Biology subjects

Kim, K.

Publications and source records attributed to Kim, K..

At least 289 records · Page 16Linked to original sources

bioRxiv2020

CRISPR/Cas9-mediated gene editing induces neurological recovery in an A53T-SNCA overexpression rat model of Parkinson's disease

To date, no publicly available disease-modifying therapy for Parkinsons disease has been developed. This can be partly attributed to the absence of techniques for in vivo deletion of the SNCA gene (encoding -synuclein), which is one of the key players in Parkinsons disease pathology. In particular, A53T-mutated SNCA (A53T-SNCA) is one of the most studied familial pathologic mutations in Parkinsons disease. Here we utilized a recently discovered genome editing technique, CRISPR/Cas9, to delete A53T-SNCA in vitro and in vivo. Among various CRISPR/Cas9 systems, SaCas9-KKH with a single guide RNA (sgRNA) targeting A53T-SNCA was packaged into adeno-associated virus. Adeno-associated virus carrying SaCas9-KKH significantly reduced A53T-SNCA levels in A53T-SNCA-overexpressed HEK293T cells, without off-target effects on wild-type SNCA. Furthermore, we tested the techniques in vivo therapeutic potential in a viral A53T-SNCA overexpression rat model of Parkinsons disease. Gene deletion of A53T-SNCA significantly prevented the overexpression of -synuclein, dopaminergic neurodegeneration, and parkinsonian motor symptoms, whereas a negative control without sgRNA did not. Our findings propose CRISPR/Cas9 system as a potential therapeutic tool for A53T-SNCA familial Parkinsons disease.

genetics↗

bioRxiv2020

Temporally-Divergent Regulatory Mechanisms Govern Neuronal Development and Diversification in the Neocortex

Mammalian neocortical neurons span one of the most diverse cell type spectra of any tissue. The regulatory strategies that neurons use during progressive development and maturation remain unclear. We present an integrated single-cell epigenomic and transcriptional analysis of individual classes of neurons from both mouse and marmoset neocortex, sampled during both early postmitotic stages of identity acquisition and later stages of neuronal plasticity and circuit integration. We find that in both species, the regulatory strategies controlling these early and late stages diverge: early postmitotic neurons use molecular regulatory programs with broader tissue distribution and greater evolutionary conservation, while programs active during later neuronal maturation implement more brain- and neuron-specific mechanisms showing greater evolutionary divergence. The data uncovers a temporally-regulated shift in regulatory choices, likely reflecting unique evolutionary constraints on distinct events of neuronal development in the neocortex.

neuroscience↗

bioRxiv2020

Targeting Vitamin-D receptor (VDR) by a small molecule antagonist MeTC7 inhibits PD-L1 but controls THMYCN neuroblastoma growth PD-L1 independently

Vitamin-D receptor (VDR) mRNA is enriched in malignant lung, ovarian and pancreatic tissues and showed poor prognoses. Calcitriol and stable or CRISPR-directed VDR upregulation increased PD-L1mRNA and protein expression in cancer cells in-vitro. A ChIP assay showed the binding of VDR with VDREPD-L1. Stattic, a STAT3 phosphorylation inhibitor blocked calcitriol or VDR overexpression induced PD-L1 upregulation. MeTC7, a VDR antagonist developed by us, reduced PD-L1 expression on macrophages, ovarian, lung, breast, and pancreatic cancer cells in-vitro. In radiotherapy inducible PD-L1 model of orthotopic MC38 murine colon cancer, MeTC7 decreased PD-L1 surface expression, suppressed inflammatory monocytes (IMs) population and increased intra-tumoral CD69+PD1+CD8+T-cells. Intriguingly, MeTC7 reduced TH-MYCN transgenic neuroblastoma tumor growth without affecting PD-L1 and tumor immune milieu. In summary, Vitamin-D/VDR drives PD-L1 expression on cancer cells via STAT-3. Inhibiting VDR exhibited anti-checkpoint effects in orthotopic colon tumors, whereas PDL1-independent and anti-VDR/MYCN effects controlled growth of transgenic neuroblastoma and xenografted tumors. SummaryVitamin-D/VDR induces PD-L1 expression on cancer cells via STAT-3; and targeting VDR by a novel small molecule antagonist MeTC7 exhibits both anti-PD-L1 and anti-VDR/MYCN effects in tumor models.

cancer biology↗

bioRxiv2020

CREB mediates the C. elegans dauer polyphenism through direct and cell-autonomous regulation of TGF-β expression

