Biology subjects

Gartrell, R. D.

Publications and source records attributed to Gartrell, R. D..

4 recordsLinked to original sources

bioRxiv2024

Combined inhibition of SHP2 overcomes adaptive resistance to type 1 BRAF inhibitors in BRAF V600E-driven high-grade glioma

BRAF-mutant gliomas can be therapeutically targeted with BRAF mutant-selective inhibitors, yet responses are often transient due to short-term adaptive or long-term treatment-emergent resistance. We hypothesized that vertical inhibition of multiple signaling nodes could improve the durability of BRAF inhibition and prevent or overcome adaptive resistance. Using human tissue samples, we identified frequent RAS pathway reactivation in gliomas resistant to BRAF inhibitors, suggesting a common escape mechanism. Using patient-derived cell lines, we observed that upregulation of RAS activity was an adaptive response to BRAFi and that knockdown of SHP2, a central regulator of RAS activity, resulted in enhanced sensitivity to BRAF or MEK inhibition. Moreover, combined small molecule inhibition with SHP2 and BRAF or MEK inhibitors increased the depth and durability of ERK pathway inhibition, as well as prevented paradoxical upregulation of RAS activity. RNA sequencing analysis revealed deeper suppression of ERK transcriptional output with combined therapy, along with decreased reactivation of EGFR. Combined SHP2/BRAF small molecule inhibitors prevented growth and induced cell death in some cell line models. In cell lines with treatment-emergent resistance, moreover, combined SHP2 and BRAF inhibition overcame resistance to BRAF inhibitor monotherapy. In vivo orthotopic and patient-derived xenograft models confirmed enhanced tumor growth inhibition with combined therapy. Together, our findings demonstrate the critical role of RAS/ERK signaling reactivation in driving resistance to BRAF inhibition in glioma, and demonstrate the potential utility for adding SHP2 inhibitors to overcome resistance in BRAF V600E mutant glioma. SignificanceThe addition of a SHP2i to BRAFi in BRAF-V600E glioma cells prevents tumor growth and can overcome resistance to BRAFi in preclinical models in vitro and in vivo.

cancer biology↗

bioRxiv2024

Elucidation and Pharmacologic Targeting of Master Regulator Dependencies in Coexisting Diffuse Midline Glioma Subpopulations

Diffuse Midline Gliomas (DMGs) are universally fatal, primarily pediatric malignancies affecting the midline structures of the central nervous system. Despite decades of clinical trials, treatment remains limited to palliative radiation therapy. A major challenge is the coexistence of molecularly distinct malignant cell states with potentially orthogonal drug sensitivities. To address this challenge, we leveraged established network-based methodologies to elucidate Master Regulator (MR) proteins representing mechanistic, non-oncogene dependencies of seven coexisting subpopulations identified by single-cell analysis--whose enrichment in essential genes was validated by pooled CRISPR/Cas9 screens. Perturbational profiles of 372 clinically relevant drugs helped identify those able to invert the activity of subpopulation-specific MRs for follow-up in vivo validation. While individual drugs predicted to target individual subpopulations--including avapritinib, larotrectinib, and ruxolitinib--produced only modest tumor growth reduction in orthotopic models, systemic co-administration induced significant survival extension, making this approach a valuable contribution to the rational design of combination therapy.

systems biology↗

bioRxiv2022

FLASH and Conventional Radiation Induce Differential Immune Responses in Diffuse Intrinsic Pontine Glioma, Highlighting Potential for Combination Immunotherapy

PurposeDiffuse Midline Glioma (DMG) is a fatal tumor traditionally treated with radiotherapy (RT) and previously characterized as having a non-inflammatory tumor immune microenvironment (TIME). FLASH is a novel RT technique using ultra-high dose rate, which is associated with decreased toxicity and effective tumor control. However, the effect of FLASH and conventional (CONV) RT on the DMG TIME have not yet been explored. MethodsHere, we perform single-cell RNA sequencing and flow cytometry on immune cells isolated from an orthotopic syngeneic murine model of brainstem DMG following the use of FLASH (90Gy/sec) or CONV (2Gy/min) dose-rate RT, and compare to unirradiated tumor (SHAM). ResultsAt day 4 post-RT, FLASH exerts similar effects as CONV in the predominant microglial (MG) population, including the presence of two activated subtypes. However, at day 10 post-RT, we observe a significant increase in type 1 interferon alpha receptor (IFNAR+) in MG in CONV and SHAM compared to FLASH. In the non-resident myeloid clusters of macrophages (MACs) and dendritic cells (DCs), we find increased type 1 interferon (IFN1) pathway enrichment for CONV compared to FLASH and SHAM by scRNA-seq. We observe this trend by flow cytometry at day 4 post-RT in IFNAR+ MACs and DCs, which equalizes by day 10 post-RT. DMG control and murine survival are equivalent between RT dose rates. ConclusionOur work is the first to map CONV and FLASH immune alterations of the DMG TIME with single-cell resolution. While DMG tumor control and survival are similar between CONV and FLASH, we find that changes in immune compartments differ over time. Importantly, while both RT modalities increase IFN1, we find that the timing of this response is cell-type and dose-rate dependent. These temporal differences, particularly in the context of tumor control, warrant further study.

immunology↗

bioRxiv2022

Radiation therapy promotes unsaturated fatty acids to maintain survival of glioblastoma

PurposeRadiation therapy (RT) is essential for the management of glioblastoma (GBM). However, GBM frequently relapses within the irradiated margins, thus suggesting that RT might stimulate mechanisms of resistance that limits its efficacy. GBM is recognized for its metabolic plasticity, but whether RT-induced resistance relies on metabolic adaptation remains unclear. MethodsWe analyzed in vitro extracellular flux and profiled targeted metabolites as well as free fatty acids in two syngenic models of glioblastomas 24hrs post RT. Metabolic adaptation of irradiated GBM were confirmed in vivo by mass spectrometry imaging. The role of the fatty acid synthase (FASN) in RT-induced lipid metabolites was assessed by genetical and pharmacological inhibition of Fasn in irradiated GBM cells. The impact of FASN-mediated lipids on endoplasmic reticulum (ER) stress and apoptosis of irradiated GBM cells were performed by transmission electronic microscopy, western blot, clonogenic assay and flow cytometry. Inhibition of FASN combined with focal RT was assessed in mice. Analysis of a public dataset of GBM patients was performed to correlate preclinical findings. ResultsHere, we show in vitro and in vivo that irradiated GBM tumors switch their metabolic program to accumulate lipids, especially unsaturated fatty acids. This resulted in an increase formation of lipid droplets to prevent ER stress. We uncovered that FASN is critical for lipid accumulation of irradiated GBM and demonstrate that genetic suppression and pharmacological inhibition of FASN lead to mitochondrial dysfunction and apoptosis. Combination of FASN inhibition with focal RT improved the median survival of GBM-bearing mice. Supporting the translational value of these findings, retrospective analysis of the GLASS consortium dataset of matched GBM patients revealed an enrichment in lipid metabolism signature in recurrent GBM compared to primary. ConclusionsOverall, these results demonstrate that RT drives GBM resistance by generating a lipogenic environment permissive to GBM survival. Targeting lipid metabolism might be required to develop more effective anti-GBM strategies.

cell biology↗