Metastatic Cancer Model Research Articles

Abstract IA013: Stromal and immune plasticity shape the metastatic microenvironment

Abstract Mortality from cancer is almost exclusively due to metastasis to distant organs. Therefore, understanding the biology of tumor metastasis is the most significant challenge in cancer research today. The tumor microenvironment is central in supporting tumor growth. However, the role of the metastatic microenvironment in the multistage process of metastasis is largely unresolved. Since each metastatic microenvironment exerts specific functions that support or oppose colonization by disseminated tumor, understanding distinct organ-specific mechanisms that enable metastatic growth is of crucial importance. Utilizing mouse models of breast cancer metastasis to lungs, brain and bone, combined with clinical data from patients with metastatic disease, we uncover the mechanisms that underlie the early events of metastasis to provide the mechanistic basis for therapeutic targeting to reduce metastatic relapse. Our findings reveal organ-specific co-evolution of stromal and immune cells at distinct metastatic niches, that lead to the formation of an immunosuppressive microenvironment, permissive to metastatic growth. Analysis of human metastasis reveal that these pathways are also operative in human disease. We show that genetic and/or pharmacological targeting of specific nodes in the interactions between stromal and immune cells results in attenuated metastasis and improved survival. Our findings imply that combinatorial therapeutic strategies to target the crosstalk between stromal and immune cells at the metastatic microenvironment may be a promising treatment strategy to inhibit metastatic relapse. Citation Format: Neta Erez. Stromal and immune plasticity shape the metastatic microenvironment [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr IA013.

Abstract PR010: TRPV1+ sensory innervation as a novel driver of ovarian cancer progression

Abstract Introduction: A primary challenge currently faced in ovarian cancer (OVCA) treatment is that patient prognoses remain poor despite current therapeutic interventions. Overall survival for OVCA patients with advanced disease is <30%, and most patients recur within 5 years. As such, there is a critical need for new therapies that can overcome OVCA chemoresistance and produce meaningful increases in patient survival. Recent efforts that have targeted the tumor microenvironment (TME), such as immune checkpoint inhibitors, have been successful in some cancers but have limited therapeutic benefit when used to treat OVCA. Thus, new TME targeting therapies must be developed to effectively treat OVCA patients. A novel TME component ripe for therapeutic targeting are the nerve fibers that infiltrate tumors. Recent studies in several cancers have shown that tumor innervation can promote tumor growth/metastasis. However, in ovarian cancer the role of innervation in promoting cancer progression remains a gap in knowledge. Here we show that Transient Receptor Potential Cation Channel Subfamily V Member 1 (TRPV1)+ sensory innervation plays a significant role in driving ovarian cancer progression. Methods: We performed IHC staining to determine the extent of TRPV1+ nerve infiltration in OVCA vs normal reproductive tissue in patients. We examined the ability of OVCA cells to promote tumor innervation using a neurite outgrowth assay. We utilized both syngeneic and patient derived xenograft (PDX) mouse models of ovarian cancer metastasis, where mouse or human OVCA cells were injected intraperitoneally, to explore the functional role of TRPV1+ sensory nerves in OVCA. TRPV1+ sensory nerves were ablated genetically using TRPV1 driven expression of diphtheria toxin alpha (DTA) or chemically using resiniferatoxin. TRPV1+ sensory nerves were activated pharmacologically using capsaicin. Results: Analysis of patient samples showed that TRPV1+ sensory innervation is much higher in ovarian tumors vs benign reproductive tissue. We found that conditioned media from OVCA cells was able to promote neurite outgrowth of dissociated mouse sensory neurons. Ablation of TRPV1+ sensory nerve fibers in vivo caused reduced tumor burden and prolonged survival in our syngeneic and PDX OVCA mouse models. Stimulation of TRPV1+ sensory nerves with capsaicin accelerated OVCA growth and decreased survival in tumor bearing mice. Conclusions: Our results establish TRPV1+ sensory innervation as a novel driver of OVCA growth/metastasis and a potential therapeutic target for OVCA treatment. Significance to tumor microenvironment field: Our data add to a growing body of evidence that sensory nerves are pro-tumorigenic in multiple cancer types and could be therapeutically targeted for cancer treatment. Citation Format: Matthew Knarr, Katherine Cummins, Dusan Racordon, Hunter Reavis, Timothy Lippert, Ryan Hausler, Priyanka Rawat, Jamie Moon, Dave S.B. Hoon, Roger Greenberg, Paola Vermeer, Ronny Drapkin. TRPV1+ sensory innervation as a novel driver of ovarian cancer progression [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr PR010.

