Identification of Markers to Predict Benefit from Trastuzumab Treatment

Despite the clinical benefit of trastuzumab, some patients do not respond to this therapy. Aims of this study are to identify new predictive biomarkers to distinguish responsive from de novo resistant tumours and to have new insights into the biological characterisation of trastuzumab activity. Whole-transcriptome analysis of primary HER2-positive breast carcinomas (BCs) treated with adjuvant trastuzumab identified a tumour subgroup, characterised by high ERBB2/low ESR1 expression, with good clinical outcome, and allowed the development of a trastuzumab risk model (TRAR) able to identify patients with high- and low-risk of relapse. Application of TRAR model to available datasets and to a new series of HER2-positive BC patients treated with neoadjuvant trastuzumab designated TRAR as predictive of response rather than associated to low aggressiveness. Our analyses showed that tumours exquisitely sensitive to treatment are addicted to HER2, enriched in immune pathways and have a peculiar circulating NK profile, characterised by high expression of the NKG2D receptor. Enrichment of immune system- and tyrosine kinase receptor signaling-related pathways was found associated also with response to trastuzumab monotherapy in clinical samples, suggesting the possibility to treat HER2-addicted tumours with trastuzumab monotherapy. Accordingly TRAR-low, high-NKG2D- and -MHC-II-expressing tumours were associated with response to one cycle of trastuzumab alone. In addition, increase of MHC-II gene expression upon a single cycle of trastuzumab characterised patients who benefit from the following combination with chemotherapy. Characterisation of biological features of TRAR-low tumours showed high infiltration of macrophages and CD8+ T cells, together with the expression of chemokines involved in their recruitment and of immune checkpoint ligands. In vitro analysis demonstrated a direct regulation of CCL2 and PD-L1 by HER2 signals. Overall, we described a tool able to identify BCs responsive to trastuzumab and understood that in these tumours HER2 is crucial for tumour growth, for infiltration of pro-trastuzumab immune cells and their suppression

Breast cancer-secreted miR-939 downregulates VE-cadherin and destroys the barrier function of endothelial monolayers.

Abstract Exosomes-secreted microRNAs play an important role in metastatic spread. During this process breast cancer cells acquire the ability to transmigrate through blood vessels by inducing changes in the endothelial barrier. We focused on miR-939 that is predicted to target VE-cadherin, a component of adherens junction involved in vessel permeability. By in silico analysis miR-939 was found highly expressed in the basal-like tumor subtypes and in our cohort of 63 triple-negative breast cancers (TNBCs) its expression significantly interacted with lymph node status in predicting disease-free survival probability. We demonstrated, in vitro , that miR-939 directly targets VE-cadherin leading to an increase in HUVECs monolayer permeability. MDA-MB-231 cells transfected with a miR-939 mimic, released miR-939 in exosomes that, once internalized in endothelial cells, favored trans-endothelial migration of MDA-MB-231-GFP cells by the disruption of the endothelial barrier. Notably, when up taken in endothelial cells exosomes caused VE-cadherin down-regulation specifically through miR-939 as we demonstrated by inhibiting miR-939 expression in exosomes-releasing TNBC cells. Together, our data indentify an extracellular pro-tumorigenic role for tumor-derived, exosome-associated miR-939 that can explain its association with worse prognosis in TNBCs

Cancer acidity: An ultimate frontier of tumor immune escape and a novel target of immunomodulation

The link between cancer metabolism and immunosuppression, inflammation and immune escape has generated major interest in investigating the effects of low pH on tumor immunity. Indeed, microenvironmental acidity may differentially impact on diverse components of tumor immune surveillance, eventually contributing to immune escape and cancer progression. Although the molecular pathways underlying acidity-related immune dysfunctions are just emerging, initial evidence indicates that antitumor effectors such as T and NK cells tend to lose their function and undergo a state of mostly reversible anergy followed by apoptosis, when exposed to low pH environment. At opposite, immunosuppressive components such as myeloid cells and regulatory T cells are engaged by tumor acidity to sustain tumor growth while blocking antitumor immune responses. Local acidity could also profoundly influence bioactivity and distribution of antibodies, thus potentially interfering with the clinical efficacy of therapeutic antibodies including immune checkpoint inhibitors. Hence tumor acidity is a central regulator of cancer immunity that orchestrates both local and systemic immunosuppression and that may offer a broad panel of therapeutic targets. This review outlines the fundamental pathways of acidity-driven immune dysfunctions and sheds light on the potential strategies that could be envisaged to potentiate immune-mediated tumor control in cancer patients

