Journal: Genes to cells : devoted to molecular & cellular mechanisms

Article Title: Identification and characterization of retinoic acid-responsive genes in mouse kidney development.

doi: 10.1111/gtc.12163

Figure Lengend Snippet: Figure 2 Pan-RAR antagonist BMS493 inhibits the ureteric bud branching morphogenesis and suppresses the expression of most of the candidates of retinoic acid (RA)-responsive genes. (A) The kidneys cultured for 48 h with 0 (DMSO), 1, 2 and 5 lM BMS493 were immunostained by anti-Six2 (red) and anti-pan cytokeratin (green). Scale bar represents 100 lm. (B) Quantification of the number of branches in (A). Values are mean SD (n ≥5). **P < 0.01, unpaired t-test. (C) The expression levels of 33 candidate genes in 0 (DMSO), 1, 2 and 5 lM BMS493 were determined by qPCR. Values are mean SD (n = 3).

Article Snippet: Kidneys were then treated with DMSO or indicated dose of each inhibitor, BMS493 (TOCRIS), BAY 11-7082 (Merck) or IKK-2 inhibitor IV (Santa Cruz) for 48 h. DMEM was supplemented 1% penicillin and streptomycin.

Techniques: Expressing, Cell Culture

Journal: Birth defects research

Article Title: Methoxyacetic acid inhibits histone deacetylase and impairs axial elongation morphogenesis of mouse gastruloids in a retinoic acid signaling-dependent manner

doi: 10.1002/bdr2.1712

Figure Lengend Snippet: Chemicals used in the present study

Article Snippet: Chemicals All chemical compounds used in the present study were commercially obtained, namely from Sigma-Aldrich (St. Louis, MO), Santa Cruz Biotechnology (Dallas, TX), and Calbiochem (San Diego, CA), and their details are described in . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Chemical name CASRN Vendor (catalog number) Stock concentration (solvent) Methoxyacetic acid 625-45-6 Sigma-Aldrich (194557) 1 M (water) Methoxyethanol 109-86-4 Sigma-Aldrich (284467) 12.7 M (undiluted) Retinoic acid 302-79-4 Sigma-Aldrich (R2625) 100 μM (DMSO) Trichostatin A 58880-19-6 Calbiochem (647925) 100 μM (DMSO) Valproic acid 1069-66-5 Sigma-Aldrich (P4543) 100 mM (water) BMS493 215030-90-3 Santa Cruz (sc-361124) 10 mM (DMSO) Open in a separate window Abbreviations: CASRN, chemical abstracts service registry number; DMSO, dimethyl sulfoxide.

Techniques: Concentration Assay, Solvent

Journal: Birth defects research

Article Title: Methoxyacetic acid inhibits histone deacetylase and impairs axial elongation morphogenesis of mouse gastruloids in a retinoic acid signaling-dependent manner

doi: 10.1002/bdr2.1712

Figure Lengend Snippet: Methoxyacetic acid (MAA) and inhibitors of histone deacetylase up-regulate Cdx1 and Hoxa1 expressions in a retinoic acid (RA) signaling-dependent manner. Vertical axis represents relative gene expression levels in monolayer culture, as measured by quantitative RT-PCR. Graphs show the relative expression levels (mean ± SD; n = 3). The control level (C) is set as one. (a) Expression levels after treatment with RA, MAA, trichostatin A (TSA), or valproic acid (VPA) for 1 day. Asterisks indicate a significant increase by a given treatment compared with the control (p < .01). (b) Expression levels after co-treatment with BMS493 (BMS) plus MAA or TSA for 1 day. Asterisks indicate a significant difference between the two groups specified by each horizontal line (p < .01)

Article Snippet: Chemicals All chemical compounds used in the present study were commercially obtained, namely from Sigma-Aldrich (St. Louis, MO), Santa Cruz Biotechnology (Dallas, TX), and Calbiochem (San Diego, CA), and their details are described in . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Chemical name CASRN Vendor (catalog number) Stock concentration (solvent) Methoxyacetic acid 625-45-6 Sigma-Aldrich (194557) 1 M (water) Methoxyethanol 109-86-4 Sigma-Aldrich (284467) 12.7 M (undiluted) Retinoic acid 302-79-4 Sigma-Aldrich (R2625) 100 μM (DMSO) Trichostatin A 58880-19-6 Calbiochem (647925) 100 μM (DMSO) Valproic acid 1069-66-5 Sigma-Aldrich (P4543) 100 mM (water) BMS493 215030-90-3 Santa Cruz (sc-361124) 10 mM (DMSO) Open in a separate window Abbreviations: CASRN, chemical abstracts service registry number; DMSO, dimethyl sulfoxide.

