Biology subjects

Wolff, S. C.

Publications and source records attributed to Wolff, S. C..

7 recordsLinked to original sources

bioRxiv2026

Non-Genetic Mechanisms of Fractional Resistance to Abemaciclib in Dedifferentiated Liposarcoma.

Dedifferentiated liposarcoma is a rare mesenchymal malignancy driven by amplification of chromosome 12q13-15, which includes the oncogenes CDK4 and MDM2. CDK4 amplification provides a rationale for targeted therapy with CDK4/6 inhibitors, and abemaciclib has shown the most durable activity reported to date in this disease. Clinical responses, however, are incomplete and often transient, and the cellular features that allow tumor cells to continue proliferating during treatment are not well understood. To address this gap, we performed multiplexed single-cell imaging to quantify 17 cell-cycle regulators in both dedifferentiated liposarcoma cell line Lipo246 and surgically resected primary human cells exposed to abemaciclib. Both models contained a subpopulation of cells that retained phosphorylated retinoblastoma protein, a marker of cell proliferation, at the highest abemaciclib doses. These fractionally resistant cells were defined by selective enrichment of cyclin-dependent kinase 2 (CDK2), cyclin B1, and phosphorylated ribosomal protein S6 (pS6), and showed enhanced sensitivity to the CDK2 inhibitor, tagtociclib. Together, these findings reveal nongenetic cell cycle plasticity as a mechanism of escape from CDK4/6 inhibition in dedifferentiated liposarcoma and nominate CDK2 and the PI3K-mTOR pathway as candidate targets for combination therapy.

cancer biology↗

bioRxiv2025

Extrachromosomal DNA Gives Cancer a New Evolutionary Pathway

During tumor progression, it has been assumed that individual cells that have acquired advantageous mutations overtake the population. Cancers driven by extrachromosomal DNA (ecDNA) do not follow this paradigm. Instead, these tumors have a spectrum of oncogene copy numbers across cells, and graded ecDNA variation may function as a form of bet-hedging that equips tumors with a broad range of phenotypes. Using imaging, single-cell multiomics, and multiplexed proteomics, we systematically characterized ecDNA levels across thousands of single cells. Higher ecDNA dosage produces proportional changes in transcript abundance, chromatin accessibility, protein levels, cell-cycle progression, and proliferation. Genes amplified on ecDNA exhibit distinct transcriptional scaling regimes that shift when the same genes are reintegrated into chromosomal homogeneous staining regions. When we experimentally disrupted the continuum of ecDNA dosage by sorting cells into low- and high-copy number states, the population rapidly recovered its original, continuous distribution. Our time-course data, live-cell imaging, and stochastic models collectively show that restoring this spectrum is an active, deterministic process rather than the passive outcome of random segregation. Together, these findings position ecDNA-mediated expression as a distinct evolutionary mechanism that endows tumors with rapid, population-level adaptability. These findings offer insight into why ecDNA-driven cancers are among the most aggressive and treatment-resistant.

genomics↗

bioRxiv2025

TRIP13 protects pancreatic cancer cells against intrinsic and therapy-induced DNA replication stress

Oncogene activation in normal untransformed cells induces DNA replication stress and creates a dependency on DNA Damage Response (DDR) mechanisms for cell survival. Different oncogenic stimuli signal via distinct mechanisms in every cancer setting. The DDR is also pathologically re-programmed and deployed in diverse ways in different cancers. Because mutant KRAS is the driver oncogene in 90% of Pancreatic Ductal Adenocarcinomas (PDAC), here we have investigated DDR mechanisms by which KRAS-induced DNA replication stress is tolerated in normal human pancreatic epithelial cells (HPNE). Using a candidate screening approach, we identify TRIP13 as a KRASG12V-induced mRNA that is also expressed at high levels in PDAC relative to normal tissues. Using genetic and pharmacological tools, we show that TRIP13 is necessary to sustain ongoing DNA synthesis and viability specifically in KRASG12V-expressing cells. TRIP13 promotes survival of KRASG12V-expressing HPNE cells in a Homologous Recombination (HR)-dependent manner. KRASG12V-expressing HPNE cells lacking TRIP13 acquire hallmark HR-deficiency (HRD) phenotypes including sensitivity to inhibitors of Trans-Lesion Synthesis (TLS) and Poly-ADP Ribose Polymerase (PARP). Established PDAC cell lines are also sensitized to intrinsic DNA damage and therapy-induced genotoxicity following TRIP13-depletion. Taken together our results expose TRIP13 as an attractive new and therapeutically-tractable vulnerability of KRAS-mutant PDAC.

cancer biology↗

bioRxiv2023

Cell cycle plasticity underlies fractional resistance to palbociclib in ER+/HER2- breast tumor cells

