Helicobacter pylori exploits host glycoconjugates to colonize the gastric niche. Infection can persist for decades promoting chronic inflammation, and in a subset of individuals lesions can silently progress to cancer. This study shows that H. pylori chronic infection and gastric tissue inflammation result in a remodeling of the gastric glycophenotype with increased expression of sialyl-Lewis a/x antigens due to transcriptional up-regulation of the B3GNT5, B3GALT5, and FUT3 genes. We observed that H. pylori infected individuals present a marked gastric local proinflammatory signature with significantly higher TNF-α levels and demonstrated that TNF-induced activation of the NF-kappaB pathway results in B3GNT5 transcriptional up-regulation. Furthermore, we show that this gastric glycosylation shift, characterized by increased sialylation patterns, favors SabA-mediated H. pylori attachment to human inflamed gastric mucosa. This study provides novel clinically relevant insights into the regulatory mechanisms underlying H. pylori modulation of host glycosylation machinery, and phenotypic alterations crucial for life-long infection. Moreover, the biosynthetic pathways here identified as responsible for gastric mucosa increased sialylation, in response to H. pylori infection, can be exploited as drug targets for hindering bacteria adhesion and counteract the infection chronicity.

Liquid biopsy represents the analysis of tumor-derived material in the blood and other body fluids of cancer patients. This portrays a minimally invasive detection tool for molecular biomarkers. Liquid biopsy has emerged as a complementary or alternative method to surgical biopsy. This non-invasive detection tool overcomes the recurrent problems in the clinical assessment of tumors that stem from the lack of accessibility to the tumor tissue and its clonal heterogeneity. Moreover, body fluid-derived components have shown to reflect the genetic profile of both primary and metastatic lesions and provide a real-time monitoring of tumor dynamics, representing a great promise for personalized medicine. This review will highlight the latest breakthroughs and the current applications of several tumor-derived biomarkers that can be found in body fluids. The authors will focus on tumor-derived exosomes, tumor-educated platelets, and circulating tumor miRNAs and mRNAs, and how these can be used for tumor detection.

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Background and Purpose: Advances in the treatment of triple-negative breast and ovarian cancer remain challenging. In particular, resistance to the available therapy, by restoring or overexpressing the DNA repair machinery, has often been reported. New strategies to improve the therapeutic outcomes of these cancers are needed.Herein, we disclose the dregamine 5-bromo-pyridin-2-ylhydrazone (BBIT20), a natural monoterpene indole alkaloid derivative, as an inhibitor of homologous DNA repair.Experimental Approach: To unveil BBIT20 antitumour activity and underlying molecular mechanism of action, two-dimensional (2D) and three-dimensional (3D) cell cultures, patient-derived cell lines and xenograft mouse models were used.Key Results: BBIT20 disrupted the BRCA1-BARD1 interaction, triggering nuclearto-cytoplasmic BRCA1 translocation, cell cycle arrest and downregulation of homologous DNA repair-related genes and proteins, with subsequent enhancement of DNA damage, reactive oxygen species generation and apoptosis, in triple-negative breast and ovarian cancer cells. BBIT20 also displayed pronounced antitumour activity in patient-derived cells and xenograft mouse models of ovarian cancer, with low toxicity in non-malignant cells and undetectable side effects

Deregulated expression of histone deacetylases (HDACs) has been implicated in tumorigenesis. Herein, we investigated class I HDACs expression in bladder urothelial cell carcinoma (BUCC), its prognostic value and biological significance. Significantly increased transcript levels of all HDACs were found in BUCC compared to 20 normal mucosas, and these were higher in lower grade and stage tumors. Increased HDAC3 levels were associated with improved patient survival. SiRNA experiments showed decrease cell viability and motility, and increased apoptosis. We concluded that class I HDACs play an important role in bladder carcinogenesis through deregulation of proliferation, migration and apoptosis, constituting putative therapeutic targets.

Background: The study of cell-free DNA (cfDNA), namely the fraction derived from tumors (ctDNA), is a clinically relevant noninvasive biomarker for cancer management. However, the intrinsic low abundance of ctDNA in plasma limits its implementation in the clinic. Aim of the study: In this study, the objective was to demonstrate that induction of apoptosisdthe major source of ctDNAdincreases ctDNA concentration, thereby increasing the sensitivity to detect clinically relevant mutations in plasma. Methods: In vitro models were used to test the effect of docetaxel on the release levels of DNA from lung cancer cells. In vivo, Rag2 À/À IL2rg À/À immunodeficient C57BL/6 xenografted mice were treated with docetaxel for 24 h or 48 h. Tumor tissue and blood were collected to evaluate the levels of apoptosis DNA release levels, respectively. Results: We observed increased levels of apoptosis in H1975 cells and a consequent increase in cfDNA released into the culture medium after docetaxel treatment. In vivo, the results show increased cfDNA concentration in plasma of xenografted mice after apoptosis stimulation. Importantly, treatment increased the sensitivity of detection of relevant cancer mutations, namely 24 h after treatment. Conclusion:This study provides new insights regarding the importance of timing for blood collection. In our experimental model, we demonstrate that blood collection should be performed 24 h after treatment (apoptosis induction), for optimal ctDNA analysis. Translating these results into

