WO2024153752A1

WO2024153752A1 - Stereoselective synthesis of intermediates and synthesis of quinagolids

Info

Publication number: WO2024153752A1
Application number: PCT/EP2024/051175
Authority: WO
Inventors: Per Ryberg; Mauro PINESCHI; Lucrezia COMPARINI
Original assignee: Ferring B.V.
Priority date: 2023-01-20
Filing date: 2024-01-18
Publication date: 2024-07-25

Abstract

Disclosed is a method for preparing compounds with improved stereoselectivities. The described compounds are useful intermediates and may find particular application in the synthesis of compounds containing an octahydrobenzoquinoline moiety (e.g. an octahydrobenzo[g]quinoline moiety), such as quinagolide and its derivatives. Also disclosed is a method for stereoselectively (e.g. enantioselectively) preparing an intermediate used in the existing manufacturing process for the synthesis of quinagolide this intermediate also finding utility in the synthesis of other compounds containing an octahydrobenzo[g]quinoline moiety, in particular those having a substituent at the 3- position on the octahydrobenzo[g]quinoline.

Description

STEREOSELECTIVE SYNTHESIS OF INTERMEDIATES AND SYNTHESIS OF QUINAGOLIDS

FIELD

The present teaching relates to novel methods for producing enantiomerically enriched (or substantially enantiopure) intermediates. These intermediates may be particularly useful in the production of compounds containing octahydrobenzo[g]-quinoline moieties, such as quinagolide and its derivatives.

BACKGROUND

Ergot alkaloids and their synthetic derivatives show a range of pharmacological activities. In particular, some types of octahydrobenzoquinolines find application as scaffolds for many medicinal drugs (e.g. octahydrobenzo[g]quinolines and octahydro-benzo[f]- quinolines). Examples of compounds containing an octahydro-benzo[f]-quinoline moiety may be found in J. Med. Chem. 1984, 27, 190; J. Med. Chem. 1991 , 34, 497; Org. Proc. Res. Dev. 1997, 1 , 395; J. Med. Chem. 1998, 41 , 4165; Biorg. Med. Chem. Lett. 2012, 20, 6366). In relation to octahydrobenzo[g]quinolines (which are synthetic analogues to the ergot family), these have been reported as having potent dopamine activity. One such synthetic analogue is quinagolide. Quinagolide (C20H33N3O3S) is a selective, D₂ receptor agonist with a molecular mass of about 395 g/mol.

Quinagolide has been described as useful in the treatment of a number of conditions including hyperprolactinaemia (Eur. J. Endocrinol February 1 , 2006 154 page 187-195) and endometriosis (US2012/0157489, US2010/0113499 and US2008/0293693). In particular, Ferring prior patent publication WO 2016/071466 describes a controlled release polymeric drug device unit comprising quinagolide intended for the treatment of endometriosis.

The racemic form of quinagolide is commercially available under the trade name NORPROLAC® and is manufactured and used in the form of the hydrochloride salt. Quinagolide hydrochloride is a white crystalline powder of high melting point (231 -237°C under decomposition), that is sparingly soluble in water.

Whilst it is currently sold in racemic form, the pharmacological activity of quinagolide is believed to reside in a single enantiomer. However, quinagolide has three stereocentres resulting in eight diastereomeric structures (as shown by compounds 1 , T, 2, 2’, 3, 3’, 4 and 4’ in Figure 1 ) and this complex stereochemistry has proven challenging when devising synthetic routes to a single enantiomeric form.

The chemical structure of the active enantiomer of quinagolide is shown below as compound 1 :

1) 3S,4aS,10aR

(1 ) (-) enantiomer

Compound 1 has the absolute configuration (3S, 4aS, 10aR) and may also be referred to as (-)-/V,/V-diethyl-/V-[(3S,4aS,10aF?)-6-hydroxy-1 -propyl-1 ,2, 3, 4, 4a, 5,10,10a- octahydrobenzo[g]quinolin-3-yl]sulfamide. In the racemic form of quinagolide, compound 1 is present in a mixture with compound 1’ in a 1 :1 ratio, wherein compound T has the absolute configuration (3R, 4aR, 10aS) (see Figure 1 ).

Nordmann et al (J. Med. Chem. 1985, 28, 367) first reported the synthesis of quinagolide in racemic form and isolated compound 1 in a subsequent pharmacological study (J. Med. Chem. 1985, 28, 1540). Banziger et al (Organic Process Research & Development, 2000, 4, 460) have described a large scale synthesis of a racemic intermediate, potentially useful in the synthesis of quinagolide and this is shown in Figure 2. More recently, Chavan et al (Org. Lett. 2018, 20, 7011 ) have described a total synthesis of racemic quinagolide using a ceric ammonium nitrate (CAN)-mediated regioselective azidoalkoxylation of an enol ether intermediate. The first formal total synthesis of the active eutomer of quinagolide has been reported by Chavan et al. (S. P. Chavan, A. L. Kadam, R. G. Gonnade, Org. Lett. 2019, 21, 9089-9094). However, the synthesis is long, makes use of expensive and toxic reagents and does not appear amenable to scale-up up to manufacturing scale.

In the manufacturing route to quinagolide shown in Figure 2, intermediates C9 and C10 have three stereocentres, meaning that there is potentially eight diastereomers (four pairs of enantiomers) at this stage in the synthesis. In fact, the reduction of the iminium ion (C8) typically favours the formation of a trans ring junction as is shown in intermediates C9 and C10 of Figure 2. However, there still remains substantially four diastereomers (two pairs of enantiomers) at this stage in the manufacturing route to quinagolide. An epimerization step (converting C13 to C14) is later used to provide a pair of enantiomers having the relative stereochemistry shown in C14 and these two enantiomers are carried through the remainder of the synthesis to provide a racemic mixture of quinagolide (see Figure 2). Thus, there remains a need to devise new enantioselective routes to quinagolide that are amenable for larger scale manufacture.

SUMMARY

The present disclosure is based on the identification of a method for preparing compounds with improved stereoselectivities. The described compounds are useful intermediates and may find particular application in the synthesis of compounds containing an octahydrobenzoquinoline moiety (e.g. an octahydrobenzo[g]quinoline moiety), such as quinagolide and its derivatives. In particular, the present disclosure relates to a method for stereoselectively (e.g. enantioselectively) preparing an intermediate used in the existing manufacturing process for the synthesis of quinagolide (this intermediate also finding utility in the synthesis of other compounds containing an octahydrobenzo[g]quinoline moiety, in particular those having a substituent at the 3- position on the octahydrobenzo[g]quinoline.

The preparation of enantiomerically enriched (or substantially enantiopure) intermediates bearing multiple stereogenic centres represents a significant synthetic challenge. For examples, compounds comprising an octahydrobenzoquinoline moiety (such as an octahydrobenzo[g]quinoline moiety) bearing multiple substituents can comprise multiple stereogenic centres. By way of example, quinagolide comprises three stereocentres and providing such moieties in an enantioselective (or substantially enantiopure fashion) is non-trivial.

The present inventors have identified that certain enzymes can be used to provide enantiomerically enriched intermediates that are useful in the synthesis of compounds containing an octahydrobenzoquinoline moiety (e.g. an octahydrobenzo[g]quinoline moiety), such as quinagolide and its derivatives. Such enzymes may facilitate and/or promote the preparation of enantiomerically enriched intermediates by way of a kinetic resolution process. The identification of the stereoselective method described herein may facilitate and/or assist in the production of quinagolide (and other compounds containing an octahydrobenzo[g]quinoline moiety) with improved stereoselectivities.

The present inventors have further identified that certain reducing agents can be used to enhance the diastereoselectivity of an iminium reduction step. The identification of this reduction step with an improved diastereoselectivity (as described herein) may facilitate and/or assist in the production of octahydrobenzoquinolines, such as quinagolide (and other compounds containing an octahydrobenzo[g]quinoline moiety, in particular those bearing a substituent at the 3-position - see, for example, Figure 1 for atom numbering in quinagolide) with improved stereoselectivities. In particular, a diastereoselective borohydride reduction in combination with an efficient deacylative enzymatic resolution of a p-aminoester precursor may find particular application in the stereoselective installation of the three chiral centres present in the (3S, 4aS, 10aR)- eutomer of quinagolide

Indeed, whilst both the kinetic resolution process and iminium reduction step can be each used independently to facilitate and/or assist in the production of quinagolide (and other compounds containing an octahydrobenzoquinoline moiety (e.g. an octahydrobenzo[g]quinoline moiety)) with improved stereoselectivities, the combined use of these steps in a multi-step synthesis is particularly effective in providing quinagolide (and other compounds containing an octahydrobenzoquinoline moiety (e.g. an octahydrobenzo[g]quinoline moiety)) with improved stereoselectivities.

As used herein, the term “quinagolide” includes all commercially available forms as well as functional derivatives and variants thereof. The term “quinagolide” also embraces all pharmaceutically acceptable (and active) salts and esters, including, for example, quinagolide hydrochloride. Consequently, it will be appreciated that the methods described herein may be applied to also the manufacture of derivatives of quinagolide in enantiomerically enriched or enantiopure form.

According to a first aspect of the disclosure there is provided a stereoselective (e.g. enantioselective) method for preparing and/or obtaining a compound of formula (II’).

The compound of formula (II’) is shown below:

wherein R¹ may be selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce alkoxy, optionally substituted Ci-Ce haloalkyl, halo, optionally substituted aryl, and optionally substituted heteroaryl, or R¹ may be absent; and

R², R³ and R⁴ are each independently selected from H, optionally substituted Ci-Ce alkyl, optionally substituted CO(Ci-Ce alkyl), optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl.

As shown in the structure above, the stereochemistry at the starred (*) position may be (R).