Techniques: Histone Deacetylase Assay, Gene Expression, Quantitative RT-PCR, Expressing, Control

Journal: Birth defects research

Article Title: Methoxyacetic acid inhibits histone deacetylase and impairs axial elongation morphogenesis of mouse gastruloids in a retinoic acid signaling-dependent manner

doi: 10.1002/bdr2.1712

Figure Lengend Snippet: The morphogenetic effect of methoxyacetic acid (MAA) is partly alleviated by the suppression of retinoic acid signaling. (a) A regimen for gastruloid treatment with MAA (4 mM) and BMS493 (BMS, 10 nM). (b) Images of Day 4 gastruloids. Scale bar = 500 μm. (c) Morphometric parameters of treated gastruloids. Graphs show the averages of relative area (left) and relative aspect ratio (AR; right) with error bars of 95% confidence interval (n = 48). Asterisks indicate a significant difference (p < .01) between the MAA and MAA + BMS groups

Article Snippet: Chemicals All chemical compounds used in the present study were commercially obtained, namely from Sigma-Aldrich (St. Louis, MO), Santa Cruz Biotechnology (Dallas, TX), and Calbiochem (San Diego, CA), and their details are described in . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Chemical name CASRN Vendor (catalog number) Stock concentration (solvent) Methoxyacetic acid 625-45-6 Sigma-Aldrich (194557) 1 M (water) Methoxyethanol 109-86-4 Sigma-Aldrich (284467) 12.7 M (undiluted) Retinoic acid 302-79-4 Sigma-Aldrich (R2625) 100 μM (DMSO) Trichostatin A 58880-19-6 Calbiochem (647925) 100 μM (DMSO) Valproic acid 1069-66-5 Sigma-Aldrich (P4543) 100 mM (water) BMS493 215030-90-3 Santa Cruz (sc-361124) 10 mM (DMSO) Open in a separate window Abbreviations: CASRN, chemical abstracts service registry number; DMSO, dimethyl sulfoxide.

Techniques:

Journal: Nature Communications

Article Title: Maternal iron deficiency perturbs embryonic cardiovascular development in mice

doi: 10.1038/s41467-021-23660-5

Figure Lengend Snippet: a Phenotypic rescue. Pregnant mice were returned to iron-replete diet between E7.5-E9.5; fed continuously on an iron and vitamin A deficient diet (ID/VAD); or fed continuously on an iron deficient diet and treated with 0.7 mg/ml BMS493 on E8.5 and E9.5 (ID/BMS). Histograms showing the percentage of embryos that were dead, abnormal or normal at E15.5. b Investigation of gene–environment interaction. Iron deficient (ID) and control (C) C57BL/6 J females were crossed with males carrying the Tbx1 null allele (T∆), or the Dp1Tyb duplication (DS+). Histograms showing the percentage of embryos that were dead, abnormal or normal at E15.5. c – f Representative images of Dp1Tyb + control ( c ), wild type control ID ( d ), Dp1Tyb + ID with holoprosencephaly ( e ), Dp1Tyb + control ( f ), E15.5 embryos. g – j Dp1Tyb + embryos from ID mothers have craniofacial defects. Representative 3D reconstructed frontal sections of HREM data from wild type ID ( g , i ) and Dp1Tyb + ID ( h , j ) E15.5 embryos showing examples of failed secondary palate fusion ( h , yellow arrow) and holoprosencephaly ( j ). g’ – j’ Sagittal sections from the same embryos showing the location of the sections in ( g – j ) (red lines). note that panels ( g ) and ( i ) are the same control embryo. T tongue. Statistical significance was tested using one-sided Fisher’s exact test (panel c ). P values: 0.0657 (ID vs E9.5); 0.1127 (ID vs E8.5); <0.0001 (ID vs E7.5); 0.0019 (ID vs ID VAD); 0.0009 (ID vs ID BMS); < 0.0001 (C vs E9.5); <0.0001 (C vs E8.5); 0.2257 (C vs E7.5); 0.0087 (C vs ID VAD); >0.9999 (C vs ID BMS); 0.4859 (DS + ID vs WT ID); <0.0001 (DS + ID vs DS + C); 0.0321 (DS + ID vs ID); 0.2494 (WT ID vs ID); (0.0006 DS + C vs ID); 0.4742 (T∆ ID vs WT ID); <0.0001 (T∆ ID vs T∆ C); 0.1000 (T∆ ID vs ID); 0.0457 (WT ID vs ID); <0.0001 (T∆ C vs ID). ns not significant, * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.Scale bar = 2 mm ( c – f ); 700 µm ( g , h ); 1.2 mm ( I , j ); 1.6 mm ( g’ – I’ ).

