Cancer invasion and metastasis: Insights from murine pubertal mammary gland morphogenesis.

Cancer invasion and metastasis accounts for the majority of cancer related mortality. A better understanding of the players that drive the aberrant invasion and migration of tumors cells will provide critical targets to inhibit metastasis. Postnatal pubertal mammary gland morphogenesis is characterized by highly proliferative, invasive, and migratory normal epithelial cells. Identifying the molecular regulators of pubertal gland development is a promising strategy since tumorigenesis and metastasis is postulated to be a consequence of aberrant reactivation of developmental stages. In this review, we summarize the pubertal morphogenesis regulators that are involved in cancer metastasis and revisit pubertal mammary gland transcriptome profiling to uncover both known and unknown metastasis genes. Our updated list of pubertal morphogenesis regulators shows that most are implicated in invasion and metastasis. This review highlights molecular linkages between development and metastasis and provides a guide for exploring novel metastatic drivers.

Molnupiravir Does Not Induce Mutagenesis in Host Lung Cells during SARS-CoV-2 Treatment.

As SARS-CoV-2 continues to evolve and spread with the emergence of new variants, interest in small molecules with broad-spectrum antiviral activity has grown. One such molecule, Molnupiravir (MOV; other names: MK-4482, EIDD-2801), a ribonucleoside analogue, has emerged as an effective SARS-CoV-2 treatment by inducing catastrophic viral mutagenesis during replication. However, there are growing concerns as MOV's potential to induce host DNA mutagenesis remains an open question. Analysis of RNA-seq data from SARS-CoV-2-infected MOV-treated golden hamster lung biopsies confirmed MOV's efficiency in stopping SARS-CoV-2 replication. Importantly, MOV treatment did not increase mutations in the host lung cells. This finding calls for additional mutation calls on host biopsies from more proliferative tissues to fully explore MOV's hypothesized mutagenic risk.

DREAM, a possible answer to the estrogen paradox of the Women's Health Initiative Trial.

Estrogen is thought to cause proliferation of all estrogen receptor positive (ER+) breast cancers. Paradoxically, in the Women's Health Initiative Trial, estrogen-only hormone replacement therapy reduced the incidence and mortality of low grade, ER+, HER2- breast cancer. We gave estradiol to 19 post-menopausal women with newly diagnosed low-grade, ER+, HER2- breast cancer in a prospective window of opportunity clinical trial and examined the changes in proliferation and gene expression before and after estradiol treatment. Ki67 decreased in 13/19 (68%) patients and 8/13 (62%) showed a decrease in Risk of Recurrence Score. We chose three prototypical estrogen responders (greatest decrease in ROR) and non-responders (no/minimal change in ROR) and applied a differential gene expression analysis to develop pre-treatment (PRESTO-30core) and post-treatment (PRESTO-45surg) gene expression profiles. The PRESTO-30core predicted adjuvant benefit in a published series of tamoxifen, the partial estrogen agonist. Of the 45 genes in the PRESTO-45surg, thirty contain the Cell cycle genes Homology Region (CHR) motif that binds the class B multi-vulva complex (MuvB) a member of the DREAM (Dimerization partner, retinoblastoma-like proteins, E2F, MuvB) complex responsible for reversible cell cycle arrest or quiescence. There was also near uniform suppression (89%) of the remaining DREAM genes consistent with estrogen induced activation of the DREAM complex to mediate cell cycle block after a short course of estrogens. To our knowledge, this is the first report to show estrogen modulation of DREAM genes and suggest involvement of DREAM pathway associated quiescence in endocrine responsive post-menopausal ER+ breast cancers.

Robust inflammatory breast cancer gene signature using nonparametric random forest analysis.

Inflammatory breast cancer (IBC) is a rare, aggressive cancer found in all the molecular breast cancer subtypes. Despite extensive previous efforts to screen for transcriptional differences between IBC and non-IBC patients, a robust IBC-specific molecular signature has been elusive. We report a novel IBC-specific gene signature (59 genes; G59) that achieves 100% accuracy in discovery and validation samples (45/45 correct classification) and remarkably only misclassified one sample (60/61 correct classification) in an independent dataset. G59 is independent of ER/HER2 status, molecular subtypes and is specific to untreated IBC samples, with most of the genes being enriched for plasma membrane cellular component proteins, interleukin (IL), and chemokine signaling pathways. Our finding suggests the existence of an IBC-specific molecular signature, paving the way for the identification and validation of targetable genomic drivers of IBC.

BAD regulates mammary gland morphogenesis by 4E-BP1-mediated control of localized translation in mouse and human models.

Elucidation of non-canonical protein functions can identify novel tissue homeostasis pathways. Herein, we describe a role for the Bcl-2 family member BAD in postnatal mammary gland morphogenesis. In Bad3SA knock-in mice, where BAD cannot undergo phosphorylation at 3 key serine residues, pubertal gland development is delayed due to aberrant tubulogenesis of the ductal epithelium. Proteomic and RPPA analyses identify that BAD regulates focal adhesions and the mRNA translation repressor, 4E-BP1. These results suggest that BAD modulates localized translation that drives focal adhesion maturation and cell motility. Consistent with this, cells within Bad3SA organoids contain unstable protrusions with decreased compartmentalized mRNA translation and focal adhesions, and exhibit reduced cell migration and tubulogenesis. Critically, protrusion stability is rescued by 4E-BP1 depletion. Together our results confirm an unexpected role of BAD in controlling localized translation and cell migration during mammary gland development.