Animals can adapt to dynamic environmental conditions by modulating their developmental programs. Understanding the genetic architecture and molecular mechanisms underlying developmental plasticity in response to changing environments is an important and emerging area of research. Here, we show a novel role of cAMP response element binding protein (CREB)-encoding crh-1 gene in developmental polyphenism of C. elegans. Under conditions that promote normal development in wild-type animals, crh-1 mutants inappropriately form transient pre-dauer (L2d) larva and express the L2d marker gene. L2d formation in crh-1 mutants is specifically induced by the ascaroside pheromone ascr#5 (asc-{omega}C3; C3), and crh-1 functions autonomously in the ascr#5-sensing ASI neurons to inhibit L2d formation. Moreover, we find that CRH-1 directly binds upstream of the daf-7 TGF-{beta} locus and promotes its expression in the ASI neurons. Taken together, these results provide new insight into how animals alter their developmental programs in response to environmental changes.

developmental biology↗

bioRxiv2020

Organoid-derived adult human colonic epithelium responds to co-culture with a probiotic strain of Bifidobacterium longum

In germ-free animals the lack of microbes within the gut results in a propensity to mucosal inflammation among other immune deficits. This suggests microbes are essential for the healthy function of the human gut, but we have lacked a reproducible mechanistic model of interactions between human colonic epithelium and the anaerobic microbes within the gut. To establish the physiological effect of a common anaerobe in the human gut, we co-cultured a probiotic strain of Bifidobacterium longum (35624) with organoid-derived adult human colonic epithelium in asymmetric gas conditions (anoxic apically, 5% oxygen basolaterally) and compared to axenic ( germ-free) epithelium. Bacteria proliferated and retained their normal cellular morphology in the presence of the human colonic epithelium. The human colonic mucosa retained trans-epithelial electrical resistance (TEER) consistent with an intact epithelium but lower than in axenic conditions. Changes in TEER corresponded to changes in Claudin-family gene expression. Inflammation was repressed in co-culture as compared to axenic, with reduced expression of executor and pyroptosis caspases; reduced expression of activators and increased expression of inhibitors of NFKB; reduced expression of toll-like-receptors (TLRs) and increased expression of TOLLIP (a negative regulator of TLRs). Consistent with the presence of actively fermenting bacteria that produce lactate and acetate but do not produce butyrate, PPARA expression was increased while PPARG expression was reduced. As in germ-free animal experiments, axenic human colonic mucosa is poised for inflammation. Co-culture with Bifidobacterium longum resolved the pro-inflammatory state while modulating barrier function (via Claudin genes) and cellular energetics (via PPARA and PPARG genes). ImportanceExperiments in animals demonstrate the importance of microbes with the gut for the health of the gut. Many of the microbes within the healthy gut are strictly anaerobic and cannot grow in the presence of oxygen; human tissues require some oxygen to live. Here we observe the effect of growing an anaerobic bacterium - Bifidobacterium longum - typically found in the human gut with human colonic tissue. We observed that the human tissue responded favorably to being cultivated with the microbe as compared to being alone. This experiment confirms human colonic tissues reduce their inflammation in the presence of a bacteria typically found in the gut.

systems biology↗

bioRxiv2020

Open Source ImmGen: network perspective on metabolic diversity among mononuclear phagocytes

We dissect metabolic variability of mononuclear phagocyte (MNP) subpopulations across different tissues through integrative analysis of three large scale datasets. Specifically, we introduce ImmGen MNP Open Source dataset that profiled 337 samples and extended previous ImmGen effort which included 202 samples of mononuclear phagocytes and their progenitors. Next, we analysed Tabula Muris Senis dataset to extract data for 51,364 myeloid cells from 18 tissues. Taken together, a compendium of data assembled in this work covers phagocytic populations found across 38 different tissues. To analyse common metabolic features, we developed novel network-based computational approach for unbiased identification of key metabolic subnetworks based on cellular transcriptional profiles in large-scale datasets. Using ImmGen MNP Open Source dataset as baseline, we define 9 metabolic subnetworks that encapsulate the metabolic differences within mononuclear phagocytes, and demonstrate that these features are robustly found across all three datasets, including lipid metabolism, cholesterol biosynthesis, glycolysis, and a set of fatty acid related metabolic pathways, as well as nucleotide and folate metabolism. We systematically define major features specific to macrophage and dendritic cell subpopulations. Among other things, we find that cholesterol synthesis appears particularly active within the migratory dendritic cells. We demonstrate that interference with this pathway through statins administration diminishes migratory capacity of the dendritic cells in vivo. This result demonstrates the power of our approach and highlights importance of metabolic diversity among mononuclear phagocytes.