Article Snippet: BMS493 (Merck) was dissolved at 10 mM in ethanol, then diluted in corn oil (Merck) and 0.7 or 3.5 mg/kg administered by oral gavage in the morning of E8.5 and E9.5.

Techniques:

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a) Schematic modeling of the RA signaling pathway, displaying known potentiators (orange) and attenuators (blue) of RA signaling. (b) Comparison of the gene-expression levels of known mediators of RA signaling between myoepithelial-like (CD49f high /KIT neg ; red) and ductal-like (CD49f low /KIT + ; green) cells isolated from bi-phenotypic ACC models. Genes identified as attenuators of RA signaling displayed preferential expression in CD49f high /KIT neg cells, while genes identified as potentiators of RA signaling displayed preferential expression in CD49f low /KIT + cells, as revealed by RNA-seq. Error bars: mean +/- standard deviation. Statistical test: Student’s t-test for paired samples (• p<0.01, *p<0.05, **p<0.01). (c) Heatmap of the mean z-score values for the expression level of genes identified as modulators of RA, as measured across the two ACC populations. (d-g) Establishment and phenotypic characterization of three-dimensional (3D) organoid cultures from human ACCs. Organoids grow as spherical structures (d) , retain cribriform histology (e) and contain both myoepithelial-like and ductal-like populations, as visualized by IHC staining for TP63 (f) and KIT (g) . (h) Analysis by flow cytometry of ACC organoids treated for one week with either DMSO (n=13), ATRA (10 µM; n=10), Bexarotene (10 µM; n=6), BMS493 (10 µM; n=10) or AGN193109 (10 µM; n=6). (i-j) Comparison of the percentage of CD49f high /KIT neg (i) and CD49f low /KIT + (j) cells found in organoids following in vitro treatment with modulators of RA signaling. Treatment with agonists of RA signaling was associated with an increase in the relative percentage of CD49f low /KIT + cells, while treatment with inhibitors of RA signaling was associated with its reduction as compared to DMSO-treated controls. Statistical test: one-way ANOVA with Dunnett’s multiple comparisons test (*p<0.05, **p<0.01, ***p<0.001). (k-p) Representative flow plots and quantification of the relative frequency of myoepithelial-like (CD49f high /KIT neg ) and ductal-like (CD49f low /KIT + ) cells in ACC organoids established from SGTX6 (k-m) or ACCX6 (n-p) PDX lines, treated for one week with either DMSO, ATRA (10 µM) or BMS493 (10 µM), confirming results previously obtained in the ACCX5M1 PDX line. Data correspond to the results of at least two independent experiments (with a minimum of 3 replicates for each treatment condition).

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: Expressing, Isolation, RNA Sequencing Assay, Standard Deviation, Immunohistochemistry, Flow Cytometry, In Vitro

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a-b) Treatment with all-trans retinoic acid (ATRA), an agonist of RA signaling, induces a dose-dependent increase in the percentage of CD49f low /KIT + cells in 3D organoid cultures established from the ACCX5M1 PDX line. The increase in the percentage of CD49f low /KIT + cells is already observed at the 0.1-1 µM concentration range (a), and becomes progressively more prominent as the concentration is raised to the 10-100 µM concentration range (b). (c-d) Treatment with either BMS493 or AGN19310, two inverse agonists (i.e., inhibitors) of RA signaling, results in a profound reduction in the percentage of CD49f low /KIT + cells in 3D organoid cultures established from either the ACCX5M1 (c; BMS43) or SGTX6 (d; AGN193109) PDX lines, even when administered at low doses (1 µM). Organoid cultures were treated for one week and analyzed by flow cytometry (n=3 wells/condition). Changes in the percentage of CD49f low /KIT + cells were tested for statistical significance using one-way ANOVA with Dunnett’s multiple comparisons test against untreated (NT) or DMSO-treated controls (ns = non-significant, *p<0.05, **p<0.01, ***p<0.001).

