Rarγ -Foxa1 signaling promotes luminal identity in prostate progenitors and is disrupted in prostate cancer.

Retinoic acid (RA) signaling is a master regulator of vertebrate development with crucial roles in directing body axis orientation and tissue differentiation, including in the reproductive system. However, a mechanistic understanding of how RA signaling promotes cell lineage identity in different tissues is often missing. Here, leveraging prostate organoid technology, we demonstrated that RA signaling orchestrates the commitment of adult mouse prostate progenitors to glandular identity, epithelial barrier integrity, and ultimately, proper specification of the prostatic lumen. Mechanistically, RA-dependent RARγ activation promotes the expression of the pioneer factor Foxa1, which synergizes with the androgen pathway for proper luminal expansion, cytoarchitecture and function. FOXA1 nucleotide variants are common in human prostate and breast cancers and considered driver mutations, though their pathogenic mechanism is incompletely understood. Combining functional genetics experiments with structural modeling of FOXA1 folding and chromatin binding analyses, we discovered that FOXA1 F254E255 is a loss-of-function mutation leading to compromised transcriptional function and lack of luminal fate commitment of prostate progenitors. Overall, we define RA as a crucial instructive signal for glandular identity in adult prostate progenitors. We propose deregulation of vitamin A metabolism as a risk factor for benign and malignant prostate disease, and identified cancer associated FOXA1 indels affecting residue F254 as loss-of-function mutations promoting dedifferentiation of adult prostate progenitors. Summary: Retinoic acid signaling orchestrates luminal differentiation of adult prostate progenitors.

Intra-epithelial non-canonical Activin A signaling safeguards prostate progenitor quiescence.

The healthy prostate is a relatively quiescent tissue. Yet, prostate epithelium overgrowth is a common condition during aging, associated with urinary dysfunction and tumorigenesis. For over thirty years, TGF-ß ligands have been known to induce cytostasis in a variety of epithelia, but the intracellular pathway mediating this signal in the prostate, and its relevance for quiescence, have remained elusive. Here, using mouse prostate organoids to model epithelial progenitors, we find that intra-epithelial non-canonical Activin A signaling inhibits cell proliferation in a Smad-independent manner. Mechanistically, Activin A triggers Tak1 and p38 ΜAPK activity, leading to p16 and p21 nuclear import. Spontaneous evasion from this quiescent state occurs upon prolonged culture, due to reduced Activin A secretion, a condition associated with DNA replication stress and aneuploidy. Organoids capable to escape quiescence in vitro are also able to implant with increased frequency into immunocompetent mice. This study demonstrates that non-canonical Activin A signaling safeguards epithelial quiescence in the healthy prostate, with potential implications for the understanding of cancer initiation, and the development of therapies targeting quiescent tumor progenitors.

ETS-related gene (ERG) undermines genome stability in mouse prostate progenitors via Gsk3ß dependent Nkx3.1 degradation.

21q22.2-3 deletion is the most common copy number alteration in prostate cancer (PCa). The genomic rearrangement results in the androgen-dependent de novo expression of ETS-related gene (ERG) in prostate cancer cells, a condition promoting tumor progression to advanced stages of the disease. Interestingly, ERG expression characterizes 5-30% of tumor precursor lesions - High Grade Prostatic Intraepithelial Neoplasia (HGPIN) - where its role remains unclear. Here, by combining organoids technology with Click-chemistry coupled Mass Spectrometry, we demonstrate a prominent role of ERG in remodeling the protein secretome of prostate progenitors. Functionally, by lowering autocrine Wnt-4 signaling, ERG represses canonical Wnt pathway in prostate progenitors, and, in turn, promotes the accumulation of DNA double strand breaks via Gsk3ß-dependent degradation of the tumor suppressor Nkx3.1. On the other hand, by shaping extracellular paracrine signals, ERG strengthens the pro-oxidative transcriptional signature of inflammatory macrophages, which we demonstrate to infiltrate pre-malignant ERG positive prostate lesions. These findings highlight previously unrecognized functions of ERG in undermining adult prostate progenitor niche through cell autonomous and non-autonomous mechanisms. Overall, by supporting the survival and proliferation of prostate progenitors in the absence of growth stimuli and promoting the accumulation of DNA damage through destabilization of Nkx3.1, ERG could orchestrate the prelude to neoplastic transformation.

