Synthesis of a Doubly Labelled Concanamycin Derivative for ATPase Binding Studies

Abstract

The synthesis of a doubly labelled concanamycin derivative for binding studies with V-and P-type ATPases is described. The starting point was 21-deoxyconcanolide A (6), which was generated from concanamycin A (1) in three steps and which exhibited the full ATPase inhibitor activity, with the advantage of a stability better than that of 1. Through use of a suitable protecting group for 6, the carbene-generating diazirine residue and 125 I were introduced regio-and stereo-

Structure–Activity Relationships of Natural and Semisynthetic Plecomacrolides Suggest Distinct Pathways for HIV-1 Immune Evasion and Vacuolar ATPase-Dependent Lysosomal Acidification

“…Based on these results, we hypothesize that hydrophobic interactions mediated by these linear substituents may be important for target binding, especially because they parallel the effects of hydrophobic substituents previously reported to play a role in V-ATPase binding. 20 A key result of our study is that CMB (22), which bears a C-8 methyl compared to a C-8 ethyl in CMA (1), represents a second lead compound for preclinical development. Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio.…”

“…Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio. While we were initially disappointed that CMB (22) did not have a correspondingly improved therapeutic ratio, further semisynthetic modification of 22 to 3′,9-diacetyl CMB (31) succeeded in reducing toxicity compared to that of the parent compound CMB (22). These results indicate that continued modification along these lines is a promising avenue for drug development.…”

“…We also made progress toward improving the characteristics of these compounds by esterifying (individually or in tandem) the C-3′-OH and C-9-OH positions, which motivated the synthesis of acetyl and diacetyl derivatives 32/33, 34/35, and 36/37 (derived from CMA (1), CMC (23), and CMF (24), respectively). The most notable compound was 3′,9-diacetyl CMA (33, Figure 8B) as this molecule showed neutralization ratios that had only been observed with CMB (22) and its derivatives, while maintaining both potent Nef inhibition and a therapeutic ratio similar to CMA (1). While a significant improvement to the therapeutic ratio was not observed in our best compound (33; slightly less than 2-fold relative to CMA), the excellent potency and potential for low-dose treatment with 33 could enable the development of a viable therapeutic.…”

“…1 Hz, 3H), 1.05 (s, 3H), 1.04 (s, 3H), 0.89 (d, J = 6.5 Hz, 3H), 0.85 (t, J = 7.6 Hz, 3H), 0.82 (d, J = 6.8 Hz, 3H). 13 NMR Data of CMB* (22). 1 H NMR (600 MHz, 278 K, CDCl 3 ): δ 6.56 (dt, J = 14.9, 9.2 Hz, 1H), 6.36 Baf A 1 Production and Purification.…”

“…In addition to CMA (1), there are other naturally occurring concanamycin analogues that can be obtained by fermentation, 26 including concanamycin B (CMB, 22) and concanamycin C (CMC, 23). CMB (22) differs from CMA (1) in that CMB (22) has a methyl instead of an ethyl group at position C-8. CMC (23) differs from CMA (1) in that it lacks the 4′-carbamoyl group on the β-D-rhamnose (Figure 5A).…”

Structure–Activity Relationships of Natural and Semisynthetic Plecomacrolides Suggest Distinct Pathways for HIV-1 Immune Evasion and Vacuolar ATPase-Dependent Lysosomal Acidification

“…Based on these results, we hypothesize that hydrophobic interactions mediated by these linear substituents may be important for target binding, especially because they parallel the effects of hydrophobic substituents previously reported to play a role in V-ATPase binding. 20 A key result of our study is that CMB (22), which bears a C-8 methyl compared to a C-8 ethyl in CMA (1), represents a second lead compound for preclinical development. Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio.…”

“…Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio. While we were initially disappointed that CMB (22) did not have a correspondingly improved therapeutic ratio, further semisynthetic modification of 22 to 3′,9-diacetyl CMB (31) succeeded in reducing toxicity compared to that of the parent compound CMB (22). These results indicate that continued modification along these lines is a promising avenue for drug development.…”

“…We also made progress toward improving the characteristics of these compounds by esterifying (individually or in tandem) the C-3′-OH and C-9-OH positions, which motivated the synthesis of acetyl and diacetyl derivatives 32/33, 34/35, and 36/37 (derived from CMA (1), CMC (23), and CMF (24), respectively). The most notable compound was 3′,9-diacetyl CMA (33, Figure 8B) as this molecule showed neutralization ratios that had only been observed with CMB (22) and its derivatives, while maintaining both potent Nef inhibition and a therapeutic ratio similar to CMA (1). While a significant improvement to the therapeutic ratio was not observed in our best compound (33; slightly less than 2-fold relative to CMA), the excellent potency and potential for low-dose treatment with 33 could enable the development of a viable therapeutic.…”

“…1 Hz, 3H), 1.05 (s, 3H), 1.04 (s, 3H), 0.89 (d, J = 6.5 Hz, 3H), 0.85 (t, J = 7.6 Hz, 3H), 0.82 (d, J = 6.8 Hz, 3H). 13 NMR Data of CMB* (22). 1 H NMR (600 MHz, 278 K, CDCl 3 ): δ 6.56 (dt, J = 14.9, 9.2 Hz, 1H), 6.36 Baf A 1 Production and Purification.…”

“…In addition to CMA (1), there are other naturally occurring concanamycin analogues that can be obtained by fermentation, 26 including concanamycin B (CMB, 22) and concanamycin C (CMC, 23). CMB (22) differs from CMA (1) in that CMB (22) has a methyl instead of an ethyl group at position C-8. CMC (23) differs from CMA (1) in that it lacks the 4′-carbamoyl group on the β-D-rhamnose (Figure 5A).…”