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: Concentration Assay, Flow Cytometry

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a-l) Organoids established from the ACCX6 PDX line were treated with DMSO, ATRA (10 µM) or BMS493 (10 µM) and subsequently analyzed by IHC. Treatment with ATRA resulted in an expansion of KIT + cells (g) , while treatment with BMS493 resulted in a complete loss of KIT expression (k) , associated with a dramatic change in the organoids’ morphology, characterized by the appearance of amorphous, eosin-rich deposits at their center (i) . Neither ATRA or BMS493 appeared to upregulate MKI67 expression in either cell population (d, h, l). Scale bars = 100 µm. (m-n) Schematic workflow of prospective experiments aimed at testing the effects of RA signaling on purified populations of myoepithelial-like and ductal-like cells: CD49f high /KIT neg and CD49f low /KIT + cells were sorted in parallel from the same ACCX5M1 tumors and cultured for one week as 2D monolayers in the presence of either ATRA (10 µM) or BMS493 (10 µM). (o) Treatment with ATRA did not reduce the viability of either CD49f high /KIT neg or CD49f low /KIT + cells (viability assay: alamarBlue; technical replicates: n=3; error bars: mean +/- standard deviation; statistical test: Student’s t-test, two-tailed; ns = non-significant, *p<0.05, **p<0.01, ***p<0.001). (p) Treatment with ATRA caused CD49f high /KIT neg cells to change phenotype and become undistinguishable from CD49f low /KIT + cells by flow cytometry. (q) Schematic modeling of the effects of RA agonism on the cell composition of ACC organoids: myoepithelial-like cells are stimulated to differentiate into ductal-like cells. (r) Treatment with BMS493 had a minor effect on the viability of CD49f high /KIT neg cells, but caused the death of the majority CD49f low /KIT + cells. (s) Upon visual inspection by conventional microscopy, CD49f low /KIT + cells treated with BMS493 appeared fragmented. Scale bar = 100 µm. (t) Schematic modeling of the effects of RA inhibition on the cell composition of ACC organoids: ductal-like cells are selectively killed.

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: Expressing, Purification, Cell Culture, Viability Assay, Standard Deviation, Two Tailed Test, Flow Cytometry, Microscopy, Inhibition

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a-o) Analysis of organoid morphology and histology, following treatment with either activators (ATRA; direct agonist) or inhibitors (BMS493; inverse agonist) of RA signaling. Organoids were established from a PDX line with bi-phenotypic histology (ACCX5M1) and treated for one week with either DMSO (a-e), 10 µM ATRA (f-j) or 10 µM BMS493 (k-o). Scale bars = 100 µm. Treatment with ATRA did not change organoid morphology (f), but increased the number of KIT + cells (i), as visualized by immunohistochemistry (IHC). Treatment with BMS493 caused a dramatic change in organoid morphology, characterized by the appearance of dense areas in the organoid centers, when observed using bright-field microscopy (k, arrowheads). When organoids were stained with hematoxylin and eosin (H&E), these areas consisted of an eosin-rich material, with apoptotic nuclei (l, arrowheads). (p-q) Analysis by flow cytometry of cell cycle distribution in organoids established from ACCX5M1 (p) and ACCX6 (q) PDX lines, following 1 week of treatment with either DMSO, ATRA (10 µM) or BMS493 (10µM). (r-s) Treatment with either ATRA or BMS493 did not increase the percentage of cells in the G2/M phase of the cell-cycle in ACCX5M1 (r) and ACCX6 (s) organoids. Experiments included at least three replicates (n=3 wells/condition). Error bars: mean +/- standard deviation. Differences in the percentage of cells in the G2/M phase were tested for statistical significance using one-way ANOVA with Dunnett’s multiple comparisons test (ns=non-significant, *p<0.05).

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: Immunohistochemistry, Microscopy, Staining, Flow Cytometry, Standard Deviation

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a-b) Analysis by flow cytometry of two PDX lines representative of human ACCs with solid histology (ACCX9, ACCX11) revealing a ductal-like, mono-phenotypic (CD49f low /KIT + ) cell composition. (c-f) IHC analysis of KIT and TP63 expression ACCX9 and ACCX11, showing ubiquitous expression of the ductal-specific marker KIT (c,e) and loss of the myoepithelial-specific marker TP63 (d,f) . Scale bars = 50 µm. (g) Principal component analysis (PCA) of RNA-seq data from human ACCs, in which results from two PDX lines with solid histology (ACCX9, ACCX11) are combined with 5 autologous pairs of CD49f high /KIT neg and CD49f low /KIT + cells populations from bi-phenotypic PDX lines (ACCX5M1, ACCX6, ACCX14, ACCX22, SGTX6). (h) Hierarchical clustering of RNA-sequencing data from human ACCs, based on the expression levels of the top 100 genes identified as differentially expressed between CD49f high /KIT neg and CD49f low /KIT + cells. Solid ACCs cluster with CD49f low /KIT + cells from bi-phenotypic tumors, irrespective of the method used to analyze their transcriptional profile. (i-j) Upon visual inspection by conventional microscopy, ACCX9 organoids cultured for one week in the presence of BMS493 (10 µM) display widespread cell fragmentation, in contrast to organoids cultured with DMSO alone. (k) Quantification of organoid viability using the alamarBlue HS assay, confirming the cytotoxic activity of BMS493 (10 µM) against PDX lines with solid histology (ACCX9, ACCX11). Error bars: mean +/- standard deviation; p-values: two-tailed Student’s t-tests; n=3 replicates/condition.