immunology↗

bioRxiv2020

Single adeno-associated virus-based multiplexed CRISPR-Cas9 system to nullify core components of the mammalian molecular clock

ABSTRACTThe mammalian molecular clock is based on a transcription-translation feedback loop (TTFL) containing Period1, 2 (Per1, 2), Cryptochrome1, 2 (Cry1, 2), and Brain and Muscle ARNT-Like 1 (Bmal1). TTFL robustness is endowed by genetic complementation between these components; therefore, multiple genes must be knocked out to physiologically investigate the molecular clock, which requires extensive research resources. To facilitate molecular clock disruption, we developed a CRISPR-Cas9-based single adeno-associated viral (AAV) system targeting the circadian clock (CSAC) for Pers, Crys, or Bmal1. First, we designed single guide RNAs (sgRNAs) targeting individual clock genes using an in silico approach and validated their efficiency in Neuro2a cells. To target multiple genes, multiplex sgRNA plasmids were constructed using Golden Gate assembly and expressed in viral vectors. CSAC efficiency was demonstrated by decreased protein expression in vitro and ablated molecular oscillation ex vivo. We also measured locomotor activity and body temperature in Cas9-expressing mice injected with CSAC at the suprachiasmatic nucleus. Circadian rhythm disruption was observed under free-running conditions, indicating that CSAC can efficiently and robustly disrupt molecular circadian clock. Thus, CSAC is a simple and powerful tool for investigating the physiological role of the molecular clock in vivo.Competing Interest StatementThe authors have declared no competing interest.View Full Text

molecular biology↗

bioRxiv2020

Supramodal executive control of attention: evidence from unimodal and crossmodal dual conflict effects

Although we have demonstrated that the executive control of attention acts supramodally as shown by significant correlation between conflict effects measures in visual and auditory tasks, no direct evidence of the equivalence in the computational mechanisms governing the allocation of executive control resources within and across modalities has been found. Here, in two independent groups of 40 participants, we examined the interaction effects of conflict processing in both unimodal (visual) and crossmodal (visual and auditory) dual-conflict paradigms (flanker conflict processing in Task 1 and then in the following Task 2) with a manipulation of the stimulus onset asynchrony (SOA). In both the unimodal and the crossmodal dual-conflict paradigms, the conflict processing of Task 1 interfered with the conflict processing of Task 2 when the SOA was short, reflecting an additive interference effect of Task 1 on Task 2 under the time constraints. These results suggest that there is a unified entity that oversees conflict processing acting supramodally by implementing comparable mechanisms in unimodal and crossmodal scenarios.

neuroscience↗

bioRxiv2020

Impaired hand dexterity function in a non-human primate model with chronic Parkinson's disease

Symptoms of Parkinsons disease (PD) caused by loss of dopaminergic neurons are accompanied by movement disorders, including tremors, rigidity, bradykinesia, and akinesia. Non-human primate (NHP) models with PD play an essential role in the analysis of PD pathophysiology and behavior symptoms. As impairments of hand dexterity function can affect activities of daily living in patients with PD, research on hand dexterity function in NHP models with chronic PD is essential. Traditional rating scales previously used in the evaluation of animal spontaneous behavior were insufficient due to factors related to subjectivity and passivity. Thus, experimentally designed applications for an appropriate apparatus are necessary. In this study, we aimed to longitudinally assess hand dexterity function using hand dexterity task (HDT) in NHP-PD models. To validate this assessment, we analyzed an alteration in Parkinsonian tremor symptoms, loss of dopaminergic neuron, and positron emission tomography (PET) imaging of dopamine transporters (DAT) in these models. HDT latency significantly increased in NHP-PD models. In addition, a significant inverse correlation between HDT and DAT was identified, but no local bias was found. The correlation with intention tremor symptoms was lower than the resting tremor. In conclusion, the evaluation of HDT may reflect behavioral symptoms of NHP-PD models. Furthermore, HDT was effectively used to experimentally distinguish intention tremors from other tremors.

neuroscience↗

bioRxiv2020

DNA micro-disk for the management of DNA-based data storage with index and write-once-read-many (WORM) memory features