As shown on formula (II’) above, when present, R¹ may be appended to the aromatic system, e.g. in some examples, R¹ may be appended to aryl ring (A). R¹ may be attached to the aryl ring by way of a covalent bond and/or may replace a hydrogen at any position on the aromatic ring.

As shown on formula (II’) above, the two pendant groups (respectively bearing OR² and NR³R⁴) are appended to the aromatic system, e.g. in some examples, these groups may be appended to aryl ring (B). These groups may each be independently attached to the aryl ring by way of a covalent bond and/or may replace a hydrogen at any position on the aromatic ring. In particular examples, these two groups may be bonded to the aryl ring B via adjacent carbon ring atoms (e.g. in a [1 ,2]- substitution relationship relative to one another).

By way of further example, the compound of formula (II’) may comprise a structure according to formula (II):

wherein R¹, R², R³, R⁴ and * are as defined above for formula (II’).

In alternative examples, the compound may comprise the structure of formula (H’a):

wherein R¹, R², R³, R⁴ and * are as defined above for formula (II’).

The method may comprise preparing and/or obtaining a compound of formula (II’) from a compound of formula (I’).

The compound of formula (I’) is shown below:

wherein R¹, R², R³ and R⁴ are as defined for formula (II’); and

R⁵ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl. In particular examples, the method may comprise preparing and/or obtaining a compound of formula (II) from a compound of formula (I).

The compound of formula (I) is shown below:

wherein R¹, R², R³, R⁴ and R⁵ are as defined for formula (I’).

In particular examples, the method may comprise preparing and/or obtaining a compound of formula (H’a) from a compound of formula (I’a).

The compound of formula (I’a) is shown below:

wherein R¹, R², R³, R⁴ and R⁵ are as defined for formula (I’).

In the structures shown above for formulae (I’), (I) and (I’a), the stereochemistry at the starred (*) position may be (R) or (S). In some cases, the compound of formula (I’), (I) or (I’a) may be provided as a mixture of the (R) and (S) enantiomer. By way of example, the compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) may be provided as a racemic mixture. In other words, the compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) may comprise equimolar amounts of each enantiomer. The method may comprise a step of contacting and/or reacting the compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) with one or more reagent(s) and/or under conditions that facilitate or promote the hydrolysis of a compound of formula (I) to provide a compound of formula (II’) (e.g. a compound of formula (II) or (I I’a)). The method may comprise a step of contacting and/or reacting the compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) with an enzyme, such as a hydrolytic enzyme. The enzyme may hydrolyse and/or catalyse the hydrolysis of the compound of formula (I’) (e.g. a compound of formula (I) or (I’a)). In some examples, the enzyme may hydrolyse and/or catalyse the hydrolysis of a pendant ester moiety on the compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) to the corresponding carboxylic acid and so provide a compound of formula (II’) (e.g. a compound of formula (II) or (ll’a)).

The method may comprise a step of kinetic resolution, in particular an enzymatic kinetic resolution. As used herein, kinetic resolution is a means of differentiating two enantiomers in a racemic mixture. An enzymatic kinetic resolution may refer to a process in which one of the two enantiomers present in a starting material mixture (e.g. a racemic mixture of the starting material) reacts preferentially with (and/or is catalysed preferentially by) an enzyme to provide an enantiomerically enriched product. By way of example, a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) having a particular stereochemistry (e.g. an R stereochemistry) may react (or be catalysed) preferentially by an enzyme to provide a product of formula (II’) (e.g. a compound of formula (II) or (ll’a)) in an enantiomerically enriched form.

As stated above, in the methods disclosed herein, compounds of formula (I’) may be generally provided in a racemic form. In other words, there may be an equimolar amount of each enantiomer present. The methods described herein (e.g. which make use of an enzymatic kinetic resolution) provide compounds of formula (II’) (e.g. a compound of formula (II) or (ll’a)) in an enantiomerically enriched form.

In some cases, enzymatic kinetic resolution may be used and/or lead to enantiomeric enrichment of the less reactive enantiomer of the starting material. By way of example, where one of the two enantiomers present in a starting material mixture (e.g. a racemic mixture of the starting material) reacts preferentially with (and/or is catalysed preferentially by) an enzyme, there is also provided an enantiomerically enriched starting material. By way of example, where a compound of formula (I’) having a particular stereochemistry (e.g. an R stereochemistry) reacts (or is catalysed) preferentially by an enzyme to provide a product of formula (II’) (e.g. a compound of formula (II) or (H’a)) in an enantiomerically enriched form, there may also be provided a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) in an enantiomerically enriched form (e.g. a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) having an (S) stereochemistry). In some cases, the enantiomerically enriched form of compounds of formula (I’) (e.g. a compound of formula (I) or (I’a)) may also have utility as useful intermediates in the synthesis of other compounds containing an octahydrobenzoquinoline moiety (e.g. an octahydrobenzo[g]quinoline moiety) (e.g. those compounds that have an opposite stereochemistry to quinagolide and its derivatives).

Accordingly, there is further provided a method of preparing and/or obtaining a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) in an enantiomerically enriched form from a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) comprising a mixture of two enantiomers (e.g. a racemic mixture of the two enantiomers). The method may comprise a step of kinetic resolution, in particular an enzymatic kinetic resolution using the enzymes and conditions such as disclosed herein.

The present inventors have particularly identified that a class of intermediates useful in the large scale synthesis of quinagolide can be produced in an enantiomerically enriched form using such an enzymatic kinetic resolution step. The provision of such enantiomerically enriched intermediates can be used to facilitate the manufacture of quinagolide in enantiomerically enriched form. Furthermore, the intermediates that undergo this kinetic resolution step are easily obtainable meaning that this route may find particular application in a larger scale synthesis of quinagolide.

In particular, the inventors have identified that the desired intermediates can unexpectedly be provided with a high degree of enantioselectivity when using the enzymatic kinetic resolution methods described herein. This means that the described processes may be practically useful in a larger scale synthesis of quinagolide and other compounds containing an octahydrobenzo[g]quinoline moiety.

The transformation and/or conversion of a compound of formula (I’) to a compound of formula (II’) is illustrated below.

wherein R¹, R², R³, R⁴ and R⁵ are as defined for formula (I’).

By way of further example, the transformation and/or conversion of a compound of formula (I) to a compound of formula (II) is illustrated below.

wherein R¹, R², R³, R⁴ and R⁵ are as defined for formula (I).

By way of further example, the transformation and/or conversion of a compound of formula (I’a) to a compound of formula (ll’a) is illustrated below.

wherein R¹, R², R³, R⁴ and R⁵ are as defined for formula (I’a).

In the described methods (and as shown above), the enzyme may catalyse the hydrolysis of the -CO2R⁵ moiety in formula (I’) (e.g. in formula (I) or formula (I’a)) to the corresponding carboxylic acid as shown in formula (II’) (e.g. in formula (II) or formula (I I’a)). In particular, the enzyme may preferentially catalyse the hydrolysis of the -CO2R⁵ moiety of compounds of formula (I’) (e.g. of formula (I’a) or (ll’a)) having an (R) stereochemistry.

Enzymes that are useful in the described methods include any enzyme that is able to hydrolyse and/or catalyse the hydrolysis of an ester group to a carboxylic acid group. Particularly useful enzymes are those which provide the carboxylic acid product in an enantiomerically enriched form.

Suitable enzymes may be selected from the group consisting of lipases and proteases. In some examples, the enzyme is a protease, e.g. an endopeptidase, such as of the serine subtype. In some examples, the enzyme may be a subtilase. For example, the enzyme may be a protease derived from Bacillus licheniformis, e.g. an endopeptidase, such as of the serine subtype derived from Bacillus licheniformis. Representative examples include, but are not limited to, Protease P4860 from Sigma Aldrich and Alcalase® from Merck-Aldrich 126741. Other examples include protease from bovine pancreas Type I. The enzyme may have an activity greater than or equal to 0.5 U/g, greater than or equal to 0.75 U/g, greater than or equal to 1 U/g, greater than or equal to 1 .5 U/g, or greater than or equal to 2 U/g. In some examples, the enzyme may have an activity of approximately greater than or equal to 2.4 U/g.

Indeed, the inventors have surprisingly identified that a protease (e.g. an endopeptidase, such as of the serine subtype) derived from Bacillus licheniformis can provide compounds of formula (II’) (e.g. of formula (II) or formula (ll’a)) with exceptionally high enantioselectivity.

In some examples, as used herein, enantiomerically enriched may mean that the enantiomeric excess (ee) of the major enantiomer is at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80% or at least about 90%. In some cases, the enantiomeric excess of the major enantiomer may be at least about 95%, at least about 97%, at least about 98%, or at least about 99%. In some cases, enantiomerically enriched may mean that the compound is substantially enantiopure. As used herein, enantiopure may mean that the compound is present in a single enantiomeric form (e.g. the (+) (dextrorotatory) or (-) (levorotatory) enantiomer).

As stated above, in the various formulae described above and herein (e.g. formulae (I’), (II’), (I), (II), (I’a) and (ll’a)), R¹ may be selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce alkoxy, optionally substituted Ci-Ce haloalkyl, halo, optionally substituted aryl, and optionally substituted heteroaryl; or R¹ may be absent. As shown in the various formulae described above, when present, R¹ is appended to the aryl ring. R¹ may be attached to the aryl ring by way of a covalent bond and/or may replace a hydrogen at any position on the aromatic ring.

In some examples, R¹ may be selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce alkoxy, halo or R¹ may be absent. In some examples, R¹ may be selected from Ci-Ce alkyl, Ci-Ce alkoxy and halo. In some examples, R¹ may be Ci-Ce alkoxy (e.g. C1-C3 alkoxy such as methoxy).

As stated above, in the various formulae described above (e.g. formulae (I’), (II’), (I), (II), (I’a) and (H’a)), R², R³ and R⁴ are each independently selected from H, optionally substituted Ci-Ce alkyl, optionally substituted CO(Ci-Ce alkyl), optionally substituted Ci- Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl.

