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Castillon, L.

Publications and source records attributed to Castillon, L..

4 recordsLinked to original sources
bioRxiv2025

Mechanisms of resistance to VHL loss-induced genetic and pharmacological vulnerabilities

The von Hippel-Lindau tumor suppressor (VHL) is a component of a ubiquitin ligase complex that normally controls cellular responses to hypoxia. Endogenous VHL is also utilized by proteolysis-targeting chimera (PROTAC) protein degraders, a promising class of anti-cancer agents. VHL is broadly essential for cell proliferation, yet it is a key tumor suppressor in renal cell carcinoma. To understand the functional consequences of VHL loss, and to identify targeted approaches for the elimination of VHL null cells, we have used genome-wide CRISPR-Cas9 screening in human renal epithelial cells. We find that, upon VHL loss, the HIF1A/ARNT complex is the central inhibitor of cellular fitness, suppressing mitochondrial respiration, and that VHL null cells show HIF1A-dependent molecular vulnerabilities that can be targeted pharmacologically. Combined VHL/HIF1A inactivation in breast and esophageal cancer cells can also provide resistance to ARV-771, a VHL-based bromodomain degrader that has anti-cancer activity. HIF1A stabilization can thus provide opportunities for early intervention in neoplastic VHL clones, and the VHL-HIF1A axis may be relevant for the development of resistance to the emerging class of PROTAC-based cancer therapies.

cancer biology↗
bioRxiv2024

Fibroblast activation protein defines aggressive EMT-associated cancer cell state and prognosis in localized ccRCC

Clear cell renal cell carcinoma (ccRCC) is among the most immune-infiltrated cancers, where high inflammation correlates with poor clinical outcomes. While epithelial-to-mesenchymal transition (EMT), immunosuppression, and cancer-associated fibroblasts (CAFs) have been linked with poor prognosis in metastatic ccRCC, their relevance and interplay in localized disease remain uncertain. To address this gap, we performed single-cell spatial profiling of highly inflamed localized tumors to characterize interactions between EMT-associated cancer cells and immune subsets, aiming to identify tumor microenvironment cell subsets and states responsible for the poor prognosis in highly inflamed ccRCC. Multiplexed immunofluorescence imaging with a 33-marker panel was applied to 1,728 tissue cores collected from tumor centers, invasive borders, and adjacent benign tissue from 435 localized ccRCC patients. Independent discovery (n=196) and validation (n=239) cohorts were used to confirm the significance of identified prognostic markers. Hierarchical clustering revealed TME subsets enriched for CD45 immune cells, CD31 endothelial cells, or fibroblasts. High CD45 cell density significantly correlated with poor recurrence-free survival across tumor regions, including tumor center, invasive border, and adjacent benign tissue. Within tumors exhibiting high CD45 infiltration, EMT-associated cancer cells strongly expressed fibroblast activation protein (FAP) at invasive borders, independently predicting worse prognosis and increased risk of liver metastasis. Furthermore, CD45high tumors with elevated FAP expression featured immunosuppressive FAP CAFs, M2-like macrophages, exhausted T cells, and regulatory T cells. Tumor-cell-specific FAP expression was validated as an independent prognostic biomarker in both early-stage localized ccRCC (pT1-2) and patients subsequently developing metastases who received sunitinib, highlighting its potential for improved patient stratification. SignificanceFAP expression identifies an aggressive, immunosuppressive EMT-associated subtype of localized ccRCC, uncovering critical interactions between cancer cells and the TME and revealing novel biological insights and therapeutic vulnerabilities driving early-stage ccRCC progression. Graphical AbstractO_ST_ABSTumor-cell FAP Expression Defines an Aggressive, Immunosuppressive EMT Subtype in Highly Inflamed Localized ccRCCC_ST_ABS O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=197 SRC="FIGDIR/small/620479v2_ufig1.gif" ALT="Figure 1"> View larger version (55K): org.highwire.dtl.DTLVardef@d25831org.highwire.dtl.DTLVardef@19c0de7org.highwire.dtl.DTLVardef@54bd9eorg.highwire.dtl.DTLVardef@16d0387_HPS_FORMAT_FIGEXP M_FIG C_FIG Localized ccRCC tumors were stratified by CD45 immune infiltration, revealing that CD45high tumors were associated with significantly poorer recurrence-free survival (5-year RFS rate: 39%). Among these, tumor cell-specific expression of fibroblast activation protein (FAP) identified a particularly aggressive subtype marked by epithelial-to-mesenchymal transition (EMT), reduced endothelial content, and an immunosuppressive tumor microenvironment enriched with M2-like macrophages, regulatory T cells, exhausted T cells, and FAP cancer-associated fibroblasts (CAFs). FAP tumors had an even lower 5-year RFS rate of 26%, compared to 50% in FAP- CD45high tumors. Tumor-cell FAP thus provides an additional layer of risk stratification within the CD45high subgroup and reflects a biologically distinct, immune-evasive tumor state. These findings highlight FAP as both a potential therapeutic target and imaging biomarker in localized ccRCC.

cancer biology↗
bioRxiv2020

High-resolution Crystal Structures of Transient Intermediates in the Phytochrome Photocycle

Phytochromes are red/far-red light photoreceptors in bacteria to plants, which elicit a variety of important physiological responses. They display a reversible photocycle between the resting (dark) Pr state and the light activated Pfr state, in which light signals are received and transduced as structural change through the entire protein to modulate the activity of the protein. It is unknown how the Pr-to-Pfr interconversion occurs as the structure of intermediates remain notoriously elusive. Here, we present short-lived crystal structures of the classical phytochrome from myxobacterium Stigmatella aurantiaca captured by an X-ray Free Electron Laser 5 ns and 33ms after light illumination of the Pr state. We observe large structural displacements of the covalently bound bilin chromophore, which trigger a bifurcated signaling pathway. The snapshots show with atomic precision how the signal progresses from the chromophore towards the output domains, explaining how plants, bacteria and fungi sense red light.

biophysics↗
bioRxiv2019

The primary structural photoresponse of phytochrome proteins captured by a femtosecond X-ray laser

Phytochrome proteins control the growth, reproduction, and photosynthesis of plants, fungi, and bacteria. Light is detected by a bilin cofactor, but it remains elusive how this leads to activation of the protein through structural changes. We present serial femtosecond X-ray crystallographic data of the chromophore-binding domains of a bacterial phytochrome at delay times of 1 ps and 10 ps after photoexcitation. The structures reveal a twist of the D-ring, which lead to partial detachment of the chromophore from the protein. Unexpectedly, the conserved so-called pyrrole water is photodissociated from the chromophore, concomitant with movement of the A-ring and a key signalling aspartate. The changes are wired together by ultrafast backbone and water movements around the chromophore, channeling them into signal transduction towards the output domains. We suggest that the water dissociation is key to the phytochrome photoresponse, explaining the earliest steps of how plants, fungi and bacteria sense red light.

biophysics↗