The CDK4/6 inhibitor palbociclib blocks cell cycle progression in ER+/HER2- breast tumor cells. Although these drugs have significantly improved patient outcomes in metastatic breast cancers, a small percentage of tumor cells continues to divide in the presence of palbociclib--a phenomenon we refer to as fractional resistance. It is critical to understand the cellular mechanisms underlying fractional resistance because the precise percentage of resistant cells in patient tissue is a strong predictor of clinical outcome. Here, we hypothesize that fractional resistance arises from cell-to-cell differences in core cell cycle regulators that allow a subset of cells to escape CDK4/6 inhibitor therapy. We used multiplex, single-cell imaging to identify fractionally resistant tumor cells both in a cell culture model of ER+/HER2- breast cancer as well as live primary tumor cells resected from a patient. We found that tumor cells capable of proliferating in the presence of palbociclib showed both expected (e.g., CDK2, E2F1) and unexpected (e.g., Cdt1, p21, cyclin B1) shifts in core cell cycle regulators. Notably, resistant cells in both tumor models showed premature enrichment of the G1 regulators E2F1 and CDK2 protein and, unexpectedly, the G2/M regulator cyclin B1 just before cell cycle entry, suggesting that resistant cells may use noncanonical mechanisms to overcome CDK4/6 inhibition. Using computational data integration and trajectory inference approaches, we show how plasticity in cell cycle regulators gives rise to alternate cell cycle "paths" that allow individual ER+/HER2- tumor cells to escape palbociclib treatment. Understanding drivers of cell cycle plasticity, and how to eliminate resistant cell cycle paths, could lead to improved cancer therapies targeting fractionally resistant cells to improve patient outcomes.

cancer biology↗

bioRxiv2022

Single-cell dynamics of core pluripotency factors in human pluripotent stem cells

The human transcription factors OCT4, SOX2, and NANOG form a core signaling network critical for maintaining stem cell pluripotency and self-renewal potential. The spatiotemporal expression dynamics of these pluripotency factors throughout differentiation is unclear, limiting our understanding of stem cell fate decisions. Here, we combined CRISPR/Cas9-mediated gene editing with microraft array technology to generate human embryonic stem cell lines with endogenously tagged fluorophores for OCT4, SOX2, and NANOG. Fluorescence time-lapse imaging revealed that pluripotent stem cells show gastrulation-like patterning without direct chemical induction. Directed differentiation to the three primary germ layers--endoderm, mesoderm, and ectoderm--revealed distinct spatiotemporal patterns of SOX2 and NANOG expression in single cells. Finally, we captured dynamic changes in cell morphology during ectoderm differentiation corresponding to the formation of neural rosettes. This study provides a robust method for generating live-cell reporters in human stem cells and describes the single-cell dynamics of human pluripotency factors during differentiation.

developmental biology↗

bioRxiv2017

The Cell Cycle Browser: an interactive tool for visualizing, simulating, and perturbing cell cycle progression

SUMMARYThe cell cycle is driven by precise temporal coordination among many molecular activities. To understand and explore this process, we developed the Cell Cycle Browser (CCB), an interactive web interface based on real-time reporter data collected in proliferating human cells. This tool facilitates visualizing, simulating, and predicting the outcomes of perturbing cell cycle parameters. Time-series traces from individual cells can be combined to build a multi-layered timeline of molecular activities. Users can simulate the cell cycle using computational models that capture the dynamics of molecular activities and phase transitions. By adjusting individual expression levels and strengths of molecular relationships, users can predict effects on the cell cycle. Virtual assays, such as growth curves and flow cytometry, provide familiar outputs to compare cell cycle behaviors for data and simulations. The CCB serves to unify our understanding of cell cycle dynamics and provides a platform for generating hypotheses through virtual experiments.\n\nHIGHLIGHTSO_LIUsers can stack and align single-cell traces for different molecular reporters\nC_LIO_LIComputational models with adjustable parameters simulate cell cycle progression\nC_LIO_LIVirtual growth curves and flow cytometry assays predict cell cycle behaviors\nC_LI

systems biology↗

bioRxiv2017

Ratiometric Distribution Of OCT4 During Stem Cell Division Controls The Balance Between Self-Renewal And Differentiation

Clonal cells can make different fate decisions, but it is often unclear to what extent these decisions are autonomous or predetermined. Here, we introduce a live-cell reporter for the pluripotency factor OCT4 into human embryonic stem cells to understand how they choose between self-renewal and differentiation. By tracing the histories of individual cells over multiple generations, we found that cells whose offspring were destined to differentiate showed decreased expression of OCT4 long before exposure to the differentiation stimulus. OCT4 levels were lineage-dependent; however, during cell division, mother cells distributed OCT4 asymmetrically to daughters. The resulting ratio of OCT4 between sister cells--established within minutes of mitosis--was predictive of downstream fates: cells receiving a greater ratio of maternal OCT4 showed sustained OCT4 levels and a reduced capacity to differentiate. Our observations imply that the choice between two developmental fates is almost entirely predetermined at the time of cell birth through inheritance of a pluripotency factor.

systems biology↗