Showing papers by "Matilde Todaro published in 2022"

Effective targeting of breast cancer stem cells by combined inhibition of Sam68 and Rad51

Abstract: Breast cancer (BC) is the second cause of cancer-related deceases in the worldwide female population. Despite the successful treatment advances, 25% of BC develops resistance to current therapeutic regimens, thereby remaining a major hurdle for patient management. Current therapies, targeting the molecular events underpinning the adaptive resistance, still require effort to improve BC treatment. Using BC sphere cells (BCSphCs) as a model, here we showed that BC stem-like cells express high levels of Myc, which requires the presence of the multifunctional DNA/RNA binding protein Sam68 for the DNA-damage repair. Analysis of a cohort of BC patients displayed that Sam68 is an independent negative factor correlated with the progression of the disease. Genetic inhibition of Sam68 caused a defect in PARP-induced PAR chain synthesis upon DNA-damaging insults, resulting in cell death of TNBC cells. In contrast, BC stem-like cells were able to survive due to an upregulation of Rad51. Importantly, the inhibition of Rad51 showed synthetic lethal effect with the silencing of Sam68, hampering the cell viability of patient-derived BCSphCs and stabilizing the growth of tumor xenografts, including those TNBC carrying BRCA mutation. Moreover, the analysis of Myc, Sam68 and Rad51 expression demarcated a signature of a poor outcome in a large cohort of BC patients. Thus, our findings suggest the importance of targeting Sam68-PARP1 axis and Rad51 as potential therapeutic candidates to counteract the expansion of BC cells with an aggressive phenotype.

Dual Inhibition of Myc Transcription and PI3K Activity Effectively Targets Colorectal Cancer Stem Cells

Abstract: Simple Summary Compelling evidence has shown that cancer stem cells (CSCs) are responsible for high resistance to conventional anti-cancer therapies. Here, we demonstrate that the tumor microenvironment protects CR-CSCs from EGFR/HER2, BRAF and PI3K targeting, promoting CD44v6 and Myc expression. Alternatively, as a substitution for HER2 and BRAF, the Myc transcription inhibitor can overcome the protective effects of microenvironmental cytokines, impairing the survival of CR-CSCs. These data highlight the targeting of Myc and PI3K activity as a novel therapeutic strategy against advanced colorectal cancer. Abstract Despite advances in the curative approach, the survival rate of advanced colorectal cancer (CRC) patients is still poor, which is likely due to the emergence of cancer cell clones resistant to the available therapeutic options. We have already shown that CD44v6-positive CRC stem cells (CR-CSCs) are refractory toward standard anti-tumor therapeutic agents due to the activation of the PI3K pathway together with high HER2 expression levels. Tumor microenvironmental cytokines confer resistance to CR-CSCs against HER2/PI3K targeting by enhancing activation of the MAPK pathway. Here, we show that the CSC compartment, spared by BRAF inhibitor-based targeted therapy, is associated with increased expression levels of CD44v6 and Myc and retains boosted clonogenic activity along with residual tumorigenic potential. Inhibition of Myc transcription, downstream of the MAPK cascade components, and PI3K pathway activity was able to overcome the protective effects of microenvironmental cytokines, affecting the survival and the clonogenic activity of CR-CSCs, regardless of their mutational background. Likewise, the double targeting induced stabilization of mouse tumor avatars. Altogether, these data outline the rationale for dual kinase targeting of CR-CSCs to prevent their adaptive response, which would lead to disease progression.

Targeting of the Peritumoral Adipose Tissue Microenvironment as an Innovative Antitumor Therapeutic Strategy

Abstract: The tumor microenvironment (TME) plays a key role in promoting and sustaining cancer growth. Adipose tissue (AT), due to its anatomical distribution, is a prevalent component of TME, and contributes to cancer development and progression. Cancer-associated adipocytes (CAAs), reprogrammed by cancer stem cells (CSCs), drive cancer progression by releasing metabolites and inflammatory adipokines. In this review, we highlight the mechanisms underlying the bidirectional crosstalk among CAAs, CSCs, and stromal cells. Moreover, we focus on the recent advances in the therapeutic targeting of adipocyte-released factors as an innovative strategy to counteract cancer progression.

Cancer cell targeting by CAR-T cells: A matter of stemness

Abstract: Chimeric antigen receptor (CAR)-T cell therapy represents one of the most innovative immunotherapy approaches. The encouraging results achieved by CAR-T cell therapy in hematological disorders paved the way for the employment of CAR engineered T cells in different types of solid tumors. This adoptive cell therapy represents a selective and efficacious approach to eradicate tumors through the recognition of tumor-associated antigens (TAAs). Binding of engineered CAR-T cells to TAAs provokes the release of several cytokines, granzyme, and perforin that ultimately lead to cancer cells elimination and patient’s immune system boosting. Within the tumor mass a subpopulation of cancer cells, known as cancer stem cells (CSCs), plays a crucial role in drug resistance, tumor progression, and metastasis. CAR-T cell therapy has indeed been exploited to target CSCs specific antigens as an effective strategy for tumor heterogeneity disruption. Nevertheless, a barrier to the efficacy of CAR-T cell-based therapy is represented by the poor persistence of CAR-T cells into the hostile milieu of the CSCs niche, the development of resistance to single targeting antigen, changes in tumor and T cell metabolism, and the onset of severe adverse effects. CSCs resistance is corroborated by the presence of an immunosuppressive tumor microenvironment (TME), which includes stromal cells, cancer-associated fibroblasts (CAFs), tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), and immune cells. The relationship between TME components and CSCs dampens the efficacy of CAR-T cell therapy. To overcome this challenge, the double strategy based on the use of CAR-T cell therapy in combination with chemotherapy could be crucial to evade immunosuppressive TME. Here, we summarize challenges and limitations of CAR-T cell therapy targeting CSCs, with particular emphasis on the role of TME and T cell metabolic demands.