TLR Agonist Nano Immune Therapy Clears Peritoneal and Systemic Ovarian Cancer.

Intraperitoneal (IP) administration of immunogenic mesoporous silica nanoparticles (iMSN) in a mouse model of metastatic ovarian cancer promotes the development of tumor-specific CD8+ T cells and protective immunity. IP delivery of iMSN functionalized with the Toll-like receptor (TLR) agonists polyethyleneimine (PEI), CpG oligonucleotide, and monophosphoryl lipid A (MPLA) stimulated rapid uptake by all peritoneal myeloid subsets. Myeloid cells quickly transported iMSN to milky spots and fat-associated lymphoid clusters (FALCs) present in tumor-burdened adipose tissues, leading to a reduction in suppressive T cells and an increase in activated memory T cells. Two doses of iMSN cleared or reduced ovarian and colorectal cancer and protected against future tumor engraftment. In contrast, subcutaneous (SC) and intravenous (IV) delivery of iMSN were without therapeutic effect in mice with peritoneal metastases, supporting the need for activation of regional immune cells. Remarkably, intraperitoneal delivery of iMSN cleared subcutaneously implanted ovarian cancer, supporting homing of antigen specific T cells to extraperitoneal tumor sites.

Mitochondrial-targeted plastoquinone therapy ameliorates early onset muscle weakness that precedes ovarian cancer cachexia in mice.

Cancer cachexia, and the related loss of muscle and strength, worsens quality of life and lowers overall survival. Recently, a novel 'pre-atrophy' muscle weakness was identified during early-stage cancer. While mitochondrial stress responses are associated with early-stage pre-atrophy weakness, a causal relationship has not been established. Using a robust mouse model of metastatic epithelial ovarian cancer (EOC)-induced cachexia, we found the well-established mitochondrial-targeted plastoquinone SkQ1 partially prevents pre-atrophy weakness in the diaphragm. Furthermore, SkQ1 improved force production during atrophy without preventing atrophy itself in the tibialis anterior and diaphragm. EOC reduced flexor digitorum brevis (FDB) force production and myoplasmic free calcium ([Ca 2+ ] i ) during contraction in single muscle fibers, both of which were prevented by SkQ1. Remarkably, changes in mitochondrial reactive oxygen species and pyruvate metabolism were heterogeneous across time and between muscle types which highlights a considerable complexity in the relationships between mitochondria and muscle remodeling throughout EOC. These discoveries identify that muscle weakness can occur independent of atrophy throughout EOC in a manner that is linked to improved calcium handling. The findings also demonstrate that mitochondrial-targeted therapies exert a robust effect in preserving muscle force during the early pre-atrophy period and in late-stage EOC once cachexia has become severe.

7065 Eradication Of Metastatic ER-Positive Breast, Ovarian, Endometrial And Lung Cancer In Mouse Xenografts And Patient Derived Organoids

