Litterature Meeting Enantioselective Total Synthesis of Avrainvillamide and Stephacidins A and B Aspergillus ochraceus

Aspergillus: A source of complexe prenylated indole alkaloids - Isolation from a fungal species found in an Indian clay sample (Sirsaganj, Uttar Pradesh, India) - Sources: 1/ Marine fungal strain Aspergillus: Fenical and coworkers 2/ Fermentation broth of Aspergillus ochraceus: 2001 – Sugie and coworkers - Isolation from Aspergillus ochraceus WC76466: 2002 – Bristol Myers Squibb -In vitro citotoxic activity (human tumor cell lines) ⇒ SPC B: 5-30 fold more active than SPC A (testosterone-dependent prostate LNCaP cell line: IC 50 =0.06 µM)

Biosynthesis of Stephacidin B: a lesson for the chemist [O] Prenylation Reverse Prenylation 2 [O] [O] 2 [O] Diels-Alder * bicyclo[2.2.2]diazaoctane * Birch and coworkers, J. Chem. Soc. Perkin I, 1974, 50. Sammes and coworkers Chem. Comm., 1970,

Presumed biosynthesis of Stephacidins A and B [O] Prenylation Intramolecular Diels-Alder [O]

Synthesis of Stephacidin A: formation of the bicyclo[2.2.2]diazaoctane nucleus Williams’ approaches NaH SN 2 ’ Diels-Alder J. Am. Chem. Soc. 1990, 112, 808. Acc. Chem. Res. 2003, 36, 127. Tetrahedron Lett. 2004, 45, 4489.

Synthesis of the bicyclo[2.2.2]diazaoctane by SN 2 ’ approach Seebach and coworkers, J. Am. Chem. Soc. 1983, 105, Somei and coworkers, Heterocycles 1981, 16, 941.

Solvent Temperature (°C) Base Ratio anti:syn Yield (%) Benzene80NaH3:9782 DMF85NaH2:163 Benzene25NaH/18-crown-66:114 Benzene80NaH/18-crown-63.9:156 Synthesis of the bicyclo[2.2.2]diazaoctane by SN 2 ’ approach (2) Brevianamide B

Synthesis of the bicyclo[2.2.2]diazaoctane by SN 2 ’ approach (2) Tight ion pair

Synthesis of the bicyclo[2.2.2]diazaoctane by Diels-Alder approach 2.5:1

Synthesis of the bicyclo[2.2.2]diazaoctane by Diels-Alder Approach 90 % Williams et al. Bioorg. Med. Chem., 1998, 6, S R

Synthesis of the bicyclo[2.2.2]diazaoctane by Diels-Alder Approach R R "EXO""ENDO" S S 90 %

William’s synthesis of bicyclo[2.2.2]diazaoctane nucleus NaH SN 2 ’ Diels-Alder  16 steps in 12 % yield overall  High stereoselectivity of alkylation based on the presence or absence of metal chelation  4 steps in 17 % yield overall from and  Medium stereoselectivity of cycloaddition based on steric effects

Synthesis of Stephacidin A: formation of the bicyclo[2.2.2]diazaoctane nucleus Liebscher’ approach Diels-Alder AcCl Based on intermolecular Diels-Alder model reactions ⇒ acidic conditions such as HCl and BF 3.OEt 2 not as effective as AcCl or HCO 2 H ⇒ high pression and temperature ⇒ slow rates (6-20 days) + + major Liebscher and coll. J. Org. Chem. 2001, 66, 3984.

Synthesis of Stephacidin A: formation of the bicyclo[2.2.2]diazaoctane nucleus Liebscher’ approach (2) + Lieberknecht and coll. Tetrahedron Lett. 1987, 28, Williams and coll. Tetrahedron Lett. 2005, 46, Z-Admpa

Synthesis of Stephacidin A: formation of the bicyclo[2.2.2]diazaoctane nucleus tBuOK 78 % Diels-Alder AcCl 48 % rt, 20 days one stereoisomer ! Liebscher’ approach (3) R minimal steric repulsion defavoring steric repulsion R S

Liebscher’s synthesis of bicyclo[2.2.2]diazaoctane nucleus Diels-Alder AcCl  2 steps in 37 % yield overall from and  Stereospecificity of cycloaddition based on steric effects due to presence of acetoxy group BUT Cycloaddition step achieved in 20 days and in only 48 % yield !!

