Drug Screening System Research Articles

Cryptococcus neoformans Prp8 Intein: An In Vivo Target-Based Drug Screening System in Saccharomyces cerevisiae to Identify Protein Splicing Inhibitors and Explore Its Dynamics.

Inteins are genetic mobile elements that are inserted within protein-coding genes, which are usually housekeeping genes. They are transcribed and translated along with the host gene, then catalyze their own splicing out of the host protein, which assumes its functional conformation thereafter. As Prp8 inteins are found in several important fungal pathogens and are absent in mammals, they are considered potential therapeutic targets since inhibiting their splicing would selectively block the maturation of fungal proteins. We developed a target-based drug screening system to evaluate the splicing of Prp8 intein from the yeast pathogen Cryptococcus neoformans (CnePrp8i) using Saccharomyces cerevisiae Ura3 as a non-native host protein. In our heterologous system, intein splicing preserved the full functionality of Ura3. To validate the system for drug screening, we examined cisplatin, which has been described as an intein splicing inhibitor. By using our system, new potential protein splicing inhibitors may be identified and used, in the future, as a new class of drugs for mycosis treatment. Our system also greatly facilitates the visualization of CnePrp8i splicing dynamics in vivo.

Simultaneous augmentation of muscle and bone by locomomimetism through calcium-PGC-1α signaling

Impaired locomotion has been extensively studied worldwide because those afflicted with it have a potential risk of becoming bedridden. Physical exercise at times can be an effective remedy for frailty, but exercise therapy cannot be applied in all clinical cases. Medication is safer than exercise, but there are no drugs that reinforce both muscle and bone when administered alone. Multiple medications increase the risk of adverse events; thus, there is a need for individual drugs targeting both tissues. To this end, we established a novel sequential drug screening system and identified an aminoindazole derivative, locamidazole (LAMZ), which promotes both myogenesis and osteoblastogenesis while suppressing osteoclastogenesis. Administration of this drug enhanced locomotor function, with muscle and bone significantly strengthened. Mechanistically, LAMZ induced Mef2c and PGC-1α in a calcium signaling–dependent manner. As this signaling is activated upon physical exercise, LAMZ mimics physical exercise. Thus, LAMZ is a promising therapeutic drug for locomotor diseases, including sarcopenia and osteoporosis.

Studying the Inflammatory Responses to Amyloid Beta Oligomers in Brain-Specific Pericyte and Endothelial Co-culture from Human Stem Cells.

Recently, the in vitro blood brain barrier (BBB) models derived from human pluripotent stem cells have been given extensive attention in therapeutics due to the implications it has with the health of the central nervous system. It is essential to create an accurate BBB model in vitro in order to better understand the properties of the BBB and how it can respond to inflammatory stimulation and be passed by targeted or non-targeted cell therapeutics, more specifically extracellular vesicles. Brain-specific pericytes (iPCs) were differentiated from iPSK3 cells using dual SMAD signaling inhibitors and Wnt activation plus fibroblast growth factor 2 (FGF-2). The derived cells were characterized by immunostaining, flow cytometry and RT-PCR. In parallel, blood vessels organoids were derived using Wnt activation, BMP4, FGF2, VEGF and SB431542. The organoids were replated and treated with retinoic acid to enhance the blood brain barrier (BBB) features in the differentiated brain endothelial cells (iECs). Co-culture was performed for the iPCs and iECs in transwell system and 3-D microfluidics channels. The derived iPCs expressed common markers PDGFRb and NG2, as well as brain-specific genes FOXF2, ABCC9, KCNJ8, and ZIC1. The derived iECs expressed common endothelial cell markers CD31, VE-cadherin, as well as BBB-associated genes BRCP, GLUT-1, PGP, ABCC1, OCLN, SLC2A1. The co-culture of the two cell types responded to the stimulation of amyloid β42 oligomers by the upregulation of expression of TNFa, IL6, NFKB, Casp3, SOD2 and TP53. The co-culture also showed the property of trans-endothelial electrical resistance. The proof-of-concept vascularization strategy was demonstrated in a 3-D microfluidics-based device. The derived iPCs and iECs have brain-specific properties and the co-culture of iPCs and iECs provides an in vitro BBB model that show inflammatory response. This study has significance in establishing micro-physiological systems for neurological disease modeling and drug screening.