Abstract Disclosure: D.J. Shapiro: None. D. Duraki: None. S. Ghosh: None. J. Zhu: None. C. Mao: None. J. Kim: None. M. Jabeen: None. M.W. Boudreau: None. M.P. Mulligan: None. R. Romero: None. Y. Han: None. J. Zhang: None. E.R. Nelson: None. O. Olopade: None. G. Chang: None. P.J. Hergenrother: Advisory Board Member; Self; Systems Oncology. We described the tumor protective estrogen-ER activated anticipatory Unfolded Protein Response (a-UPR) pathway. Here we report a previously undescribed multistep pathway by which very rapid estrogen-ER activation of the a-UPR authorizes and regulates subsequent transcription of the estrogen-ER transcriptome. We repurposed the tumor protective a-UPR through identification and optimization of the small molecule ErSO and its family of agents. ErSO acts non-competitively with estrogen to induce lethal hyperactivation of the a-UPR. In orthotopic mouse xenografts containing wild type or mutant ER, ErSO often induced complete regression without recurrence of large primary breast tumors and of most lung, bone and liver metastases, near complete regression of normally lethal brain metastases and complete destruction of patient derived organoids (PDOs) in Matrigel. Six research teams in four locations have confirmed the remarkable effectiveness of ErSO in ER-positive breast cancer. We next evaluated ErSO in diverse ER containing cancers in which estrogen is not the primary driver of proliferation. ErSO often induced complete regression in multiple orthotopic xenograft models of primary and metastatic ovarian cancer. Using fresh ovarian cancer organoids immediately after removal from patients by paracentesis, ErSO destroyed the organoids from some patients with advanced stage III and stage IV disease. ErSO was effective in a mouse xenograft model of endometrial cancer and, surprisingly, even in ERalpha positive lung cancer. Our ongoing studies demonstrate ErSO induces complete, or near complete regression, of primary and metastatic lung cancer in mice. Studies of ErSO’s mechanism of action using genome-wide CRISPR screens with positive selection, and other techniques, unveil a novel death pathway in which ErSO activation of the a-UPR triggers rapid calcium release into the cell body. The calcium opens the plasma membrane TRPM4 sodium channel, leading to an influx of extracellular sodium. This swells the cells, resulting in osmotic stress that sustains lethal a-UPR activation. A little-studied cytoskeleton regulator we identified, FGD3, plays a pivotal role in whether the cells rupture their membranes, inducing necrotic cell death. Notably, medium from diverse cancer cells killed by ErSO-induced necrosis robustly activates human macrophage and induces their migration - potentially facilitating immunotherapy. These studies describe a pathway from a multiprotein ER-SRC-PLC complex, through the a-UPR, to a novel cell swelling and necrosis pathway, that we are exploiting to target diverse, therapeutically challenging, ER-positive human cancers. Presentation: 6/2/2024

Prognostic model for second progression-free survival and overall survival in patients with high-risk metastatic hormone-sensitive prostate cancer treated with abiraterone acetate and androgen deprivation therapy.

To develop and validate a prognostic risk model for high-risk metastatic hormone-sensitive prostate cancer (mHSPC) patients treated with upfront abiraterone acetate (ABI). This retrospective multicenter study involved 233 high-risk mHSPC patients who received upfront ABI, developed by three academic centers. The model was externally validated with an independent cohort of 282 patients. To identify independent prognostic factors for second progression-free survival (PFS2) and develop the best-fitted model, Cox proportional hazards regression, followed by the Akaike information criterion, was used. Patients were categorized into three groups based on their risk scores. PFS2 and overall survival (OS) were evaluated according to the risk groups in the discovery and validation cohorts. The median age was 72 (range 51-89) years, with a median follow-up duration of 27 months. Independent factors linked to PFS2 included an Eastern Cooperative Oncology Group performance status ≥2, a primary Gleason score of 5, an extent of disease score of ≥3 or liver metastasis, and lactate dehydrogenase >220 U/L. Median PFS2 for favorable-, intermediate-, and poor-risk groups were not reached, 43 months, and 16 months, respectively. The median OS was 29 months in the poor-risk group, whereas it was not reached in the favorable- and intermediate-risk groups. The 2-year OS rates in the favorable-, intermediate- and poor-risk groups were 94.5%, 80.1%, and 60.3%, respectively. The validation cohort confirmed the risk model's relationship with PFS2 and OS. The median PFS2 and OS in the high-risk group were 21 months and 32 months, respectively. Our prognostic model, including five clinical factors, is useful for patient care and treatment selection in high-risk mHSPC patients treated with ADT plus ABI. The developed model could provide more accurate information, guide treatment decisions, or classify patients in future clinical trials.

