1. Volume 2, Issue 6, Pages 1492-1497 (December 2012)
The Sirtuin 2 Inhibitor AK-7 Is Neuroprotective in Huntington’s Disease Mouse Models 
Vanita Chopra, Luisa Quinti, Jinho Kim, Lorraine Vollor, K. Lakshmi Narayanan, Christina Edgerly, Patricia M. Cipicchio, Molly A. Lauver, Soo Hyuk Choi, Richard B. Silverman, Robert J. Ferrante, Steven Hersch, Aleksey G. Kazantsev 
Cell Reports 
Volume 2, Issue 6, Pages (December 2012)
DOI: /j.celrep
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2. Cell Reports 2012 2, 1492-1497DOI: (10.1016/j.celrep.2012.11.001)
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3. Figure 1 Efficacy of AK-7 in R6/2 Mouse Model of HD
(A) AK-7 ameliorates motor deficits in HD mice. Latency to fall was increased by 37% at 11 weeks of age (vehicle treated mice = 81 s, AK-7 (10 mg/kg) treated mice = 129 s). Values represent geometric mean ± 95% confidence interval. n = 10–15; ∗ p ≤ 0.05; ∗∗, p ≤ 0.01.
(B and C) Kaplan–Meier probability of survival analysis (B) and the column graph (C) shows that AK-7 treatment significantly extends the survival of R6/2 mice by 13.2%; n = 10–11. Data were analyzed using Gehan–Wilcoxon test with Graph Pad prism (B) (p = ) and GLM procedure in SAS software (C) (p = ).
(D and E) Stereological quantification of total striatal volume and neuronal cell body volume. AK-7 treatment (20 mg/kg/12 h) improves the total striatal volume of HD mice by 9% (p = 0.044) and rescues the shrinkage of striatal neuronal cell body volume by 15% (p = 0.033). Data represent mean ± SEM, n = 8–10; ∗∗∗, p ≤ 0.001.
(F) Thionin-stained sections of neostriatum from a wild-type littermate mouse (right), a placebo-treated R6/2 mouse (center), and an AK-7-treated R6/2 mouse (left) at 12 weeks of age. There is evidence of neuronal atrophy in the untreated R6/2 mouse with relative preservation of neuronal atrophy in the AK-7-treated mouse. The arrowheads represent healthy neurons in wild-type, shrunken neurons in untreated R6/2 mice, and relatively preserved neurons in AK-7-treated R6/2 mice (scale bars, 25 μm).
See also Figures S1 and S2.
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4. Figure 2 Compound Treatment-Mediated Changes in R6/2 Mouse Brain
(A) AK-7 decreases the size of striatal neuronal intranuclear huntingtin aggregates. Immunodetection of intraneuronal huntingtin aggregates by using EM48 antibody (arrows) in the neostriatum of 12-week-old R6/2 mice with and without AK-7 treatment (scale bars, 25 μm).
(B) Stereological counts of striatal intraneuronal aggregates in R6/2 mice. AK-7 treatment significantly reduced (35%) the striatal nuclear aggregate volume in R6/2 mice.
(C and D) AK-7 treatment had no effect on mean aggregate count or levels of soluble mutant huntingtin exon 1 fragments, as detected by highly sensitive HTRF method. Error bars indicate SEM; ∗∗∗, p ≤ 0.001; n = 8–10.
See also Figures S1 and S2.
Cell Reports 2012 2, DOI: ( /j.celrep )
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5. Figure 3 Efficacy of AK-7 in 140 CAG Knockin Htt Mouse Model
(A and B) Behavioral test. Open field analysis of AK7 administration in 140 CAG mice shows a significant improvement at all the doses in (A) distance traveled and (B) resting time, with 20 mg/kg as the most efficacious dose. Symbols: filled circles, untreated wild-type; filled diamond, untreated CAG 140; filled squares, 10 mg/kg/12h; open triangles, 20 mg/kg/12h; and open circles, 30 mg/kg/12h.
(C and D) Aggregation. There was a significant reduction in huntingtin aggregates (untreated 140 CAG mice: 4.31 × 106 ± 1.07, AK-7 20 mg/kg-treated 140 CAG mice: 2.01 × 106 ± 0.63; p < 0.01). Error bars indicate SEM. Arrowheads represent intraneuronal huntingtin aggregates.