Improving Drug Sensitivity of HIV-1 Protease Inhibitors by Restriction of Cellular Efflux System in a Fission Yeast Model.

Fission yeast can be used as a cell-based system for high-throughput drug screening. However, higher drug concentrations are often needed to achieve the same effect as in mammalian cells. Our goal here was to improve drug sensitivity so reduced drugs could be used. Three different methods affecting drug uptakes were tested using an FDA-approved HIV-1 protease inhibitor (PI) drug Darunavir (DRV). First, we tested whether spheroplasts without cell walls increase the drug sensitivity. Second, we examined whether electroporation could be used. Although small improvements were observed, neither of these two methods showed significant increase in the EC50 values of DRV compared with the traditional method. In contrast, when DRV was tested in a mutant strain PR836 that lacks key proteins regulating cellular efflux, a significant increase in the EC50 was observed. A comparison of nine FDA-approved HIV-1 PI drugs between the wild-type RE294 strain and the mutant PR836 strain showed marked enhancement of the drug sensitivities ranging from an increase of 0.56 log to 2.48 logs. Therefore, restricting cellular efflux through the adaption of the described fission yeast mutant strain enhances the drug sensitivity, reduces the amount of drug used, and increases the chance of success in future drug discovery.

Replacement of Mouse Microglia With Human Induced Pluripotent Stem Cell (hiPSC)-Derived Microglia in Mouse Organotypic Slice Cultures.

Microglia, the major immune cells in the brain, are reported to differ in gene expression patterns among species. Therefore, it would be preferable in some cases to use human microglia rather than mouse microglia in microglia-targeted disease research. In the past half a decade, researchers have developed in vivo transplantation methods in which human induced pluripotent stem cell-derived microglia (hiPSC-MG) are transplanted into a living mouse brain. However, in vivo transplantation methods are not necessarily accessible to all researchers due to the difficulty of obtaining the materials needed and the transplantation technique itself. In addition, for in vivo systems for microglia-targeted drug screening, it is difficult to control the pharmacokinetics, especially blood-brain barrier permeability. Therefore, in addition to existing in vivo transplantation systems, the development of an ex vivo transplantation system would help to further evaluate the properties of hiPSC-MG. In this study, we aimed to establish a method to efficiently transplant hiPSC-MG into cultured mouse hippocampal slices. We found that approximately 80% of the total microglia in a cultured slice were replaced by hiPSC-derived microglia when innate microglia were pharmacologically removed prior to transplantation. Furthermore, when neuronal death was induced by applying Kainic acid (KA) to slice cultures, transplanted hiPSC-MG changed their morphology and phagocytosed cell debris. Thus, this study provides a method to transplant hiPSC-MG into the mouse hippocampal slice cultures with a high replacement rate. Because the transplanted microglia survived and exerted phagocytic functions, this method will be useful for evaluating the properties of hiPSC-MG ex vivo.

3D printed human organoids: High throughput system for drug screening and testing in current COVID-19 pandemic.