HI-Net: A novel histopathologic image segmentation model for metastatic breast cancer via lightweight dataset construction

Since 2020, breast cancer has remained the most prevalent cancer worldwide and the World Health Organisation projects significant increases by 2040, with new cases expected to exceed 3 million annually (a 40% increase) and deaths to surpass 1 million (a 50% increase), highlighting the urgent need for advancements in detection and treatment. Current detection of metastasis is highly dependent on labour-intensive and error-prone pathological examination of large-scale biotissue. Given the high-resolution (100,000 × 100,000 gigapixels) but limited quantity of open-source pathological slide datasets, existing deep learning models face preprocessing challenges. This paper introduces HI-Net, a high-speed panoramic feature-extraction pyramid network for rapid and accurate detection of metastatic breast cancer, balancing panoramic segmentation and local attention. Additionally, a lightweight pathological slide dataset optimised for 512 x 512-pixel resolution, derived from downsampled and reassembled competitive datasets, accelerates training and reduces computational costs. HI-Net demonstrates superior performance on existing medical imaging competition datasets and our lightweight dataset, evidencing its effectiveness across datasets and potential for contributing to the generalisation of intelligent diagnostics.

Effects of HyaRegen gel on tumour proliferation of colorectal peritoneal metastases.

Pressurized intraperitoneal aerosol chemotherapy (PIPAC) is a valuable therapeutic alternative for patients with peritoneal metastases. PIPAC uses a hyaluronic acid-based gel to reduce surgically induced adhesions. The aim of this study was to evaluate the effects of the hyaluronic acid-based gel on tumor dissemination. First, we explored whether the survival of CT26 luciferase-expressing murine colonic tumor cells was correlated with the dose of HyaRegen® Gel, and we determined the half-maximal inhibitory concentration (the IC50) of the gel. Next, we performed an in vitro study of cell survival rates after gel application on day 0 (D0) and day 1 (D1). Finally, we intraperitoneally administered the gel to mice with immunocompetent BALB/c colonic peritoneal metastases (on D0, D5, D10, D14, and D18). Tumor growth was regularly monitored using a bioluminescence assay (on D11, D17, and D21). After all mice had been sacrificed on D21, the body weights and the volumes of intraperitoneal ascites were measured; the Peritoneal Carcinosis Index (PCI) and Ki-antigen 67 scores were calculated. The IC50 value was 70 μL of gel in a total volume of 100 μL. The cell survival rates on D4 were identical in the control group and the two groups that had been treated with gel on D0 and D1. The bioluminescence levels over time were similar in the gel and control groups. The PCI scores were 35.5 ± 2.89 for the control group and 36 ± 2.45 for the gel group (p = 0.8005). The mean Ki-67 index percentages were 37.28 ±1 1.75 for the control group and 34.03 ± 8.62 for the gel group (p = 0.1971). This in vitro and in vivo study using a mouse model of immunocompetent metastatic peritoneal cancer did not reveal any pro- or anti-tumoral effect of HyaRegen® Gel. These findings indicate that the gel can be used to treat PIPACs with minimal apprehension.

SSPH I, A Novel Anti-cancer Saponin, Inhibits EMT and Invasion and Migration of NSCLC by Suppressing MAPK/ERK1/2 and PI3K/AKT/ mTOR Signaling Pathways.

Saponin of Schizocapsa plantaginea Hance I (SSPH I).a bioactive saponin found in Schizocapsa plantaginea, exhibits significant anti-proliferation and antimetastasis in lung cancer. To explore the anti-metastatic effects of SSPH I on non-small cell lung cancer (NSCLC) with emphasis on epithelial-mesenchymal transition (EMT) both in vitro and in vivo. The effects of SSPH I at the concentrations of 0, 0.875,1.75, and 3.5 μM on A549 and PC9 lung cancer cells were evaluated using colony formation assay, CCK-8 assay, transwell assay and wound-healing assay. The actin cytoskeleton reorganization of PC9 and A549 cells was detected using the FITC-phalloidin fluorescence staining assay. The proteins related to EMT (N-cadherin, E-cadherin and vimentin), p- PI3K, p- AKT, p- mTOR and p- ERK1/2 were detected by Western blotting. A mouse model of lung cancer metastasis was established by utilizing 95-D cells, and the mice were treated with SSPH I by gavage. The results suggested that SSPH I significantly inhibited the migration and invasion of NSCLC cells under a non-cytotoxic concentration. Furthermore, SSPH I at a non-toxic concentration of 0.875 μM inhibited F-actin cytoskeleton organization. Importantly, attenuation of EMT was observed in A549 cells with upregulation in the expression of epithelial cell marker E-cadherin and downregulation of the mesenchymal cell markers vimentin as well as Ncadherin. Mechanistic studies revealed that SSPH I inhibited MAPK/ERK1/2 and PI3K/AKT/mTOR signaling pathways. SSPH I inhibited EMT, migration, and invasion of NSCLC cells by suppressing MAPK/ERK1/2 and PI3K/AKT/mTOR signaling pathways, suggesting that the natural compound SSPH I could be used for inhibiting metastasis of NSCLC.