Lipid Droplets Fuel Small Extracellular Vesicle Biogenesis

Abstract: Despite an increasing gain of knowledge regarding small extracellular vesicle (sEV) composition and functions in cell-cell communication, the mechanism behind their biogenesis remains unclear. Here, we revealed for the first time that the sEV biogenesis and release into the microenvironment are tightly connected with another important organelle: Lipid Droplets (LD). We have observed this correlation using different human cancer cell lines as well as patient-derived colorectal cancer stem cells (CR-CSCs). Our results showed that the use of external stimuli such as radiation, pH, hypoxia, or lipid interfering drugs, known to affect the LD content, had a similar effect in terms of sEV secretion. Additional validations were brought using multiple omics data, at the mRNA and protein levels. Altogether, the possibility to fine-tune sEV biogenesis by targeting LDs, could have a massive impact on the amount, the cargos and the properties of those sEVs, paving the way for new clinical perspectives. Significance Statement

1430 The immunological and genomic profiling of colorectal and breast cancer cell lines can distinguish stem like from differentiated tumor cells: implications for cancer immunotherapy

Abstract:

Background

Rare cells endowed with “stemness” and tumor initiating properties, cancer stem cells (CSCs), are considered responsible of metastatization and resistance to therapy, including immunotherapy. The aim of this study is to identify the molecular mechanisms regulating the immunological properties of CSCs isolated from solid tumors.

Methods

CRC (N=15) and BC (N=21) cell lines), including differentiated tumor cells and CSCs, and, for BC cell lines, selected in vitro for radioresistance or invasiveness were used for this study. The expression of HLA molecules and the components involved in the antigen processing machinery (APM) was assessed through flow cytometry. DNA and RNA were isolated from these cell lines. The nCounter platform (Nanostring) was utilized to assess the hybridization with 800 probes for miRNAs and the RNA seq-based transcriptomic profile was also assessed. The methylation profiling of cancer cell lines was investigated through Infinium EPIC arrays (Illumina). In addition, the co-culture of tumor cells with HLA-matched lymphocytes, either with or without the treatment with immunomodulatory or epigenetic agents, was performed to assess the ability of the CRC and BC cells to elicit antigen-specific T cell responses.

Results

A general impairment of the expression of HLA and APM (e.g., LMPs, TAP and tapasin) molecules was observed in CRC and BC lines. The down-modulation of the expression of HLA and APM was superior in “stem-like” cells as compared to the differentiated tumor cells. The treatment of these cells with immunomodulating or epigenetic agents could only partially restore the expression of these molecules. The induction in vitro of anti-tumor T cell responses through the co-culture of tumor cells with PBMCs, was suboptimal and with variability depending on the subtype of cells and the pre-treatment or not of tumor cells with either immunomodulating or epigenetic agents. Differential miRNAs, transcriptomic and methylation profiles (p<0.05) were identified in either CRC or BC cells with stemness properties vs. differentiated cells, and in different subtypes of cells. These includes genes and their regulators involved in immunological functions. The integration analyses among methylation, miRNAs and transcriptomic profiles is under investigations, although preliminary results are available showing the differential involvement in CSCs vs. differentiated tumor cells of immune related pathways.

Conclusions

The immunological and genomic profiles of CRC and BC are associated with cell subtypes. These investigations will contribute to understand the mechanisms regulating the biological and immunological properties of tumor cell endowed with variable cell fate and their susceptibility to immunotherapy.

Acknowledgements

This study was funded by QNRF; grant # NPRP10-0129-170277

Ethics Approval

The study obtained ethics approval from both Sidra Medicine #1805024172, and Hamad Medical Corporation #MRC-03-17-150 institutional review boards; participants gave informed consent before taking part to the study.

Destroying the Shield of Cancer Stem Cells: Natural Compounds as Promising Players in Cancer Therapy

Abstract: In a scenario where eco-sustainability and a reduction in chemotherapeutic drug waste are certainly a prerogative to safeguard the biosphere, the use of natural products (NPs) represents an alternative therapeutic approach to counteract cancer diseases. The presence of a heterogeneous cancer stem cell (CSC) population within a tumor bulk is related to disease recurrence and therapy resistance. For this reason, CSC targeting presents a promising strategy for hampering cancer recurrence. Increasing evidence shows that NPs can inhibit crucial signaling pathways involved in the maintenance of CSC stemness and sensitize CSCs to standard chemotherapeutic treatments. Moreover, their limited toxicity and low costs for large-scale production could accelerate the use of NPs in clinical settings. In this review, we will summarize the most relevant studies regarding the effects of NPs derived from major natural sources, e.g., food, botanical, and marine species, on CSCs, elucidating their use in pre-clinical and clinical studies.