Biology subjects

Cook, K. L.

Publications and source records attributed to Cook, K. L..

4 recordsLinked to original sources

bioRxiv2025

Proton FLASH Exposure Preserves Gut Commensal Microbiomes and Spares Intestinal Stem Cells

Emerging evidence shows Proton FLASH radiotherapy can spare normal tissues while maintaining anti-tumor efficacy. However, its impact on intestinal stem cell (ISC) populations and the gut microbiome remains unclear. This is critical, as the gut microbiome influences ISC radiosensitivity. In a mouse model of radiation-induced gastrointestinal syndrome, FLASH-irradiated mice exhibited better survival and less crypt-villus damage compared to mice exposed to conventional proton irradiation. Using scRNA-sequencing, we demonstrated that proton FLASH exposure using pulsed pencil beam scanning spares two distinct ISC populations--Lgr5+ CBCs and a Clu+, Mif+, Fabp2+, Anxa2+ revival stem cell (revSC) population--by modulating oxidative stress and cell cycle progression. Analysis of alpha and beta diversity demonstrated that FLASH modulates gut microbiota composition without compromising overall species richness. Notably, FLASH-irradiated mice had higher abundances of Alistipes sp. and Akkermensia sp., both known for protective effects on ISCs and the intestinal mucosa. The critical role of microbiome in FLASH-mediated sparing effect against radiation toxicity was further confirmed by fecal microbiota transplantation, where FLASH-donor microbiota demonstrated reduced lethality in recipients exposed to proton irradiation with conventional dose rate. Our findings highlight the crucial role of the microbiome in the FLASH-mediated sparing of the mucosal epithelium.

bioengineering↗

bioRxiv2023

Anti-CD47 immunotherapy as a therapeutic strategy for the treatment of breast cancer brain metastasis

The presence of cell surface protein CD47 allows cancer cells to evade innate and adaptive immune surveillance resulting in metastatic spread. CD47 binds to and activates SIRP on the surface of myeloid cells, inhibiting their phagocytic activity. On the other hand, CD47 binds the matricellular protein Thrombospondin-1, limiting T-cell activation. Thus, blocking CD47 is a potential therapeutic strategy for preventing brain metastasis. To test this hypothesis, breast cancer patient biopsies were stained with antibodies against CD47 to determine differences in protein expression. An anti-CD47 antibody was used in a syngeneic orthotopic triple-negative breast cancer model, and CD47 null mice were used in a breast cancer brain metastasis model by intracardiac injection of the E0771-Br-Luc cell line. Immunohistochemical staining of patient biopsies revealed an 89% increase in CD47 expression in metastatic brain tumors compared to normal adjacent tissue (p [≤] 0.05). Anti-CD47 treatment in mice bearing brain metastatic 4T1br3 orthotopic tumors reduced tumor volume and tumor weight by over 50% compared to control mice (p [≤] 0.05) and increased IBA1 macrophage/microglia marker 5-fold in tumors compared to control (p [≤] 0.05). Additionally, CD47 blockade increased the M1/M2 macrophage ratio in tumors 2.5-fold (p [≤] 0.05). CD47 null mice had an 89% decrease in metastatic brain burden (p [≤] 0.05) compared to control mice in a brain metastasis model. Additionally, RNA sequencing revealed several uniquely expressed genes and significantly enriched genes related to tissue development, cell death, and cell migration tumors treated with anti-CD47 antibodies. Thus, demonstrating that CD47 blockade affects cancer cell and tumor microenvironment signaling to limit metastatic spread and may be an effective therapeutic for triple-negative breast cancer brain metastasis.

cancer biology↗

bioRxiv2023

Absence of CD47 in the tumor microenvironment modulates tumor metabolism and immunosuppressive signatures limiting breast cancer progression.

