Orthotopic Xenograft Mouse Model Research Articles

Dual rectification of metabolism abnormality in pancreatic cancer by a programmed nanomedicine

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive and lethal malignancy characterized by dysregulated energy and stromal metabolism. It is strongly supported by activated pancreatic stellate cells (PSC) which drive excessive desmoplasia and tumor growth via metabolic crosstalk. Herein, a programmed nanosystem is designed to dual rectify the metabolism abnormalities of the PDAC cells, which overexpress glucose transporter 1(GLUT1) and CD71, and the PSC for oncotherapy. The nanosystem is based on a tumor microenvironment-responsive liposome encapsulating an NF-κB inhibitor (TPCA-1) and a CD71 aptamer-linked Glut1 siRNA. TPCA-1 reverses the activated PSC to quiescence, which hampers metabolic support of the PSC to PDAC cells and bolsters the PDAC cell-targeting delivery of the siRNA. Aerobic glycolysis and the following enhancement of oxidative phosphorylation are restrained by the nano-modulation so as to amplify anti-PDAC efficacy in an orthotopic xenograft mouse model, which implies more personalized PDAC treatment based on different energy metabolic profiles.

Longikaurin A induces ferroptosis and inhibits glioblastoma progression through DNA methylation - Mediated GPX4 suppression.

Glioblastoma (GBM) is the most common primary intracranial tumor highly resistant to conventional clinical chemotherapy. Recently, the induction of ferroptosis is emerging as a putative strategy to treat various tumors. However, the identification of the effective and applicable tumor ferroptosis-inducing agents remains challenging. In this study, we showed that longikaurin A (LK-A), a natural diterpenoid isolated from the medicinal plant Isodon ternifolius with strong anti-GBM capacities, induced remarkable GBM cell ferroptosis along with suppressing the key anti-ferroptosis factor glutathione peroxidase 4 (GPX4). GPX4 promoter contains conserved CpG islands. The LK-A-induced GPX4 suppression coincided with the inhibition of ten-eleven translocation 2 (TET2), a key DNA demethylation enzyme and an increase in the hypermethylation of the GPX4 promoter. Further, LK-A promoted the GBM ferroptotic alterations and inhibited GBM progression in both subcutaneous and orthotopic xenograft mouse models, whereas GPX4 overexpression largely abrogated its anti-GBM effects both in vitro and in vivo, suggesting that LK-A inductions of the DNA methylation-incurred GPX4 suppression and ferroptosis are crucial for its anti-GBM functions. Together, our study has elaborated an important epigenetic pathway of GBM ferroptosis and uncovered a critical pharmacological property of LK-A for treating GBM patients.

Self-assembling nanoparticles for delivery of miR-603 and miR-221 in glioblastoma as a new strategy to overcome resistance to temozolomide

Glioblastoma (GBM) is a highly aggressive brain cancer with poor clinical outcome. Unfortunately, chemotherapy with temozolomide (TMZ) has a limited efficacy due to resistance mainly attributed to O6-methylguanine methyl transferase (MGMT) activity. Recently, miR-603 and miR-221 have been identified to target MGMT, thus improving the efficacy of temozolomide (TMZ) in the treatment of GBM. Previously, self-assembling nanoparticles (SANPs) have been proposed to deliver miRNAs into the brain. Here, SANP co-encapsulating miRNA-603 (miR-603) and miRNA-221 (miR-221) have been developed to enhance the efficacy of TMZ in the treatment of GBM by preventing the occurrence of chemoresistance. Preliminarily, SANPs encapsulating miRNAs were optimized in terms of lipid composition to assure physical stability and no hemolytic activity. Subsequently, SANPs with the lowest cytotoxicity and excellent internalization efficiency of miRNAs were selected through MTT assay and real-time PCR, respectively. To evaluate a potential synergistic effect between TMZ and miRNAs, MTT and clonogenic assays were performed. In our biological model, miRNA delivery via SANPs in combination with TMZ treatment strongly reduced cell viability and tumorigenic potential. Finally, in vivo assays were carried out on orthotopic xenograft mouse models. The treatment with SANPs encapsulating both miRNAs in combination with TMZ greatly decreased tumour growth, and even more significantly increased animal survival. In conclusion, this strategy provides the rationale for the development of new therapeutic approaches based on SANP technology to deliver miRNAs that play a key role in suppressing tumour.

