Brain Tumor Model Research Articles

3D Reconstruction Methods Purporting 3D Visualization and Volume Estimation of Brain Tumors

This work proposes the Crust algorithm for 3D reconstruction of brain tumor, an effective mechanism in the visualization of tumors for presurgical planning, radiation dose calculation. Despite the promising performance of Crust algorithm in reconstruction of Stanford models, it has not yet been considered in 3D reconstruction of brain tumor. Validation of the results is done using the comparison of the 3D models from two cutting edge techniques namely the Marching Cube and the Alpha shape algorithm. The obtained result shows that Crust algorithm provides the brain tumor model with an average quality of triangle meshes ranging from 0.85 to 0.95. Concerning the visual realism, the quality of Crust algorithm models is higher on comparison to the other models. Precision of tumor volume measurement by convex hull method is analysed by repeatability and reproducibility. The standard deviations of repeatability were between 2.03 % and 3.97 %. The experimental results show that Linear Crust algorithm produces high quality meshes with average quality of equilateral triangles close to 1.

Analysis of Fluorescence Decay Kinetics of Indocyanine Green Monomers and Aggregates in Brain Tumor Model In Vivo.

Spectroscopic approach with fluorescence time resolution allows one to determine the state of a brain tumor and its microenvironment via changes in the fluorescent dye’s fluorescence lifetime. Indocyanine green (ICG) is an acknowledged infra-red fluorescent dye that self-assembles into stable aggregate forms (ICG NPs). ICG NPs aggregates have a tendency to accumulate in the tumor with a maximum accumulation at 24 h after systemic administration, enabling extended intraoperative diagnostic. Fluorescence lifetime analysis of ICG and ICG NPs demonstrates different values for ICG monomers and H-aggregates, indicating promising suitability for fluorescent diagnostics of brain tumors due to their affinity to tumor cells and stability in biological tissue.

Boron neutron capture therapy using dodecaborated albumin conjugates with maleimide is effective in a rat glioma model

IntroductionBoron neutron capture therapy (BNCT) is a biologically targeted, cell-selective particle irradiation therapy that utilizes the nuclear capture reaction of boron and neutron. Recently, accelerator neutron generators have been used in clinical settings, and expectations for developing new boron compounds are growing.Methodsand Results In this study, we focused on serum albumin, a well-known drug delivery system, and developed maleimide-functionalized closo-dodecaborate albumin conjugate (MID-AC) as a boron carrying system for BNCT. Our biodistribution experiment involved F98 glioma-bearing rat brain tumor models systemically administered with MID-AC and demonstrated accumulation and long retention of boron. Our BNCT study with MID-AC observed statistically significant prolongation of the survival rate compared to the control groups, with results comparable to BNCT study with boronophenylalanine (BPA) which is the standard use of in clinical settings. Each median survival time was as follows: untreated control group; 24.5days, neutron-irradiated control group; 24.5days, neutron irradiation following 2.5h after termination of intravenous administration (i.v.) of BPA; 31.5days, and neutron irradiation following 2.5 or 24h after termination of i.v. of MID-AC; 33.5 or 33.0days, respectively. The biological effectiveness factor of MID-AC for F98 rat glioma was estimated based on these survival times and found to be higher to 12. This tendency was confirmed in BNCT 24h after MID-AC administration. ConclusionMID-AC induces an efficient boron neutron capture reaction because the albumin contained in MID-AC is retained in the tumor and has a considerable potential to become an effective delivery system for BNCT in treating high-grade gliomas.

Acquisition and quantification pipeline for in vivo hyperpolarized 13 C MR spectroscopy.

