Translation into Russian:  Tomasetti, C. , Li, L. , & Vogelstein, B.  (2017).  Stem cell divisions, somatic mutations, cancer etiology, and cancer prevention . Science ,  355 (6331), 1330-1334. DOI:  10.1126/science.aaf9011

Stem cell divisions, somatic mutations, cancer etiology, and cancer prevention

ABSTRACT Introduction Esophageal cancer (EC) is a worldwide healthcare concern and represents an aggressive malignancy. Squamous cell carcinoma (ESCC) and adenocarcinoma (EAC) are the two primary histological subtypes but have yet to vastly differ in management. Outcomes remain poor with current treatment approaches; however, recent progress is focused on distinguishing separate targets based on thistology. Areas covered Here we provide an overview of EC management via a historical review and recent discoveries. As noted in this review, targeted therapy has lagged behind other solid tumors. Over the previous decade, for EACs there were only two targeted therapies used in the advanced setting with limited benefits. ESCC progress was rather non-existent. We present current ongoing advancements that have occurred in the realm of immunotherapy and emerging new agents. Expert opinion It is becoming clearer that segregating these two histological subtypes in trials should be the goal of future trial designs. ESCC appears to be more amenable to immune modulation than EAC; however, we are navigating in exciting times as molecular interrogations of EC has expanded with the hope of making more rapid progress. There is still hard work ahead of us to painfully define subsets representing heterogeneity and then finding appropriate agents.

Esophageal cancer: emerging therapeutics

Esophageal cancer is a lethal malignancy with a high mortality rate, while the molecular mechanisms underlying esophageal cancer pathogenesis are still poorly understood. Here, we found that the N6-methyladenosine (m6A) methyltransferase-like 3 (METTL3) is significantly upregulated in esophageal squamous cell carcinoma (ESCC) and associated with poor patient prognosis. Depletion of METTL3 results in decreased ESCC growth and progression in vitro and in vivo. We further established ESCC initiation and progression models using Mettl3 conditional knockout mouse and revealed that METTL3-mediated m6A modification promotes ESCC initiation and progression in vivo. Moreover, using METTL3 overexpression ESCC cell model and Mettl3 conditional knockin mouse model, we demonstrated the critical function of METTL3 in promoting ESCC tumorigenesis in vitro and in vivo. Mechanistically, METTL3-catalyzed m6A modification promotes NOTCH1 expression and the activation of the Notch signaling pathway. Forced activation of Notch signaling pathway successfully rescues the growth, migration, and invasion capacities of METTL3-depleted ESCC cells. Our data uncovered important mechanistical insights underlying ESCC tumorigenesis and provided molecular basis for the development of novel strategies for ESCC diagnosis and treatment.

METTL3-mediated m6A mRNA modification promotes esophageal cancer initiation and progression via Notch signaling pathway.

Comparative genomic analysis of esophageal squamous cell carcinoma and adenocarcinoma: New opportunities towards molecularly targeted therapy

Esophageal squamous cell carcinoma (ESCC) is a deadly disease that requires extensive research on its mechanisms, prevention, and therapy. Recent studies have shown that NOTCH mutations are commonly seen in human ESCC. This chapter summarizes our current understanding of the NOTCH pathway in normal esophagus and in ESCC. In normal esophagus, NOTCH pathway regulates the development of esophageal squamous epithelium, in particular, squamous differentiation. Exposure to extrinsic and intrinsic factors, such as gastroesophageal reflux, alcohol drinking, and inflammation, downregulates the NOTCH pathway and thus inhibits squamous differentiation of esophageal squamous epithelial cells. In ESCC, NOTCH plays a dual role as both a tumor suppressor pathway and an oncogenic pathway. In summary, further studies are warranted to develop NOTCH activators for the prevention of ESCC and NOTCH inhibitors for targeted therapy of a subset of ESCC with activated NOTCH pathway.

NOTCH and Esophageal Squamous Cell Carcinoma

Alcohol drinking has been established as a major risk factor for esophageal diseases. Our previous study showed that ethanol exposure inhibited PAX9 expression in human esophageal squamous epithelial cells in vitro and in vivo. In this study, we aimed to investigate the molecular pathways through which alcohol drinking suppresses PAX9 in esophageal squamous epithelial cells. We first demonstrated the inhibition of NOTCH by ethanol exposure in vitro. NOTCH regulated PAX9 expression in KYSE510 and KYSE410 cells in vitro and in vivo. RBPJ and NOTCH intracellular domain (NIC) D1 ChIP-PCR confirmed Pax9 as a direct downstream target of NOTCH signaling in mouse esophagus. NOTCH inhibition by alcohol drinking was further validated in mouse esophagus and human tissue samples. In conclusion, ethanol exposure inhibited NOTCH signaling and thus suppressed PAX9 expression in esophageal squamous epithelial cells in vitro and in vivo. Our data support a novel mechanism of alcohol-induced esophageal injury through the inhibition of NOTCH-PAX9 signaling. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Alcohol drinking inhibits NOTCH-PAX9 signaling in esophageal squamous epithelial cells.

