Thorne, C. | Search bioRxiv

bioRxiv2021

Molecular Drivers of Tumor Progression in Microsatellite Stable APC Mutation-Negative Colorectal Cancers

BackgroundThe tumor suppressor gene adenomatous polyposis coli (APC) is the initiating mutation in approximately 80% of all colorectal cancers (CRC), underscoring the importance of aberrant regulation of intracellular WNT signaling in CRC development. Recent studies have found that early-onset CRC exhibits an increased proportion of tumors lacking an APC mutation. We set out to identify mechanisms underlying APC mutation-negative (APCmut-) CRCs. MethodsWe analyzed data from The Cancer Genome Atlas to compare clinical phenotypes, somatic mutations, copy number variations, gene fusions, RNA expression, and DNA methylation profiles between APCmut- and APC mutation-positive (APCmut+) microsatellite stable CRCs. ResultsTranscriptionally, APCmut- CRCs clustered into two approximately equal groups. Cluster One was associated with enhanced mitochondrial activation. Cluster Two was strikingly associated with genetic inactivation or decreased RNA expression of the WNT antagonist RNF43, increased expression of the WNT agonist RSPO3, activating mutation of BRAF, or increased methylation and decreased expression of AXIN2. APCmut- CRCs exhibited evidence of increased immune cell infiltration, with significant correlation between M2 macrophages and RSPO3. ConclusionsAPCmut- CRCs comprise two groups of tumors characterized by enhanced mitochondrial activation or increased sensitivity to extracellular WNT, suggesting that they could be respectively susceptible to inhibition of these pathways.

cancer biology↗

bioRxiv2020

Paradoxical effects of cigarette smoke and COPD on SARS-CoV2 infection and disease

IntroductionHow cigarette smoke (CS) and chronic obstructive pulmonary disease (COPD) affect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and severity is controversial. We investigated the protein and mRNA expression of SARS-CoV-2 entry receptor ACE2 and proteinase TMPRSS2 in lungs from COPD patients and controls, and lung tissue from mice exposed acutely and chronically to CS. Also, we investigated the effects of CS exposure on SARS-CoV-2 infection in human bronchial epithelial cells. MethodsIn Cohort 1, ACE2-positive cells were quantified by immunostaining in FFPE sections from both central and peripheral airways. In Cohort 2, we quantified pulmonary ACE2 protein levels by immunostaining and ELISA, and both ACE2 and TMPRSS2 mRNA levels by RT-qPCR. In C57BL/6 WT mice exposed to air or CS for up to 6 months, pulmonary ACE2 protein levels were quantified by triple immunofluorescence staining and ELISA. The effects of CS exposure on SARS-CoV-2 infection were evaluated after 72hr in vitro infection of Calu-3 cells. After SARS-CoV-2 infection, the cells were fixed for IF staining with dsRNA-specific J2 monoclonal Ab, and cell lysates were harvested for WB of viral nucleocapsid (N) protein. Supernatants (SN) and cytoplasmic lysates were obtained to measure ACE2 levels by ELISA. ResultsIn both human cohorts, ACE2 protein and mRNA levels were decreased in peripheral airways from COPD patients versus both smoker and NS controls, but similar in central airways. TMPRSS2 levels were similar across groups. Mice exposed to CS had decreased ACE2 protein levels in their bronchial and alveolar epithelia versus air-exposed mice exposed to 3 and 6 months of CS. In Calu3 cells in vitro, CS-treatment abrogated infection to levels below the limit of detection. Similar results were seen with WB for viral N protein, showing peak viral protein synthesis at 72hr. ConclusionsACE2 levels were decreased in both bronchial and alveolar epithelial cells from uninfected COPD patients versus controls, and from CS-exposed versus air-exposed mice. CS-pre-treatment did not affect ACE2 levels but potently inhibited SARS-CoV-2 replication in this in vitro model. These findings urge to further investigate the controversial effects of CS and COPD on SARS-CoV2 infection.

immunology↗