The goal of this study was to combine a specialized acquisition method with a new quantification pipeline to accurately and efficiently probe the metabolism of hyperpolarized 13 C-labeled compounds in vivo. In this study, we tested our approach on [2-13 C]pyruvate and [1-13 C]α-ketoglutarate data in rat orthotopic brain tumor models at 3T. We used a multiband metabolite-specific radiofrequency (RF) excitation in combination with a variable flip angle scheme to minimize substrate polarization loss and measure fast metabolic processes. We then applied spectral-temporal denoising using singular value decomposition to enhance spectral quality. This was combined with LCModel-based automatic 13 C spectral fitting and flip angle correction to separate overlapping signals and rapidly quantify the different metabolites. Denoising improved the metabolite signal-to-noise ratio (SNR) by approximately 5. It also improved the accuracy of metabolite quantification as evidenced by a significant reduction of the Cramer Rao lower bounds. Furthermore, the use of the automated and user-independent LCModel-based quantification approach could be performed rapidly, with the kinetic quantification of eight metabolite peaks in a 12-spectrum array achieved in less than 1 minute. The specialized acquisition method combined with denoising and a new quantification pipeline using LCModel for the first time for hyperpolarized 13 C data enhanced our ability to monitor the metabolism of [2-13 C]pyruvate and [1-13 C]α-ketoglutarate in rat orthotopic brain tumor models in vivo. This approach could be broadly applicable to other hyperpolarized agents both preclinically and in the clinical setting.

BIOM-10. PRECLINICAL PLATFORM FOR THE IDENTIFICATION OF DEUTERIUM MAGNETIC RESONANCE SPECTROSCOPY-BASED BIOMARKERS OF BRAIN TUMOR METABOLISM

Abstract Metabolic reprogramming is a fundamental hallmark of cancer, which can be exploited for non-invasive tumor imaging. Deuterium magnetic resonance spectroscopy (2H-MRS) recently emerged as a novel, translational method of interrogating flux from 2H-labeled substrates to metabolic products. However, to date, preclinical studies have been performed in vivo, an endeavor which suffers from low-throughput and potential wastage of animal life, especially when considering studies of treatment response. Developing in vitro assays for monitoring metabolism of 2H-labeled substrates will enhance throughput, lead to the rapid evaluation of new 2H-based probes and enable identification of treatment response biomarkers, thereby allowing the best 2H-based probes to be translated for further in vivo assessment. The goal of this study was to develop a preclinical cell-based platform for quantifying metabolism of 2H-labeled probes in brain tumor models. Since the Warburg effect, which is characterized by elevated glycolytic production of lactate, is a metabolic phenotype of cancer, including brain tumors, we examined metabolism of 2H-glucose or 2H-pyruvate in patient-derived glioblastoma (GBM6) and oligodendroglioma (BT88) cells and compared to normal human astrocytes (NHACONTROL). Following incubation in media containing [6,6’-2H]glucose or [U-2H]pyruvate, 2H-MR spectra obtained from live cell suspensions showed elevated 2H-lactate production in GBM6 and BT88 cells relative to NHACONTROL. Importantly, 2H-lactate production from [6,6’-2H]glucose or from [U-2H]pyruvate was reduced in GBM6 or BT88 cells subjected to irradiation and temozolomide, which is standard of care for glioma patients, pointing to the utility of this method for detecting response to therapy. Collectively, we have, for the first time, demonstrated the ability to quantify metabolism of 2H-MRS probes in live cell suspensions and validated the utility of our assay for differentiating tumor from normal cells and assessing response to therapy. Our studies will expedite the identification of novel 2H-MRS probes for imaging brain tumors and potentially other types of cancer.

CTIM-27. LOCOREGIONAL B7-H3-SPECIFIC CAR T CELLS FOR CHILDREN AND YOUNG ADULTS WITH DIPG: INTERIM REPORT OF BRAINCHILD-03 ARM C

