Biology subjects

Simeone, D. M.

Publications and source records attributed to Simeone, D. M..

5 recordsLinked to original sources

bioRxiv2025

Clonal Heterogeneity in Human Pancreatic Ductal Adenocarcinoma and Its Impact on Tumor Progression

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer characterized by profound desmoplasia and cellular heterogeneity, which cannot be fully resolved using traditional bulk sequencing approaches. To understand the contribution of this heterogeneity to PDAC biology, we analyzed a large cohort of primary human PDAC samples (n = 62), profiling 443,451 single cells and 53,236 spatial transcriptomic spots using a combined single-cell RNA sequencing and spatial transcriptomics approach. Our analysis revealed significant intratumoral heterogeneity, with multiple genetically distinct neoplastic clones co-existing within individual tumors. These clones exhibited diverse transcriptional states and subtype profiles, challenging the traditional binary classification of PDAC into basal and classical subtypes; instead, our findings support a transcriptional continuum influenced by clonal evolution and spatial organization. Additionally, these clones each interacted uniquely with surrounding cell types in the tumor microenvironment. Phylogenetic analysis uncovered a rare but consistent classical-to-basal clonal transition associated with MYC amplification and immune response depletion, which were validated experimentally, suggesting a mechanism driving the emergence of a more aggressive basal clonal phenotype. Spatial analyses further revealed dispersed clones enriched for epithelial-to-mesenchymal transition (EMT) activity and immune suppression, correlating with metastatic potential and colonization of lymph node niches. These dispersed clones tended to transition toward a basal phenotype, contributing to disease progression. Our findings highlight the critical role of clonal diversity, transcriptional plasticity, and TME interactions in shaping human PDAC biology. This work provides new insights into the molecular and spatial heterogeneity of PDAC and offers potential avenues for therapeutic intervention targeting clonal evolution and the mechanisms driving metastasis.

cancer biology↗

bioRxiv2022

Single-Cell RNA Sequencing Reveals the Effects of Chemotherapy on Human Pancreatic Adenocarcinoma and its Tumor Microenvironment

The tumor microenvironment (TME) in pancreatic ductal adenocarcinoma (PDAC) is a complex ecosystem that drives tumor progression; however, in-depth single cell characterization of the PDAC TME and its role in response to therapy is lacking. We performed single-cell RNA sequencing on freshly collected human PDAC samples either before or after chemotherapy. Overall, we found a heterogeneous mixture of basal and classical cancer cell subtypes, along with distinct cancer-associated fibroblast and macrophage subpopulations. Strikingly, classical and basal-like cancer cells exhibited similar transcriptional responses to chemotherapy, and did not demonstrate a shift towards a basal-like transcriptional program among treated samples. We observed decreased ligand-receptor interactions in treated samples, particularly TIGIT on CD8+ T cells and its receptor on cancer cells, and identified TIGIT as the major inhibitory checkpoint molecule of CD8+ T cells. Our results suggest that chemotherapy profoundly impacts the PDAC TME and may promote resistance to immunotherapy.

cancer biology↗

bioRxiv2021

Ultra-low Input Circulating Tumor DNA Detection by MED-Amp in Early-Stage Pancreatic Cancer