Synthesis of Stephacidin A: formation of the bicyclo[2.2.2]diazaoctane nucleus Myers’ approach Acyl radical approach Abrams and coll. Tetrahedron 1991, 47, J. Am. Chem. Soc. 2005, 127, 5342.

Formation of the bicyclo[2.2.2]diazaoctane nucleus: Myers’ approach Corey and coworkers, Tetrahedron Lett. 1991, 32, Corey E. J., Bakshi R. K., Shibata S. J. Am. Chem. Soc. 1987, 109, 5551.

Formation of the bicyclo[2.2.2]diazaoctane nucleus: Myers’ approach S R

Ghaffar T., Parkins A. W. J. Mol. Cat. A 2000, 160, 249. Formation of the bicyclo[2.2.2]diazaoctane nucleus: Myers’ approach X 7-membered ring ! S = H 2 O

Formation of the bicyclo[2.2.2]diazaoctane nucleus: Myers’ approach Jackson L. V., Walton J. C. Chem. Commun. 2000, 2327.

Formation of the bicyclo[2.2.2]diazaoctane nucleus: Myers’ approach BUT 62 %

Myers’ synthesis of bicyclo[2.2.2]diazaoctane nucleus Enantioselective synthesis of the desired nucleus 12 steps in 19 % yield overall from and Product used as precursor for synthesis of Stephacidin B

Synthesis of Stephacidin A: formation of the bicyclo[2.2.2]diazaoctane nucleus Baran’ s approach Three steps: 1/ Synthesis of a model of the bicyclo[2.2.2]diazaoctane nucleus 2/ Application of the strategy to a functionalized system for eventual elaboration into Stephacidin A 3/ Formation of Stephacidin A J. Am. Chem. Soc. 2006, 128, 8678.

- Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus - Intramolecular Diels-Alder Intramolecular vinyl radical cyclisation Intramolecular oxidative enolate heterocoupling

- Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus - First strategy: Ring closure by intramolecular Diels-Alder reaction Dehydrogenation Peptide coupling N-Boc-L-Trp Dehydrogenation

First strategy: Ring closure by intramolecular Diels-Alder reaction (2) Dehydrogenation: - Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus - Yamamoto and coll. J. Am. Chem. Soc. 2004, 126, % ⇒ Study of direct dehydrogenation of simplified Trp derivatives

First strategy: Ring closure by intramolecular Diels-Alder reaction (3) X X - Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus -

Second strategy: Ring closure by intramolecular vinyl radical cyclization X - Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus -

Third strategy: Ring closure by intramolecular oxidative enolate coupling Intramolecular Oxidative Coupling - Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus - Baran and coll. Angew. Chem. Int. Ed. 2005, 44, 609.

Third strategy: Ring closure by intramolecular oxidative enolate coupling Diastereoselectivity Mechanism ? - Baran’s Synthesis of Stephacidin A – - First step: Preparation of a model of the bicyclo[2.2.2]diazaoctane nucleus R6R 7

- Baran’s Synthesis of Stephacidin A – - Second step: Application to the elaboration of a suitable functionalized system - Benzopyran Tryptophan Synthesis: Amide bond formation Reider and coll. J. Org. Chem. 1997, 62, 2676.

- Baran’s Synthesis of Stephacidin A – - Third step: Final formation of Stephacidin A - Benzopyran Tryptophan Synthesis (2): Proline Synthesis:

- Baran’s Synthesis of Stephacidin A – - Third step: Final formation of Stephacidin A - Union of Tryptophan and Proline Fragments Ohfune and coll. J. Org. Chem. 1990, 55, 870.

- Baran’s Synthesis of Stephacidin A – - Third step: Final formation of Stephacidin A - Union of Tryptophan and Proline Fragments (2) Yield: 4.5 % from 1 in 8 steps Comparison with natural Stephacidin A (spectra and optical data)

- Baran’s Synthesis of Stephacidin A – - Third step: Final formation of Stephacidin A - Determination of absolute configuration + + Stephacidin A R S  1 H and 13 C NMR: identical in all respects to natural Stephacidin A  Optical properties ? R S

Synthesis of Stephacidin B DIMERIZATION Stephacidin A a b Double Michael addition pathway c d c d Cationic pathway

Synthesis of Stephacidin B Myers’ approach: Three steps: 1/ Preparation and reactivity study of a model of Avrainvillamide 2/ Enantioselective synthesis of Avrainvillamide from bicyclodiazaoctane nucleus 3/ Formation of Stephacidin B 2 X Oxidation J. Am. Chem. Soc. 2005, 127, J. Am. Chem. Soc. 2003, 125,

-Myer’s Synthesis of Stephacidin B – - First step: Preparation and reactivity study of a model of avrainvillamide - Oxidative addition Formation of aryl copper derivative 1,1-reductive elimination Shimizu and coworkers, Tetrahedron Lett. 1993, 34, 3421.