Abstract Following preclinical optimization of B7-H3-specific CAR T cells against pediatric brain tumor models, we opened BrainChild-03 (NCT04185038), a phase 1 clinical trial of repeatedly dosed, outpatient, locoregional B7-H3-specific CAR T cells for children with recurrent/refractory central nervous system (CNS) tumors or diffuse intrinsic pontine glioma (DIPG). Here, we report the interim findings from patients enrolled on Arm C, dedicated to DIPG. The primary endpoints are feasibility and safety, with secondary endpoints of disease response. We utilize second-generation CAR T cells with a 4-1BB costimulatory domain and a methotrexate-resistant human DHFR mutein (huDHFRFS; L22F,F31S), allowing for methotrexate selection. We do not deliver conditioning chemotherapy. The first three evaluable patients with DIPG all met feasibility for generating a balanced CD4:CD8 CAR T cell product, with 3.85x109 CAR T cells generated for S005, 4.29x109 for S008, and 2.45x109 for S012, allowing for greater than 6 months of biweekly dosing for each patient. All subjects were treated at Dose Level 1 (1x107 CAR T cells). S005 received 10 doses before clinical progression greater than 2 years from diagnosis, S008 has received 10 doses and continues on therapy with decreased tumor volume, and S012 has received 5 doses and continues on study with stable disease. There have been no dose limiting toxicities (DLT). 3/3 patients exhibited post infusion fever, headache, and elevated serum CRP but no evidence of cytokine release syndrome (CRS) or systemic CAR T cells. 0/3 patients required PICU admissions. In the cerebrospinal fluid (CSF), 2/3 patients have had elevations of cytokines such as CXCL10 and CCL2, as well as circulating CSF CAR T cells. Advanced serial patient CSF proteomic and transcriptomic profiling are underway. Ultimately, this report provides preliminary evidence that outpatient locoregional B7-H3 CAR T cells for children with DIPG may be feasible and tolerable.

CSIG-30. TARGETING TELOMERASE VIA MEK INHIBITION IN AGGRESSIVE TERT PROMOTER MUTATED BRAIN TUMORS

Abstract Activating point mutations within the TERT promoter (C228T or C250T) account for the most frequent alteration in aggressive brain tumors. Presence of these alterations results in the generation of binding sites for E-twenty-six (ETS) transcription factors accompanied by enhanced TERT expression. Accordingly, TERT promoter mutations foster cellular immortalization and subsequently tumor aggressiveness. Due to the limitation of treatment options in aggressive brain tumors, including glioblastoma and medulloblastoma, new therapeutic targets need to be discovered. As we previously described a strong interaction of oncogenic MEK/ETS signaling and TERT promoter mutations, we hypothesize that inhibition of these factors halters cell immortalization in TERT-driven brain tumors. Our study included three TERT promoter wild-type (TERTwt), six mutated (TERTmut) glioblastoma and three TERTmut medulloblastoma cell models and tested the effect of MEK inhibitors (U0126 and trametinib) and the ETS inhibitor YK-4-279 on cell viability and clone formation. Cellular senescence upon treatment was evaluated by beta-galactosidase assays. Impact on TERT mRNA expression and TERT promoter activity were analyzed by quantitative real-time PCR and luciferase reporter assays, respectively. Furthermore, the effects on MAPK- and PI3K pathway activation were evaluated by Western blot. Amongst the investigated inhibitors, tumor cells harboring C228T mutation were distinctly more sensitive against trametinib as compared to TERTwt and C250T TERTmut cells. Similar effects were observed on clonogenicity upon long-term exposure to this inhibitor. Regarding MAPK signaling activation, trametinib treatment completely blocked ERK phosphorylation in every cell model, while activation of ETS1 was more effectively reduced in C228T TERTmutcells. Accordingly, exposure to trametinib reduced TERT expression and promoter activity accompanied by induction of cellular senescence in cells with C228T mutation. Impact of trametinib is currently investigated in preclinical experiments using TERTmut brain tumor models. Summarizing, MEK inhibition represents a novel strategy to overcome cell immortalization especially in C228T TERTmut brain tumors.