Development of targeted therapy of NRF2high esophageal squamous cell carcinoma.

Paired box 9 (PAX9) is a transcription factor of the PAX family functioning as both a transcriptional activator and repressor. Its functional roles in the embryonic development of various tissues and organs have been well studied. However, its roles and molecular mechanisms in cancer development are largely unknown. Here, we review the current understanding of PAX9 expression, upstream regulation of PAX9, and PAX9 downstream events in cancer development. Promoter hypermethylation, promoter SNP, microRNA, and inhibition of upstream pathways (e.g., NOTCH) result in PAX9 silencing or downregulation, whereas gene amplification and an epigenetic axis upregulate PAX9 expression. PAX9 may contribute to carcinogenesis through dysregulation of its transcriptional targets and related molecular pathways. In summary, extensive studies on PAX9 in its cellular and tissue contexts are warranted in various cancers, in particular, HNSCC, ESCC, lung cancer, and cervical SCC.

https://www.mdpi.com/1422-0067/23/10/5589/pdf?version=1652781026

PAX9 in Cancer Development

Objective NRF2 is a master transcription factor that regulates the stress response. NRF2 is frequently mutated and activated in human esophageal squamous cell carcinoma (ESCC), which drives resistance to chemotherapy and radiation therapy. Therefore, a great need exists for NRF2 inhibitors for targeted therapy of NRF2high ESCC. Design We performed high-throughput screening of two compound libraries from which hit compounds were further validated in human ESCC cells and a genetically modified mouse model. The mechanism of action of one compound was explored by biochemical assays. Results Using high-throughput screening of two small molecule compound libraries, we identified 11 hit compounds as potential NRF2 inhibitors with minimal cytotoxicity at specified concentrations. We then validated two of these compounds, pyrimethamine and mitoxantrone, by demonstrating their dose- and time-dependent inhibitory effects on the expression of NRF2 and its target genes in two NRF2Mut human ESCC cells (KYSE70 and KYSE180). RNAseq and qPCR confirmed the suppression of global NRF2 signaling by these two compounds. Mechanistically, pyrimethamine reduced NRF2 half-life by promoting NRF2 ubiquitination and degradation in KYSE70 and KYSE180 cells. Expression of an Nrf2E79Q allele in mouse esophageal epithelium (Sox2CreER;LSL-Nrf2E79Q/+) resulted in an NRF2high phenotype, which included squamous hyperplasia, hyperkeratinization, and hyperactive glycolysis. Treatment with pyrimethamine (30mg/kg/day, p.o.) suppressed the NRF2high esophageal phenotype with no observed toxicity. Conclusion We have identified and validated pyrimethamine as an NRF2 inhibitor that may be rapidly tested in the clinic as a radiation and chemotherapy sensitizer for NRF2high ESCC. Summary What is already known on this topic – summarise the state of scientific knowledge on this subject before you did your study and why this study needed to be done Mutational activation of the NRF2 transcription factor drives ESCC progression and therapeutic resistance. Targeted therapies to block NRF2 have not yet been realized, despite great needs. What this study adds – summarise what we now know as a result of this study that we did not know before A screen of >35,000 small molecules identified eleven potential NRF2 inhibitors. Pyrimethamine and mitoxantrone were validated to inhibit the expression of NRF2 and NQO1 in human ESCC cells in both dose- and time-dependent manners. Pyrimethamine enhanced NRF2 protein ubiquitination and degradation, resulting a decreased half-life. A genetically modified mouse model was established to express the Nrf2E79Q mutant allele in the mouse esophageal epithelium upon tamoxifen induction. Pyrimethamine suppressed the NRF2high esophageal phenotype induced by the mutant allele. How this study might affect research, practice or policy – summarise the implications of this study As an FDA-approved drug, Pyrimethamine has the potential for immediate translation to a clinical trial on NRF2high ESCC in humans. Further exploration of its mechanisms of action may lead to more potent NRF2 inhibitors for future use.

Yahui Li

Papers

NOTCH and Esophageal Squamous Cell Carcinoma

Alcohol drinking inhibits NOTCH-PAX9 signaling in esophageal squamous epithelial cells.

Development of targeted therapy of NRF2high esophageal squamous cell carcinoma.

PAX9 in Cancer Development

Small Molecule Screen Identifies Pyrimethamine as an Inhibitor of NRF2-driven Esophageal Hyperplasia