TMET-29. TARGETING IDH1-MUTATED OLIGODENDROGLIOMA WITH ACID CERAMIDASE INHIBITORS

Abstract BACKGROUND Currently, an estimated 14,950 people lives with oligodendroglioma in the United States. Oligodendroglioma is genetically defined as a tumor harboring isocitrate dehydrogenase 1 or 2 mutations (IDH1mut/IDH2mut). Previously, we reported that in IDH1mut gliomas, D-2HG, the product of IDH1 mutant enzyme produces an increase in monounsaturated fatty acid levels that are incorporated into ceramides, tilting the S1P-to-ceramide rheostat toward apoptosis. Herein, we exploited this imbalance to further induce and IDHmut-specific glioma cell death. METHODS TCGA and CCGA data were used to explore the association between acid ceramidase (AC) gene expression and patient survival. CCK8 viability assays were carried out to measure the sensitivity of glioma cell lines to acid ceramidase inhibitors. Western blot and liquid chromatography-mass spectrometry analyses were performed for studying the mechanism of action of acid ceramidase inhibitors. The in vivo efficacy of the SABRAC drug, an irreversible inhibitor of AC, was evaluated in an orthotopic oligodendroglioma xenograft mice model using TS603 patient-derived oligodendroglioma cells. RESULTS. We report for the first time that the inhibition of acid ceramidase (AC) induces apoptosis and increases the survival of mice with IDH1mut oligodendroglioma. We demonstrated an IDH1mut-specific cytotoxicity of SABRAC, in patient-derived oligodendroglioma cells. Exploring the mechanism of action of this drug, we found that SABRAC activates both extrinsic and intrinsic apoptosis in an ER stress-independent manner, pointing to a direct action of AC-related ceramides in mitochondria permeability. The activation of apoptosis detected under SABRAC treatment was associated with up to a 30-fold increase in some ceramide levels and its derivatives from the salvage pathway. CONCLUSIONS We propose AC as a novel therapeutic target in oligodendroglioma with IDH1mut and we are conducting additional preclinical testing.

EXTH-17. THERAPEUTIC EFFICACY OF B7-H3 CAR T CELLS IN PEDIATRIC HIGH-GRADE GLIOMAS WITH H3G34R/V MUTATION

Abstract Pediatric high-grade gliomas (pHGGs) are aggressive brain tumors responsible for significant childhood mortality. These tumors often carry mutations in the H3F3A gene, which encodes histone H3.3 proteins. However, current treatment options are limited, underscoring the need for innovative approaches. Chimeric antigen receptor (CAR) therapy, which was successful in treating certain childhood cancers, holds promise for pHGGs with H3G34R/V mutation. In this study, we examined the expression of B7-H3 in patient tumor specimens and various pHGG cell lines with H3G34R/V mutation. Data revealed substantial B7-H3 expression, confirming its potential as a therapeutic target. Co-culturing B7-H3 CAR T cells with H3G34R/V mutated KNS42 and SJ-HGG-X42 cells led to significant increase in the expression of markers associated with T cell activation (IL2, IL6, IF-γ, TNF-α) and immune responses involving cytolytic cells (Granzyme A, B, Perforin) and elevated levels of exhaustion markers (2B4, PD1, Tim3, Lag3) in comparison to mock-treated cells. We evaluated the capability of B7-H3 CAR T cells for cytolytic activity against KNS-42 and SJ-HGG-X42 cells through in vitro testing. Our results after 72 hours of co-culture revealed a strong killing potential of B7-H3 CARs against B7-H3 positive KNS-42 and SJ-HGG-X42 cells, while minimal or no killing was observed on B7-H3KO KNS-42 and B7-H3 negative MDA MD 453 cells. In vivo efficacy of B7-H3 CAR T cells in an orthotopic mouse xenograft model implanted with KNS-42 cells. B7-H3 CAR T cell treated mice showed a marked reduction in tumor burden and a substantial improvement in survival in contrast of CD19-CAR T cell treated control group. Lastly, we have developed a novel In-Utero Electroporation (IUE) immune-competent mouse model of pHGGs with H3G34R/V mutation, that enables exploration of the impact of immune suppressive tumor microenvironment on CAR T cells efficacy. In conclusion, B7-H3 emerges as a promising therapeutic target, and CAR T cells display encouraging functionality within the context of pHGGs with H3G34R/V model.