TAMI-43. HYPOXIC REGULATION OF GLIOBLASTOMA TUMOR GROWTH THROUGH L(3)MBTL1

Abstract Glioblastoma (GBM) is the deadliest and most common of all primary brain tumors. Drosophila brain tumor models have uncovered signaling pathways regulating tumor growth that are highly conserved in GBM. Our search for a novel tumor suppressor using Drosophila led to Lethal (3) malignant brain tumor [l(3)mbt], temperature-sensitive mutants of which cause neuroepithelial tumor-like overproliferation in optic lobes. dL(3)MBT and its human orthologs L3MBTL1-L3MBTL4 all harbor Malignant Brain Tumor (MBT) domains that recognize methylated lysines on histone tails. Like dL(3)MBT, hL(3)MBTL1 acts as a chromatin compaction factor that represses transcription and inhibits cytokinesis in GBM cell lines. The highly hypoxic tumor microenvironment (TME) in GBM drives its progression, recurrence, and therapeutic resistance. However, it remains unclear if L(3)MBTL1 is regulated by TME cues to promote GBM growth. Based on this knowledge gap and our preliminary data, we hypothesize that hypoxia directly regulates L(3)MBTL1 in favor of GBM growth. Analysis of TCGA data for IDH-wildtype gliomas revealed that L3MBTL1 gene expression is downregulated in GBM, which are necrotic and severely hypoxic, compared to histologic grade 2/3 gliomas, which do not contain necrosis, indicating that hypoxia could potentially suppress L3MBTL1 to enhance glioma progression. TCGA data also revealed a number of HIF pathway and hypoxia-inducible genes strongly correlating with L3MBTL1 expression, including HIF1a and VHL. Using patient-derived GBM neurosphere cultures, we exposed glioma cells to hypoxia (1% O2 for 24hrs) and found that L3MBTL1 protein levels were suppressed compared to normoxia (21%). Under these same conditions, we found more rapid cell proliferation under hypoxia. Exploration of hypoxic TME regulation of the novel tumor suppressor L3MBTL1 in glioma progression has the potential to uncover novel mechanisms involving epigenetic modulation and potentially new therapeutic strategies.

Eliciting an immune-mediated antitumor response through oncolytic herpes simplex virus-based shared antigen expression in tumors resistant to viroimmunotherapy

BackgroundOncolytic virotherapy (OV) is an immunotherapy that incorporates viral cancer cell lysis with engagement of the recruited immune response against cancer cells. Pediatric solid tumors are challenging targets because they...

Adenosine A2A Receptor Activation Enhances Blood-Tumor Barrier Permeability in a Rodent Glioma Model.

The blood-tumor barrier (BTB) limits the entry of effective chemotherapeutic agents into the brain for treatment of malignant tumors like glioblastoma. Poor drug entry across the BTB allows infiltrative glioma stem cells to evade therapy and develop treatment resistance. Regadenoson, an FDA-approved adenosine A2A receptor (A2AR) agonist, has been shown to increase drug delivery across the blood-brain barrier in non-tumor-bearing rodents without a defined mechanism of enhancing BTB permeability. Here, we characterize the time-dependent impact of regadenoson on brain endothelial cell interactions and paracellular transport, using mouse and rat brain endothelial cells and tumor models. In vitro, A2AR activation leads to disorganization of cytoskeletal actin filaments by 30 minutes, downregulation of junctional protein expression by 4 hours, and reestablishment of endothelial cell integrity by 8 hours. In rats bearing intracranial gliomas, regadenoson treatment results in increase of intratumoral temozolomide concentrations, yet no increased survival noted with combined temozolomide therapy. These findings demonstrate regadenoson's ability to induce brain endothelial structural changes among glioma to increase BTB permeability. The use of vasoactive mediators, like regadenoson, which transiently influences paracellular transport, should further be explored to evaluate their potential to enhance central nervous system treatment delivery to aggressive brain tumors. IMPLICATIONS: This study provides insight on the use of a vasoactive agent to increase exposure of the BTB to chemotherapy with intention to improve glioma treatment efficacy.

Profile and management of pediatric brain tumors: A single–center experience

Context: Pediatric brain tumors are the most common solid tumors and the leading cause of death among children worldwide. Although there are some published studies from many countries regarding the pathological model of pediatric brain tumors, comprehensive clinical pathology studies conducted at Vietnam are still lacking. Aims: To explore the clinical and histopathological features of pediatric brain tumors at a central hospital in Vietnam. Methods: This study used a retrospective epidemiological approach. Children aged <17 years were selected according to their profiles registered in our department during 2016–2020 when they were diagnosed with primary brain tumors. Data of patients’ age, sex, clinical features, and histology were collected. The 2016 World Health Organization classification of tumors was adopted. Results: A total of 34 patients with brain tumors were identified, with a males/females ratio of 2.8/1. Patients’ age ranged from 3 months to 16 years, with a median age of 6 years. The most common presentation of brain tumor in our series was headache (47.1%). According to the location of involvement, 51.5% of the tumors were in supratentorial region. Moreover, astrocytes (55.9%) were the most common type of pediatric brain tumors. Surgery was the main treatment, and chemotherapy was administered to 14.7% of patients. Conclusions: This study addresses the issue of sensitivity but not that of specificity of clinical features to the presence of an underlying brain tumor. Histopathology analysis is necessary to develop further treatment after neurosurgery.