DNA-based data storage has attracted attention because of its higher physical density of the data and longer retention time than those of conventional digital data storage1-7. However, previous DNA-based data storage lacked index features and the data quality of storage after a single access is not preserved, obstructing its industrial use. Here, we propose DNA micro-disks, quick response (QR)-coded micro-sized disks that harbour data-encoded DNA molecules for the efficient management of DNA-based data storage. We demonstrate the two major features that previous DNA-based data storage studies could not achieve. One feature is accessing data items efficiently by indexing the data-encoded DNA library. Another is achieving write-once-read-many (WORM) memory through the immobilization of DNA molecules on the disk and their enrichment through in situ DNA production. Through these features, the reliability of DNA-based data storage was increased by allowing multiple accession of data-encoded DNA without data loss.

bioengineering↗

bioRxiv2020

Haplotracker: a web application for simple and accurate mitochondrial haplogrouping using short DNA fragments

Mitochondrial DNA (mtDNA) haplogrouping is widely used in population genetics, forensics, and medical research to study evolutionary questions in human populations, examine degraded remains, and search for mtDNA-associated diseases. Next-generation sequencing methods have become a revolutionary tool for mtDNA analysis, particularly for degraded DNA, but they remain costly, laborious, and time-consuming. If an accurate and simple haplogroup-tracking tool were available, haplogrouping could be performed easily and rapidly. Here, we present Haplotracker, a web application for highly accurate and simple haplogroup tracking using a few sequence fragments. Haplotracker offers a unique user-friendly interface and estimates highly probable haplogroups from control-region sequences using a novel algorithm based on Phylotree Build 17 and our haplotype database (n=118,869). Haplotracker provides a simple, novel HG tracking solution, which was established through repeated blind simulation tests. It narrows down potential haplogroups and identifies their differential coding-region variants to confirm the haplogroups or to track sub-haplogroups. Haplotracker gives detailed information on sample variants, including their frequency in a large mtGenome database, which may give researchers an insight into common, rare, and potentially pathogenic mutations. It also offers a conserved region mapping tool for PCR primer design for successful tracking. Haplotracker produced high haplogroup prediction accuracy using 8,216 control-region Phylotree-provided sequences. It estimated top-ranking haplogroups with a higher concordance rate (56.6%, p<0.0001) than the similar tools MitoTool (29.4%) and HaploGrep 2 (33.9%). The significance persisted up to Rank 30. Haplotracker accurately estimated super-HGs from 94% of the control-region sequences at Rank 1. Further evaluation of the accuracy with 46,322 control-region sequences was significant. Laboratory application of Haplotracker to an ancient DNA extract demonstrated its practical usefulness. These results highlight the potential for the use of our web application as an alternative to full genome sequencing for easy haplogrouping, which may be useful in related fields. Free access: https://haplotracker.cau.ac.kr. Author summaryMitochondrial haplogroup (HG) classification is required in the search for answers to evolutionary questions about human populations, in the investigation of forensic samples, and in the search for disease-associated mutations. The sequencing of mitochondrial DNA (mtDNA) at the genome level is now frequently used due to the falling cost of next-generation sequencing. It offers accurate and detailed analysis of mtDNA variation, but it is still costly, laborious, and time-consuming. If an accurate and simple HG tracking tool were available, haplogrouping could be performed easily and rapidly. Here, we developed Haplotracker, a simple and accurate HG tracking web application with a novel algorithm and a novel tracking tool. The highly accurate prediction performance of Haplotracker was demonstrated in a series of tests. Using only control-region sequences, it accurately predicted HGs in more than half of the total Phylotree-provided mtGenome sequences and approximately 80% up to Rank 5; for super-HGs, it accurately predicted 94% at Rank 1 and more than 98% up to Rank 3. We demonstrated simple tracking for HG confirmation using Haplotracker. These results show that mtDNA haplogrouping with our web application may be useful in related fields.

bioinformatics↗

bioRxiv2020

The relative densities of cell cytoplasm, nucleoplasm, and nucleoli are robustly conserved during cell cycle and drug perturbations

The cell nucleus is a compartment in which essential processes such as gene transcription and DNA replication occur. While the large amount of chromatin confined in the finite nuclear space could install the picture of a particularly dense organelle surrounded by less dense cytoplasm, recent studies have begun to report the opposite. However, the generality of this newly emerging, opposite picture has so far not been tested. Here, we used combined optical diffraction tomography (ODT) and epi-fluorescence microscopy to systematically quantify the mass densities of cytoplasm, nucleoplasm, and nucleoli of human cell lines, challenged by various perturbations. We found that the nucleoplasm maintains a lower mass density than cytoplasm during cell cycle progression by scaling its volume to match the increase of dry mass during cell growth. At the same time, nucleoli exhibited a significantly higher mass density than the cytoplasm. Moreover, actin and microtubule depolymerization and changing chromatin condensation altered volume, shape, and dry mass of those compartments, while the relative distribution of mass densities was generally unchanged. Our findings suggest that the relative mass densities across membrane-bound and membraneless compartments are robustly conserved, likely by different as of yet unknown mechanisms, which hints at an underlying functional relevance. This surprising robustness of mass densities contributes to an increasing recognition of the importance of physico-chemical properties in determining cellular characteristics and compartments.

