The immune system is a dynamic feature of each individual and a footprint of our unique internal and external exposures. Indeed, the type and level of exposure to physical and biological agents shape the development and behavior of this complex and diffuse system. Many pathological conditions depend on how our immune system responds or does not respond to a pathogen or a disease or on how the regulation of immunity is altered by the disease itself. T-cells are important players in adaptive immunity and, together with B-cells, define specificity and monitor the internal and external signals that our organism perceives through its specific receptors, TCRs and BCRs, respectively. Today, high-throughput sequencing (HTS) applied to the TCR repertoire has opened a window of opportunity to disclose T-cell repertoire development and behavior down to the clonal level. Although TCR repertoire sequencing is easily accessible today, it is important to deeply understand the available technologies for choosing the best fit for the specific experimental needs and questions. Here, we provide an updated overview of TCR repertoire sequencing strategies, providers and applications to infectious diseases and cancer to guide researchers’ choice through the multitude of available options. The possibility of extending the TCR repertoire to HLA characterization will be of pivotal importance in the near future to understand how specific HLA genes shape T-cell responses in different pathological contexts and will add a level of comprehension that was unthinkable just a few years ago.

Circulating tumor cells (CTCs) are a rare population of cells representing a key player in the metastatic cascade. They are recognized as a validated tool for the identification of patients with a higher risk of relapse, including those diagnosed with breast cancer (BC). However, CTCs are characterized by high levels of heterogeneity that also involve copy number alterations (CNAs), structural variations associated with gene dosage changes. In this study, single CTCs were isolated from the peripheral blood of 11 early-stage BC patients at different time points. A label-free enrichment of CTCs was performed using OncoQuick, and single CTCs were isolated using DEPArray. Libraries were prepared from single CTCs and DNA extracted from matched tumor tissues for a whole-genome low-coverage next-generation sequencing (NGS) analysis using the Ion Torrent S5 System. The analysis of the CNA burden highlighted that CTCs had different degrees of aberration based on the time point and subtype. CTCs were found even six months after surgery and shared CNAs with matched tumor tissue. Tumor-associated CNAs that were recurrent in CTCs were patient-specific, and some alterations involved regions associated with BC and survival (i.e., gains at 1q21-23 and 5p15.33). The enrichment analysis emphasized the involvement of aberrations of terms, associated in particular with interferon (IFN) signaling. Collectively, our findings reveal that these aberrations may contribute to understanding the molecular mechanisms involving CTC-related processes and their survival ability in occult niches, supporting the goal of exploiting their application in patients’ surveillance and follow-up.

CDK4/6 inhibitors (CDK4/6i) are recommended in patients with estrogen receptor (ER)-positive, HER2-negative advanced breast cancer (ABC). Up to now, no prognostic biomarkers have been identified in this setting. We retrospectively analyzed the expression of progesterone receptor (PR) and Ki67, assessed by immunohistochemistry, in 71 ABC patients treated with CDK4/6i and analyzed the impact of these markers on progression-free survival (PFS). The majority of patients 63/71 (88.7%) received palbociclib, 4 (5.6%) received ribociclib, and 4 (5.6%) received abemaciclib. A higher median value of Ki67 was observed in cases undergoing second-line treatment (p = 0.047), whereas the luminal B subtype was more prevalent (p = 0.005). In the univariate analysis of the first-line setting, luminal A subtype showed a trend towards a correlation with a longer PFS (p = 0.053). A higher continuous Ki67 value led to a significantly shorter PFS. When the interaction between pathological characteristics and line of treatment was considered, luminal B subtype showed a significantly (p = 0.043) worse outcome (Hazard Ratio (HR) 2.84; 1.03–7.82 95% Confidence Interval (CI)). PFS in patients undergoing endocrine therapy plus CDK4/6i was inversely correlated with Ki67 expression but not with PR, suggesting that tumor proliferation has a greater impact on cell cycle inhibitors combined with endocrine therapy than PR expression.

Androgen receptor (AR) is widely expressed in breast cancer (BC) but its role in estrogen receptor (ER)-positive tumors is still controversial. The AR/ER ratio has been reported to impact prognosis and response to antiestrogen endocrine therapy (ET). We assessed whether AR in primary tumors and/or matched metastases is a predictor of efficacy of first-line ET in advanced BC. Patients who had received first-line ET (2002–2011) were recruited, while those given concomitant chemotherapy or trastuzumab or pretreated with > 2 lines of chemotherapy were excluded. ER, progesterone receptor (PgR), Ki67 and AR expression were assessed by immunohistochemistry, and HER2 mainly by fluorescent in-situ hybridization. Cut-offs of 1 and 10% immunostained cells were used to categorize AR expression. Among 102 evaluable patients, biomarkers were assessed in primary tumors in 70 cases and in metastases in 49, with 17 patients having both determinations. The overall concordance rate between primary tumors and metastases was 64.7% (95% CI 42%-87.4%) for AR status. AR status did not affect TTP significantly, whereas PgR and Ki67 status did. AR/PgR ≥0.96 was associated with a significantly shorter TTP (HR = 1.65, 95% CI 1.05-2.61, p = 0.028). AR status in primary tumors or metastases was not associated with progressive disease (PD) as best response. In contrast, Ki67 ≥ 20% and PgR < 10% showed a statistically significant association with PD as best response. AR expression does not appear to be useful to predict the efficacy of ET in advanced BC, whereas Ki67 and PgR exert a greater impact on its efficacy.

Background: General anesthesia (GA) is the most commonly used anesthesiological technique for radical mastectomy operations and can be associated with loco-regional anesthesia techniques. The aim of our study, carried out on 51 patients, was to assess the effectiveness of thoracic paravertebral block (TPVB) associated with GA, or as a sole anesthesiological technique for postoperative pain control and for the reduction of intra and postoperative opioids consumption. Materials and methods: Fifty-one patients with neoplastic breast disease and elected as candidates for radical mastectomy were included in the study. The primary outcomes for this study were intra and postoperative opioid consumption and postoperative pain intensity. In 37 patients, TPVB was associated with GA while in 14 patients it was used as the sole anesthesiological technique. Data are reported as mean with standard deviation median with interquartile range, number, and percentage, depending on the underlying distribution. Results: We did not use intra or postoperative opioids for any patient and the Numeric Rate Scale, assessed at time 0, at the end of the surgery, and 2, 6, 12, and 24 hrs after surgery, was >3 in seven patients only. Conclusions: This study aims to show how TPVB can be used to carry out radical mastectomy procedures so that intra and postoperative opioids use can be avoided. In our study, TPVB was used in total mastectomy procedures in association with GA or as the sole anesthesiological technique, without the intra and postoperative use of opioids and with a significant reduction of local anesthetic dosages compared to those reported in the existing literature.

Breast cancer (BC) is the most common diagnosed cancer and the second leading cause of cancer-associated death in women, with the triple negative (TNBC) subtype being characterized by the poorest p...