In some examples, R² may be selected from H, optionally substituted Ci-Ce alkyl, and optionally substituted Ci-Ce haloalkyl. In some examples, R² may be selected from optionally substituted Ci-Ce alkyl (e.g. C1-C3 alkyl such as methyl).

In some examples, R³ and R⁴ may each be independently selected from H, optionally substituted Ci-Ce alkyl, optionally substituted CO(Ci-Ce alkyl), and optionally substituted Ci-Ce haloalkyl. In some examples, at least one of R³ and R⁴ is H. In some examples, both R³ and R⁴ are H. In some examples, R³ may be H and R⁴ may be CO(Ci-Ce alkyl) (e.g. COCH3).

As stated above, the various formulae described above (e.g. formulae (I’), (I) and (I’a)), R⁵ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl. In some examples, R⁵ may be selected from optionally substituted Ci-Ce alkyl and optionally substituted Ci-Ce haloalkyl. In some examples, R⁵ may be optionally substituted Ci-Ce alkyl (e.g. methyl).

In particular examples of the formulae (I’), (I) and (I’a) described above:

R¹ may be selected from Ci-Ce alkyl, Ci-Ce alkoxy and halo;

R² may be selected from Ci-Ce alkyl;

R³ and R⁴ may each be H; and

R⁵ may be Ci-Ce alkyl (e.g. methyl). In particular examples of the formulae (II’), (II) and (H’a) described above:

R¹ may be selected from Ci-Ce alkyl, Ci-Ce alkoxy and halo;

R² may be selected from Ci-Ce alkyl; and

R³ and R⁴ may each be H.

In some examples, the compound that undergoes the kinetic resolution step may comprise a structure according to formula (la):

wherein R^1A is Ci-Csalkoxy;

R^2A is Ci-C₃alkyl;

R^3A and R^4A are each H; and

R^5A is Ci-C₃alkyl.

In a yet further example, the compound may comprise a structure according to formula (lb):

In the structures of formulae (I’), (I), (I’a), (la) and (lb) above, the stereochemistry at the starred (*) position may be (R) or (S). In some cases, the compound of any one of formulae (I’), (I), (I’a), (la), and (lb) may be provided as a mixture of the (R) and (S) enantiomer. By way of example, the compound of any one of formulae (I’), (I), (I’a), (la) and (lb) may be provided as a racemic mixture. The compound of formula (I) may be methyl 3-amino-2-((3,8-dimethoxynaphthalen-2- yl)methyl)propanoate.

In some examples, the compounds that are prepared in the described reactions (e.g. that are formed following the kinetic resolution step) have a structure according to formula (Ila):

wherein R^1A is Ci-Csalkoxy; and R^2A is Ci-C₃alkyl.

In a yet further example, the compound that may be prepared in the described reactions has a structure according to formula (lib):

In the structures of formulae (II), (Ila) and (lib), the stereochemistry at the starred (*) position may be (R).

The compound of formula (II) may be fR)-3-amino-2-((3,8-dimethoxynaphthalen-2- yl)methyl)propanoic acid.

The methods described herein may be carried out under any suitable conditions that allow the enzyme to hydrolyse and/or catalyse the hydrolysis of the pendant -CO2R⁵ (or- COsR^5A) group to the corresponding carboxylic acid group. By way of example, the method may be carried out in any suitable solvent that allows the enzyme to hydrolyse and/or catalyse the hydrolysis of the pendant -CO2R⁵ (or- CC>2R^5A) group to the corresponding carboxylic acid group. The solvent may be or comprise an organic solvent and/or an aqueous solvent. Representative examples of suitable solvents include, but are not limited to, acetone, acetonitrile, dichloromethane (DCM), dimethylformamide (DMF), dimethyl sulfoxide (DMSO), 1 ,4-dioxane, ethanol, ethyl acetate (AcOEt or EtOAc), methanol, pyridine, tetrahydrofuran (THE), toluene, water etc. The hydrolysis reaction may take place in an aqueous solution.

The solvent may be or comprise a buffer (e.g. a buffered aqueous solution). Representative examples of suitable buffers include, but are not limited to, phosphate- based buffers, citrate-based buffers, tris-based buffers and acetate-based buffers. In particular, the buffer may be a phosphate-based buffer.

The method may be carried out at any temperature that allows the enzyme to hydrolyse and/or catalyse the hydrolysis of the pendant -CO2R⁵ (or-CC>2R^5A) group to the corresponding carboxylic acid group. For example, the method may be carried out at a temperature below the denaturation temperature of the enzyme. By way of example only, the method may be carried out at a temperature between about 10 °C and 100 °C, between about 20 °C and 65 °C, or between about 25 °C and 50 °C, for example between 30 °C and 40 °C. In some examples, the method may be carried out between about 32 °C and 39 °C, such as approximately 35 °C or 37 °C.

The method may be carried out at any pH that allows the enzyme to hydrolyse and/or catalyse the hydrolysis of the pendant -CO2R⁵ (or-CC>2R^5A) group to the corresponding carboxylic acid group. By way of example only, the method may be carried out at a pH range between about pH 4 and pH 11 , or between about pH 6 and pH 10, or between about pH 6.5 and pH 9.5. In some examples, the method may be carried out at about pH 7

The inventors have also identified that, in some examples, it is not necessary to maintain a constant pH value throughout the duration of the hydrolysis reaction. As such, in some examples, the method does not comprise adding a basic solution (such as sodium hydroxide) to the reactants to maintain a constant pH. In other words, in some examples, there is no requirement to add a base to maintain a constant pH and the reaction proceeded effectively without the addition of a basic solution. In such examples, during the reaction, the pH may decrease to around pH 5. As such, in some examples, the method may be carried out at a pH range between about 5 and 7.

As stated previously, the enantiomerically enriched compounds of formula (II) may find particular application in the synthesis of quinagolide. In other words, the enantiomerically enriched compounds of formula (II) may be useful intermediates in the synthesis of an enantiomerically enriched quinagolide (e.g. an enantiomerically enriched form of (3S,4aS,1 OaR-quinagolide).

As such, according to a further aspect of the present disclosure there is provided a use of compounds according to formulae (I’) and (II’) in the synthesis of a compound comprising an octahydrobenzoquinoline moiety.

By way of particular example, there is provided a use of compounds according to formulae (I), (la), (lb), (II), (Ila) and (lib) in the synthesis of quinagolide and/or in the synthesis of a compound comprising an octahydrobenzo[g]quinoline moiety.

By way of further example, there is provided a use of compounds according to formulae (I’a) and (H’a) in the synthesis of a compound comprising an octahydrobenzo[f]quinoline moiety.

There is further provided a method for preparing a compound containing an octahydrobenzoquinoline moiety in an enantiomerically enriched form which comprises a step of converting a compound of formula (I’) to a compound of formula (II’) e.g. by way of the enzymatic hydrolytic kinetic resolution processes as described herein.

By way of particular example, there is further provided a method for preparing a compound containing an octahydrobenzo[g]quinoline moiety (e.g. quinagolide) in an enantiomerically enriched form which comprises a step of converting a compound of formula (I) to a compound of formula (II) e.g. by way of the enzymatic hydrolytic kinetic resolution processes as described herein.

By way of further example, there is further provided a method for preparing a compound containing an octahydrobenzo[f]quinoline moiety in an enantiomerically enriched form which comprises a step of converting a compound of formula (I’a) to a compound of formula (H’a) e.g. by way of the enzymatic hydrolytic kinetic resolution processes as described herein. By way of example only, an enantiomerically enriched compound of formula (II) can be used in place of the racemic C6 intermediate in all or part of the synthesis shown on Figure 2. In this way, the enantiomerically enriched compound of formula (II) can be used to provide an enantiomerically enriched quinagolide and/or an enantiomerically enriched compound containing an octahydrobenzo[g]quinoline moiety (such as an enantiomerically enriched C14 intermediate (as shown in Figure 2)). In particular, the present inventors have identified that subsequent steps that are performed on the enantiomerically enriched C6 intermediate (similar to those used in the existing manufacture of quinagolide) can proceed diastereoselectively and favour the desired diastereomer.

Accordingly, the described methods can assist in and/or facilitate the provision of enantiomerically enriched intermediates (such as intermediates C9, C10, C1 1 , C13 and C14 as shown in Figure 2) that may be useful in the synthesis of compounds containing an octahydrobenzo[g]quinoline moiety, such as quinagolide.

A general overview of such a method is illustrated in Figure 3. In particular, the general method shown in Figure 3 shows how enantiomerically enriched compounds of formula (II) can be used to provide enantiomerically enriched intermediates (III), (IV), (V), (VI) and (VII). Whilst these steps are all shown as part of a general method, it will be appreciated that each one of the intermediates (III) to (VII) may find use in the synthesis of compounds comprising an octahydrobenzo[g]quinoline moiety in an enantiomerically enriched form. Thus, the present disclosure further provides methods for preparing each one of the compounds (III), (IV), (V), (VI) and (VII) in an enantiomerically enriched form.

Accordingly, there is provided a method of preparing a compound of formula (III):

In the iminium ions of formula (III), R¹ may be as is defined above for formulae (I) and (II) (or R¹ may be as is defined for R^1A in formulae (la) and (Ila)). X- may be a counter ion. For example, X- may be selected from halide (e.g. chloride, fluoride, bromide or iodide), sulfate, phosphate, organic carboxylate (e.g. citrate, acetate, lactate, pyruvate, oxalate, etc) and organic sulfonate (e.g. p-toluenesulfonate, methanesulfonate, ethanesulfonate, benzenesulfonate, etc). In some examples, X- may be chloride.

In formula (III), the wavy line bond indicates that the stereochemistry is either (R) or (S).

