4. Hemicalide: Conclusions and Future Work
4.2. Future work
The successful synthesis of the C1-C28 truncate, while validating and confirming several key steps important in the planned total synthesis of the natural product, marks a significant milestone towards the end goal of a synthesis-enabled stereochemical elucidation of hemicalide. As such, this section will detail the immediate future work pertaining to the C1-C28 truncate, as well as how key lessons in this synthesis can inform how best to tackle the remaining regions of the molecule.
4.2.1. Future work in the C1-C28 fragment
The inability to remove all silyl protecting groups in one operation merits further attention in the continued strategy evolution. The peculiar reactivity may be partly attributable to the hindered nature of the TES groups in the dihydroxylactone compared to ones on the C1-C15 region. While a degree of chemical orthogonality could be useful for the site-selective incorporation of covalent modification, such as linkers in an ADC context, the lack of a common set of deprotection conditions across all silyl groups, especially right at the end of a synthetic campaign, only serves to hinder efforts in total synthesis. To this end, an immediate improvement that may benefit the desilylation efficiency could be the incorporation of more labile TMS groups (as exemplified in 139) rather than TES groups in this region (Scheme 23a). The strategic use of TMS groups has proven instrumental in the context of several total syntheses, such as leiodermatolide70 and rhizopodin,79 and perhaps would allow the facile deprotection at the end of the synthesis. Here, the judicious choice of protecting groups is a particularly important consideration in light of Ardisson/Cossy’s recent failures at the global deprotection of the full skeleton of hemicalide.47
The stereochemical elucidation of the C1-C28 truncate marks an important proof-of-concept study for the ability to assign the relative configuration of hemicalide from only the listed 1H and 13C NMR chemical shifts. However, for a full elucidation of its stereochemistry as well as looking ahead towards analogue design, the absolute configuration of hemicalide needs to be determined. Given that direct NMR techniques are unable to ascertain a molecule’s absolute configuration, the only piece of information that could aid determining the absolute configuration of hemicalide is its biological activity. As the C1-C28 fragment constitutes over 60% of the natural product, it is hypothesised that this truncate may exhibit activity in cancer cell lines. Here, the synthesis of the enantiomeric 13,18-syn C1-C28 truncate ent-103 would allow a comparison with 103. By identifying the more active truncate from enantiomers of the C1-C28 region, this can give an indication towards which enantiomeric series to pursue for a total synthesis (Scheme 23b). If these truncates prove to be active, 103 and ent-103 can also serve as probes towards ascertaining the as-yet unknown biological mode of action and begin to develop a comprehensive SAR study for this otherwise
Scheme 23. a) Replacing the inert C21 and C22 TES ethers for more labile TMS ethers in 139 should allow for a more facile global deprotection. b) Assessing the biological activity of truncates 103 and its enantiomer ent-103 can help illuminate the likely correct
enantiomer for hemicalide, as well as revealing needed data on how hemicalide may exert its mechanism of action
1
Requires TASF Requires HF/py
Opportunity for site-selective modification by using the appropriate desilylation conditions
TES to TMS:
(by TMS protection of diol ent-52) Minimises late-stage manipulatuons
Predicted: One enantiomer more active than other Activity-guided rationale for pursuing one enantiomeric
series over another
Opportunities for:
Protein binding studies in silico docking studies Preliminary SAR studies
Focus total synthesis campaign to one enantiomer of the C1-C28 fragment
134
4.2.2. Beyond the C1-C28 region: towards the stereochemical elucidation of hemicalide
The C32-C46 region of hemicalide contains a hydroxylactone motif that is structurally analogous to the synthesised C16-C25 fragment. Therefore, intelligence gathered from exploratory studies during the synthesis of the C16-C25 dihydroxylactone was anticipated to be broadly useful towards developing a robust synthesis of the terminal C35-C46 hydroxylactone.
During the synthesis of model truncate 102 and 103, exploratory studies were conducted to see what the best method was to configure C27 relative to the dihydroxylactone. Unsurprisingly, enantioselective methods (e.g. MeCBS-catalysed borane reductions) proved ineffective owing to the poor steric differentiation around the prochiral carbonyl. This outcome was deemed unimportant as the resulting C27 stereocentre was planned to be set via an enantioselective aldol reaction, but the same process cannot be applied to the analogous C45 stereocentre. The steric hypothesis led to a test scale CBS reduction on enone 127, posited to be a better substrate owing to the ‘larger’ alkene unit compared to the methyl group, which delivered the allylic alcohol 140 in useful selectivities (5:1 dr) (Scheme 24a). This outcome resulted in the overall retrosynthetic analysis for the C35-C46 region (141), where the distal C45 alcohol can be set in either configuration via a CBS reduction using the appropriate catalyst (Scheme 24b). While it was originally thought that a directed hydrogenation mediated by a suitable homogenous catalyst may allow the selective removal of the allylic olefin to give 142, test scale studies on model substrates failed to deliver any activity.