Cell Reports 2012 2, DOI: ( /j.celrep )
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6. Figure S1
Levels of SIRT2 in Brain Tissues of R6/2 and Knockin HD Mice Targeted by SIRT2 Inhibitor AK-7, Related to Figures 1, 2, and 3
(A) Levels of SIRT2 and GAPDH proteins in cortices of age-matched wild-type and R6/2 transgenic HD mice at 8 weeks of age as detected by Western blots.
(B) Levels of SIRT2 and GAPDH proteins in cortices of age-matched wild-type and CAG-140 knock-in HD mice at 18 months of age as detected by Western blots. Quantification of SIRT2 isoforms (not shown) shows no dramatic difference in SIRT2 expression between HD and wild-type animals with exception of SIRT2.1, in which levels were slightly reduced in R6/2 mice. There were no progressive disease changes detected in either SIRT2 isoform levels in R6/2 or 140CAG mice as they age. SIRT2.1 and SIRT2.2 isoforms, visualized as double bands by immunostaining, and SIRT2.3 isoform are marked -SIRT2.1, -SIRT2.2, and -SIRT2.3, respectively. The expression pattern of the SIRT2 isoforms in striata of wild-type, R6/2, and 140CAG knock-in mice was identical to that in cortices and therefore not shown.
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7. Figure S2
Synthesis, Purification, and Analysis of 3-(Azepan-1-Ylsulfonyl)-N-(3-Bromophenyl) Benzamide, Compound AK-7, Related to Figures 1, 2, and 3
(A) To a solution of 3-(chlorosulfonyl)benzoic acid (1, 1.0 equiv, ca. 0.1 M) in dichloromethane, was slowly added hexamethyleneimine (3.1 equiv) at 0°C with stirring. The mixture was stirred at room temp for 12 hr. The solvent was removed by a stream of nitrogen gas, and 1 M aqueous NaOH (>10 equiv) was added to the mixture, which was extracted with ether three times. The aqueous fraction was then acidified with 3 M aqueous HCl to ∼pH 1. The mixture turned cloudy during the addition. The resulting precipitate was collected by suction filtration on a Buchner funnel, washed with distilled water, and dried overnight at reduced pressure to give 3-(azepan-1-ylsulfonyl)benzoic acid (2, 50%–60%), which was used without further purification. To a solution of 2 (1.0 equiv, ca. 0.1 M), 3-bromoaniline (1.1 equiv), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI, 1.3 equiv), and 4-dimethylaminopyridine (DMAP, 0.1 equiv) were added. The mixture was stirred at room temp for 12 hr, and then diluted with excess EtOAc. The organic mixture was washed with 1 M aqueous HCl (2 times), 1 M aqueous NaHCO3 (2 times), and brine. The organic layer was dried over MgSO4, filtered, and concentrated in vacuo to yield a crude product, which was purified by silica gel column chromatography to give a white solid (AK-7, 60%–70%); mp 157°C.
(B) 1H NMR (500 MHz, CDCl3) δ 8.22 (t, J = 1.6 Hz, 1H), (m, 2H), 7.97 (t, J = 1.8 Hz, 1H), 7.95 (dt, J = 7.7, 1.3 Hz, 1H), 7.66 (t, J = 7.8 Hz, 1H), 7.58 (d, J = 8.1 Hz, 1H), 7.32 (d, J = 8.1 Hz, 1H), 7.26 (t, J = 8.0 Hz, 1H), 3.30 (t, J = 5.9 Hz, 4H), 1.72 (br s, 4H), (m, 4H).
(C) 13C NMR (125.8 MHz, CDCl3) δ , , , , , , , , , , , , , 48.34, 29.11,
(D) HRMS (ESI) m/z [M+H]+ calcd for C19H22BrN2O3S , , found , Anal. (C19H22BrN2O3S) calcd C 52.18, H 4.84, N 6.41, found C 52.28, H 4.92, N 6.33.
Cell Reports 2012 2, DOI: ( /j.celrep )
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