The method of preparing a compound of formula (III) is illustrated on Figure 3 as “step b”.

The method of preparing a compound of formula (III) may comprise a step of reducing a compound of formula (II).

The step of reducing a compound of formula (II) may be carried out by any suitable reagents such as are known in the art. By way of example only, the reduction may be a Birch reduction and/or may comprise contacting a compound of formula (II) with a metal in ammonia (e.g. lithium in ammonia). The method may further comprise an acidification step to provide the iminium ion of formula (III).

In a compound of formula (III), the stereochemistry at the 3-position (the pendant carboxylic acid group) may predominantly be in the R-configuration via the use of an enantiomerically enriched starting material (e.g. the compound of formula (II)). As shown by the wavy line in formula (III) above, the stereochemistry at the 4a position of the octahydrobenzo[g]quinoline moiety may be either S or R. In some cases, there may be a mixture of the two diastereomers, e.g. an equimolar amount of the two diastereomers (e.g. (3R, 4aS) and (3R, 4aR)).

By way of example, a compound of formula (II) may undergo a reduction followed by an acidic treatment to provide the iminium salt as shown in formula (III). Compounds of formula (II) (which are present in an enantiomerically enriched form) selectively provide two diastereomers (a single enantiomer of each), as the stereochemistry of the pendant carboxylic acid group may have been essentially fixed by the enzymatic kinetic resolution step. This is in comparison to the known manufacturing process of quinagolide which is carried out on racemic C6 intermediate (as shown in Figure 2). In the existing process, a mixture of four stereoisomers is typically produced (e.g. a pair of enantiomers for each diastereomer). In the existing manufacturing process of quinagolide, the next step of the synthesis involves a reduction of an iminium ion (see, for example, steps going from intermediate C8 to C9 to C10 on Figure 2). The step of reducing the iminium ion intermediate essentially installs the stereochemistry of the two remaining stereocentres.

Thus, according to the disclosure, there is further provided a method of preparing a compound of formula (IV):

wherein R¹ is as is defined above for formula (I’), (I), (I’a), (II’), (II) and (H’a) (or R¹ may be as is defined for R^1A in formulae (la) and (Ila)); and

R⁶ may be selected from H, optionally substituted Ci-Ce alkyl, optionally substituted Ci- Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl. In some cases, R¹ may C1-C3 alkoxy (e.g. methoxy) and R⁶ may be H or Ci-Ce alkyl (e.g. methyl).

The method for preparing a compound of formula (IV) may comprise reacting and/or converting a compound of formula (III). In particular, the method may comprise reducing the iminium ion of formula (III). The method may further comprise acidifying and/or esterifying the reduced iminium ion to provide the compound of formula (IV).

The method of preparing a compound of formula (IV) is illustrated on Figure 3 as “step c”.

The method for preparing a compound of formula (IV) may comprise drying a compound of formula (III) prior to subjecting the compound of formula (III) to the reduction (and esterification) shown in step c of Figure 3. The step of drying compound (III) prior to the reduction (and then esterification) may assist in providing a reproducible reaction. In some examples, the method may further comprise drying a compound of formula (III) by heating and/or under reduced pressure (e.g. under a vacuum). In some examples, the compound of formula (III) may be dried at a temperature between about 25 and 75 °C, or between about 30 and 60 °C, e.g. about 50 °C (optionally under vacuum). The compound of formula (III) may be dried until only a residual amount of amount of water remains, e.g. between about 0 and 20% w/w of water, between about 5 and 15% w/w of water, or between about 7 to 10% w/w of water.

As noted previously, in the manufacturing route to quinagolide shown in Figure 2, the corresponding intermediates C9 and C10 are typically obtained as four diastereomers (two pairs of enantiomers). In contrast, using the methods described herein, and particularly starting from a compound of formula (II) (which is present in an enantiomerically enriched form), substantially only two diastereomers may be obtained at this stage of the process (a single enantiomer of each). This can greatly reduce the complexities associated with later synthetic steps by reducing the number of diastereomers present in the material.

As illustrated in Figure 5, the step of reducing (and then esterifying) the iminium ion of formula (III) may provide two diastereomers (e.g. compound of formula (IV) and a compound of formula (VIII)).

The compound of formula (VIII) is shown below:

wherein R¹ is as is defined above for formula (I) and (II) (or R¹ may be as is defined for R^1A in formulae (la) and (Ila)); and

R⁶ may be selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl. In some cases, R¹ may C1-C3 alkoxy (e.g. methoxy) and R⁶ may be H or Ci-Ce alkyl (e.g. methyl).

In some examples, compounds of formula (IV) may be separated and/or substantially separated from compounds of formula (VIII) using methods as known in the art (e.g. crystallization, chromatography (such as silica column chromatography) etc). In some examples, the step of reducing the iminium ion of formula (III) may be diastereoselective. For example, the step of reducing the iminium ion of formula (III) may provide compounds of formula (IV) in a diastereomerically enriched form. In other words, the step of reducing the iminium ion of formula (III) may provide compounds of formula (IV) in a greater molar amount than compounds of formula (VIII) and/or may favour the formation of compounds of formula (IV).

In particular, the present inventors have identified that the use of particular metal borohydride reducing agents and/or the presence of additional metal salts can unexpectedly lead to higher levels of diastereoselectivity (favouring the formation of compounds of formula (IV)). Suitable metal borohydrides that may be used in the reduction step include but are not limited to, alkali metal borohydrides (such as lithium borohydride and sodium borohydride).

Where certain types of metal borohydride are used as the reducing agent (e.g. sodium borohydride), the inventors have unexpectedly identified that the presence of additional metal salts can further enhance the diastereoselectivity. In particular, the presence of magnesium salts such as magnesium halides (e.g. magnesium chloride and magnesium bromide) and magnesium carbonate and/or lithium salts such as lithium halide (e.g. lithium chloride) may increase the diastereoselectivity of this reduction step. Thus, in some examples, the additional metal salts may be selected from magnesium halide, magnesium carbonate and lithium halide. In particular examples, the additional metal salts may be selected from magnesium salts, such as magnesium halides.

In other examples, the inventors have identified that the use of lithium borohydride (LiBH₄) as the reducing agent can lead to good levels of diastereoselectivity with good levels of reproducibility.

Without being bound by theory, the inventors hypothesize that one of the two diastereomers of the C8 intermediate (see for example, Figure 2 of the manufacture of quinagolide) is more reactive and that reduction occurs from the same side of the carboxylic moiety e.g. by means of a metal chelation. The inventors further hypothesize that the less reactive diastereomer may undergo a partial equilibration to the more reactive diastereomer via epimerization at the 4a position (e.g. imine-enamine tautomerism). Such mechanism may account for the high levels of diastereoselectivity observed in this reaction step (see also Figure 8). Thus, according to a further aspect of this disclosure there is provided a method of reducing an iminium ion of formula (III) by contacting the iminium salt with a reducing agent selected from lithium borohydride and sodium borohydride, wherein when sodium borohydride is used as the reducing agent an additional metal salt is present.

As stated above, the step may proceed diastereoselectively to favour the formation of intermediate (IV).

The method may further comprise acidifying and/or esterifying the intermediate to provide compounds of formula (IV) and (VIII). For example, as would be appreciated by the skilled person, following acidification R⁶ of compounds of formula (IV) would be H and following esterification (e.g. using a Ci-Ce alcohol), R⁶ of compounds of formula (IV) would be Ci-Ce alkyl.

The reducing agent may be added in any suitable amount to facilitate and/or promote the reduction of the iminium ion of formula (III). For example, the reducing agent may be added in an amount between about 1 and 10 equivalents, such as between about 1 and 8 equivalents, between about 1 and 5 equivalents or about 1.5 equivalents (based on the molar amount of the starting material e.g. the iminium compound of formula (III)).

Where an additional metal salt is present, the amount of additional metal salt may be between about 0.0001 equivalents and 1 equivalent based on the molar amount of the starting material (e.g. the iminium ion of formula (III)), such as between about 0.005 and 0.75 equivalents, between about 0.01 equivalents and 0.5 equivalents, or between about 0.10 equivalents and 0.25 equivalents. In some examples, the amount of additional metal salt may be about 0.2 equivalents (based on the molar amount of the starting material (e.g. the iminium ion of formula (III)).

In some examples, the additional metal salt may be present in the metal borohydride as a contaminant and/or a manufacturing impurity. Thus, the reducing agent may be a metal borohydride of greater than or equal to about 90%, about 95% or about 98% purity (and optionally wherein the balance by weight is comprised of additional metal salts).

The reaction may take place in any suitable solvent. In some examples, the reaction takes place in an anhydrous solvent. In yet further examples, the reaction may be conducted in an anhydrous alcohol solvent (e.g. anhydrous methanol). In some examples, the reaction is conducted in (and/or the reactants are added to) a solvent containing between about 0 ppm and 100000 ppm, or between about 0.1 ppm and 100000 ppm, such as between about 1 ppm and 10000 ppm of water. In some examples, the reaction is conducted in (and/or the reactants are added to) a solvent containing between about 300 and 900 ppm of water, or between about 400 and 800 ppm of water.

The reaction may take place at any suitable temperature. For example, in some cases, the reducing agent may be added at a temperature between 0 °C and -150 °C, such as between -20 °C and -100 °C, or between -40 °C and -80 °C. In some examples, the reducing agent may be added at a temperature of about -75 °C (e.g. between about -70 °C and -78 °C).

As used herein, a diastereomerically enriched form may mean that the diastereomeric excess (d e) of the major diastereomer is at least about 5%, at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80% or at least about 90%. In some cases, the diastereomeric excess of the major diastereomer may be at least about 95%, at least about 97%, at least about 98%, or at least about 99%.