biophysics↗

bioRxiv2020

Acetylcholine is released in the basolateral amygdala in response to predictors of reward and enhances learning of cue-reward contingency

The basolateral amygdala (BLA) is critical for associating initially neutral cues with appetitive and aversive stimuli and receives dense neuromodulatory acetylcholine (ACh) projections. We measured BLA ACh signaling and principal neuron activity in mice during cue-reward learning using a fluorescent ACh sensor and calcium indicators. We found that ACh levels and activity of nucleus basalis of Meynert (NBM) cholinergic terminals in the BLA (NBM-BLA) increased sharply in response to reward-related events and shifted as mice learned the tone-reward contingency. BLA principal neuron activity followed reward retrieval and moved to the reward-predictive tone after task acquisition. Optical stimulation of cholinergic NBM-BLA terminal fibers during cue-reward learning led to more rapid learning of the cue-reward contingency. These results indicate that BLA ACh signaling carries important information about salient events in cue-reward learning and provides a framework for understanding how ACh signaling contributes to shaping BLA responses to emotional stimuli.

neuroscience↗

bioRxiv2020

Gait analysis of healthy rhesus monkeys using a pressure-sensing walkway system

Gait analysis in non-human primate models has been performed to elucidate the neural systems involved in controlling quadrupedal locomotor behavior. This study aimed to use a pressure-sensing walkway to identify characteristics of species-specific quadrupedal locomotion in rhesus monkeys. A total of nine healthy adult female rhesus monkeys (Macaca mulatta) were used for gait analysis. We measured the temporo-spatial and kinetic parameters of quadrupedal gait using a custom-built pressure-sensing walkway and compared the left- and right-side parameters to assess the symmetry of the gait pattern. All temporo-spatial and kinetic parameter values showed no significant differences among the nine rhesus monkeys for both the forelimbs and hindlimbs. However, significant differences were observed between forelimb and hindlimb kinetic parameters such as peak vertical force, vertical impulse, and the percentage of body weight distribution. All kinetic parameter values were higher for the forelimbs than for the hindlimbs. These data indicated that the center of gravity in healthy rhesus monkeys is located at the forelimbs rather than at the hindlimbs while walking. Furthermore, the symmetry indices considered for symmetric gait pattern showed a low variability. Most median symmetry index values were nearly zero, indicating no difference between the right and left sides. This study described valid methods for assessing gait parameters and demonstrated rhesus-specific characteristics of quadrupedal locomotion, providing a basis for the assessment of gait normality in rhesus monkeys.

animal behavior and cognition↗

bioRxiv2020

Improved Parameterization of Protein-DNA Interactions for Molecular Dynamics Simulations of PCNA Diffusion on DNA

Here, we quantitatively evaluate the accuracy of the protein-DNA interactions in AMBER and CHARMM force fields by comparing experimental and simulated diffusion coefficients of proliferating cell nuclear antigen (PCNA). We find that both force fields underestimate diffusion coefficients by at least an order of magnitude because the interactions between basic amino acids and DNA phosphate groups are too attractive. Then, we propose Lennard-Jones parameters optimized using the experimental osmotic pressure data of model chemicals, by using which one can reproduce the experimental diffusion coefficients. Newly optimized parameters will have a broad impact on general protein-DNA interactions.

biophysics↗

bioRxiv2020

Increases in ambient temperature progressively disassemble Arabidopsis phytochrome B from individual photobodies with distinct thermostabilities