Resistance to treatment is a major clinical problem and a major cause of cancer-related deaths. Understanding the biological basis of resistance acquisition is of utmost importance to improve the clinical management of cancer patients. NGS analysis of human lung cancer (LC) tumors from patients that relapsed after treatment with EGFR-tyrosine kinase inhibitors (TKI), revealed that the p.T790M resistance mutation is not present in all the relapsing tumor cells, suggesting that LC cells can become resistant even if not carrying the p.T790M mutation. Using in vitro treatments with conditioned medium (CM) and in vivo co-inoculation experiments, we show that LC cells sensitive to EGFR-TKIs (S cells) acquire resistance faster when treated with CM from LC cells resistant to EGFR-TKIs (R cells) or when co-inoculated with R cells in opposite flanks of the same animal. Importantly, we show that acquisition of resistance is not due to the emergence of subpopulations of cancer cells with new resistance mutations. Using transcriptomics, we show that acquisition of resistance is associated with upregulation of genes involved in endocytosis, namely caveolins CAV1 and CAV2. These findings were validated in human clinical samples, where an increase in CAV1 and CAV2 expression was associated with tumor relapse after treatment with EGFR-TKIs. Our results suggest that acquisition of resistance to targeted therapies results from the combined effect of selection of cells harboring specific resistance mutations and horizontal transfer of the resistance phenotype. These findings may pave the way to bring intercellular communication into the realm of cancer treatment.

Targeted therapies almost universally fail due to the development of resistance. EGFR-mutant non-small cell lung cancer (NSCLC) is a well characterized model of such mechanism. In this model, treatment with anti-EGFR therapy leads to resistance driven by the presence of the EGFR T790M mutation in about 50% of the cases. However, clinical data shows the EGFR resistance mutation is present only in a subpopulation of the relapsing cells. How cancer cells devoid of the EGFR T790M mutation become resistant is not known. The aim of this study is to explore if T790M-mediated resistance to anti-EGFR therapy can spread horizontally, through a mitosis-independent mechanism, among NSCLC cells. Immunodeficient mice (Rag2-/-Il2rg-/-) were engrafted with NSCLC HCC827 cells, harboring the Erlotinib-sensitive mutation EGFR E746-A750del, and H1975 cells, carrying the EGFR T790M Erlotinib-resistant mutation. These animals received a single inoculation of HCC827 cells in a single flank (n=10), or dual inoculation of HCC827 and H1975 cells in opposite flanks (n=20). Erlotinib was orally administered three times a week (25 mg/Kg) and tumors were collected at humane endpoint for genetic and transcriptomic analysis by next generation sequencing and digital PCR. T790M-driven resistance to Erlotinib was transferred in vivo from resistant to sensitive cells in dual inoculated animals, resulting in a significant decrease of relapse-free survival. Short tandem repeat profiling of the tumors showed absence of H1975 cells in sensitive tumors in dual inoculated animals, ruling out cell migration as the mechanism of resistance. This result can also not be explained by transfer of mutated DNA from resistant to sensitive cells because high-end sequencing detected the presence of the T790M mutation only in few sensitive tumors from dual inoculated mice and with an extremely low allele frequency (<0,1%). These results support our contention that anti-EGFR therapy resistance does not imply the presence of the T790M mutation in every resistant cell. RNA sequencing revealed differences in transcriptomic profiles from sensitive tumors between single and dual inoculated mice. Gene ontology analysis identified several pathways, such as endocytosis, lysosome biogenesis and RNA transport, as segregators of sensitive tumors that relapsed in the presence or absence of resistant tumors. Our findings suggest that mutation-driven resistance to anti-EGFR can spread horizontally among lung cancer cells and results in alteration of the transcriptional landscape of sensitive cells. Our results question the established concept that acquisition of resistance to targeted therapies relies solely on the selection of tumor cells carrying a specific resistance mutation, which is then passed on to its progeny. PhD Scholarship: SFRH/BD/115099/2016; FEDER funds: COMPETE 2020—(POCI), Portugal 2020; FCT: PTDC/DTP-PIC/2500/2014, NORTE 2020: NORTE- 01-0145-FEDER-000029. Citation Format: Susana Junqueira-Neto, Ana Rita Oliveira, Joana Marques, Sónia Melo, José Luís Costa, José Carlos Machado. Resistance to anti-EGFR therapy can spread horizontally among cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1099.