The majority of breast cancers are generally considered immune-deprived tumors. This lack of immunogenicity severely hinders effectiveness of current immunotherapy approaches limiting therapeutic options to control disease. Therefore, we need new biomarkers to determine and enhance immune responses to improve the outcome of cancer patients experiencing invasive disease. Our data in matched human patient biopsies show that CD47 expression increases from primary to metastatic tumors. CD47 is an integral membrane protein that impairs antitumor immunosurveillance and influences normal tissue metabolism. However, whether CD47 plays a role in regulating tumor bioenergetics is unknown. A carcinogen-induced mouse mammary carcinogenesis model demonstrates that the absence of CD47 reduces tumor burden, which is associated with a distinct metabolic signature compared to WT tumors. Depletion of several lipid metabolites was observed in the absence of CD47, and metabolic dependency experiments suggest that anti-sense blockade of CD47 limits reliance on fatty acid oxidation as a fuel supporting cellular respiration on cancer cells. Our global metabolomics analysis also implicated the absence of CD47 in downregulation of immunosuppressive metabolites of the tryptophan and prostaglandin pathways. Spatial proteomic analysis revealed increased immune infiltrate and substantial reduction in immunosuppressive immune checkpoint proteins in the absence of CD47 with the highest reduction in intra-tumoral PD-L1 expression. Since anti-PD-L1 therapy is used in the current strategy to treat triple-negative breast cancer (TNBC), we targeted CD47 in an EMT-6 syngeneic TNBC model. The in vivo knockdown of CD47 sensitized tumors to anti-PD-L1 therapy to decrease tumor burden and increase intratumoral cytotoxic T cells. Therefore, targeting CD47 may be a suitable immunotherapeutic option to limit immunosuppression and enhance the efficacy of immune checkpoint blockade.

cancer biology↗

bioRxiv2021

A Novel Model of Estrogen Receptor-Positive Breast Cancer Bone Metastasis with Antiestrogen Responsiveness

Most women diagnosed with breast cancer (BC) have estrogen receptor alpha positive (ER+) disease. ER+ BC preferentially metastasizes to bone; at which time it is considered incurable. Treatments for bone metastasis have not advanced in decades, in part due to a lack of appropriate ER+ BC bone metastasis models. We developed an immunocompetent ER+ BC murine model with spontaneous bone metastasis and antiestrogen responsiveness. To do this, we transduced triple-negative (TN) bone-tropic murine BC cell lines 4T1.2 and E0771/Bone to express ER. These cells were then injected into the mammary fat pads of Balb/c (n=21) or C57Bl/6 (n=27), respectively. Once tumors established, mice were treated with either the selective estrogen receptor modulator (SERM) tamoxifen (TAM), the selective estrogen receptor degrader (SERD) ICI 182,780 (ICI, Faslodex, fulvestrant), or vehicle control for 21 days. Tumor volumes and weights significantly decreased in the ER+ groups treated with TAM and ICI compared with ER+ vehicle-treated groups. Staining for immune profiles and total RNA sequencing demonstrated modified immune cell infiltration between TN and ER-derived tumors. Approximately 25% of the mice with ER+ 4T1.2 tumors developed metastases to long bones while none of the mice with TN tumors developed metastases. This immunocompetent ER+ 4T1.2 BC model may allow for further exploration of ER+ BC bone metastasis mechanisms and for the development of new therapeutics for women diagnosed with bone metastasis from ER+ BC. Simple SummaryEstrogen receptor alpha positive (ER+) breast cancer is the most common subtype of breast cancer. When it metastasizes to bone, it becomes incurable. Little advancement has occurred in the treatment of bone metastasis from ER+ breast cancer, partly due to the lack of animal models. To establish an animal model of ER+ BC, we genetically modified two triple-negative breast cancer cell lines to express ER and injected the cell lines into murine mammary glands. Mice were treated with standard antiestrogen therapies, the selective estrogen receptor modulator tamoxifen or the selective estrogen receptor degrader ICI 182,780. We found that compared to mice with triple-negative breast cancer, mice with ER+ breast cancer developed bone metastases and were responsive to antiestrogen therapy. This model allows for further exploration of bone metastasis mechanisms and for the development of new therapeutics, translating into improved clinical outcomes for women with bone metastasis from ER+ breast cancer.

cancer biology↗