In some examples, the diastereoselectivity of a step may be expressed as a diastereoselective ratio (e.g. a ratio of a first diastereomer to a second diastereomer). In some examples, the reduction of the iminium compound of formula (III) followed by an acidification and/or esterification step may provide compounds of formula (IV) in a diastereomeric ratio of at least about 3:1 , at least about 4:1 , at least about 5:1 , or at least about 9:1 (e.g. as the ratio of compound (IV) to the compound (VIII) which is the nondesired diastereomer in the context of a quinagolide synthesis).

It is noted that the diastereoselective reduction using the metal borohydride (as described above) is described in the context of a specific enantiomer (e.g. that obtained via the earlier described kinetic resolution step). However, the diastereoselective reduction using metal borohydrides (e.g. lithium borohydride or sodium borohydride with additional metal salts) may also find application in the reduction of racemic mixtures of compounds of formula (III).

In alternative examples, but in a similar fashion, an enantiomerically enriched compound of formula (H’a) can be used to provide an enantiomerically enriched compound containing an octahydrobenzo[f]quinoline moiety. A general overview of such a method is illustrated in Figure 4. In particular, the general method shown in Figure 4 shows how enantiomerically enriched compounds of formula (H’a) can be used to provide enantiomerically enriched intermediates (I H’a), and (IV’a). Thus, the present disclosure further provides methods for preparing each one of the compounds (lll’a) and (IV’a) in an enantiomerically enriched form.

In such examples, there is provided a method of preparing a compound of formula (lll’a):

wherein R¹ and X- is as defined above for formula (III).

The method of preparing a compound of formula (lll’a) is illustrated on Figure 4 as “step b”. The method of preparing a compound of formula (lll’a) may comprise a step of reducing a compound of formula (H’a). The method may further comprise an acidification step to provide the iminium ion of formula (lll’a).

Suitable conditions for reducing a compound of formula (H’a) and/or subsequent acidification may be those as described above in relation to the preparation of a compound of formula (HI) from a compound of formula (II).

In these examples, a compound of formula (H’a) may undergo a reduction followed by an acidic treatment to provide the iminium salt as shown in formula (lll’a). Compounds of formula (H’a) (which are present in an enantiomerically enriched form) selectively provide two diastereomers (a single enantiomer of each), as the stereochemistry of the pendant carboxylic acid group may have been essentially fixed by the enzymatic kinetic resolution step.

There is further provided a method of preparing a compound of formula (IV’a):

wherein R¹ and R⁶ are as defined for formula (IV).

In these examples, the method for preparing a compound of formula (IV’a) may comprise reacting and/or converting a compound of formula (I H’a). In particular, the method may comprise reducing the iminium ion of formula (lll’a). The method may further comprise acidifying and/or esterifying the reduced iminium ion to provide the compound of formula (IV’a). The method of preparing a compound of formula (IV’a) is illustrated on Figure 4 as “step c”.

Suitable conditions for reducing the iminium ion of formula (lll’a) and/or acidifying and/or esterifying the reduced iminium ion to provide a compound of formula (IV’a) may be as described above in relation to the conversion of a compound of formula (III) to a compound of formula (IV). In some examples, the step may proceed diastereoselectively to favour the formation of intermediate (IV’a).

Whilst compounds of formula (Vl’a) may have utility in the manufacture of other compounds containing an octahydrobenzo[f]quinoline moiety and compounds of formula (VIII) may have utility in the manufacture of other compounds containing an octahydrobenzo[g]quinoline moiety (in particular those comprising a substituent at the 3- position), it is compounds of formula (IV) that are of interest in the synthesis of quinagolide and its derivatives. In particular, where the stereocentre of position 3 has been effectively fixed by the earlier kinetic resolution step and the resultant iminium compound has been subjected to a diastereoselective reduction, it is noted that compounds of formula (IV) have the correct absolute and relative stereochemistry to provide the active enantiomer of quinagolide (when subjected to further manufacturing steps e.g. such as those shown in Figure 2). Thus, the provision of compounds of formula (IV) in a substantially enantiopure or enantiomerically enriched form can facilitate the manufacture of quinagolide in a substantially enantiopure or enantiomerically enriched form.

There is further provided a method of preparing a compound of formula (V):

Wherein R¹ is as defined in formulae (I) and (II) (or as defined for R^1A in formulae (la) and (Ila));

R⁶ is as defined for formula (IV) (such as C1-C3 alkyl, e.g. methyl); and

Y is a counterion (such as tosylate).

By way of example, the method may comprise forming a salt of the amine of formula (IV) (e.g. an acid addition salt), wherein the salt is a compound of formula (V).

Suitable acid addition salts may be formed with organic carboxylic acids such as acetic, lactic, tartaric, maleic, citric, pyruvic, oxalic, fumaric, oxaloacetic, isethionic, lactobionic and succinic acids; organic sulfonic acids such as methanesulfonic, ethanesulfonic, benzenesulfonic and p-toluenesulfonic acids and inorganic acids such as hydrochloric, sulfuric, phosphoric and sulfamic acids.

For example, the method may comprise forming a p-toluenesulfonate salt (e.g. wherein

Y is p-toluenesulfonate). The step of forming the acid addition salt may assist in providing a compound of formula (V) in a diastereomerically enriched form.

The disclosure further provides a method of providing a compound of formula (VI):

Wherein R¹ is as defined in formula (I) (or as defined for R^1A in formulae (la) and (Ila)); R⁶ is as defined in formula (VI) (such as C1-C3 alkyl, e.g. methyl); and

R⁷ is selected from optionally substituted Ci-Ce alkyl. In some examples, R⁷ may be propyl (e.g. n-propyl).

In particular, the method for providing a compound of formula (VI) may comprise derivatising and/or reacting the amine group on the octahydrobenzo[g]quinoline moiety of a compound of formula (V). For example, the method may comprise alkylating the amine group on a compound of formula (V) to provide a compound of formula (VI).

Such a method is illustrated by step e on Figure 3.

The disclosure further provides a method of preparing a compound of formula (VII):

wherein R¹ is as is defined in formulae (I) and (II) (or as defined for R^1A in formulae (la) and (Ila)); and

R⁷ is as defined in formula (VI).

The method may comprise epimerizing a compound of formula (VI) to provide a compound of formula (VII).

The epimerization step may be used to convert the pendant ester group (CO2R⁶) to the desired stereochemistry (e.g. a pendant methyl ester group to the desired stereochemistry).

As used herein, an epimer is one of a pair of diastereomers, wherein the pair of diastereomers has an opposite stereochemistry at only one stereocentre (e.g. in formula (VII), there is an opposite stereochemistry at only one stereocentre out of the three stereocentres). As used herein, “epimerization” may mean a reaction or chemical transformation in which an epimer is converted into another. By way of example only, an epimer of formula (VI) may be converted into an epimer of formula (VII). Such a step is illustrated as step f in Figure 3.

In view of the above, the skilled person will appreciate that compounds of formula (III), (IV), (V), (VI) and (VII) may be provided in an enantiomerically enriched form from compounds of formula (II) that are themselves provided in an enantiomerically enriched form.

As explained previously, such methods and intermediates are especially useful in the synthesis of quinagolide. In particular, an example synthesis of an enantiomerically enriched form of quinagolide is illustrated in Figure 6. In particular, the synthesis involves the use of an enzymatically catalyzed hydrolysis of compound (la) to provide an enantiomerically enriched form of compound (Ila). Compound (Ila) may be further reacted and/or converted to an iminium compound of formula (Illa) by way of reduction and acidification steps. Compound (Illa) may then be reduced and esterified to provide a compound of formula (IVa). It is noted that the reduction of compound (Illa) may proceed diastereoselectively to favour a compound of formula (IVa) (which has the absolute stereochemistry shown in this figure). Alternatively, in some cases, where the undesired diastereomer is also obtained, this can be removed from the desired diastereomer using methods known in the art (e.g. chromatographic separation, crystallization etc).

Thus, the disclosed methods, which can be used to provide an enantiomerically enriched form of compound of (Ila), can be used to provide compounds of formulae (Illa) and (IVa) in an enantiomerically enriched form.

As illustrated on Figure 6, this enantiomerically enriched material can be carried through the rest of the known manufacturing procedure (such as is illustrated in Figure 2) to provide an enantiomerically enriched form of quinagolide, in particular the active enantiomer 3S, 4aS, 10aR-quinagolide. In some cases, the described methods can be used to provide a substantially enantiopure form of this active enantiomer.

These steps are generally illustrated on Figure 6. Thus, the method of preparing quinagolide in an enantiomerically enriched form may further comprise: formation of a tosylate salt (step d), alkylation of an amino group (step e), an epimerization of the pendant methyl ester group (step f), hydrazide formation (step g), formation of a quaternary ammonium salt (step h), sulfonamide formation (step i), a demethylation (step j), and optionally the formation of a hydrochloride salt (step k).

In particular, a method of preparing quinagolide in an enantiomerically enriched form may further comprise reacting and/or converting the compound according to formula (Vila) to provide an enantiomerically enriched form of (3S,4aS,10aR)-quinagolide using the following steps and/or transformations:

(i) converting and/or reacting the methyl ester of compound (Vila) to form a hydrazide of formula (IXa):

(ii) converting and/or reacting the hydrazide of formula (IXa) to form a quaternary ammonium salt of formula

(iii) converting and/or reacting the quaternary ammonium salt of formula (Xa) to form a sulfonamide salt of formula (Xia)

(iv) demethylating the methoxy group of the sulfonamide salt compound of formula (Xia) to provide (3S,4aS,10aR)-quinagolide in an enantiomerically enriched form.

The method may further converting the enantiomerically enriched (3S,4aS,10aR)- quinagolide into a hydrochloride salt.

As will be appreciated by the skilled person, these steps may be carried out using reagents and procedures known in the art. By way of example only, suitable reagents and conditions are illustrated in Figure 1 and may also be found in Banziger et al Organic Process Research & Development, 2000, 4, 460, the contents of which are herein incorporated by reference.