Role of Three-dimensional Printing in Neurosurgery: An Institutional Experience.

Background:Recent advancements in three-dimensional (3D) printing technology in the field of neurosurgery have given a newer modality of management for patients. In this article, we intend to share our institutional experience regarding the use of 3D printing in three modalities, namely, cranioplasty using customized 3D-printed molds of polymethylmethacrylate, 3D-printed model-assisted management of craniovertebral (CV) junction abnormalities, and 3D model-assisted management of brain tumors.Materials and Methods:A total of 55 patients were included in our study between March 2017 and December 2019 at S. M. S Medical College, Jaipur, India. 3D-printed models were prepared for cranioplasty in 30 cases, CV junction anomalies in 18 cases, and brain tumors in 7 cases. Preoperative and postoperative data were analyzed as per the diagnosis.Results:In cranioplasty, cranial contour and approximation of the margins were excellent and esthetic appearance improved in all patients. In CV junction anomalies, neck pain and myelopathy were improved in all patients, as analyzed using the visual analog scale and the Japanese Orthopedic Association Scale score, respectively. Our questionnaire survey revealed that 3D models for brain tumors were useful in understanding space interval and depth intraoperatively with added advantage of patient education.Conclusion:Rapid prototyping 3D-printing technologies provide a practical and anatomically accurate means to produce patient-specific and disease-specific models. These models allow for surgical planning, training, simulation, and devices for the assessment and treatment of neurosurgical disease. Expansion of this technology in neurosurgery will serve practitioners, trainees, and patients.

Cytoskeleton-associated protein 4 (CKAP4) promotes malignant progression of human gliomas through inhibition of the Hippo signaling pathway.

Gliomas are the most common and aggressive malignant brain tumors and are associated with high mortality and incidence in humans. Despite rigorous multi-modal therapy, including surgery, chemotherapy and radiotherapy, patients with malignant glioma survive an average of 12-15months following primary diagnosis. Therefore, new molecular biomarkers are urgently needed for diagnosis and targeted therapy. Here, we find that suppression of CKAP4 might inhibit glioma growth through regulation of Hippo signaling. We examined the expression levels of CKAP4 through analysis of RNA sequencing data from GEPIA and CGGA databases. Then, Lentivirus was used toconstruct stable cell lines with knockout or overexpression of CKAP4. Next, the function of CKAP4 on glioma was investigated in vitro and in an orthotopic brain tumor model in mice. Lastly, luciferase reporter assay, immunofluorescence and immunoblotting were performed to explore the potential mechanism of how CKAP4 affects gliomas. CKAP4 is highly upregulated in glioma and high CKAP4 expressing tumors were associated with poor patient survival. And CKAP4 promotes malignant progression of gliomas via inhibiting Hippo signaling. CKAP4 has potential as a promising biomarker and can predict the prognosis of patients with gliomas. And targeting CKAP4 expression may be an effective therapeutic strategy for the treatment of human gliomas.

A mathematical model and numerical solution for brain tumor derived using fractional operator

In this paper, we present a mathematical model of brain tumor. This model is an extension of a simple two-dimensional mathematical model of glioma growth and diffusion which is derived from fractional operator in terms of Caputo which is called the fractional Burgess equations (FBEs). To obtain a solution for this model, a numerical technique is presented which is based on operational matrix. First, we assume the solution of the problem under the study is as an expansion of the Bernoulli polynomials. Then with combination of the operational matrix based on the Bernoulli polynomials and collocation method, the problem under the study is changed to a system of nonlinear algebraic equations. Finally, the proposed technique is simulated and tested on three types of the FBEs to confirm the superiority and accuracy.