Preclinical assessment of MAGMAS inhibitor as a potential therapy for pediatric medulloblastoma.

DAOY (SHH driven/tp53 mutant) and D425 (non-SHH group 3) were treated with BT9. For in vitro analysis, cell proliferation, death, migration, invasion, and metabolic activity were assessed using MTT assay, TUNEL staining, scratch wound assay, Matrigel invasion chambers, and seahorse assay, respectively. A D425 orthotopic xenograft mouse model was used to evaluate BT9 efficacy in vivo. BT9 treatment resulted in a significant decrease in cell proliferation (DAOY, 24 hours IC50: 3.6 μM, 48 hours IC50: 2.3 μM, 72 hours IC50: 2.1 μM; D425 24 hours IC50: 3.4 μM, 48 hours IC50: 2.2 μM, 72 hours IC50: 2.1 μM) and a significant increase in cell death (DAOY, 24 hours p = 0.0004, 48 hours p<0.0001; D425, 24 hours p = 0.0001, 48 hours p = 0.02). In DAOY cells, 3 μM BT9 delayed migration and significantly reduced DAOY and D425 cell invasion (p < 0.0001). It also modified mitochondrial respiratory function in both medulloblastoma cell lines. Compared to control, however, BT9 administration did not improve survival in a D425 orthotopic xenograft mouse model. Our in vitro data showed BT9 antitumor efficacy in DAOY and D425 cell lines, suggesting that BT9 may represent a promising targeted therapeutic in pediatric medulloblastoma. These data, however, need to be further validated in animal models.

Tumor-intrinsic role of ICAM-1 in driving metastatic progression of triple-negative breast cancer through direct interaction with EGFR

Triple-negative breast cancer (TNBC), the most aggressive subtype, presents a critical challenge due to the absence of approved targeted therapies. Hence, there is an urgent need to identify effective therapeutic targets for this condition. While epidermal growth factor receptor (EGFR) is prominently expressed in TNBC and recognized as a therapeutic target, anti-EGFR therapies have yet to gain approval for breast cancer treatment due to their associated side effects and limited efficacy. Here, we discovered that intercellular adhesion molecule-1 (ICAM-1) exhibits elevated expression levels in metastatic breast cancer and serves as a pivotal binding adaptor for EGFR activation, playing a crucial role in malignant progression. The activation of EGFR by tumor-expressed ICAM-1 initiates biased signaling within the JAK1/STAT3 pathway, consequently driving epithelial-to-mesenchymal transition and facilitating heightened metastasis without influencing tumor growth. Remarkably, ICAM-1-neutralizing antibody treatment significantly suppressed cancer metastasis in a breast cancer orthotopic xenograft mouse model. In conclusion, our identification of ICAM-1 as a novel tumor intrinsic regulator of EGFR activation offers valuable insights for the development of TNBC-specific anti-EGFR therapies.

Tumor-Secreted Extracellular Vesicles Counteract Therapy Response by Triggering Inflammatory Mesenchymal Stem Cell Development.

Therapy resistance is a major clinical hurdle in bone cancer treatment and seems to be largely driven by poorly understood microenvironmental factors. Recent evidence suggests a critical role for a unique subpopulation of mesenchymal stem cells with inflammatory features (iMSC), though their origin and function remained unexplored. We demonstrate that cancer-secreted extracellular vesicles (EV) trigger the development of iMSCs, which hinder therapy response in vivo, and set out to identify strategies to counteract their function. The role of iMSCs in therapy resistance was evaluated in an orthotopic xenograft mouse model of osteosarcoma. EV-induced alterations of the MSC transcriptome were analyzed and compared with single-cell RNA sequencing data of biopsies from patients with osteosarcoma and multiple myeloma. Functional assays identified EV components driving iMSC development. We assessed the efficacy of clinical drugs in blocking iMSC-induced resistance in vivo. We found that iMSCs are induced by interaction with cancer EVs and completely abrogate the antimetastatic effect of TGFβ signaling inhibition. Importantly, EV-induced iMSCs faithfully recapitulate the inflammatory single-cell RNA signature of stromal cells enriched in biopsies from patients with multiple myeloma and osteosarcoma. Mechanistically, cancer EVs act through two distinct mechanisms. EV-associated TGFβ induces IL6 production, whereas the EV-RNA cargo enhances TLR3-mediated chemokine production. We reveal that simultaneous blockade of downstream EV-activated pathways with ladarixin and tocilizumab disrupts metastasis formation and overcomes iMSC-induced resistance. Our observations establish iMSCs as major contributors to drug resistance, reveal EVs as triggers of iMSC development, and highlight a promising combination strategy to improve therapy response in patients with bone cancer.