-Myer’s Synthesis of Stephacidin B – - First step: Preparation and reactivity study of a model of avrainvillamide (2) - Identification of the Mickael acceptor group A B T = 23 °C A:B = 2:1 T = -20 °C A:B = 10:1

-Myer’s Synthesis of Stephacidin B – - First step: Preparation and reactivity study of a model of avrainvillamide (2) - X !!!

-Myer’s Synthesis of Stephacidin B – - Second step: Synthesis of Avrainvillamide from bicyclodiazaoctane nucleus - Knochel and coll. Angew. Chem. Int. Ed. 2002, 41, 1610.

-Myer’s Synthesis of Stephacidin B – - Second step: Synthesis of Avrainvillamide from bicyclodiazaoctane nucleus - Avrainvillamide Nicolaou and coll. Angew. Chem. Int. Ed. 2005, 44, 3736.

-Myer’s Synthesis of Stephacidin B - - Third Step: Final Formation of Stephacidin B - Avrainvillamide Optical property: Synthetic  D 25 = -35,1 (c 1,0; CHCl 3 ) Natural  D 25 = + 10,6 (c 1,0; CHCl 3 ) Comparison 1 H and 13 C NMR spectra: 1 H NMR: lack of correspondence in the region  C NMR: identical spectra Optical property: Synthetic  D 25 = +91,0 (c 1,0; CHCl 3 ) Natural  D 25 : unknown Comparison 1 H and 13 C NMR spectra: ⇒ Exact correspondence Stephacidin B Interconversion in various solvent-acetonitrile systems: T = 38 °C AVR : SPC B = 2 : 1 T = 23 °C AVR : SPC B = 1 : 2 after 48h

-Synthesis of Stephacidin B - Baran’s approach: Increasing Oxidation State J. Am. Chem. Soc. 2006, 128, 8678.

-Synthesis of Stephacidin B - Baran’s approach: X X X

-Synthesis of Stephacidin B - Baran’s approach: 1/ Initial oxidation studies performed on simplified Stephacidin A models 2/ Total synthesis of Stephacidin B starting from Stephacidin A via Avrainvillamide 3/ Biological evaluation of Avrainvillamide and simplified mimics J. Am. Chem. Soc. 2006, 128, Angew. Chem. Int. Ed. 2005, 44, 3892.

-Synthesis of Stephacidin B - - First Step: Initial Oxidation Studies performed on Simplified Stephacidin A models - Stephacidin A model Synthesis of a Stephacidin A model :

-Synthesis of Stephacidin B - - First Step: Initial Oxidation Studies performed on Simplified Stephacidin A models - Oxidation of Stephacidin A models:

- Synthesis of Stephacidin B - - Second Step: Formation of Stephacidin B starting from Stephacidin A via Avrainvillamide - Synthetic CompoundNatural Compound 3 [  ] D = +11 (c 0.1, CHCl 3 )[  ] D = (c 0.1, CHCl 3 ) 4 [  ] D = -33 (c 0.1, MeCN)[  ] D = (c 0.19, CDCl 3 ) Identical in all respects to the natural Stephacidin B:  LCMS  TLC in several solvent mixtures  1 H NMR  Optical rotation X

- Synthesis of Stephacidin B - - Third Step: Biological Evaluation of Avrainvillamide and Simplified Mimics Biological assays of simplified analogues using the human colon HCT-116 cell line (+)-Stephacidin A (±)-Stephacidin B Model (-)-Stephacidin B Stephacidin A Model (±)-Avrainvillamide Model (+)-Avrainvillamide Activity (µg/mL) no significant activity 10.4 Activity (µg/mL) Essential for anti-cancer activity Low activity Activity restored Best candidate for in vivo studies

- Conclusions - Avrainvillamide (2)Stephacidin B (3) Myers - 17 steps 4.2 % overall 1 step from 2 95 % Baran 8 steps 4.5 % overall 3 steps from 1 26 % overall 1 step from 2 15 – 95 % Stephacidin A (1)

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