BIK drives an aggressive breast cancer phenotype through sublethal apoptosis and predicts poor prognosis of ER-positive breast cancer.

Apoptosis is fundamental to normal animal development and is the target for many anticancer therapies. Recent studies have explored the consequences of "failed apoptosis" where the apoptotic program is initiated but does not go to completion and does not cause cell death. Nevertheless, this failed apoptosis induces DNA double-strand breaks generating mutations that facilitate tumorigenesis. Whether failed apoptosis is relevant to clinical disease is unknown. BCL-2 interacting killer (BIK) is a stress-induced BH3-only protein that stimulates apoptosis in response to hormone and growth factor deprivation, hypoxia, and genomic stress. It was unclear whether BIK promotes or suppresses tumor survival within the context of breast cancer. We investigated this and show that BIK induces failed apoptosis with limited caspase activation and genomic damage in the absence of extensive cell death. Surviving cells acquire aggressive phenotypes characterized by enrichment of cancer stem-like cells, increased motility and increased clonogenic survival. Furthermore, by examining six independent cohorts of patients (total n = 969), we discovered that high BIK mRNA and protein levels predicted clinical relapse of Estrogen receptor (ER)-positive cancers, which account for almost 70% of all breast cancers diagnosed but had no predictive value for hormone receptor-negative (triple-negative) patients. Thus, this study identifies BIK as a biomarker for tumor recurrence of ER-positive patients and provides a potential mechanism whereby failed apoptosis contributes to cancer aggression.

A glucose-dependent spatial patterning of exocytosis in human ß-cells is disrupted in type 2 diabetes.

Impaired insulin secretion in type 2 diabetes (T2D) is linked to reduced insulin granule docking, disorganization of the exocytotic site, and an impaired glucose-dependent facilitation of insulin exocytosis. We show in ß-cells from 80 human donors that the glucose-dependent amplification of exocytosis is disrupted in T2D. Spatial analyses of granule fusion, visualized by total internal reflection fluorescence (TIRF) microscopy in 24 of these donors, demonstrate that these are non-random across the surface of ß-cells from donors with no diabetes (ND). The compartmentalization of events occurs within regions defined by concurrent or recent membrane-resident secretory granules. This organization, and the number of membrane-associated granules, is glucose-dependent and notably impaired in T2D ß-cells. Mechanistically, multi-channel Kv2.1 clusters contribute to maintaining the density of membrane-resident granules and the number of fusion 'hotspots', while SUMOylation sites at the channel N- (K145) and C-terminus (K470) determine the relative proportion of fusion events occurring within these regions. Thus, a glucose-dependent compartmentalization of fusion, regulated in part by a structural role for Kv2.1, is disrupted in ß-cells from donors with type 2 diabetes.

Non-canonical BAD activity regulates breast cancer cell and tumor growth via 14-3-3 binding and mitochondrial metabolism.

The Bcl-2-associated death promoter BAD is a prognostic indicator for good clinical outcome of breast cancer patients; however, whether BAD affects breast cancer biology is unknown. Here we showed that BAD increased cell growth in breast cancer cells through two distinct mechanisms. Phosphorylation of BAD at S118 increased S99 phosphorylation, 14-3-3 binding and AKT activation to promote growth and survival. Through a second, more prominent pathway, BAD stimulated mitochondrial oxygen consumption in a novel manner that was downstream of substrate entry into the mitochondria. BAD stimulated complex I activity that facilitated enhanced cell growth and sensitized cells to apoptosis in response to complex I blockade. We propose that this dependence on oxidative metabolism generated large but nonaggressive cancers. This model identifies a non-canonical role for BAD and reconciles BAD-mediated tumor growth with favorable outcomes in BAD-high breast cancer patients.

Ligand-induced growth and compaction of CD36 nanoclusters enriched in Fyn induces Fyn signaling.

Nanoclustering is an emerging organizational principle for membrane-associated proteins. The functional consequences of nanoclustering for receptor signaling remain largely unknown. Here, we applied quantitative multi-channel high- and super-resolution imaging to analyze the endothelial cell surface receptor CD36, the clustering of which upon binding to multivalent ligands, such as the anti-angiogenic factor thrombospondin-1 (TSP-1), is thought to be crucial for signaling. We found that a substantial fraction of unligated CD36 exists in nanoclusters, which not only promote TSP-1 binding but are also enriched with the downstream effector Fyn. Exposure to multivalent ligands (TSP-1 or anti-CD36 IgM) that result in larger and denser CD36 clusters activates Fyn. Conversely, pharmacological perturbations that prevent the enhancement of CD36 clustering by TSP-1 abrogate Fyn activation. In both cases, there is no detectable change in Fyn enrichment at CD36 nanoclusters. These observations reveal a crucial role for the basal organization of a receptor into nanoclusters that are enriched with the signal-transducing downstream effectors of that receptor, such that enhancement of clustering by multivalent ligands is necessary and sufficient to activate the downstream effector without the need for its de novo recruitment.