Biology subjects

Oldham, M.

Publications and source records attributed to Oldham, M..

4 recordsLinked to original sources

bioRxiv2025

An Investigation of mini-GRID Radiation therapy with Immune Checkpoint Blockade in a Murine Tumor Model

Spatially fractionated radiotherapy has shown potential to improve therapeutic outcomes possibly with an immunogenic mechanistic component. Here we report on in vivo mouse studies investigating mini-GRID pencil-beam radiotherapy combined with anti-PD-1 immune checkpoint blockade. Methods: GRID therapy was delivered at 225kV using the XStrahl Small Animal Radiation Research Platform with two custom lead mini-GRIDs, each consisting of an array of equally spaced holes: 1 mm diameter with 1mm spacing and 254 {micro}m diameter with 508 {micro}m spacing. GRID dosimetry was characterized using EBT3 film to determine peak-to-valley dose ratios and output. Two studies were performed with C57BL/6J mice bearing subcutaneous LLC1 flank tumors. In the first, mice (n=5/group) were treated in 3 groups with a single fraction: 15 Gy open field, 15 Gy 1 mm GRID, or 24 Gy 1 mm GRID. In the second, mice (n=6-7/group) were treated with fractionated GRID radiotherapy in 5 groups: 15 Gy open field x 3 fractions, 15 Gy hemi-irradiation x 3 fractions, (15 Gy 1 mm GRID x 3 fractions, or 15 Gy 254 {micro}m GRID x 3 fractions. All mice were treated with 200 g anti-PD-1 antibody on days 0, 3, and 6, then weekly until humane endpoint (tumor >15 mm in any dimension or ulceration). Results: Peak to valley ratios were 24.5 {+/-} 0.6 and 19.8 {+/-} 0.7 for the 1 mm and 254 {micro}m GRIDs, respectively. Tumor growth and mean survival times in both studies were significantly shorter for all non-open field arms (p < 0.05; Log Rank for survival; 2-way ANOVA for tumor growth). Conclusions: Two novel mini-GRIDs were characterized and tested in combination with anti-PD-1 therapy. In this study, neither single dose nor fractionated GRID therapy with anti-PD-1 improved tumor growth delay or survival. Similarly, hemi-irradiation resulted in worse tumor control compared to conventional open field radiotherapy.

cancer biology↗

bioRxiv2025

Investigating Rat-Brain Normal Tissue and Tumor FLASH Effects with a Novel Very High Energy Electron Beam

PurposeUltra-high dose rate (FLASH) irradiation is reported to reduce normal tissue toxicity while maintaining tumor control, however mechanism(s) remain obscure. To study FLASH mechanisms in brain tissue, we developed a novel experimental platform featuring a unique high-energy electron linear accelerator (High Intensity Gamma Source, HIGS) paired with an organotypic ex vivo brain metastasis model. MethodsWe varied inter-pulse spacing to modulate the mean dose rate (MDR) of our unique 35 MeV electron beam, while maintaining extremely high instantaneous dose rate (IDR), and used film dosimetry to characterize dosimetry and targeting accuracy. We combined this HIGS-FLASH beam with an organotypic rat brain slice/breast carcinoma co-culture model of brain metastasis to assess effects on normal and neoplastic tissues. Live cell and bioluminescence imaging demonstrated cancer cell growth effects, while normal tissue responses and immune activation were assessed with live cell imaging, cytokine profiles, and confocal microscopy. We performed comparison experiments with 20 MeV electrons from a Varian clinical linear accelerator (VCLA) operating at conventional dose rate. ResultsThe highest IDR of the HIGS-FLASH beam to-date was 20.7 {+/-} 0.6 MGy/s, with maximum MDR of 20.7 MGy/s (1 s pulse of 20.7 Gy). Beam targeting was accurate to < 1 mm and reproducible. HIGS-FLASH and VCLA dose rates equivalently decreased cancer cell growth. HIGS-FLASH irradiation significantly increased TNF and fractalkine levels and confocal microscopy revealed distinct changes in microglial morphology in normal brain slices, suggesting microglia activation following HIGS-FLASH irradiation. ConclusionsOur novel experimental platform produces extremely high dose rates and rapid normal/neoplastic tissue readouts for mechanistic research into the effects of FLASH radiation on the brain. HIGS-FLASH irradiation induces comparable cancer cell growth inhibition but differential effects on cytokines and microglial morphology, suggesting that acute innate immune responses may be involved in FLASH normal tissue effects in the brain.

cancer biology↗

bioRxiv2025

Cortical organoids reveal human-specific roles of METTL5 in neurodevelopment via regulation of CHCHD2

METTL5 is a conserved methyltransferase responsible for catalyzing N6-methyladenosine (m6A) modification on 18S ribosomal RNA. Human patients with mutations in METTL5 show severe microcephaly and intellectual disability, which has not been fully recapitulated in animal models. Given its emerging role in neurodevelopment, we sought to investigate METTL5 function in a human-specific context using cortical forebrain organoids derived from human induced pluripotent stem cells. We generated METTL5 knockout organoids and observed a marked delay in neural stem cell proliferation and the timing of neuronal differentiation, suggesting a critical role for METTL5 in the temporal regulation of neurogenesis. Though METTL5 methylates rRNA near the decoding center, the mechanism of this methylation remains highly contested. In our cortical organoids, broad translational changes mirror stress response rather than transcript-specific regulation of translation. Transcriptomic analysis further revealed a significant downregulation of CHCHD2, a nuclear-encoded mitochondrial gene linked to cellular energy metabolism and neurodevelopmental processes. Overexpression of CHCHD2 rescued proliferation defects of METTL5-KO neural progenitor cells, suggesting dysregulation of CHCHD2 is heavily involved in the phenotypes of METTL5 mutant patient brains. These findings highlight a previously uncharacterized link between METTL5, ribosomal RNA modification, and cellular metabolism essential for proper human brain development.

molecular biology↗

bioRxiv2022

Atm loss does not radiosensitize a primary mouse model of Pten-deleted brainstem glioma

Diffuse midline gliomas arise in the brainstem and other midline brain structures and cause a large proportion of childhood brain tumor deaths. Radiation therapy is the most effective treatment option, but these tumors ultimately progress. Inhibition of the phosphoinositide-3-kinase (PI3K)-like kinase ataxia telangiectasia mutated (ATM), which orchestrates the cellular response to radiation-induced DNA damage, may enhance the efficacy of radiation therapy. Diffuse midline gliomas in the brainstem contain loss-of-function mutations in the tumor suppressor PTEN, or functionally similar alterations in the phosphoinositide-3-kinase (PI3K) pathway, at moderate frequency. Here, we sought to determine if Atm inactivation could radiosensitize a primary mouse model of brainstem glioma driven by Pten loss. Using Cre/loxP recombinase technology and the RCAS/TVA retroviral gene delivery system, we established a mouse model of brainstem glioma driven by Pten deletion. We find that Pten-null brainstem gliomas are relatively radiosensitive at baseline. In addition, we show that deletion of Atm in the tumor cells does not extend survival of mice bearing Pten-null brainstem gliomas after focal brain irradiation. These results characterize a novel primary mouse model of PTEN-mutated brainstem glioma and provide insights into the mechanism of radiosensitization by Atm deletion, which may guide the design of future clinical trials. Brief SummaryWe develop a mouse model of PTEN-mutated brainstem glioma and find that perturbation of the ATM does not enhance radiation efficacy in this model.

cancer biology↗