Methods for intratumoral microdialysis probe targeting and validation in murine brain tumor models

BackgroundMicrodialysis is a well validated sampling technique that can be used for pharmacokinetic studies of oncological drugs targeting the central nervous system. This technique has also been applied to evaluate tumor metabolism and identify pharmacodynamic biomarkers of drug activity. Despite the potential utility of microdialysis for therapeutic discovery, variability in tumor size and location hamper routine use of microdialysis as a preclinical tool. Quantitative validation of microdialysis membrane location relative to radiographically evident tumor regions could facilitate rigorous preclinical studies. However, a widely accessible standardized workflow for preclinical catheter placement and validation is needed. New methodWe provide methods for a workflow to yield tailored placement of microdialysis probes within a murine intracranial tumor and illustrate in an IDH1-mutant patient-derived xenograft (PDX) model. This detailed workflow uses a freely available on-line tool built within 3D-slicer freeware to target microdialysis probe placement within the tumor core and validate probe placement fully within the tumor. ResultsWe illustrate use of this workflow to validate microdialysis probe location relative to implanted IDH1-mutant PDXs, using the microdialysis probes to quantify levels of extracellular onco-metabolite D-2 hydroxyglutarate. Comparison with existing methodsPrevious methods have used 3D slicer to reliably measure tumor volumes. Prior microdialysis studies have targeted expected tumor locations without validation. ConclusionsThe new method offers a streamlined and freely available workflow in 3D slicer to optimize and validate microdialysis probe placement within a murine brain tumor.

Machine learning for real-time optical property recovery in interstitial photodynamic therapy: a stimulation-based study.

With the continued development of non-toxic photosensitizer drugs, interstitial photodynamic therapy (iPDT) is showing more favorable outcomes in recent clinical trials. IPDT planning is crucial to further increase the treatment efficacy. However, it remains a major challenge to generate a high-quality, patient-specific plan due to uncertainty in tissue optical properties (OPs), µ a and µ s . These parameters govern how light propagates inside tissues, and any deviation from the planning-assumed values during treatment could significantly affect the treatment outcome. In this work, we increase the robustness of iPDT against OP variations by using machine learning models to recover the patient-specific OPs from light dosimetry measurements and then re-optimizing the diffusers' optical powers to adapt to these OPs in real time. Simulations on virtual brain tumor models show that reoptimizing the power allocation with the recovered OPs significantly reduces uncertainty in the predicted light dosimetry for all tissues involved.

Liposomal IR-780 as a Highly Stable Nanotheranostic Agent for Improved Photothermal/Photodynamic Therapy of Brain Tumors by Convection-Enhanced Delivery.

Simple SummaryTo improve the use of hydrophobic photosensitizer IR-780 in photothermal/photodynamic therapy (PTT/PDT), we entrap IR-780 within the lipid bilayer of liposomes (ILs). Compared to free IR-780, ILs showed well-preserved photothermal response by maintaining the photostability of IR-780 from repeated near infrared (NIR) laser exposure both in vitro and in vivo. Combined with fast endocytosis by human glioblastoma cells, ILs demonstrated enhanced cytotoxicity and induced higher cell apoptosis rate toward human glioblastoma cells over free IR-780, due to PTT with overexpression of heat shock protein and PDT with generation of intracellular reactive oxygen species. To overcome the blood–brain barrier, we used convection enhanced delivery (CED) for specific delivery of ILs to brain tumors in intracranial glioma xenograft. Upon three successive NIR laser irradiations, the liposomal IR-780 could significantly improve the anti-cancer efficacy in glioma treatment, leading to diminished intracranial tumor size and prolonged animal survival time.As a hydrophobic photosensitizer, IR-780 suffers from poor water solubility and low photostability under near infrared (NIR) light, which severely limits its use during successive NIR laser-assisted photothermal/photodynamic therapy (PTT/PDT). To solve this problem, we fabricate cationic IR-780-loaded liposomes (ILs) by entrapping IR-780 within the lipid bilayer of liposomes. We demonstrate enhanced photostability of IR-780 in ILs with well-preserved photothermal response after three repeated NIR laser exposures, in contrast to the rapid decomposition of free IR-780. The cationic nature of ILs promotes fast endocytosis of liposomal IR-780 by U87MG human glioblastoma cells within 30 min. For PTT/PDT in vitro, ILs treatment plus NIR laser irradiation leads to overexpression of heat shock protein 70 and generation of intracellular reactive oxygen species by U87MG cells, resulting in enhanced cytotoxicity and higher cell apoptosis rate. Using intracranial glioma xenograft in nude mice and administration of ILs by convection enhanced delivery (CED) to overcome blood-brain barrier, liposomal IR-780 could be specifically delivered to the brain tumor, as demonstrated from fluorescence imaging. By providing a highly stable liposomal IR-780, ILs significantly improved anti-cancer efficacy in glioma treatment, as revealed from various diagnostic imaging tools and histological examination. Overall, CED of ILs plus successive laser-assisted PTT/PDT may be an alternative approach for treating brain tumor, which can retard glioma growth and prolong animal survival times from orthotopic brain tumor models.