Synergistic combination of perphenazine and temozolomide suppresses patient-derived glioblastoma tumorspheres.

Glioblastoma (GBM), a primary malignant brain tumor, has a poor prognosis, even with standard treatments such as radiotherapy and chemotherapy. In this study, we explored the anticancer effects of the synergistic combination of perphenazine (PER), a dopamine receptor D2/3 (DRD2/3) antagonist, and temozolomide (TMZ), a standard treatment for GBM, in patient-derived human GBM tumorspheres (TSs). The biological effects of the combination of PER and TMZ in GBM TSs were assessed by measuring cell viability, ATP, stemness, invasiveness, and apoptosis. Changes in protein and mRNA expression were analyzed using western blotting and RNA sequencing. Co-administration of PER and TMZ was evaluated in vivo using a mouse orthotopic xenograft model. The Severance dataset showed that DRD2 and DRD3 expression was higher in tumor tissues than in the tumor-free cortex of patients with GBM. DRD2/3 knockout by CRISPR/Cas9 in patient-derived human GBM TSs inhibited cell growth and ATP production. The combined treatment with PER and TMZ resulted in superior effects on cell viability and ATP assays compared to those in single treatment groups. Flow cytometry, western blotting, and RNA sequencing confirmed elevated apoptosis in GBM TSs following combination treatment. Additionally, the combination of PER and TMZ downregulated the expression of protein and mRNA associated with stemness and invasiveness. In vivo evaluation showed that combining PER and TMZ extended the survival period of the mouse orthotopic xenograft model. The synergistic combination of PER and TMZ has potential as a novel combination treatment strategy for GBM.

Engineered extracellular vesicles for combinatorial TNBC therapy: SR-SIM-guided design achieves substantial drug dosage reduction

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer that has no therapeutic targets, relies on chemotherapeutics for treatment, and is in dire need of novel therapeutic approaches for improved patient outcomes. Extracellular vesicles (EVs) serve as intercellular communicators and have been proposed as ideal drug delivery vehicles. Here, EVs were engineered with RNA nanotechnology to develop TNBC tumor inhibitors. Using super resolved-structured illumination microscopy, EVs were optimized for precise Survivin small interfering RNA (siRNA) conjugated to chemotherapeutics loading and CD44 aptamer ligand decoration, thereby enhancing specificity toward TNBC cells. Conventional treatments typically employ chemotherapy drugs gemcitabine (GEM) and paclitaxel (PTX) at dosages on the order of mg/kg respectively, per injection (intravenous) in mice. In contrast, engineered EVs encapsulating these drugs saw functional tumor growth inhibition at significantly reduced concentrations: 2.2μg/kg for GEM or 5.6μg/kg for PTX, in combination with 21.5μg/kg survivin-siRNA in mice. The result is a substantial decrease in the chemotherapeutic dose required, by orders of magnitude, compared with standard regimens. Invivo and invitro evaluations in a TNBC orthotopic xenograft mouse model demonstrated the efficacy of this decreased dosage strategy, indicating the potential for decreased chemotherapy-associated toxicity.

LSD1 and CoREST2 Potentiate STAT3 Activity to Promote Enteroendocrine Cell Differentiation in Mucinous Colorectal Cancer.