In the current pandemic, scenario the world is facing a huge shortage of effective drugs and other prophylactic medicine to treat patients which created havoc in several countries with poor resources. With limited demand and supply of effective drugs, researchers rushed to repurpose the existing approved drugs for the treatment of COVID-19. The process of drug screening and testing is very costly and requires several steps for validation and treatment efficacy evaluation ranging from in-vitro to in-vivo setups. After these steps, a clinical trial is mandatory for the evaluation of treatment efficacy and side effects in humans. These processes enhance the overall cost and sometimes the lead molecule show adverse effects in humans and the trial ends up in the final stages. Recently with the advent of three-dimensional (3D) organoid culture which mimics the human tissue exactly the process of drug screening and testing can be done in a faster and cost-effective manner. Further 3D organoids prepared from stems cells taken from individuals can be beneficial for personalized drug therapy which could save millions of lives. This review discussed approaches and techniques for the synthesis of 3D-printed human organoids for drug screening. The key findings of the usage of organoids for personalized medicine for the treatment of COVID-19 have been discussed. In the end, the key challenges for the wide applicability of human organoids for drug screening with prospects of future orientation have been included.

Abstract 200: Microtube Array Membrane-Hollow Fiber Assay (MTAM-HFA): A combination of immunocheckpoint blocker (ICBs) and secondary drug anti-cancer drug screening system with significant clinical value propositions for personalized medicine and pre-clinical drug development

Abstract Despite modern advances in technology, significant challenges still plague the development of new cancer drugs and cancer care. To address these issue, we are introducing our novel platform technology- Microtube Array Membrane-Hollow Fiber Assay (MTAM-HFA) that is based on combination of the patented MTAM membrane with unique microstructures; replacing the role of traditional hollow fibers enabled the entire HFA assay to be significantly improved in terms of clinical relevance, accuracy, reliability and cost resulting in unprecedented value propositions as an anti-cancer drug screening platform. In the current work, we have clinically demonstrated in several cases where the MTAM-HFA system is capable of accurately predicting clinical response in several Cancers-Lung, Breast and Colon cancers; with unprecedented flexibility of being screened against multiple drug class (including anti-PD1/PDL1) in monotherapy and combination therapy settings. Despite being a phenotypic based assay, the test cycle of MTAM-HFA is only 7-10 days while maintaining a very low cost of up to 60% savings when compared to traditional models such as Patient Derived Xenografts (PDX). When screening ICBs, without the need to use Genetically Engineered Mice (GEM) translates to a cost savings of 98% just for the animal model alone. The combined value proposition of MTAM-HFA makes is a suitable system fulfilling the needs of multiple stakeholders. Citation Format: Wan-Ting Huang, Chee Ho Chew, Amanda Chen, Kang-Yuan Lee, Hsing-Lun Lee, Po-Hao Feng, Jeng-Fong Chiou, Kuan-Chou Lin, Po-Li Wei, Chien-Chung Chen. Microtube Array Membrane-Hollow Fiber Assay (MTAM-HFA): A combination of immunocheckpoint blocker (ICBs) and secondary drug anti-cancer drug screening system with significant clinical value propositions for personalized medicine and pre-clinical drug development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 200.

AGE/RAGE axis regulates reversible transition to quiescent states of ALK-rearranged NSCLC and pancreatic cancer cells in monolayer cultures

Cancer recurrence due to tumor cell quiescence after therapy and long-term remission is associated with cancer-related death. Previous studies have used cell models that are unable to return to a proliferative state; thus, the transition between quiescent and proliferative states is not well understood. Here, we report monolayer cancer cell models wherein the human non-small cell lung carcinoma cell line H2228 and pancreatic cancer cell line AsPC-1 can be reversibly induced to a quiescent state under hypoxic and serum-starved (HSS) conditions. Transcriptome and metabolome dual-omics profiles of these cells were compared with those of the human lung adenocarcinoma cell line A549, which was unable to enter a quiescent state under HSS conditions. The quiescence-inducible cells had substantially lower intracellular pyruvate and ATP levels in the quiescent state than in the proliferative state, and their response to sudden demand for energy was dramatically reduced. Furthermore, in quiescence-inducible cells, the transition between quiescent and proliferative states of these cells was regulated by the balance between the proliferation-promoting Ras and Rap1 signaling and the suppressive AGE/RAGE signaling. These cell models elucidate the transition between quiescent and proliferative states, allowing the development of drug-screening systems for quiescent tumor cells.