Dual Adjuvant-Loaded Peptide Antigen Self-Assembly Potentiates Dendritic Cell-Mediated Tumor Immunotherapy.

Clinical translation of current cancer vaccine research has been hampered by limited antitumor immune responses due to inefficient antigen delivery and presentation, suboptimal DC and T cell activation. Biomaterial-based nanovaccine offers targeted antigen delivery, protection from degradation in vivo, and prolonged tumor therapeutic efficacy. This study introduces a lipid-coated deoxycholic acid-survivin nanoassembly (DA-L-DSA). Survivin, overexpressed in several cancer cells and involved in cancer cell growth and immune evasion, is selected as a tumor-associated antigen. An major histocompatibility complex class I binding epitope of survivin is engineered into the nanoassembly. R848, TLR 7/8 agonist, and SD-208, TGF-beta receptor1 kinase inhibitor, are coencapsulated into the nanoassembly as potent adjuvants to boost DC maturation and enhance antigen presentation. The DA-L-DSA effectively stimulates the maturation of dendritic cells, migrates into lymph nodes, and enhances T-cell activation and Th1 response. A substantial influx of cytotoxic T lymphocytes into primary tumors is observed in a murine melanoma model and demonstrates anti-metastatic effects in a spontaneous breast cancer metastasis model. Furthermore, DA-L-DSA exhibits a remarkable synergistic effect in the combination therapy with immune checkpoint inhibitors alleviating immunosuppressive tumor microenvironment. Taken together, these findings suggest DA-L-DSA as a promising immuno-therapeutic platform that could be applicable to diverse intractable cancers.

Extracellular vesicle surface display enhances the therapeutic efficacy and safety profile of cancer immunotherapy

Immunotherapy has emerged as a mainstay in cancer therapy, yet its efficacy is constrained by the risk of immune-related adverse events. In this study, we present a nanoparticle-based delivery system that enhances the therapeutic efficacy of immunomodulatory ligands while concurrently limiting systemic toxicity. We demonstrate that extracellular vesicles (EVs), lipid bilayer enclosed particles released by cells, can be efficiently engineered via iEDDA-mediated conjugation to display multiple immunomodulatory ligands on their surface. Display of immunomodulatory ligands on the EV surface conferred substantial enhancements in signaling efficacy, particularly for tumor necrosis factor receptor superfamily (TNFRSF) agonists, where EV surface display served as an alternative FcγR-independent approach to induce ligand multimerization and efficient receptor crosslinking. EVs displaying a complementary combination of immunotherapeutic ligands were able to shift the tumor immune milieu towards an anti-tumorigenic phenotype and significantly suppress tumor burden and increase survival in multiple models of metastatic cancer to a greater extent than an equivalent dose of free ligands. In summary, we present an EV-based delivery platform for cancer immunotherapeutic ligands that facilitates superior anti-tumor responses at significantly lower doses with less side-effects than is possible with conventional delivery approaches.

A laponite-based immunologically active gel delivery system for long-acting tumor vaccine

Traditional bolus vaccines typically require multiple doses, which complicates the vaccination process and may cause missed shots, leading to sub-optimal immunity and reduced vaccine effectiveness. Herein, a gel-based long-acting vaccine system with self-adjuvant properties based on laponite was constructed to simplify vaccination procedures and improve vaccine effectiveness. Firstly, the gel system could recruit multiple types of immune cells to form immune niches. Secondly, it could achieve sustained delivery of antigens to lymph nodes by active transport and passive drainage. Then, the gel system triggered the formation of a large number of germinal centers, which elicited enhanced and durable humoral immune responses, as well as strong cellular immune responses. As a result, it eventually showed good prophylactic and therapeutic effects in a variety of tumor models including melanoma, colorectal cancer and peritoneal metastasis models. By further combining the immunoadjuvant CpG ODN and cytokine IL-12, the effect of the gel-vaccine could be further enhanced. In a murine peritoneal metastasis model of colorectal carcinoma, a single administration of the gel-vaccine resulted in complete tumor eradication in 8/9 mice. In summary, this study developed an immunologically active gel-vaccine system. And as a robust and versatile vaccine platform, by loading different antigens and adjuvants, this gel-vaccine system is expected to realize its better therapeutic potential.