Chloride intracellular channel protein 2 is secreted and inhibits MMP14 activity, while preventing tumor cell invasion and metastasis

The abilities to invade surrounding tissues and metastasize to distant organs are the most outstanding features that distinguish malignant from benign tumors. However, the mechanisms preventing the invasion and metastasis of benign tumor cells remain unclear. By using our own rat distant metastasis model, gene expression of cells in primary tumors was compared with that in metastasized tumors. Among many distinct gene expressions, we have focused on chloride intracellular channel protein 2 (CLIC2), an ion channel protein of as-yet unknown function, which was predominantly expressed in the primary tumors. We created CLIC2 overexpressing rat glioma cell line and utilized benign human meningioma cells with naturally high CLIC2 expression. CLIC2 was expressed at higher levels in benign human brain tumors than in their malignant counterparts. Moreover, its high expression was associated with prolonged survival in the rat metastasis and brain tumor models as well as with progression-free survival in patients with brain tumors. CLIC2 was also correlated with the decreased blood vessel permeability likely by increased contents of cell adhesion molecules. We found that CLIC2 was secreted extracellularly, and bound to matrix metalloproteinase (MMP) 14. Furthermore, CLIC2 prevented the localization of MMP14 in the plasma membrane, and inhibited its enzymatic activity. Indeed, overexpressing CLIC2 and recombinant CLIC2 protein effectively suppressed malignant cell invasion, whereas CLIC2 knockdown reversed these effects. Thus, CLIC2 suppress invasion and metastasis of benign tumors at least partly by inhibiting MMP14 activity.

Abstract 634: Targeting metastatic brain germinoma using PDOX tumor models with sequential metastases