According to further aspects there are provided intermediates that may be useful in the synthesis of compounds comprising an octahydrobenzo[g]quinoline core in an enantiomerically enriched form.

These intermediates may be or comprise any of the compounds described herein.

In particular, these intermediates may be compounds of formulae (I) and (II) as described herein. By way of example only, there is provided compounds having the following structures in an enantiomerically enriched form:

According to a further aspect of the disclosure there is provided a method for screening for suitable enzymes for use in preparing a compound of formula (II’) (e.g. a compound of formula (II) or formula (I I’a)) (or for use in preparing a compound of formula (I’), (I) or (I’a)) in an enantiomerically enriched form.

In particular, the described screening methods may be utilised to identify enzymes that can promote, facilitate and/or catalyse the hydrolysis of a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) to a compound of formula (II’) (e.g. a compound of formula (II) or (I I’a)). In other words, the methods may be used to identify enzymes that can facilitate and/or promote the hydrolysis in a kinetic resolution process.

The method may comprise contacting a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) with a candidate enzyme. The compound of formula (I’) may be contacted with a candidate enzyme under such conditions that allow a hydrolysis reaction to take place (e.g. including any of the conditions (e.g. solvents, temperatures, pH etc) described above in relation to the earlier aspects of the disclosure). The method may comprise determining if the candidate enzyme promotes, facilitates and/or catalyses the conversion (e.g. hydrolysis) of a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) to a compound of formula (II’) (e.g. a compound of formula (II) or (H’a)).. The method may comprise detecting and/or monitoring a level of starting material (e.g. a compound of formula (I’)) and/or a level of product (e.g. a compound of formula (II’)) over a period of time.

Where a level of starting material decreases over the period of time and/or the level of product increases over the period of time, the candidate enzyme may be determined to be suitable for use in preparing a compound of formula (II’) (e.g. a compound of formula (II) or (H’a)) or a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) by way of the described methods. By way of example only, where the level of product is determined to be at least about 10%, at least about 20%, at least about 30%, at least about 40% or about 50% conversion from the starting material, the candidate enzyme may be determined to be suitable for use in preparing a compound of formula (II’) (e.g. a compound of formula (II) or (H’a)) or a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) by way of the described methods.

Additionally or alternatively, the method may comprise determining if the candidate enzyme promotes, facilitates and/or catalyses the conversion in an enantioselective manner.

The method may comprise detecting a level of enantiomeric excess of the product (e.g. a compound of formula (H’) (e.g. a compound of formula (II) or (H’a)) and/or the starting material (e.g. a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)). Where the level of enantiomeric excess that is detected is greater than or equal to a reference level, the candidate enzyme may be determined to be suitable for use in preparing a compound of formula (H’) (e.g. a compound of formula (II) or (H’a) or a compound of formula (I’) (e.g. a compound of formula (I) or (I’a)) in an enantiomerically enriched form.

In other words, the method may comprise detecting a level of enantiomeric excess of the product (e.g. a compound of formula (H’)) and/or the starting material (e.g. a compound of formula (I’)). Where the level of enantiomeric excess that is detected is greater than or equal to a reference level, the candidate enzyme may be determined to be suitable for use in preparing a compound of formula (H’) (e.g. a compound of formula (II) or (H’a) or a compound of formula (I’) in an enantiomerically enriched form. By way of example only, the reference level may be greater than or equal to about 5% (e.g. greater than or equal to about 10%, greater than or equal to about 20%, greater than or equal to about 50%, greater than or equal to about 75%, greater than or equal to about 97%, or greater than or equal to about 99%.

The step of monitoring the reaction may be carried out using any methods that are known in the art e.g. NMR, LC-MS, HPLC, GC analysis.

Isotopically-labelled compounds

The disclosure also encompasses various deuterated forms of the compounds of any of the formulae disclosed herein, including formulae (I) to (XI) (including corresponding subgeneric formulae defined herein) or a pharmaceutically acceptable salt and/or a corresponding tautomer form thereof (including subgeneric formulae, as defined above) of the present disclosure. Each available hydrogen atom attached to a carbon atom may be independently replaced with a deuterium atom. A person of ordinary skill in the art will know how to synthesize deuterated forms of the compounds of any of the formulae disclosed herein, including those referred to above. For example, deuterated materials, such as alkyl groups may be prepared by conventional techniques (see for example: methyl-cfe -amine available from Aldrich Chemical Co., Milwaukee, Wl, Cat. No.489, 689- 2).

The disclosure also includes isotopically-labelled compounds which are identical to those recited in any of the formulae disclosed herein, including formulae (I) to (XI) (including corresponding subgeneric formulae defined herein) or a pharmaceutically acceptable salt and/or a corresponding tautomer form thereof (including subgeneric formulae, as defined above) of the present disclosure, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number most commonly found in nature. Examples of isotopes that can be incorporated into compounds of the disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, fluorine, iodine and chlorine such as ³H, ¹¹C, ¹⁴C, ¹⁸F, ¹²³l or ¹²⁵L Compounds of the present disclosure and pharmaceutically acceptable salts of said compounds that contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of the present disclosure. Isotopically labelled compounds of the present disclosure, for example those into which radioactive isotopes such as ³H or ¹⁴C have been incorporated, are useful in drug and/or substrate tissue distribution assays. Tritiated, i.e. ³H, and carbon-14, i.e. ¹⁴C, isotopes are particularly preferred for their ease of preparation and detectability. ¹¹C and ¹⁸F isotopes are particularly useful in PET (positron emission tomography).

Definitions

In the present disclosure, reference is made to a number of terms, which are to be understood to have the meanings provided below, unless a context indicates to the contrary. The nomenclature used herein for defining compounds, in particular the compounds described herein, is intended to be in accordance with the rules of the International Union of Pure and Applied Chemistry (IUPAC) for chemical compounds, specifically the “IUPAC Compendium of Chemical Terminology (Gold Book)” (see A. D. Jenkins et al., Pure & AppL Chem., 68, 2287-2311 (1996)). For the avoidance of doubt, if an IUPAC rule is contrary to a definition provided herein, the definition herein is to prevail.

As used herein, the term “alkyl” refers to a straight or branched chain hydrocarbyl group. The chain may be saturated or unsaturated, e.g. in some cases the chain may contain one or more double or triple bonds.

As used herein, “Ci-C_nalkyl” may be selected from straight or branched chain hydrocarbyl groups containing from 1 to n carbon atoms. For example, “Ci-Cealkyl” may be selected from straight or branched chain hydrocarbyl groups containing from 1 to 6 carbon atoms and Ci-Csalkyl may be selected from straight or branched chain hydrocarbyl groups containing from 1 to 3 carbon atoms. Representative examples are methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, isohexyl, neohexyl, etc. When a Ci-C_n alkyl group is substituted, any hydrogen atom(s), CH₃, CH₂ or CH group(s) may be replaced with the substituent(s), providing valencies are satisfied.

As used herein, an alkoxy refers to an alkyl group, as defined above, appended to the parent molecular moiety through an oxy group, -O-. As used herein, a Ci-C_nalkoxy refers to a Ci-C_n alkyl group (as defined above), appended to the parent molecular moiety through a oxy group, -O-, e.g. a Ci-Cealkoxy refers to a Ci-Cealkyl group (as defined above), appended to the parent molecular moiety through a oxy group, -O-. Representative examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy, 2-propoxy, butoxy, tert-butoxy, pentyloxy, hexyloxy etc. As used herein, “haloalkyl” may be an alkyl group as defined above, in which one or more hydrogen atoms thereon have been replaced with a halogen atom. By way of a representative example, a Ci-Ce haloalkyl may be a haloalkyl containing from 1 to 6 carbon atoms. The haloalkyl may be a fluoroalkyl, such as trifluoromethyl (-CF₃) or 1 , 1 - difluoroethyl (-CH₂CHF₂).

As used herein, a “halo” group may be F, Cl, Br, or I, typically F.

As used herein, the term "aryl" refers to a mono- or polycyclic aromatic hydrocarbon system having 6 to 14 carbon atoms, in some cases having 6 to 10 carbon atoms. Representative examples of suitable "aryl" groups include, but are not limited to, phenyl, biphenyl, naphthyl, 1 -naphthyl, 2-naphthyl and anthracenyl. As used herein, “substituted aryl” refers to an aryl group as defined herein which comprises one or more substituents on the aromatic ring. When an aryl group is substituted, any hydrogen atom(s) may be replaced with the substituent(s), providing valencies are satisfied.

As used herein, “heteroaryl” may be a single or fused ring system having one or more aromatic rings containing 1 or more O, N and/or S heteroatoms. The term “heteroaryl” may refer to a mono- or polycyclic heteroaromatic system having 5 to 10 ring atoms. A C_n-Cn heteroaryl is a heteroaryl containing n to n’ carbon atoms in the ring, where n and n’ are integers. Representative examples of heteroaryl groups may include, but are not limited to, pyrrolyl, furanyl, thiophenyl, pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, pyridinyl, pyrimidinyl, pyridazinyl, pyrazinyl, indolyl, benzofuranyl, benzothiazolyl, benzimidazolyl, indazolyl, benzoxazolyl, benzisoxazolyl etc. As used herein, “substituted heteroaryl” refers to a heteroaryl group as defined herein which comprises one or more substituents on the heteroaromatic ring.

As used herein, the term “optionally substituted” means that the moiety may comprise one or more substituents.