A Diet Lacking Selenium, but Not Zinc, Copper or Manganese, Induces Anticancer Activity in Mice with Metastatic Cancers.

Selenium, zinc, copper, and manganese are essential components of antioxidant enzymes involved in the elimination of reactive oxygen species (ROS). Given that cancer cells produce high levels of ROS and the accumulation of ROS can lead to cell death, cancer cells may be susceptible to strategies that reduce ROS elimination. In this work, we prepared several artificial diets that contained normal carbohydrate, protein, and lipid levels but lacked selenium, zinc, copper, or manganese. The anticancer activity of these diets was examined in a metastatic ovarian cancer model, established by injecting ID8 Trp53-/- murine ovarian cancer cells into the peritoneal cavity of C57BL/6JRj mice. Treatments started 15 days later and consisted of replacing a normal diet with one of the artificial diets for several weeks. A significant improvement in mice survival was observed when the normal diet was replaced with the selenium-free diet. Diets lacking zinc, copper, or manganese showed no significant impact on mice survival. All diets were very well tolerated. The anticancer efficacy of a diet lacking selenium was confirmed in mice with metastatic colon cancer and in mice with metastatic triple-negative breast cancer. These results suggest that diets lacking selenium hold potential for the treatment of metastatic cancers.

Accumulation of liposomes in metastatic tumor sites is not necessary for anti-cancer drug efficacy

BackgroundThe tumor microenvironment is profoundly heterogeneous particularly when comparing sites of metastases. Establishing the extent of this heterogeneity may provide guidance on how best to design lipid-based drug delivery systems to treat metastatic disease. Building on our previous research, the current study employs a murine model of metastatic cancer to explore the distribution of ~ 100 nm liposomes. MethodsFemale NCr nude mice were inoculated with a fluorescently labeled, Her2/neu-positive, trastuzumab-resistant breast cancer cell line, JIMT-1mkate, either in the mammary fat pad to create an orthotopic tumor (OT), or via intracardiac injection (IC) to establish tumors throughout the body. Animals were dosed with fluorescent and radio-labeled liposomes. In vivo and ex vivo fluorescent imaging was used to track liposome distribution over a period of 48 h. Liposome distribution in orthotopic tumors was compared to sites of tumor growth that arose following IC injection. ResultsA significant amount of inter-vessel heterogeneity for DiR distribution was observed, with most tumor blood vessels showing little to no presence of the DiR-labelled liposomes. Further, there was limited extravascular distribution of DiR liposomes in the perivascular regions around DiR-positive vessels. While all OT tumors contained at least some DiR-positive vessels, many metastases had very little or none. Despite the apparent limited distribution of liposomes within metastases, two liposomal drug formulations, Irinophore C and Doxil, showed similar efficacy for both the OT and IC JIMT-1mkate models. ConclusionThese findings suggest that liposomal formulations achieve therapeutic benefits through mechanisms that extend beyond the enhanced permeability and retention effect.

Prioritizing drug targets by perturbing biological network response functions.

Therapeutic interventions are designed to perturb the function of a biological system. However, there are many types of proteins that cannot be targeted with conventional small molecule drugs. Accordingly, many identified gene-regulatory drivers and downstream effectors are currently undruggable. Drivers and effectors are often connected by druggable signaling and regulatory intermediates. Methods to identify druggable intermediates therefore have general value in expanding the set of targets available for hypothesis-driven validation. Here we identify and prioritize potential druggable intermediates by developing a network perturbation theory, termed NetPert, for response functions of biological networks. Dynamics are defined by a network structure in which vertices represent genes and proteins, and edges represent gene-regulatory interactions and protein-protein interactions. Perturbation theory for network dynamics prioritizes targets that interfere with signaling from driver to response genes. Applications to organoid models for metastatic breast cancer demonstrate the ability of this mathematical framework to identify and prioritize druggable intermediates. While the short-time limit of the perturbation theory resembles betweenness centrality, NetPert is superior in generating target rankings that correlate with previous wet-lab assays and are more robust to incomplete or noisy network data. NetPert also performs better than a related graph diffusion approach. Wet-lab assays demonstrate that drugs for targets identified by NetPert, including targets that are not themselves differentially expressed, are active in suppressing additional metastatic phenotypes.