Calcium cytotoxicity sensitizes prostate cancer cells to standard-of-care treatments for locally advanced tumors.

Therapy resistance is a major roadblock in oncology. Exacerbation of molecular dysfunctions typical of cancer cells have proven effective in twisting oncogenic mechanisms to lethal conditions, thus offering new therapeutic avenues for cancer treatment. Here, we demonstrate that selective agonists of Transient Receptor Potential cation channel subfamily M member 8 (TRPM8), a cation channel characteristic of the prostate epithelium frequently overexpressed in advanced stage III/IV prostate cancers (PCa), sensitize therapy refractory models of PCa to radio, chemo or hormonal treatment. Overall, our study demonstrates that pharmacological-induced Ca2+ cytotoxicity is an actionable strategy to sensitize cancer cells to standard therapies.

The Organoid Era Permits the Development of New Applications to Study Glioblastoma.

Glioblastoma (GB) is the most frequent and aggressive type of glioma. The lack of reliable GB models, together with its considerable clinical heterogeneity, has impaired a comprehensive investigation of the mechanisms that lead to tumorigenesis, cancer progression, and response to treatments. Recently, 3D cultures have opened the possibility to overcome these challenges and cerebral organoids are emerging as a leading-edge tool in GB research. The opportunity to easily engineer brain organoids via gene editing and to perform co-cultures with patient-derived tumor spheroids has enabled the analysis of cancer development in a context that better mimics brain tissue architecture. Moreover, the establishment of biobanks from GB patient-derived organoids represents a crucial starting point to improve precision medicine therapies. This review exemplifies relevant aspects of 3D models of glioblastoma, with a specific focus on organoids and their involvement in basic and translational research.

Screen-based identification and validation of four new ion channels as regulators of renal ciliogenesis.

To investigate the contribution of ion channels to ciliogenesis, we carried out a small interfering RNA (siRNA)-based reverse genetics screen of all ion channels in the mouse genome in murine inner medullary collecting duct kidney cells. This screen revealed four candidate ion channel genes: Kcnq1, Kcnj10, Kcnf1 and Clcn4. We show that these four ion channels localize to renal tubules, specifically to the base of primary cilia. We report that human KCNQ1 Long QT syndrome disease alleles regulate renal ciliogenesis; KCNQ1-p.R518X, -p.A178T and -p.K362R could not rescue ciliogenesis after Kcnq1-siRNA-mediated depletion in contrast to wild-type KCNQ1 and benign KCNQ1-p.R518Q, suggesting that the ion channel function of KCNQ1 regulates ciliogenesis. In contrast, we demonstrate that the ion channel function of KCNJ10 is independent of its effect on ciliogenesis. Our data suggest that these four ion channels regulate renal ciliogenesis through the periciliary diffusion barrier or the ciliary pocket, with potential implication as genetic contributors to ciliopathy pathophysiology. The new functional roles of a subset of ion channels provide new insights into the disease pathogenesis of channelopathies, which might suggest future therapeutic approaches.

DNA replication stress underlies renal phenotypes in CEP290-associated Joubert syndrome.

Juvenile ciliopathy syndromes that are associated with renal cysts and premature renal failure are commonly the result of mutations in the gene encoding centrosomal protein CEP290. In addition to centrosomes and the transition zone at the base of the primary cilium, CEP290 also localizes to the nucleus; however, the nuclear function of CEP290 is unknown. Here, we demonstrate that reduction of cellular CEP290 in primary human and mouse kidney cells as well as in zebrafish embryos leads to enhanced DNA damage signaling and accumulation of DNA breaks ex vivo and in vivo. Compared with those from WT mice, primary kidney cells from Cep290-deficient mice exhibited supernumerary centrioles, decreased replication fork velocity, fork asymmetry, and increased levels of cyclin-dependent kinases (CDKs). Treatment of Cep290-deficient cells with CDK inhibitors rescued DNA damage and centriole number. Moreover, the loss of primary cilia that results from CEP290 dysfunction was rescued in 3D cell culture spheroids of primary murine kidney cells after exposure to CDK inhibitors. Together, our results provide a link between CEP290 and DNA replication stress and suggest CDK inhibition as a potential treatment strategy for a wide range of ciliopathy syndromes.