Taxanes enhance trastuzumab-mediated ADCC on tumor cells through NKG2D-mediated NK cell recognition

Recent clinical data indicate a synergistic therapeutic effect between trastuzumab and taxanes in neoadjuvantly treated HER2-positive breast cancer (BC) patients. In HER2+ BC experimental models and patients, we investigated whether this synergy depends on the ability of drug-induced stress to improve NK cell effectiveness and thus trastuzumab-mediated ADCC. HER2+ BC cell lines BT474 and MDAMB361 treated with docetaxel showed up-modulation of NK activator ligands both in vitro and in vivo, accompanied by a 15-40% increase in in vitro trastuzumab-mediated ADCC; antibodies blocking the NKG2D receptor significantly reduced this enhancement. NKG2D receptor expression was increased by docetaxel treatment in circulating and splenic NK cells from mice xenografted with tumor cells, an increase related to expansion of the CD11b+Ly6G+ cell population. Accordingly, NK cells derived from HER2+ BC patients after treatment with taxane-containing therapy expressed higher levels of NKG2D receptor than before treatment. Moreover, plasma obtained from these patients recapitulated the modulation of NKG2D on healthy donors' NK cells, improving their trastuzumab-mediated activity in vitro. This enhancement occurred mainly using plasma from patients with low NKG2D basal expression. Our results indicate that taxanes increase tumor susceptibility to ADCC by acting on tumor and NK cells, and suggest that taxanes concomitantly administered with trastuzumab could maximize the antibody effect, especially in patients with low basal immune effector cytotoxic activit

Early immune modulation by single-agent trastuzumab as a marker of trastuzumab benefit

BACKGROUND: Optimising the selection of HER2-targeted regimens by identifying subsets of HER2-positive breast cancer (BC) patients who need more or less therapy remains challenging. We analysed BC samples before and after treatment with 1 cycle of trastuzumab according to the response to trastuzumab. METHODS: Gene expression profiles of pre- and post-treatment tumour samples from 17 HER2-positive BC patients were analysed on the Illumina platform. Tumour-associated immune pathways and blood counts were analysed with regard to the response to trastuzumab. HER2-positive murine models with differential responses to trastuzumab were used to reproduce and better characterise these data. RESULTS: Patients who responded to single-agent trastuzumab had basal tumour biopsies that were enriched in immune pathways, particularly the MHC-II metagene. One cycle of trastuzumab modulated the expression levels of MHC-II genes, which increased in patients who had a complete response on treatment with trastuzumab and chemotherapy. Trastuzumab increased the MHC-II-positive cell population, primarily macrophages, only in the tumour microenvironment of responsive mice. In patients who benefited from complete trastuzumab therapy and in mice that harboured responsive tumours circulating neutrophil levels declined, but this cell subset rose in nonresponsive tumours. CONCLUSIONS: Short treatment with trastuzumab induces local and systemic immunomodulation that is associated with clinical outcomes

CSF β-amyloid predicts prognosis in patients with multiple sclerosis

Background: The importance of predicting disease progression in multiple sclerosis (MS) has increasingly been recognised, hence reliable biomarkers are needed. Objectives: To investigate the prognostic role of cerebrospinal fluid (CSF) Amyloid beta1-42 (A) levels by the determination of a cut-off value to classify patients in slow and fast progressors. To evaluate possible association with white (WM) and grey matter (GM) damage at early disease stages. Methods: Sixty patients were recruited and followed-up for three to five years. Patients underwent clinical assessment, CSF analysis to determine Aβ levels, and brain MRI (at baseline and after 1 year). T1-weighted volumes were calculated. T2-weighted scans were used to quantify WM lesion loads. Results: Lower CSF Aβ levels were observed in patients with a worse follow-up EDSS (r=−0.65, p0.05). Conclusions: Low CSF Aβ levels may represent a predictive biomarker of disease progression in MS

Whole-transcriptome analysis links trastuzumab sensitivity of breast tumors to both HER2 dependence and immune cell infiltration.