Development of Traceable Mouse Models of Advanced and Metastatic Bladder Cancer.

Bladder cancer (BC) is the fourth most common cancer in men, with a poor patient prognosis for advanced disease. The poor survival of patients with muscle-invasive bladder cancer (MIBC) and metastatic status emphasizes the urgent need to develop new therapies. Lacking in the field of BC is the availability of relevant advanced BC mouse models, especially metastatic ones, that accurately recapitulate the complexities of human pathology to test and study new therapeutic strategies. Addressing this need, we developed a traceable mouse model of BC that expresses tumor-associated antigens within the context of advanced muscle-invasive BC. This novel system was achieved through the deletion of the tp53 and pten genes, alongside the incorporation of the fusion construct of Firefly luciferase (Luc) and the SIYRYYGL (SIY) T-cell antigen. We validate that the presence of the transgene did not impact on the development of the tumors while allowing us to measure tumor progression by bioluminescence. We show that the transgene did not influence the composition of the immune tumor microenvironment. More importantly, we report that this model was unresponsive to anti-PD-1 treatment, as in the majority of patients with BC. We also develop a new model based on the orthotopic injection of BC clonal cell lines derived from our first model. We demonstrate that this new model invades the muscle layer and has a metastasis development rate of 83%. The advantage of this model is that we can visualize tumor growth and metastasis development in vivo. These mouse models' characteristics, displaying many similarities with the human pathology, provide a valuable tool for tracking tumor progression, metastasis spread in vivo, and treatment resistance, as well as exploring fundamental and translational aspects of BC biology. This work contributes to the improvement in the landscape of mouse models of advanced BC for testing new therapeutic strategies.

Antibody-activation of connexin hemichannels in bone osteocytes with ATP release suppresses breast cancer and osteosarcoma malignancy

Bone tissue represents the most frequent site of cancer metastasis. We developed a hemichannel-activating antibody, Cx43-M2. Cx43-M2, directly targeting osteocytes in situ, activates osteocytic hemichannels and elevates extracellular ATP, thereby inhibiting the growth and migration of cultured breast and osteosarcoma cancer cells. Cx43-M2 significantly decreases breast cancer metastasis, osteosarcoma growth, and osteolytic activity, while improving survival rates in mice. The antibody's inhibition of breast cancer and osteosarcoma is dose dependent in both mouse and human cancer metastatic models. Furthermore, Cx43-M2 enhances anti-tumor immunity by increasing the population and activation of tumor-infiltrating immune-promoting effector T lymphocytes, while reducing immune-suppressive regulatory Tcells. Our results suggest that the Cx43-M2 antibody, by activating Cx43 hemichannels and facilitating ATP release and purinergic signaling, transforms the cancer microenvironment from a supportive to a suppressive state. Collectively, our study underscores the potential of Cx43-M2 as a therapeutic for treating breast cancer bone metastasis and osteosarcoma.

Partial hepatectomy accelerates colorectal metastasis by priming an inflammatory premetastatic niche in the liver.

Resection of colorectal liver metastasis is the standard of care for patients with Stage IV CRC. Despite undoubtedly improving the overall survival of patients, pHx for colorectal liver metastasis frequently leads to disease recurrence. The contribution of this procedure to metastatic colorectal cancer at a molecular level is poorly understood. We designed a mouse model of orthograde metastatic colorectal cancer (CRC) to investigate the effect of partial hepatectomy (pHx) on tumor progression. CRC organoids were implanted into the cecal walls of wild type mice, and animals were screened for liver metastasis. At the time of metastasis, 1/3 partial hepatectomy was performed and the tumor burden was assessed longitudinally using MRI. After euthanasia, different tissues were analyzed for immunological and transcriptional changes using FACS, qPCR, RNA sequencing, and immunohistochemistry. Mice that underwent pHx presented significant liver hypertrophy and an increased overall metastatic load compared with SHAM operated mice in MRI. Elevation in the metastatic volume was defined by an increase in de novo liver metastasis without any effect on the growth of each metastasis. Concordantly, the livers of pHx mice were characterized by neutrophil and bacterial infiltration, inflammatory response, extracellular remodeling, and an increased abundance of tight junctions, resulting in the formation of a premetastatic niche, thus facilitating metastatic seeding. Regenerative pathways following pHx accelerate colorectal metastasis to the liver by priming a premetastatic niche.