Abstract Introduction. Intracranial germ cell tumor is an Asian-centric tumor, showing great variation in geographical distribution, with an incidence 5-8 fold greater in Asia/East Asia, than Western countries. Germinoma is one histological subtype of intracranial germ cell tumors. Current standard of care for germinoma, is chemoradiotherapy. Metastatic/recurrent germinoma have poor prognosis. There is currently no targeted therapy. Purpose: To create patient-derived orthotopic xenograft (PDOX) models of distant metastases in brain germinoma beyond brain boundaries to recapitulate sequential metastases of original patient tumors. These tumor models will serve as clinically accurate metastatic models for preclinical testing, replicating patient's pattern of tumor progression/metastases. There is a lack of such metastatic brain tumor models in the field. c-KIT is the most common mutation in germinoma. We evaluated preclinical efficacy of a c-KIT inhibitor. Experimental Procedures. Most PDOX brain tumor models are created by implanting patient-derived tumor cells in cerebrum/cerebellum. Germinoma originates from pineal gland/suprasellar region. We implanted germinoma cells nearer location of tumor origin. We were successful and achieved consistent tumor implantation into this location (100% post-surgery survival, 0% surgical mortality, 0% post-surgical neurological deficits). Summary. We successfully established two models with sequential metastases. Model 1 was created from metastatic cells from spinal recurrence (Patient 1) post-chemo-radiotherapy. This model faithfully created skull base metastases in 100% of murine replicas (n=9). Both tumor cells from brain and skull base metastases (n=7) were serially transplantable. Our findings were replicated (n=5) and consistent. Model 2 was created from metastatic cells of cerebrospinal fluid dissemination (Patient 2). Brain surface metastases were formed recapitulating patient tumor. Both models recapitulated a similar path of metastases in craniospinal axis, corresponding to respective patients. c-KIT inhibitor, was effective in significantly prolonging animal survival in Model 2 (p=0.0046) but not in Model 1 (p=0.0541). In Model 2, c-kit inhibition in-vivo was efficacious and resulted in 3-fold effect, (1) Significantly improved animal survival, (2) Macroscopically reduced frequency, severity of brain surface metastases (Controls=50%, Treated=14%, lesser in severity), (3) Microscopically reduced tumor burden to minimal residual disease limited to cerebrospinal fluid outline. Control mice brains were largely replaced by tumor bulk, while treated mice had minimal residual tumor burden. Conclusions. Understanding metastatic patterns in such aggressive models is relevant in biologic targeting for preclinical studies and clinical decision to extend radiotherapy field in recurrent disease. Citation Format: Shiying Huang, Sekar Karthik, Jack M Su, Qi Lin, YuChen Du, Ching C Lau, Adesina Adekunle, Angela Major, Kam-Man Hui, M. Tarek Elghetany, Xiaonan Li, Wan-Yee Teo. Targeting metastatic brain germinoma using PDOX tumor models with sequential metastases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 634.

Abstract 1474: Antitumor activity of belvarafenib in melanoma brain metastasis and NRAS mutation melanoma models

Abstract Over the past decade, BRAF inhibitors, either alone or in combination with MEK inhibitors, have set a new milestone in the treatment of patients with BRAF V600 mutation melanoma, which accounts for about 50% of melanomas. More recently, anti-PD-(L)1 therapies such as nivolumab or pembrolizumab and atezolizumab have been approved as very useful treatments for patients with melanoma. Nevertheless, metastases to the brain in melanoma patients, which are highly aggressive and associated with poor outcomes, still require better treatment. Also, to date, no targeted therapy has been developed for patients with NRAS mutations, which account for about 20% of melanoma. Previously, we reported that belvarafenib (HM95573 or GDC5573) strongly inhibits the growth of various cancer cell lines harboring BRAF, NRAS or KRAS mutations and demonstrates remarkable anticancer efficacy in preclinical animal models. Belvarafenib, currently in Phase 1 clinical trials, is a potent and selective pan-RAF kinase inhibitor showing strong inhibition activities for BRAF WT, BRAF V600E and CRAF. Here, we present the results of preclinical model studies suggesting that belvarafenib could be effective in treating melanoma patients with metastases to the brain as well as NRAS mutation melanoma patients. First, belvarafenib was found to be able to accumulate in the brain of mice and rats following oral administration. The exposure of belvarafenib in the brain was similar to that in plasma (approximately 100% brain to plasma ratio). This level of brain penetration of belvarafenib differs significantly from currently approved BRAF inhibitors that are known to have low brain penetration. Belvarafenib showed excellent anti-tumor activity in an orthotopic brain tumor model using melanoma cells. Belvarafenib significantly increased the overall survival of mice implanted intracranially with BRAF V600E A375SM melanoma cells. Overall, our data demonstrated that belvarafenib has therapeutic potential to treat NRAS mutation melanoma patients. In fact, in the NRAS G13D K1735 murine melanoma cell syngeneic mice model, the combination of belvarafenib with atezolizumab (anti-PD-L1) significantly inhibited tumor growth and effectively induced infiltration of cytotoxic T-cells into tumor tissues. These effects of belvarafenib identified in the above preclinical studies need to be immediately warranted through appropriate clinical trials. Citation Format: Inhwan Bae, Yu-Yon Kim, Jisook Kim, Hyunjin Park, Taehyun Song, Joo-Yun Byun, Semi Lim, Young Hoon Kim, Young Gil Ahn, Kwee Hyun Suh. Antitumor activity of belvarafenib in melanoma brain metastasis and NRAS mutation melanoma models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1474.