This led to the incorporation of a benzyl protecting group at C35, anticipated to be concomitantly cleaved alongside with the reduction of the C43 double bond over Pd/C under hydrogenative conditions. To this end, Stockdale has obtained promising results validating this route towards the C32-C46 hydroxylactone (see Scheme 7 in section 2.3.3) and has highlighted how the observations made during the synthesis of the C16-C25 fragment have assisted us in the strategy development for the C35-C46 hydroxylactone.
O
I O
TESO
OTES O
127
(R)-MeCBS, BH3•SMe2, THF, −78 ˚C
80%
ca 5:1 dr
O
I O
TESO
OTES
140
OH O
I O
TESO
OTES
142 OH
RhCl(PPh3)3 or [Irpy(cod)PCy3]+
H2 a)
b)
O O TESO
H O
OBn O 35
O TESO
46 H
141
OH TESO
CBS reduction TES Protection Hydrogenation (Pd/C, H2)
HWE olefination
O O TESO
H OBn
PMBO
The configurationally elusive C27-C32 polyacetate region, containing four stereocentres, ideally needs to be elucidated before coupling with either the C1-C25 region, or the C35-C46 hydroxylactone. To this end, Han has completed preliminary efforts towards building a library of model compounds for spectroscopic comparisons with the natural product (Figure 27), which, in comparison with the data presented by Cossy (see Figure 14 in section 2.2.3), should allow for the clear elimination of unlikely diastereomers in this region. Here, even eliminating a few possibilities drastically reduces the number of diastereomeric possibilities to consider.
Figure 27. Preliminary efforts by Han in generating a library of diastereomers for the C27-C32 region, compared with
1
Hemicalide (1); Unknown stereocentres marked in orange
OMe OMe
OH Generate library
of eight diastereomers
TBSO TBSO OH OH
Evan-Tischenko
reduction Narasaka reduction OH
OH
TBSO TBSO OH OH
bis-methylation by Ardisson/Cossy in 23 likely bearing incorrect stereochemistry
1
Greyed structure highlight unlikely structures based on the large deviations reported by Ardisson and Cossy in 23
likely syn likely anti
Deprotect and compare with hemicalide: Ascertain relative configuration within C27-C32 region
To probe the diastereomeric relationship between the C27-C32 and the C35-C46 regions, a modular approach can be developed by employing a cross-metathesis disconnection across C34-C35, which thermodynamically favours the formation of E-olefins (Scheme 25).80 In doing so, this allows the facile appendage of alkene 103a with candidate C27-C32 truncates, deemed to contain the most probable configuration in hemicalide. This can help rapidly narrow down the possibilities for the remaining segment of hemicalide down to a more tractable number. Once the likely diastereomers for the C27-C46 region are identified, a focused synthesis of the C29-C46 aldehyde can take place. In this case, the cross-metathesis partner – alkene 143 – contains two stereocentres, that can be set up by an enantioselective crotylation of a suitably protected aldehyde 144, followed by a hydroboration/oxidation/Wittig olefination sequence.
Alkene 143 can then engage in a cross-metathesis reaction with the C35-C46 alkene 103a, of which the resulting truncate can be treated under Swern conditions, anticipated to result in the concomitant primary TES group cleavage and oxidation81 to give the required C29-C46 aldehyde 145 in anticipation for the planned endgame sequence.
The C1-C28 ketone 146 and the C29-C46 aldehyde 145 can then be subjected to the enantioselective DIPCl-mediated aldol reaction to configure C29, followed by either a 1,3-syn or 1,3-anti reduction to configure C27 – marking the assembly of all 21 stereocentres in hemicalide. Bis-methylation of the resulting diol gives the fully protected skeleton of hemicalide 147. A global deprotection mediated by HF·py/py would then remove all silyl protecting groups, including the now more labile C21 and C22 TMS ethers, followed by methyl ester hydrolysis to give diastereomeric candidates of hemicalide.
At this point, detailed NMR comparisons with hemicalide can then be undertaken to further refine the number of diastereomeric candidates. Any particularly close matches with the natural product can be tested in cancer cell lines, which alongside with the biological data obtained for enantiomers of the C1-C28 fragment, can help point towards the likely absolute stereochemistry of hemicalide.
The present chapter detailing the synthesis-enabled stereochemical elucidation of the C1-C28 truncate of hemicalide, while representing a milestone towards the overall narrative, exemplifies the power of synthesis and spectroscopic analysis in discerning a molecule’s relative configuration. These preliminary results give confidence that a full stereochemical assignment of a molecule as complex as hemicalide, from sparse initial NMR data, is indeed possible.
Scheme 25. Illustration of the proposed workflow for the stereochemical elucidation of hemicalide (1), and a proposal for the endgame of the total synthesis. BAIB = bisacetoxyiodobenzene
O 35
1) NMR comparisons with hemicalide 2) Reduce number of candidate C27-C46 diastereomers
Focused synthesis of C27-C46 aldehyde
O 35 between C27-C32 and C36-C42
fragments
Hemicalide (1 or ent-1): 24 steps LLS
C1-C28 Fragment 146 or ent-146 1) 9-BBN-H then
1) NMR comparisons to identify correct diastereomer 2) Biological studies to confirm absolute configuration
85 103a
147