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: Flow Cytometry, Expressing, Marker, RNA Sequencing Assay, Microscopy, Cell Culture, Activity Assay, Standard Deviation, Two Tailed Test

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a) Individual growth curves of ACCX9 tumors treated with DMSO (gray) or BMS493 (magenta). Cross symbols (†) denote two animals who were sacrificed early, due to deterioration of their health condition. (b) Growth rates of ACCX9 tumors treated with either DMSO (n=6) or BMS493 (n=7). Two treatment cohorts are identified by different symbols (circles = cohort 1; triangles = cohort 2). Differences in mean growth rates were statistically significant (Welch’s t-test; **p<0.01). Black symbols denote the two animals who were sacrificed because of a deterioration of their health condition. (c) Individual animal weights over the course of in vivo treatment with BMS493 (†: animals sacrificed due to deterioration of health conditions). (d) Individual growth curves of ACCX11 tumors treated with DMSO (gray) or BMS493 (magenta). (e) Growth rates of ACCX11 tumors treated with either DMSO (n=4) or BMS493 (n=5). Differences in mean growth rates were statistically significant (Welch’s t-test; **p<0.01). (f) Individual animal weights over the course of in vivo treatment with BMS493. (g) Individual growth curves of ACCX5M1 tumors treated with DMSO (gray) or BMS493 (magenta). Cross symbols (†) denote two animals who either were sacrificed early due to deterioration of their general health or who died at the final time point. (h) Growth rates of ACCX5M1 tumors treated with either DMSO (n=6) or BMS493 (n=6). Two treatment cohorts are identified by different symbols (circles = cohort 1; triangles = cohort 2). Differences in mean growth rates were statistically significant (Welch’s t-test; *p<0.05). Black symbols denote the two animals who either were sacrificed early due to deterioration of their general health or died at the final time point. (i) Individual animal weights over the course of treatment (†: animals sacrificed or found dead).

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: In Vivo

Journal: bioRxiv

Article Title: Phenotypic dissection of epithelial lineages and therapeutic manipulation of differentiation programs in human Adenoid Cystic Carcinomas (ACCs)

doi: 10.1101/2022.01.19.476843

Figure Lengend Snippet: (a ) Schematic of the BMS493 dosing regimen utilized for the in vivo treatment of solid ACC models (40 mg/kg doses, i.p., 3 times/week x 3 weeks). Schematic was created with BioRender.com. (b-e) Comparison of tumor growth kinetics, expressed as either fold-increases in mean tumor volumes or growth rates, between mice treated with BMS493 and mice treated with the drug’s vehicle alone (DMSO), following subcutaneous engraftment with solid ACC models (ACCX9: b-c ; ACCX11: d-e ). (f) Schematic of the BMS493 dosing regimen utilized for the in vivo treatment of a bi-phenotypic ACC model (40 mg/kg doses, i.p., 4 times/week x 3 weeks). (g-h) Comparison of tumor growth kinetics, expressed as either fold-increases in tumor volumes or growth rates, between mice treated with BMS493 and mice treated with the drug’s vehicle alone (DMSO), following subcutaneous engraftment with a bi-phenotypic ACC model (ACCX5M1). Differences between mean fold-increases in tumor volume were tested for statistical significance using a two-tailed Student’s t-test (ns = non-significant, *p<0.05, **p<0.01, ***p<0.001). Differences between mean growth rates were tested for statistical significance using a two-tailed Welch’s t-test. Growth rates were calculated assuming exponential kinetics. Error bars: mean +/- standard deviation.

Article Snippet: On the day of in vivo administration, single-use aliquots were thawed, and BMS493 was further diluted to a concentration of 2 mg/mL in DPBS supplemented with 0.15M hydroxypropyl β-cyclodextrin (HP-β-CD; Cayman Chemicals 16169), for a total volume of 0.5 mL per dose (1 mg/dose).

Techniques: In Vivo, Two Tailed Test, Standard Deviation