Intrinsic epigenetic control of angiogenesis in induced pluripotent stem cell-derived endothelium regulates vascular regeneration

Human-induced pluripotent stem cell-derived endothelial cells (iECs) provide opportunities to study vascular development and regeneration, develop cardiovascular therapeutics, and engineer model systems for drug screening. The differentiation and characterization of iECs are well established; however, the mechanisms governing their angiogenic phenotype remain unknown. Here, we aimed to determine the angiogenic phenotype of iECs and the regulatory mechanism controlling their regenerative capacity. In a comparative study with HUVECs, we show that iECs increased expression of vascular endothelial growth factor receptor 2 (VEGFR2) mediates their highly angiogenic phenotype via regulation of glycolysis enzymes, filopodia formation, VEGF mediated migration, and robust sprouting. We find that the elevated expression of VEGFR2 is epigenetically regulated via intrinsic acetylation of histone 3 at lysine 27 by histone acetyltransferase P300. Utilizing a zebrafish xenograft model, we demonstrate that the ability of iECs to promote the regeneration of the amputated fin can be modulated by P300 activity. These findings demonstrate how the innate epigenetic status of iECs regulates their phenotype with implications for their therapeutic potential.

Cell fiber-based 3D tissue array for drug response assay

For the establishment of a reproducible and sensitive assay system for three-dimensional (3D) tissue-based drug screening, it is essential to develop 3D tissue arrays with uniform shapes and high cell numbers that prevent cell death in the center of the tissue. In recent years, 3D tissue arrays based on spheroids have attracted increased attention. However, they have only been used in specific tissues with hypoxic regions, such as cancer tissues, because nutrient deprivation and hypoxic regions are formed in the core as spheroids grow. Herein, we propose a method to array cell-encapsulated tube-like tissue (cell fiber (CF)) with diameters < 150 μm to prevent nutrient deprivation and hypoxia using a device that can fix the CFs, section them in uniform sizes, and transfer them to a 96-well plate. We fabricated the arrays of CF fragments from cell lines (GT1-7), cancer cells (HeLa), mouse neural stem cells (mNSCs) and differentiated mNSCs, and performed drug response assays. The array of CF fragments assessed the drug response differences among different cell types and drug responses specific to 3D tissues. The array of CF fragments may be used as a versatile drug screening system to detect drug sensitivities in various types of tissues.

The application and research advances of organoids in clinical medicine

<p indent="0mm">Three-dimensional cultured organoids recapitulate histopathologic and molecular characteristics of the primary tissues. The rise and development of organoids provide an important platform for clinical medicine research. Patient-derived organoids could be used as models to help clinicians to better understand the pathophysiology of related diseases and formulate corresponding treatment strategies; Organoids derived from tumors could be utilized as a drug screening system for precision medicine, helping clinicians to select the most appropriate regimen for cancer patients and to improve the clinical outcome. This review would summarize the application and research advances of organoids in clinical medicine.

Identifying Acute Cardiac Hazard in Early Drug Discovery Using a Calcium Transient High-Throughput Assay in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