Neuroendocrine cells have been implicated in therapeutic resistance and worse overall survival in many cancer types. Mucinous colorectal cancer (mCRC) is uniquely enriched for enteroendocrine cells (EECs), the neuroendocrine cell of the normal colon epithelium, as compared to non-mCRC. Therefore, targeting EEC differentiation may have clinical value in mCRC. Here, single cell multi-omics uncovered epigenetic alterations that accompany EEC differentiation, identified STAT3 as a regulator of EEC specification, and discovered a rare cancer-specific cell type with enteric neuron-like characteristics. Furthermore, LSD1 and CoREST2 mediated STAT3 demethylation and enhanced STAT3 chromatin binding. Knockdown of CoREST2 in an orthotopic xenograft mouse model resulted in decreased primary tumor growth and lung metastases. Collectively, these results provide rationale for developing LSD1 inhibitors that target the interaction between LSD1 and STAT3 or CoREST2, which may improve clinical outcomes for patients with mCRC.

Energy and endoplasmic reticulum stress induction by gold(III) dithiocarbamate and 2-deoxyglucose synergistically trigger cell death in breast cancer

The elusiveness of triple-negative breast cancer from targeted therapy has redirected focus towards exploiting the metabolic shortcomings of these highly metastatic subtypes of breast cancer. Cueing from the metabolic heterogeneity of TNBC and the exposition of the dual dependence of some TNBCs on OXPHOS and glycolysis for ATP, we herein report the efficacy of cotreatment of TNBCs with an OXPHOS inhibitor, 2a and 2DG, a potent glycolysis inhibitor. 2a-2DG cotreatment inhibited TNBC cell proliferation with IC50 of ∼5 to 36 times lower than that of 2a alone and over 5000 times lower than IC50 of 2DG alone. 2a-2DG cotreatment suppressed mitochondrial ATP production and significantly induced AMPK activation. Mechanistic studies revealed the distinct yet synergistic contributions of 2a and 2DG to the antiproliferative effect of the cotreatment. While 2a induced apoptotic cell death, 2DG sensitized TNBCs to the antiproliferative effects of 2a via endoplasmic reticulum stress induction. Strikingly, the combination of 2a-2DG ablated SUM159 tumors in an orthotopic xenograft mouse model. This study highlights the synergistic effect of a gold-based complex with 2DG and the potential benefit of multi-metabolic pathways targeting as an effective therapeutic strategy against TNBCs.

Bismuth-based mesoporous nanoball carrying sorafenib for synergistic photothermal and molecularly-targeted therapy in an orthotopic hepatocellular carcinoma xenograft mouse model

Sorafenib (SOR), a multi-kinase inhibitor for advanced hepatocellular carcinoma (HCC), has limited clinical application due to severe side effects and drug resistance. To overcome these challenges, we developed a bismuth-based nanomaterial (BOS) for thermal injury-assisted continuous targeted therapy in HCC. Initially, the mesoporous nanomaterial was loaded with SOR, forming the BOS@SOR nano-carrier system for drug delivery and controlled release. Notably, compared to targeted or photothermal therapy alone, the combination therapy using this nano-carrier system significantly impaired cell proliferation and increased apoptosis. In vivo efficacy evaluations demonstrated that BOS@SOR exhibited excellent biocompatibility, confirmed through hemolysis and biochemical analyses. Additionally, BOS@SOR enhanced contrast in computed tomography, aiding in the precise identification of HCC size and location. The photothermal therapeutic properties of bismuth further contributed to the synergistic anti-tumor activity of BOS@SOR, significantly reducing tumor growth in an orthotopic xenograft HCC model. Taken together, encapsulating SOR within a bismuth-based mesoporous nanomaterial creates a multifunctional and environmentally stable nanocomposite (BOS@SOR), enhancing the therapeutic effect of SOR and presenting an effective strategy for HCC treatment.

