-enriched clusters. However, due to epithelial-to-mesenchymal transition, resorting to marker-independent approaches can improve CTC-cluster detection. Blood samples collected from healthy donors and spiked-in with tumor mammospheres, or from BC patients, were processed for CTC-cluster detection with 3 technologies: CellSearch^®, CellSieve™ filters, and ScreenCell^® filters. In spiked-in samples, the 3 technologies showed similar recovery capability, whereas, in 19 clinical samples processed in parallel with CellSearch^® and CellSieve™ filters, filtration allowed us to detect more CTC-clusters than CellSearch^® (median number = 7 versus 1, p = 0.0038). Next, samples from 37 early BC (EBC) and 23 metastatic BC (MBC) patients were processed using ScreenCell^® filters for attaining both unbiased enrichment and marker-independent identification (based on cytomorphological criteria). At baseline, CTC-clusters were detected in 70% of EBC cases and in 20% of MBC patients (median number = 2, range 0–20, versus 0, range 0–15, p = 0.0015). Marker-independent approaches for CTC-cluster assessment improve detection and show that CTC-clusters are more frequent in EBC than in MBC patients, a novel finding suggesting that dissemination of CTC-clusters is an early event in BC natural history.

Multiple myeloma (MM) cells consume huge amounts of glutamine and, as a consequence, the amino acid concentration is lower-than-normal in the bone marrow (BM) of MM patients. Here we show that MM-dependent glutamine depletion induces glutamine synthetase in stromal cells, as demonstrated in BM biopsies of MM patients, and reproduced in vitro by co-culturing human mesenchymal stromal cells (MSCs) with MM cells. Moreover, glutamine depletion hinders osteoblast differentiation of MSCs, which is also severely blunted by the spent, low-glutamine medium of MM cells, and rescued by glutamine restitution. Glutaminase and the concentrative glutamine transporter SNAT2 are induced during osteoblastogenesis in vivo and in vitro, and both needed for MSCs differentiation, pointing to enhanced the requirement for the amino acid. Osteoblastogenesis also triggers the induction of glutamine-dependent asparagine synthetase (ASNS), and, among non-essential amino acids, asparagine rescues differentiation of glutamine-starved MSCs, by restoring the transcriptional profiles of differentiating MSCs altered by glutamine starvation. Thus, reduced asparagine availability provides a mechanistic link between MM-dependent Gln depletion in BM and impairment of osteoblast differentiation. Inhibition of Gln metabolism in MM cells and supplementation of asparagine to stromal cells may, therefore, constitute novel approaches to prevent osteolytic lesions in MM.