While results thus far demonstrate the clinical benefit of trastuzumab, some patients do not respond to this therapy. To identify a molecular predictor of trastuzumab benefit, we conducted whole-transcriptome analysis of primary HER2+ breast carcinomas obtained from patients treated with trastuzumab-containing therapies and correlated the molecular portrait with treatment benefit. The estimated association between gene expression and relapse-free survival allowed development of a trastuzumab risk model (TRAR), with ERBB2 and ESR1 expression as core elements, able to identify patients with high and low risk of relapse. Application of the TRAR model to 24 HER2+ core biopsies from patients treated with neo-adjuvant trastuzumab indicated that it is predictive of trastuzumab response. Examination of TRAR in available whole-transcriptome datasets indicated that this model stratifies patients according to response to trastuzumab-based neo-adjuvant treatment but not to chemotherapy alone. Pathway analysis revealed that TRAR-low tumors expressed genes of the immune response, with higher numbers of CD8-positive cells detected immunohistochemically compared to TRAR-high tumors. The TRAR model identifies tumors that benefit from trastuzumab-based treatment as those most enriched in CD8-positive immune infiltrating cells and with high ERBB2 and low ESR1 mRNA levels, indicating the requirement for both features in achieving trastuzumab response

CSF β-amyloid and white matter damage: a new perspective on Alzheimer's disease

Objective: To assess the connection between amyloid pathology and white matter (WM) macro- and micro-structural damage in demented patients compared with controls. Methods: Eighty-five participants were recruited: 65 with newly diagnosed Alzheimer’s disease (AD), non-AD dementia or mild cognitive impairment (MCI), and 20 age- and sex-matched heatlhy controls. β-amyloid1-42 (Aβ) levels were determined in cerebrospinal fluid (CSF) samples from all patients and 5 controls. Among patients, 42 had pathological CSF Aβ levels (Aβ+), while 23 had normal CSF Aβ levels (Aβ-). All participants underwent neurological examination, neuropsychological testing and brain magnetic resonance imaging (MRI). We used T2-weighted scans to quantify white matter (WM) lesion loads (LL), and diffusion weighted images (DWI) to assess their microstructural substrate. Non-parametric statistical tests were used for between-group comparisons and multiple regression analyses. Results: We found an increased WM-LL in Aβ(+) compared to both, healthy controls (p=0.003) and Aβ(-) patients (p=0.02). Interestingly, CSF Aβ concentration was the best predictor patients’ WM-LL (r=-0.30, p<0.05) when using age as a covariate. Lesion apparent diffusion coefficient (ADC) value was higher in all patients than in controls (p=0.0001), and correlated with WM-LL (r=0.41, p=0.001). In Aβ(+), WM-LL correlated with WM microstructural damage in the left peritrigonal WM (p<0.0001). Conclusions: WM damage is crucial in Alzheimer’s disease (AD) pathogenesis. The correlation between CSF Aβ levels and WM-LL suggests a direct link between amyloid pathology and WM macro- and microstructural damage

Antibiotic-induced disturbances of the gut microbiota result in accelerated breast tumor growth

The gut microbiota's function in regulating health has seen it linked to disease progression in several cancers. However, there is limited research detailing its influence in breast cancer (BrCa). This study found that antibiotic-induced perturbation of the gut microbiota significantly increases tumor progression in multiple BrCa mouse models. Metagenomics highlights the common loss of several bacterial species following antibiotic administration. One such bacteria, Faecalibaculum rodentium, rescued this increased tumor growth. Single-cell transcriptomics identified an increased number of cells with a stromal signature in tumors, and subsequent histology revealed an increased abundance of mast cells in the tumor stromal regions. We show that administration of a mast cell stabilizer, cromolyn, rescues increased tumor growth in antibiotic treated animals but has no influence on tumors from control cohorts. These findings highlight that BrCa-microbiota interactions are different from other cancers studied to date and suggest new research avenues for therapy development