As used herein, a “substituent” may include, but is not limited to, hydroxyl, thiol, carboxyl, cyano (CN), nitro (NO₂), halo, haloalkyl (e.g. a Ci to Ce haloalkyl), an alkyl group (e.g. Ci to C or Ci to Ce), aryl (e.g. phenyl and substituted phenyl for example benzyl or benzoyl), alkoxy group (e.g. Ci to Ce alkoxy) or aryloxy (e.g. phenoxy and substituted phenoxy), thioether (e.g. Ci to Ce alkyl or aryl), keto (e.g. Ci to Ce keto), ester (e.g. Ci to Ce alkyl or aryl, which may be present as an oxyester or carbonylester on the substituted moiety), thioester (e.g. Ci to Ce alkyl or aryl), alkylene ester (such that attachment is on the alkylene group, rather than at the ester function which is optionally substituted with a Ci to Ce alkyl or aryl group), amine (including a five- or six-membered cyclic alkylene amine, further including a Ci to Ce alkyl amine or a Ci to Ce dialkyl amine which alkyl groups may be substituted with one or two hydroxyl groups), amido (e.g. which may be substituted with one or two Ci to Ce alkyl groups (including a carboxamide which is optionally substituted with one or two Ci to Ce alkyl groups), alkanol (e.g. Ci to Ce alkyl or aryl), or carboxylic acid (e.g. Ci to Ce alkyl or aryl), sulfoxide, sulfone, sulfonamide, and urethane (such as -O-C(O)-NR₂ or-N(R)-C(0)-0-R, wherein each R in this context is independently selected from Ci to Ce alkyl or aryl).

It should be noted that throughout this specification the term “comprising” is used to denote that embodiments of the invention “comprise” the noted features and as such, may also include other features. However, in the context of this invention, the term “comprising” may also encompass embodiments in which the invention “consists essentially of’ the relevant features or “consists of’ the relevant features.

As used herein, the term isomers may refer to compounds having the same number and type of atoms and hence the same molecular weight, but differing with respect to the arrangement or configuration of the atoms. It will be appreciated however, that some isomers or racemates or others mixtures of isomers may exhibit more activity than others. As used herein, stereoisomers may refer to isomers that differ only in the arrangement of the atoms in space. As used herein, diastereoisomer may refer to stereoisomers with two or more stereocentres that are not mirror images of each other. As used herein, the term enantiomer refers to stereoisomers that are non- superimposable mirror images of one another.

DETAILED DESCRIPTION

The present disclosure will now be described, by way of example only, with reference to the following Figures which show:

Figure 1 : The eight diastereomers of quinagolide including their absolute configurations.

Figure 2: Manufacturing route to quinagolide (in a racemic form).

Figure 3: General reaction scheme showing how compounds of formula (II)

(prepared in an enantiomerically enriched form) may be used to provide intermediates (III) to (VI) that have utility in the stereoselective synthesis of compounds comprising an octahydrobenzo[g]quinoline moiety.

Figure 4: General reaction scheme showing how compounds of formula (H’a) (prepared in an enantiomerically enriched form) may be used to provide intermediates (lll’a) to (IV’a) that have utility in the stereoselective synthesis of compounds comprising an octahydrobenzo[f]quinoline moiety.

Figure 5: Reaction scheme showing the two possible diastereomers that are obtained following a reduction (and subsequent esterification) of an iminium ion of formula (III).

Figure 6: General reaction scheme showing how compounds of formula (Ila)

(prepared in an enantiomerically enriched form) may be used to provide intermediates (Illa) to (Via) and also (3S, 4aS, 10aR)-quinagolide in an enantiomerically enriched form according to an example of the disclosure.

Figure 7a: Synthesis of racemic p-amino acid 7 and kinetic resolution of its methyl ester derivatives 8a, b according to an example of the disclosure.

Figure 7b: Formal asymmetric synthesis of (-)-quinagolide according to an example of the disclosure.

Figure 8: DFT (Density Functional Theory) study of thermodynamic equilibria of reactive species during borohydride reduction.

Experimental methods and results

Materials and Methods

All reagents were purchased from commercially available sources. Solvents for extraction and chromatography were distilled before use. Anhydrous MeOH was obtained by distillation on Mg turnings. Protease from Bacillus licheniformis (P4860 >2.4 U/g) has been purchased by Sigma-Aldrich. A particular kind of NaBH₄ powder (>98% purity) purchased by Sigma-Aldrich was used for entry 13 of Table 1. Analytical TLC were performed on silica gel on TLC Al foils (Sigma-Aldrich) with detection by exposure to ultraviolet light (254 nm) and/or by immersion in an acidic staining solution of vanillin in EtOH. Merck silica gel 60 (230-400 mesh) was used for flash chromatography. Semipreparative TLC were performed on Merck PLC silica gel 60. Molecular sieves AW- 300 (Alfa Aesar), 4 A (SigmaAldrich) used were activated under vacuum by heating with a drying pistol. All compounds have been characterized using one-dimensional and bidimensional NMR spectroscopic techniques, respectively ¹H, ¹³C and HSQC, HMBC, NOESY/ROESY and COSY. NMR spectra were recorded on Broker Avance II 400 spectrometer, dissolving the compounds in appropriate deuterated solvents. ¹H NMR spectra were recorded on Broker Avance II 400 MHz spectrometer. Chemical shifts are reported in ppm downfield from tetramethylsilane with the solvent resonance as the internal standard (chloroform-d: 5 7.26). Signal patterns are indicated as follows: br s, broad singlet; s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet. Coupling constants (J) are given in hertz (Hz). ¹³C NMR spectra were recorded at 101 MHz with complete proton decoupling. Chemical shifts are reported in ppm downfield from tetramethylsilane with the solvent resonance as the internal standard (chloroform-d: 5 77.16). Melting points were determined on a Kofler apparatus and are uncorrected. HRESIMS were acquired in positive ion mode with Orbitrap high-resolution mass spectrometer (Thermo, San Jose, CA, USA), equipped with H-ESI source. Analytical high performance liquid chromatography (HPLC) was performed on a Waters 600E equipped with Varian Prostar 325 detector using a Daicel® Chiralpak AD-H (250 X 4.6 mm) columns with detection at 220 nm.

Part A - Investigations into reaction conditions for the (asymmetric) reduction of cyclic iminium ion 2.

When looking to devise suitable manufacturing routes to provide quinagolide in an enantiomerically enriched form, initial efforts focussed on identifying a suitable set of reaction conditions to facilitate the asymmetric reduction of the cyclic iminium ion 2 (essentially corresponding to intermediate C8 as shown in the existing synthesis of quinagolide, see, for example, Figure 2). It was hoped that this route could provide the desired enantiomer with good levels of enantioselectivity.

The screening conditions that were employed for these investigations are illustrated below in scheme 1 .

(R)-RUCY®-XylBINAP

(6g)

(5.5)-6a: Ar = mesitylene, R = 4 (>S,5)-6b: Ar =p-cymene, R = 4-

(5.5)-6c: Ar =p-cymene, R = C₆

(S’, S)-6f: M = Ir

Scheme 1 - screening conditions for asymmetric reduction of cyclic iminium ion 2 (essentially corresponding to intermediate C8 as shown in the existing synthesis of quinagolide, see, for example, Figure 2).

The structures of compounds 3a, ent-3a, 3b and ent-3b are shown below.

The results of these screening reactions are shown in Table 1 below.

^a> Unless noted otherwise, all ATH (asymmetric transfer hydrogenation) reactions were carried out in MeOH with 6.0 equiv. of NFUCOOH or NaCOOH at 60 °C for 3.5 hours ^fa) NMR yield using 1 -bromo-3-chlorobenzene as internal standard. ^c) Values determined by HPLC on Daicel Chiralcel AD-H on the corresponding acetamide. ^d Reaction carried out at 2 bar of H2 at 60 °C in MeOH for 15h. ^e Reaction carried out in THF with methyl ester of compound 2 at 0° C for 1 h. ^f Technical grade NaBH4. ⁹ NaBH4 (granules >99.99% purity) in anhydrous MeOH. ^h NaBH4 powder (>98% purity), Aldrich SKU 452882. ' Conditions g + MgCl2 (20 mol%). / Conditions g + LiCI (20 mol%). ^k LiBH4 (1 .5 equiv) in anhydrous MeOH. Na = not applicable

Table 1 - Investigations into the reduction of cyclic iminium ion 2

In preliminary experiments, the reduction of iminium salt (compound 2) was carried out in the presence of Noyori’s ruthenium catalysts 6a-d (2 mol%) with ammonium formate (6 equivalents) as a hydrogen donor in MeOH at 60 °C (Table 1 , entries 1 -4). The reduction occurred with low to moderate diastereo- and enantioselectivity. The best diastereoselectivity (d.r = 3.5) was obtained with Ru-TsDpen catalyst 6b having p- cymene as ancillary ligand (entry 2).

A modest increase in the enantioselectivity was found using catalysts 6c, d (entries 3 and 4). Partially aromatized compound 5a and totally aromatized 5b were obtained as major products by means of expensive ruthenium phosphine catalyst 6g (entry 5). Rhodium and iridium catalysts 6e and 6f afforded a complex mixture of aromatized products (entries 6 and 7). The use of HCOONa in combination with catalyst 6b reduced both collateral aromatization and the diastereoselectivity of the reaction (entry 8). Conducting the reaction under 2 bar of hydrogen gas in the presence of catalyst 6b an equimolar mixture of racemic 3a and 3b was cleanly obtained without traces of aromatized products (entry 9). On the basis of these data it seems that metal -catalyzed asymmetric transfer hydrogenation (ATH) with HCOONH4 promoted two main competitive processes: the first is a partial re-aromatization and the second is the reduction of the iminium salt. In view of the known affinity of ruthenium for carboxylic acids, it was hypothesized that the two diastereoisomers of compound 2 could exhibit a divergent reactivity. Thus, it is plausible that the major trans diastereoisomer 3a is generated by intramolecular delivery of the hydride from the same side of the carboxylic acid functionality starting from (4aR*, 3R*)- 2, whereas diastereoisomeric (4aS*, 3R*)-2 undergoes a preferential aromatization. The inventors also considered the possibility to use the asymmetric reduction of the methyl ester of iminium salt 2 with (S)-CBS catalyst in combination with BH₃ (1.1 equivalents) (entry 10). The very low enantio- and diastereoselectivities together with the difficult attainment of the methyl ester of compound 2 discouraged the exploration of further chiral boranes.