Warm temperature is postulated to induce plant thermomorphogenesis through a signaling mechanism similar to shade, as both destabilize the active form of the photoreceptor and thermosensor phytochrome B (phyB). At the cellular level, shade antagonizes phyB signaling by triggering phyB disassembly from photobodies. Here we report temperature-dependent photobody localization of fluorescent protein-tagged phyB (phyB-FP) in the epidermal cells of Arabidopsis hypocotyl and cotyledon. Our results demonstrate that warm temperature operates through different photobody dynamics from shade. Increases in temperature from 12{degrees}C to 27{degrees}C incrementally reduce photobody number by stimulating phyB-FP disassembly from selective thermo-unstable photobodies. The thermostability of photobodies relies on phyBs photosensory module. Surprisingly, elevated temperatures inflict opposite effects on phyBs functions in the hypocotyl and cotyledon despite inducing similar photobody dynamics, indicative of tissue/organ-specific temperature signaling circuitry downstream of photobody dynamics. Our results thus provide direct cell biology evidence supporting an early temperature signaling mechanism via dynamic assembly/disassembly of individual photobodies possessing distinct thermostabilities.

plant biology↗

bioRxiv2020

Reliable data collection in participatory trials to assess digital healthcare apps.

The number of digital healthcare mobile apps on the market is increasing exponentially owing to the development of the mobile network and widespread usage of smartphones. However, only a few of these apps have undergone adequate validation. As with many mobile apps, healthcare apps are generally considered safe to use, making them easy for developers and end-users to exchange them in the marketplace. The existing platforms are not suitable to collect reliable data for evaluating the effectiveness of the apps. Moreover, these platforms only reflect the perspectives of developers and experts, not of end-users. For instance, data collection methods typical of clinical trials are not appropriate for participant-driven assessment of healthcare apps because of their complexity and high cost. Thus, we identified a need for a participant-driven data collection platform for end-users that is interpretable, systematic, and sustainable --as a first step to validate the effectiveness of the apps. To collect reliable data in the participatory trial format, we defined distinct stages for data preparation, storage, and sharing. Interpretable data preparation consists of a protocol database system and semantic feature retrieval method to create a protocol without professional knowledge. Collected data reliability weight calculation belongs to the systematic data storage stage. For sustainable data collection, we integrated the weight method and the future reward distribution function. We validated the methods through statistical tests conducted on 718 human participants. The validation results demonstrate that the methods have significant differences in the comparative experiment and prove that the choice of the right method is essential for reliable data collection. Furthermore, we created a web-based system for our pilot platform to collect reliable data in an integrated pipeline. We validate the platform features with existing clinical and pragmatic trial data collection platforms. In conclusion, we show that the method and platform support reliable data collection, forging a path to effectiveness validation of digital healthcare apps.

bioinformatics↗

bioRxiv2019

Chlorotoxin Conjugated with Saporin Reduces Viability of ML-1 Thyroid Cancer Cells In Vitro

BackgroundAlthough differentiated thyroid cancer has good prognosis, radioactive iodine (RAI) resistant thyroid cancer is difficult to treat. Current therapies for progressive RAI resistant thyroid cancer are not very effective. There is an unmet need for better therapeutic agents in this scenario. Studies have shown that aggressive thyroid cancers express matrix metalloproteinase -2 (MMP-2). Chlorotoxin is a selective MMP-2 agonist. Given that Saporin is a well-known ribosome-inactivating protein used for anti-cancer treatment, we hypothesized that Chlorotoxin-conjugated Saporin (CTX-SAP) would inhibit the growth of aggressive thyroid cancer cell lines expressing MMP-2. MethodsThe ML-1 thyroid cancer cell line was used for this study because it is known to express MMP-2. ML-1 cells were treated with a toxin consisting of biotinylated Chlorotoxin bonded with a secondary conjugate of Streptavidin-ZAP containing Saporin (CTX-SAP) from 0 to 600 nM for 72 hours. Then, cell viability was measured via XTT assay at an absorbance of A450-630. Control experiments were set up using Chlorotoxin and Saporin individually at the same varying concentrations. ResultsAfter 7 hours of incubation, there was a statistically significant reduction in cell viability with increasing concentrations of the CTX-SAP conjugate (F=4.286, p=0.0057). In particular, the cell viability of ML-1 cells was decreased by 49.77% with the treatment of 600 nM of CTX-SAP (F=44.24), and the reduction in cell viability was statistically significant (Dunnetts test p<0.0001). In contrast, individual Chlorotoxin or Saporin in increasing concentrations had no significant effect on cell viability using similar assay. ConclusionThis in vitro study demonstrated the efficacy of a CTX-SAP conjugate in reducing the viability of ML-1 thyroid cancer cells in a dose dependent manner. Further studies are needed to delineate the effectiveness of CTX-SAP in the treatment of aggressive thyroid cancer. Our study points towards MMP-2 as a potential target for RAI-resistant thyroid cancer.

cancer biology↗