Association of gut microbe and stress-related cancer metastasis and oleic acid degradation.

e15517 Background: Cancer patients often face psychological distress. Cancer metastasis is critical for prognosis. Recent research highlights the link between chronic stress and metastasis. However, the underlying mechanisms is far from known. Methods: Animal models of cancer metastasis were used to explore the effect of chronic stress in promoting tumor metastasis. A combination of antibiotic cocktail treatment, fecal microbiota transplantation and germ-free mice were carried out to confirm whether the pro-metastatic ability of chronic stress was dependent on gut microbiota. 16S rDNA sequencing and liquid chromatography mass spectrometry were parallelly performed on mice stools to investigate alterations in microbiota and metabolites. Additionally, we assessed hormone changes post-stress and demonstrated the correlation between glucocorticoid elevation and gut microbiota through in vivo and in vitro experiments. Finally, we validated our hypothesis using a prospective colorectal cancer cohort. Results: Mice undergone chronic stress had significantly increased cancer metastasis compared with stress-free control mice. Gut microbial dysbiosis was observed in mice undergone chronic stress, with Bifidobacterium being significantly depleted. Stress hormone, corticosterone could alter intestinal epithelium metabolism, and thus inhibit the growth of Bifidobacterium. Metabolomic analysis showed increased Oleic Acid (OA) in the colonic contents from chronic stress-treated mice, and the pro-metastatic role of OA were examined in vitro and in vivo experiments. Supplement of Bifidobacterium animalis (isolated from feces of the stress-free control mice) could decrease the level of OA and inhibit pro-metastasis effect of stress in mice. OhyA which encoded oleate hydratase was identified in Bifidobacterium animalis by whole genome sequencing. Heterogeneous expression of OhyA in Escherichia coli BL21 showed that supplement of OhyA could decrease the level of OA and abrogate the pro-metastatic effect of chronic stress. We found that glucocorticoid secretion increases during chronic stress, promoting the nuclear translocation of glucocorticoid receptors in epithelial cells and activating the transcription of downstream target genes. Through integrated metabolomic analysis, we demonstrated that glucocorticoids degrade guanine, reducing its concentration in the intestine and thereby inhibiting the growth of Bifidobacterium. Furthermore, our investigation into clinical cohorts revealed that patients with higher stress scores are more prone to tumor metastasis, accompanied by a decrease in the abundance of Bifidobacterium in the gut and an increase in serum and fecal oleic acid levels. Conclusions: Chronic phycological stress promotes cancer metastasis by inhibiting the growth of probiotic microbe Bifidobacterium through modulating epithelium purine metabolism and increasing the level of OA.

Clinically Relevant Humanized Mouse Models of Metastatic Prostate Cancer Facilitate Therapeutic Evaluation.

There is tremendous need for improved prostate cancer models. Anatomically and developmentally, the mouse prostate differs from the human prostate and does not form tumors spontaneously. Genetically engineered mouse models lack the heterogeneity of human cancer and rarely establish metastatic growth. Human xenografts are an alternative but must rely on an immunocompromised host. Therefore, we generated prostate cancer murine xenograft models with an intact human immune system (huNOG and huNOG-EXL mice) to test whether humanizing tumor-immune interactions would improve modeling of metastatic prostate cancer and the impact of androgen receptor-targeted and immunotherapies. These mice maintain multiple human immune cell lineages, including functional human T-cells and myeloid cells. Implications: To the best of our knowledge, results illustrate the first model of human prostate cancer that has an intact human immune system, metastasizes to clinically relevant locations, responds appropriately to standard-of-care hormonal therapies, and can model both an immunosuppressive and checkpoint-inhibition responsive immune microenvironment.