Introduction: Early identification of cardiac risk is essential for reducing late-stage attrition in drug development. We adapted the previously published cardiac hazard risk-scoring system using a calcium transient assay in human stem cell-derived CMs for the identification of cardiac risks recorded from the new hiPSC-CM line and investigated its predictivity and translational value based on the screening of a large number of reference and proprietary compounds. Methods: Evaluation of 55 reference drugs provided the translation of various pharmacological effects into a single hazard label (no, low, high, or very high hazard) using a Ca2+-sensitive fluorescent dye assay recorded by -by FDSS/µCell Functional Drug Screening System (Hamamatsu on hiPSC-CM line (FCDI iCell Cardiomyocytes2). Results: Application of the adapted hazard scoring system in the Ca2+ transient assay, using a second hiPS-CM line, provided comparable scoring results and predictivity of hazard, to the previously published scoring approach, with different pharmacological drug classes, as well as screening new chemical entities (NCE’s) using a single hazard label from four different scoring levels (no, low, high, or very high hazard). The scoring system results also showed minimal variability across three different lots of hiPSC-CMs, indicating good reproducibility of the cell line. The predictivity values (sensitivity and specificity) for drug-induced acute cardiac risk for QT-interval prolongation and Torsade de pointes (TdPs) were >95% and statistical modeling confirmed the prediction of proarrhythmic risk. The outcomes of the NCEs also showed consistency with findings in other well-established in vitro and in vivo cardiac risk assays. Conclusion: Evaluation of a large list of reference compounds and internal NCEs has confirmed the applicability of the adaptations made to the previously published novel scoring system for the hiPSC-CMs. The validation also established the predictivity for drug-induced cardiac risks with good translation to other established preclinical in vitro and in vivo assays, confirming the application of this novel scoring system in different stem cell-CM lines for early cardiac hazard identification.

A novel cell tool for α2δ-1-NMDAR target-based analgesic drug discovery

The α2δ-1 protein coded by Cacna2d1 is dramatically up-regulated in dorsal root ganglion (DRG) neurons and spinal dorsal horn following sensory nerve injury in various animal models of neuropathic pain. Cacna2d1 overexpression potentiates presynaptic and postsynaptic NMDAR activity of spinal dorsal horn neurons to cause pain hypersensitivity. The α2δ-1-NMDAR interaction promotes surface trafficking and synaptic targeting of NMDARs in neuropathic pain caused by chemotherapeutic agents and peripheral nerve injury, as well as in other pathological conditions such as in the paraventricular nucleus (PVN) with neurogenic hypertension and in the brain with ischemic stroke. The lentiviral transfection method was used to construct a human embryonic kidney HEK293T cell line that could stably express α2δ-1-NMDAR complex. A stably transfected cell line was observed by florescence microscope, and identified by RT-qPCR and Western blotting. The results showed that the HEK293T cell line was successfully transfected and the genes could be stably expressed. Subsequently, the transfected cell line was successfully developed into a target drug screening system using patch clamp techniques. It provides a promising cell model for further research on the interaction mechanism of α2δ-1-NMDAR complex and drug screening for chronic pain and related diseases with low side effects.

Organoid screening reveals epigenetic vulnerabilities in human colorectal cancer.

Precision oncology presumes an accurate prediction of drug response on the basis of the molecular profile of tumors. However, the extent to which patient-derived tumor organoids recapitulate the response of in vivo tumors to a given drug remains obscure. To gain insights into the pharmacobiology of human colorectal cancer (CRC), we here created a robust drug screening platform for patient-derived colorectal organoids. Application of suspension culture increased organoid scalability, and a refinement of the culture condition enabled incorporation of normal and precursor organoids to high-throughput drug screening. Drug screening identified bromodomain and extra-terminal (BET) bromodomain protein inhibitor as a cancer-selective growth suppressor that targets genes aberrantly activated in CRC. A multi-omics analysis identified an association between checkpoint with forkhead and ring finger domaines (CHFR) silencing and paclitaxel sensitivity, which was further validated by gene engineering of organoids and in xenografts. Our findings highlight the utility of multiparametric validation in enhancing the biological and clinical fidelity of a drug screening system.

Otic Organoids Containing Spiral Ganglion Neuron-like Cells Derived from Human-induced Pluripotent Stem Cells as a Model of Drug-induced Neuropathy.