Abstract A073: Desmoglein-2 is a regulator of pancreatic ductal adenocarcinoma progression

Abstract Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies, with a dismal 5-year survival rate of &amp;lt;13%. We have discovered that desmoglein-2 (DSG2), a cell surface protein, is highly expressed on PDAC cells. In silico data indicate that elevated levels of DSG2 is associated with PDAC progression and poor patient survival. Immunohistochemistry (IHC) analysis of a tissue microarray of 302 patient samples showed that DSG2 was expressed by &amp;gt;90% of PDAC patients. We hypothesized that DSG2 is an under-appreciated contributor to PDAC progression and that DSG2 can be targeted. The main aims of this study were to: 1) investigate the role of DSG2 in PDAC cell function in vitro, 2) investigate the molecular biology underpinning the role of DSG2 in PDAC, and 3) to use murine PDAC tumor models to evaluate the contribution of DSG2 to cancer progression in vivo. Our in vitro data showed that reducing the expression of DSG2 (via small interfering RNA (siRNA)) significantly decreased PDAC cell proliferation, migration, and invasion. Moreover, signaling analysis of PDAC cells with or without DSG2 (via siRNA) using a reverse-phase protein array and cytokine array revealed that DSG2 modulates pro-tumorigenic and pro-metastatic cellular pathways, through reducing the effects of certain proteins. Particular alterations were observed in pathways involving gene regulation (e.g., beta-catenin), migration (e.g., epidermal growth factor receptor and integrin-beta1), and cytokine release (e.g., serpine-1). Next, in an orthotopic xenograft mouse model, BxPC-3 PDAC cells with a stable knockdown of DSG2 (via short-hairpin RNA) were injected into NSG (NOD scid gamma) mice, where we observed a significant reduction in tumor burden and liver metastasis when compared to control mice. IHC staining of tumor sections revealed that the DSG2 knockdown tumors contained reduced levels of collagen, tumor vasculature and cancer associated fibroblasts. Thus, suggesting that DSG2 also has a modulatory effect on the tumor microenvironment (TME). A similar reduction in tumor burden, cancer progression and changes to the TME were observed when a neutralizing anti-DSG2 monoclonal antibody was administered intraperitoneally twice a week at 5mg/kg to BxPC-3 tumor bearing mice. Finally, to investigate the immune cell landscape, we used an orthotopic syngeneic mouse model, whereby Dsg2 was knocked out of KPC (LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre) mouse PDAC cells via clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9. This mouse model also showed that loss of Dsg2 resulted in a significant reduction in tumor burden, cancer cell metastasis and changes to the TME. In summary, we have identified a potential new target for the treatment of PDAC, namely DSG2, and future experiments are pursuing clinical opportunities to improve PDAC patient survival. Citation Format: Charlie B Ffrench, Kay K Myo Min, Mark DeNichilo, Michaelia P Cockshell, Emma L Dorward, Emma J Thompson, Michael Ortiz, Michael S Samuel, Savio George Barreto, Claudine S Bonder. Desmoglein-2 is a regulator of pancreatic ductal adenocarcinoma progression [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A073.

Three-dimensional analysis of mitochondria in a patient-derived xenograft model of triple negative breast cancer reveals mitochondrial network remodeling following chemotherapy treatments.

Mitochondria are hubs of metabolism and signaling and play an important role in tumorigenesis, therapeutic resistance, and metastasis in many cancer types. Various laboratory models of cancer demonstrate the extraordinary dynamics of mitochondrial structure, but little is known about the role of mitochondrial structure in resistance to anticancer therapy. We previously demonstrated the importance of mitochondrial structure and oxidative phosphorylation in the survival of chemotherapy-refractory triple negative breast cancer (TNBC) cells. As TNBC is a highly aggressive breast cancer subtype with few targeted therapy options, conventional chemotherapies remain the backbone of early TNBC treatment. Unfortunately, approximately 45% of TNBC patients retain substantial residual tumor burden following chemotherapy, associated with abysmal prognoses. Using an orthotopic patient-derived xenograft mouse model of human TNBC, we compared mitochondrial structures between treatment-naïve tumors and residual tumors after conventional chemotherapeutics were administered singly or in combination. We reconstructed 1,750 mitochondria in three dimensions from serial block-face scanning electron micrographs, providing unprecedented insights into the complexity and intra-tumoral heterogeneity of mitochondria in TNBC. Following exposure to carboplatin or docetaxel given individually, residual tumor mitochondria exhibited significant increases in mitochondrial complexity index, area, volume, perimeter, width, and length relative to treatment-naïve tumor mitochondria. In contrast, residual tumors exposed to those chemotherapies given in combination exhibited diminished mitochondrial structure changes. Further, we document extensive intra-tumoral heterogeneity of mitochondrial structure, especially prior to chemotherapeutic exposure. These results highlight the potential for structure-based monitoring of chemotherapeutic responses and reveal potential molecular mechanisms that underlie chemotherapeutic resistance in TNBC.