Structure–Activity Relationships of Natural and Semisynthetic Plecomacrolides Suggest Distinct Pathways for HIV-1 Immune Evasion and Vacuolar ATPase-Dependent Lysosomal Acidification

“…Based on these results, we hypothesize that hydrophobic interactions mediated by these linear substituents may be important for target binding, especially because they parallel the effects of hydrophobic substituents previously reported to play a role in V-ATPase binding. 20 A key result of our study is that CMB (22), which bears a C-8 methyl compared to a C-8 ethyl in CMA (1), represents a second lead compound for preclinical development. Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio.…”

“…Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio. While we were initially disappointed that CMB (22) did not have a correspondingly improved therapeutic ratio, further semisynthetic modification of 22 to 3′,9-diacetyl CMB (31) succeeded in reducing toxicity compared to that of the parent compound CMB (22). These results indicate that continued modification along these lines is a promising avenue for drug development.…”

“…We also made progress toward improving the characteristics of these compounds by esterifying (individually or in tandem) the C-3′-OH and C-9-OH positions, which motivated the synthesis of acetyl and diacetyl derivatives 32/33, 34/35, and 36/37 (derived from CMA (1), CMC (23), and CMF (24), respectively). The most notable compound was 3′,9-diacetyl CMA (33, Figure 8B) as this molecule showed neutralization ratios that had only been observed with CMB (22) and its derivatives, while maintaining both potent Nef inhibition and a therapeutic ratio similar to CMA (1). While a significant improvement to the therapeutic ratio was not observed in our best compound (33; slightly less than 2-fold relative to CMA), the excellent potency and potential for low-dose treatment with 33 could enable the development of a viable therapeutic.…”

“…1 Hz, 3H), 1.05 (s, 3H), 1.04 (s, 3H), 0.89 (d, J = 6.5 Hz, 3H), 0.85 (t, J = 7.6 Hz, 3H), 0.82 (d, J = 6.8 Hz, 3H). 13 NMR Data of CMB* (22). 1 H NMR (600 MHz, 278 K, CDCl 3 ): δ 6.56 (dt, J = 14.9, 9.2 Hz, 1H), 6.36 Baf A 1 Production and Purification.…”

“…In addition to CMA (1), there are other naturally occurring concanamycin analogues that can be obtained by fermentation, 26 including concanamycin B (CMB, 22) and concanamycin C (CMC, 23). CMB (22) differs from CMA (1) in that CMB (22) has a methyl instead of an ethyl group at position C-8. CMC (23) differs from CMA (1) in that it lacks the 4′-carbamoyl group on the β-D-rhamnose (Figure 5A).…”

Structure–Activity Relationships of Natural and Semisynthetic Plecomacrolides Suggest Distinct Pathways for HIV-1 Immune Evasion and Vacuolar ATPase-Dependent Lysosomal Acidification

“…Based on these results, we hypothesize that hydrophobic interactions mediated by these linear substituents may be important for target binding, especially because they parallel the effects of hydrophobic substituents previously reported to play a role in V-ATPase binding. 20 A key result of our study is that CMB (22), which bears a C-8 methyl compared to a C-8 ethyl in CMA (1), represents a second lead compound for preclinical development. Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio.…”

“…Although CMB ( 22) has a 10-fold lower potency for Nef inhibition compared to that of CMA (1), it has an even lower potency for V-ATPase inhibition as assessed by Lysotracker fluorescence, resulting in an expanded neutralization ratio. While we were initially disappointed that CMB (22) did not have a correspondingly improved therapeutic ratio, further semisynthetic modification of 22 to 3′,9-diacetyl CMB (31) succeeded in reducing toxicity compared to that of the parent compound CMB (22). These results indicate that continued modification along these lines is a promising avenue for drug development.…”

“…We also made progress toward improving the characteristics of these compounds by esterifying (individually or in tandem) the C-3′-OH and C-9-OH positions, which motivated the synthesis of acetyl and diacetyl derivatives 32/33, 34/35, and 36/37 (derived from CMA (1), CMC (23), and CMF (24), respectively). The most notable compound was 3′,9-diacetyl CMA (33, Figure 8B) as this molecule showed neutralization ratios that had only been observed with CMB (22) and its derivatives, while maintaining both potent Nef inhibition and a therapeutic ratio similar to CMA (1). While a significant improvement to the therapeutic ratio was not observed in our best compound (33; slightly less than 2-fold relative to CMA), the excellent potency and potential for low-dose treatment with 33 could enable the development of a viable therapeutic.…”

“…1 Hz, 3H), 1.05 (s, 3H), 1.04 (s, 3H), 0.89 (d, J = 6.5 Hz, 3H), 0.85 (t, J = 7.6 Hz, 3H), 0.82 (d, J = 6.8 Hz, 3H). 13 NMR Data of CMB* (22). 1 H NMR (600 MHz, 278 K, CDCl 3 ): δ 6.56 (dt, J = 14.9, 9.2 Hz, 1H), 6.36 Baf A 1 Production and Purification.…”

“…In addition to CMA (1), there are other naturally occurring concanamycin analogues that can be obtained by fermentation, 26 including concanamycin B (CMB, 22) and concanamycin C (CMC, 23). CMB (22) differs from CMA (1) in that CMB (22) has a methyl instead of an ethyl group at position C-8. CMC (23) differs from CMA (1) in that it lacks the 4′-carbamoyl group on the β-D-rhamnose (Figure 5A).…”