The spiral ganglion of the cochlea is essential for hearing and contains primary bipolar neurons that relay action potentials generated by mechanosensory hair cells. Injury to spiral ganglion neurons (SGNs) causes permanent hearing loss because these cells have limited regenerative capacity. Establishment of human cell-derived inner ear tissue in vitro could facilitate the development of treatments for hearing loss. Here, we report a stepwise protocol for differentiating human-induced pluripotent stem cells (hiPSCs) into otic organoids that contain SGN-like cells and demonstrate that otic organoids have potential for use as an experimental model of drug-induced neuropathy. Otic progenitor cells (OPCs) were created by 2D culture of hiPSCs for 9 days. Otic spheroids were formed after 2D culture of OPCs for 2 days in a hypoxic environment. Otic organoids were generated by 3D culture of otic spheroids under hypoxic conditions for 5 days and normoxic conditions for a further 30 days or more. The protein expression profile, morphological characteristics, and electrophysiological properties of SGN-like cells in otic organoids were similar to those of primary SGNs. Live-cell imaging of AAV-syn-EGFP-labeled neurons demonstrated temporal changes in cell morphology and revealed the toxic effects of ouabain (which causes SGN-specific damage in animal experiments) and cisplatin (a chemotherapeutic drug with ototoxic adverse effects). Furthermore, a cyclin-dependent kinase-2 inhibitor suppressed the toxic actions of cisplatin on SGN-like cells in otic organoids. The otic organoid described here is a candidate novel drug screening system and could be used to identify drugs for the prevention of cisplatin-induced neuropathy.

Development of in vitro three-dimensional drug screening system for obesity-related metabolic syndrome

Metabolic syndrome is increasingly common, and closely related with overweight or obesity. In the obese state, macrophages infiltrate to the adipose tissue (AT), resulting in chronic inflammation and insulin resistance in the AT cells. Recently, attention has been paid to the role of AT macrophages in metabolic disorders should be applied to the initial drug screening step, but it was difficult to mimic the inflammatory adipocytes using the traditional 2-dimensional (2D) culture. In this study, we developed the 3-dimensional (3D) culture system to overcome this limitation. After adipogenic differentiation, lipid droplets were highly accumulated in cells, and differentiation of preadipocytes was not declined by macrophage co-culture. However, only co-cultured cells expressed the insulin resistance features. Compare to mono-cultured adipocytes, co-cultured adipocytes showed reduced glucose uptake and GLUT4 did not translocated to cell membrane even though treatment of high concentration of insulin. Using 3D co-culture model, we develop a microwell-scale drug screening protocol to test anti-obesity effect. 3D cultured cells reacted more sensitive to drugs, and PPARγ antagonist GW9662 (10, 20 μM) repressed adipogenic differentiation in a concentration-dependent manner in 3D co-cultured cells.

Personalized Treatment of Advanced Gastric Cancer Guided by the MiniPDX Model.

Background The morbidity and mortality of gastric cancer are high in China. There are challenges to develop precise and individualized drug regimens for patients with gastric cancer after a standard treatment. Choosing the most appropriate anticancer drug after a patient developing drug resistance is very important to improve the patient's prognosis. MiniPDX has been widely used as a new and reliable preclinical research model to predict the sensitivity of anticancer drugs. Methods The OncoVee® MiniPDX system developed by Shanghai LIDE Biotech Co., Ltd. was used to establish the MiniPDX models using specimens of patients with gastric cancer. The cancer tissues were biopsied under endoscopy, and then, the tumor cell suspension was prepared for a drug sensitivity test by subcutaneously implanting into Balb/c-nude mice. The selected optimal regimen obtained from the MiniPDX assay was used to treat patients with drug-resistant gastric cancer. Results We successfully established an individualized and sensitive drug screening system for four patients from January 2021 to July 2021. MiniPDX models identified potentially effective drugs for these four patients, with partial remission in two of the patients after treatment and disease progression in the remaining of two patients. Severe side effects from chemotherapy or targeted therapy were not observed in all patients. Conclusion Establishing a personalized drug screening system for patients with drug-resistant gastric cancer can guide the selection of clinical drugs, improve the clinical benefit of patients, and avoid ineffective treatments. It can be an effective supplement for treatment options.