Prediction of the response to antiangiogenic sunitinib therapy by non-invasive hybrid diffuse optics in renal cell carcinoma.

In this work, broadband diffuse reflectance spectroscopy (DRS) and diffuse correlation spectroscopy (DCS) were used to quantify deep tissue hemodynamics in a patient-derived orthotopic xenograft mouse model of clear cell renal cancer undergoing antiangiogenic treatment. A cohort of twenty-two mice were treated with sunitinib and compared to thirteen control untreated mice, and monitored by DRS/DCS. A reduction in total hemoglobin concentration (THC, p = 0.03), oxygen saturation (SO2, p = 0.03) and blood flow index (BFI, p = 0.02) was observed over the treatment course. Early changes in tumor microvascular blood flow and total hemoglobin concentration were correlated with the final microvessel density (p = 0.014) and tumor weight (p = 0.024), respectively. Higher pre-treatment tumor microvascular blood flow was observed in non-responder mice with respect to responder mice, which was statistically predictive of the tumor intrinsic resistance (p = 0.01). This hybrid diffuse optical technique provides a method for predicting tumor intrinsic resistance to antiangiogenic therapy and could be used as predictive biomarker of response to antiangiogenic therapies in pre-clinical models.

Lemur tail kinase 3 serves as a predictor of patient outcomes and a target for the treatment of ovarian cancer

Lemur tail kinase 3 (LMTK3) belongs to a family of tyrosine kinases that are known to correlate with tumor grade and patient survival in some cancers. Here, we validated LMTK3 as a specific target and a prognostic biomarker in ovarian cancer (OC). In samples from 204 stage I-II OC patients, immunohistochemical studies revealed a higher cytoplasmic-to-nuclear staining intensity of LMTK3, which correlated with worse overall survival (p<0.001). Efficacy studies utilizing novel LMTK3 binding peptides (LMTK3BPs) showed that all chemosensitive and chemoresistant OC cells were killed without affecting normal cells (p<0.005), with synergistic effects shown following cisplatin and docetaxel treatment. In an orthotopic xenograft mouse model of OC, we saw a 35% tumor reduction in response to intravenous injections of 2mg/kg LMTK3BP given three times a week for 3weeks. Furthermore, invivo safety studies showed no signs of toxicity after LMTK3BP treatment, even at doses as high as 40mg/kg. This study highlights LMTK3 as a predictor of patient clinical outcomes. More importantly, novel LMTK3BPs represent potential safe treatment options, either alone or in combination with therapies, for OC.

Osteosarcoma patient-derived orthotopic xenograft (PDOX) models for identification of novel and effective therapeutics.

228 Background: Osteosarcoma is the most common malignant primary tumor of bone and mainly occurs in young generations. Due to the heterogeneity, rarity, poor response rate to systemic therapy, and metastatic potential of osteosarcoma, individualized precision medicine and novel drug discovery are greatly needed. Toward this goal, we have established the patient-derived orthotopic xenograft (PDOX) mouse model with surgical orthotopic implantation for all major cancers. The PDOX models recapitulate human tumors better than subcutaneous-transplanted xenografts including patient-derived xenograft (PDX). Metastasis is observed to a greater extent in PDOX models due to the intact histology and correct-organ tumor micro-environment of the orthotopically implanted tissue. Therefore, the PDOX model is highly predictive of drug efficacy and useful for identifying effective drugs for tumors in which new drugs are difficult to develop. Methods: We have reported 18 osteosarcoma PDOX studies evaluating approved and experimental drugs since 2017. The present report reviews our research group’s experience with the osteosarcoma-PDOX model, and the power of the PDOX models to identify effective therapeutics. Results: Effective treatment for drug-resistant osteosarcoma includes regorafenib, as monotherapy, and temozolomide-irinotecan, trabectedin-irinotecan, sorafenib-everolimus, sorafenib-palbociclib, and olaratumab-doxorubicin-cisplatin, as combinations. Experimental therapy with recombinant methioninase, tumor-targeting S. typhimurium A1-R, or combinations of these agents and other drugs have shown surprising efficacy in the recalcitrant-osteosarcoma PDOX models. Conclusions: Owing to the high concordance of drug efficacy between patients and their corresponding PDOX models, these models provide improved and personalized treatment options for patients with osteosarcoma. The patient does not need to suffer from the potential drug toxicity and morbidity of ineffective chemotherapies. In an era of growing promise of new treatment and precision medicine, PDOX models can offer a unique opportunity to provide specific and individualized therapy and novel therapeutic options for osteosarcoma patients.