PurposeThe clinical utility of circulating tumor DNA (ctDNA) has been shown in advanced pancreatic ductal adenocarcinoma (PDA). However, diagnostic sensitivity of many ctDNA assays is low in resectable and locally advanced disease, where tumor burden is substantially lower. We have previously described Multiplex Enrichment using Droplet Pre-Amplification (MED-Amp), a multiplexed panel for the detection of the most common oncogenic KRAS mutations in PDA. In this study, we aimed to assess the diagnostic sensitivity of MED-Amp for detection of rare mutant alleles present in the plasma of patients with localized PDA. Experimental DesignWe retrospectively analyzed ninety-eight plasma samples from 51 patients with various stages of localized disease. For comparison, we measured ctDNA levels in 20 additional patients with metastatic PDA. The MED-Amp assay was used to measure the abundance of the four most common KRAS codon 12 mutations (G12C/D/R/V). We correlated the presence and quantity of ctDNA with overall survival (OS) as well as progression-free survival (PFS). Using serial plasma draws, we also assessed the relationship between changes in ctDNA allelic frequency and progression. ResultsKRAS-positive ctDNA was detected in 52.9% of localized PDA and 75% of metastatic samples tested using DNA inputs as low as 2 ng. As previously reported, the presence of KRAS mutant ctDNA was correlated with worse OS for all disease stages (p = 0.02). In patients with localized PDA high ctDNA levels also correlated with significantly worse median OS (533 days vs 1090 days) and PFS (192 days vs 787 days). We also studied a small cohort of serial plasma draws to observe the relationship between ctDNA fold change and PFS. We found 83% of patients with increased fold change in mutant KRAS experienced disease progression (n=6). In contrast, 75% (n=4) of patients with decreased fold change remained disease-free (p=0.03). ConclusionsMED-Amp is a flexible and cost-effective approach for measurement of ctDNA in patients with localized cancer. Though this study focused on KRAS mutation detection, this assay could be adapted for a number of common oncogenic alterations. Statement of translational relevanceOnly 25% of pancreatic ductal adenocarcinoma (PDA) patients with localized disease survive five years post-resection. It is hypothesized PDA undergoes dissemination at the earliest stages of tumor formation, driving formation of occult metastases which go undetected using conventional screening methods. Development of high specificity, high sensitivity biomarkers is critical to improving patient mortality. Circulating tumor DNA (ctDNA) has gained increasing acceptance as a non-invasive prognostic in metastatic disease. However, the sensitivity of most targeted ctDNA assays precludes reliable detection of localized and resected disease. Here, we present a digital droplet PCR assay for multiplexed enrichment and detection of KRAS mutations, the most commonly mutated oncogene in PDA. This assay preserves ctDNA allelic frequency in the original sample, while increasing the molecular signal over 50-fold. This study shows the ctDNA has potential diagnostic value in early-stage PDA, and that digital preenrichment of cell-free DNA increases overall assay sensitivity without sacrificing specificity.

genomics↗

bioRxiv2018

SHP2 Inhibition Abrogates MEK inhibitor Resistance in Multiple Cancer Models

Adaptive resistance to MEK inhibitors (MEK-Is) typically occurs via induction of genes for different receptor tyrosine kinases (RTKs) and/or their ligands, even in tumors of the same histotype, making combination strategies challenging. SHP2 (PTPN11) is required for RAS/ERK pathway activation by most RTKs, and might provide a common resistance node. We found that combining the SHP2 inhibitor SHP099 with a MEK-I inhibits proliferation of multiple cancer cells in vitro. PTPN11 knockdown/MEK-I had similar effects, while expressing SHP099-binding mutants conferred resistance, demonstrating that SHP099 was on-target. This combination was efficacious in xenograft and/or genetically engineered models of KRAS-mutant pancreas cancer and ovarian cancer and in wild-type RAS-expressing triple negative breast cancer. Biochemical studies show that SHP099 impedes SOS/RAS/MEK/ERK1/2 reactivation in response to MEK-Is and blocks ERK1/2-dependent transcriptional programs. SHP099 alone also inhibited RAS activation in some, but not all, KRAS-mutant lines. Hence, SHP099/MEK-I combinations could have therapeutic utility in multiple malignancies.\n\nSIGNIFICANCEMEK inhibitors have shown limited efficacy as single agents because of the rapid development of adaptive resistance. We find that combining SHP2 and MEK inhibition abrogates adaptive resistance in multiple cancer models, expressing mutant and wild-type KRAS.

cancer biology↗

bioRxiv2017

HNF1A is a Novel Oncogene and Central Regulator of Pancreatic Cancer Stem Cells

The biological properties of pancreatic cancer stem cells (PCSCs) remain incompletely defined and the central regulators are unknown. By bioinformatic analysis of a PCSC-enriched gene signature, we identified the transcription factor HNF1A as a putative central regulator of PCSC function. Levels of HNF1A and its target genes were found to be elevated in PCSCs and tumorspheres, and depletion of HNF1A resulted in growth inhibition, apoptosis, impaired tumorsphere formation, PCSC depletion, and downregulation of OCT4 expression. Conversely, HNF1A overexpression increased PCSC numbers and tumorsphere formation in pancreatic cancer cells and drove PDA cell growth. Importantly, depletion of HNF1A in primary tumor xenografts impaired tumor growth and depleted PCSCs in vivo. Finally, we established an HNF1A-dependent gene signature in PDA cells that significantly correlated with reduced survivability in patients. These findings identify HNF1A as a central transcriptional regulator of the PCSC state and novel oncogene in pancreatic ductal adenocarcinoma.

cancer biology↗