Genetic and Physiological Characterization of Fructose-1,6-Bisphosphate Aldolase and Glyceraldehyde-3-Phosphate Dehydrogenase in the Crabtree-Negative Yeast Kluyveromyces lactis

The milk yeast Kluyveromyces lactis degrades glucose through glycolysis and the pentose phosphate pathway and follows a mainly respiratory metabolism. Here, we investigated the role of two reactions which are required for the final steps of glucose degradation from both pathways, as well as for gluconeogenesis, namely fructose-1,6-bisphosphate aldolase (FBA) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In silico analyses identified one gene encoding the former (KlFBA1), and three genes encoding isoforms of the latter (KlTDH1, KlTDH2, KlGDP1). Phenotypic analyses were performed by deleting the genes from the haploid K. lactis genome. While Klfba1 deletions lacked detectable FBA activity, they still grew poorly on glucose. To investigate the in vivo importance of the GAPDH isoforms, different mutant combinations were analyzed for their growth behavior and enzymatic activity. KlTdh2 represented the major glycolytic GAPDH isoform, as its lack caused a slower growth on glucose. Cells lacking both KlTdh1 and KlTdh2 failed to grow on glucose but were still able to use ethanol as sole carbon sources, indicating that KlGdp1 is sufficient to promote gluconeogenesis. Life-cell fluorescence microscopy revealed that KlTdh2 accumulated in the nucleus upon exposure to oxidative stress, suggesting a moonlighting function of this isoform in the regulation of gene expression. Heterologous complementation of the Klfba1 deletion by the human ALDOA gene renders K. lactis a promising host for heterologous expression of human disease alleles and/or a screening system for specific drugs.

Establishment of high-throughput drug screening system using human limb bud mesenchymal cells derived from skeletal dysplasia specific iPSC

Establishment of high-throughput drug screening system using human limb bud mesenchymal cells derived from skeletal dysplasia specific iPSC

DDRE-43. SCREENING OF FDA-APPROVED COMPOUNDS FOR THE TREATMENT OF CHORDOMA, WITH IN VIVO VALIDATION USING THREE DIFFERENT XENOGRAFT MODELS, IDENTIFIES BRIGATINIB AS A POTENTIAL TREATMENT

Abstract BACKGROUND Chordoma is a rare malignant tumor with poor surgical control and no existing pharmacotherapies. Therefore, these tumors require additional research into novel therapeutics for their treatment. METHODS In this study we created a high-throughput drug screen and culture system to evaluate the efficacy of existing FDA-approved compounds in 10 chordoma cell lines and primary tumors. The cell lines were graciously donated by the Chordoma Foundation. Primary tumors were collected from our operating room. In vivo validation using three separate chordoma xenograft models was also performed through the Chordoma Foundation. One model was a primary clival pediatric tumor, the second was a metastatic sacral tumor, and the third model was a recurrent skull base tumor. RESULTS Using a 127 FDA-approved compound library, we screened 6 donated chordoma cell lines and 4 tumors resected from our institution. 5 of the chordomas were primary, 3 were recurrent, and 2 were metastatic. 6 chordoma were located in the sacrum, three were located in the mobile spine, and one was located in the clivus. Five tumors came from female patients and five came from male patients. After a single 72-hour 1um dose of brigatinib, the average tumor viability in our drug screen was reduced to 81.5% +/-9.5SD (p=1.61x10-13). In the in vivo studies, brigatinib achieved a full response in the metastatic sacral chordoma xenograft model (TGI=100%, p&amp;lt; 0.0001), a partial response in the recurrent skull base xenograft model (TGI=54%, p=0.3048), and no response in the primary clival pediatric xenograft model (TGI = 0%, p &amp;gt;0.9). CONCLUSIONS Brigatinib may be a viable treatment option for recurrent and metastatic chordomas.