Transforming Albumin into a Trojan Horse of Immunotherapy-Resistant Colorectal Cancer with a High Microsatellite Instability.

The therapeutic response of microsatellite instability-high (MSI-H) colorectal cancer (CRC) to immune checkpoint inhibitors (ICI) is indeed surprising; however, the emergence of acquired resistance poses an even greater threat to the survival of these patients. Herein, bioinformatics analysis of MSI-H CRC samples revealed that Wnt signaling pathway represents a promising target for acquired immune reactivation, while subsequent analysis and biochemical testing substantiated the inclination of Wnt-hyperactive CRC cells to engage in macropinocytosis with human serum albumin (HSA). These findings have inspired us to develop an engineered HSA that not only possesses the ability to specifically target cancer cells but also effectively suppresses the Wnt/β-catenin cascade within these malignant cells. In pursuit of this objective, a comprehensive screening of reported Wnt small-molecule inhibitors was conducted to evaluate their affinity with HSA, and it was discovered that Carnosic acid (CA) exhibited the highest affinity while simultaneously revealing multiple binding sites. Further investigation revealed that CA HSA the capability to engineer HSA into spherical and size-tunable nanostructures known as eHSA (Engineering HSA particle), which demonstrated optimized macropinocytosis-dependent cellular internalization. As anticipated, eHSA effectively suppressed the Wnt signaling pathway and reactivated the acquired immune response in vivo. Furthermore, eHSA successfully restored sensitivity to Anti-PD1's anticancer effects in both subcutaneous and orthotopic mouse homograft models of MSI-H CRC, as well as a humanized hu-PBMC patient-derived orthotopic xenograft (PDOX) mouse model of MSI-H CRC, all while maintaining a favorable safety profile. The collective implementation of this clinically viable immune reactivation strategy not only enables the delivery of Wnt inhibitors for CRC therapy, but also serves as an exemplary demonstration of precision-medicine-guided nanopharmaceutical development that effectively harnesses specific cellular indications in pathological states.

Investigation on the mechanisms of scorpion venom in hepatocellular carcinoma model mice via untargeted metabolomics profiling

Metabolic reprogramming is frequently accompanied by hepatocellular carcinoma (HCC) progression. Disrupted metabolites act as potential biomarkers and drug therapeutic targets for HCC. Peptide extract of scorpion venom (PESV) induces cytotoxic anti-proliferative effects and apoptosis in tumors. However, the action mechanisms of PESV remain unknown. This study aimed to explore the serum metabolic profiles of tumor-bearing mouse model. We generated an orthotopic HCC xenograft mouse model by implanting H22 cells into the left hepatic lobe of male C57BL/6 mice. After surgery, the mice were assigned to two groups randomly: PESV (PESV-treated 40 mg/kg daily, i.g.; n = 6) and control (treated with the solvent equally for 14 d, n = 6) groups. Based on an untargeted metabolomics approach using ultra-high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry, differential metabolites were screened via univariate and multivariate data analyses. A total of 48 differential metabolites in negative ion mode and 63 in positive ion mode were identified in the serum samples. Furthermore, metabolic pathway analysis revealed that aminoacyl-tRNA biosynthesis, amino acid pathway, glutathione metabolism, protein transports, protein digestion and absorption, and cAMP signaling pathways play vital roles in PESV-induced inhibition of tumors. These findings highlight the distinct changes in the metabolic profiles of HCC-bearing mice after PESV treatment, suggesting the potential of the identified metabolic molecules as therapeutic targets for HCC.