As the yields and enantioselectivities initially obtained were relatively unsuitable for pharmaceutical development, efforts were concentrated towards a high yielding and diastereoselective reduction step. To this end, the reduction of iminium ion 2 with NaBH₄ as reported in the manufacturing process was investigated introducing variations in the reaction conditions.

Unexpectedly it was noted that the ratio of diastereoisomeric amino esters 3a/3b varied considerably using NaBH₄ of different quality. Whereas technical grade and highly pure (granules > 99.99% purity trace metal basis) afforded almost the same diastereoisomeric ratio (entries 11 and 12, respectively), the use of a particular type of NaBH₄with a purity > 98% gave a sharp increase of diastereoselectivity (dr = 9, entry 13). It is known from the manufacturer that this particular kind of NaBH₄ contains inter alia magnesium salts (such as magnesium carbonate in amount between 0.45% and 0.70% by weight) as contaminants

(https://www.sigmaaldrich.com/catalog/product/aldrich/452882?lang=it&region=IT).

Therefore, a reaction was carried out with NaBH₄ used in entry 12 in the presence of 5% of MgCh. This again provided a significant increase of diastereoselectivity (entry 14).

Even if it is known that the methanolysis of NaBH₄ and thus the formation of different reducing species is strongly influenced by its purity (R. E. Davis, J. A. Gottbrath, J. Am. Chem. Soc. 1962, 84, 895-898), the influence of the purity of NaBH₄ in the diastereoselective outcome of a reduction step was surprising. The inventors further found some variability of the d.r. in some reactions carried out in accordance with entries 13 and 14 using MeOH with different content of water. The effects of other additives were also screened. The presence of catalytic amount (20 mol%) of LiCI increased the “basal” diastereoselectivity of high purity NaBH₄ (entry 15). The use of LiBH₄ gave a slightly lower but more reproducible diastereoselectivity (entry 16).

The reaction was repeated several times also on gram scale with minimal variations of d.r. ranging from 4 to 5.7. It was remarkable that starting from a ca. 55/45 mixture of diastereoisomers of iminiun ion 2, compound 3a (ent-3a) was obtained with a moderate to good diastereoselectivity despite the full conversion of starting material.

To rationalize this behavior it was undertaken a DFT study of a putative equilibria between the two diastereoisomers of the free bases of compound C8 during borohydride reduction (see Figure 8). Without being bound by theory, although less stable than conjugated enamines D and F, the presence of tetrasubstituted enamine intermediate of type E could epimerize at least partially the 4a position and shift the equilibria towards the more reactive diastereoisomer F.

Thus, it was hypothesised that if intermediate C8 could be provided in a highly enantioenriched form, then a moderate-to-high diastereoselectivity for the reduction of iminium salt C8 (as illustrated in Figure 2 of the existing manufacturing method of quinagolide) could be a viable route to a novel asymmetric synthesis of (3S, 4aS, 10aF?)- (-)-quinagolide and/or provide the desired eutomer of quinagolide in an enantiomerically enriched form.

Further detail of the methods used in entries 11 to 15 of table 1 is provided below. References to C8 in the description below may be taken to represent compound 2 in the screening investigations.

Entry 11 : C8 (100 mg, 0,34 mmol) was added in a Schlenk-tube in argon atmosphere and dissolved in HPLC grade MeOH (2 mL). The solution in cooled to -75°C, technical grade NaBH₄ (1.5 eq, 0.51 mmol, 19.2 mg) was portion-wise added. The reaction was left under vigorous stirring for 2h at -70°C, then the cooling bath was removed and a solution of H2SC in MeOH (0.5 mL, 0.64 M) was dropwise added to the mixture at - 20°C, the reaction was then refluxed for 3.5h. The mixture was allowed to cool down to room temperature and then was quenched with water (3 mL), NasCOs was added to reach pH>7 and then the biphasic mixture was extracted with DCM (10 mLx3). The organic phase was dried over Na2SO₄, filtered and then the solvent were evaporated under reduced pressure affording the desired product as a dark yellow oil (crude 72.3 mg, dr: 60/40).

Entry 12: C8 (100 mg, 0,34 mmol) was added in a Schlenk-tube in argon atmosphere and dissolved in HPLC grade MeOH (2 mL). The solution in cooled to -75°C, NaBFU (99.99% purity)(1 .5 eq, 0.51 mmol, 19.2 mg) was portion-wise added. The reaction was left under vigorous stirring for 2h at -70°C, then the cooling bath was removed and a solution of H2SO4 in MeOH (0.5 mL, 0.64 M) was dropwise added to the mixture at - 20°C, the reaction was then refluxed for 3.5h. The mixture was allowed to cool down to room temperature and then was quenched with water (3 mL), NasCOs was added to reach pH>7 and then the biphasic mixture was extracted with DCM (10 mLx3). The organic phase was dried over NasSOzi, filtered and then the solvent were evaporated under reduced pressure affording the desired product as a dark yellow oil (crude 79 mg, dr: 60/40).

Entry 13: C8 (100 mg, 0,34 mmol) was added in a Schlenk-tube in argon atmosphere and dissolved in HPLC grade MeOH (2 mL). The solution in cooled to -75°C, NaBH4 (98% purity, Aldrich SKU 452882. )(1 .5 eq, 0.51 mmol, 19.2 mg) was portion-wise added. The reaction was left under vigorous stirring for 2h at -70°C, then the cooling bath was removed and a solution of H2SO4 in MeOH (0.5 mL, 0.64 M) was dropwise added to the mixture at -20°C, the reaction was then refluxed for 3.5h. The mixture was allowed to cool down to room temperature and then was quenched with water (3 mL), Na2COs was added to reach pH>7 and then the biphasic mixture was extracted with DCM (10 mLx3). The organic phase was dried over Na2SO4, filtered and then the solvent were evaporated under reduced pressure affording the desired product as a dark yellow oil (crude 76 mg, dr: 90/10).

Entry 14: C8 (100 mg, 0,34 mmol) was added in a Schlenk-tube in argon atmosphere and dissolved in HPLC grade MeOH (2 mL). The solution in cooled to -75°C, NaBH4 (99.99% purity)(1 .5 eq, 0.51 mmol, 19.2 mg) was mixed with dry MgCl2 (20 mol%, 6.47 mg) and portion-wise added. The reaction was left under vigorous stirring for 2h at -70°C, then the cooling bath was removed and a solution of H2SO4 in MeOH (0.5 mL, 0.64 M) was dropwise added to the mixture at -20°C, the reaction was then refluxed for 3.5h. The mixture was allowed to cool down to room temperature and then was quenched with water (3 mL), Na2COs was added to reach pH>7 and then the biphasic mixture was extracted with DCM (10 mLx3). The organic phase was dried over Na2SO4, filtered and then the solvent were evaporated under reduced pressure affording the desired product as a dark yellow oil (crude 82 mg, dr: 80/20).

Entry 15: C8 (200 mg, 0,68 mmol) was added in a Schlenk-tube in argon atmosphere and dissolved in HPLC grade MeOH (4 mL). The solution in cooled to -75°C, NaBFU (99.99% purity)( 1 .5 eq, 1 .02 mmol, 38.5 mg) was mixed with dry LiCI (20 mol%, 5.7 mg) was portion-wise added. The reaction was left under vigorous stirring for 2h at -70°C, then the cooling bath was removed and a solution of H2SO4 in MeOH (0.5 mL, 0.64 M) was dropwise added to the mixture at -20°C, the reaction was then refluxed for 3.5h. The mixture was allowed to cool down to room temperature and then was quenched with water (3 mL), NasCOs was added to reach pH>7 and then the biphasic mixture was extracted with DCM (10 mLx3). The organic phase was dried over NasSC , filtered and then the solvent were evaporated under reduced pressure affording the desired product as a dark yellow oil (crude 1 12 mg, dr: 80/20).

Part B - Investigations into hydrolytic kinetic resolution conditions using different enzymes

Following the identification of a number of suitable diastereoselective reduction conditions, attention was focussed on identifying a suitable set of conditions to provide an enantiomerically enriched precursor.

The preparation of enantiomerically pure 3-substituted p-amino acids is an active area of research due to their unique pharmacological effects and their utility as precursors for 3-substituted p-lactams and in preparation of biologically active natural and unnatural polypeptides (see, for example, Enantioselective Synthesis of p-Amino Acids; Juaristi, E., Soloshonok, V. A., Eds., 2nd ed.; Wiley-VCH: New York, 2005; Choi, E. et al. Tetrahedron: Asymmetry 2013, 24, 1449-1452; and Cardillo, G. et al. Chem. Soc. Rev. 1996, 25, 1 17-128).

Although a number of chemical and enzymatic methods are available for the enantioselective synthesis of p-amino acids and their derivatives (Weiner, B. et al. Chem. Soc. Rev. 2010, 39, 1656-1691 ) there is still a great demand for simple approaches for the preparation of enantiomerically pure derivatives with various substitution patterns.

The inventors particularly looked to investigate a deracemization process of racemic amino ester C6a, that is an immediate precursor of cyclic iminium salt C8 in the manufacturing process of racemic quinagolide (see, for example, Figure 2). In fact, the corresponding acid C6 in racemic form can be obtained from 1 ,6-dimethoxynaphthalene in 46% yields for three steps at pilot scale (see Figure 7a).

The intermediate C6a is a relatively complex substrate containing a functionalized naphthalene moiety which could impact the efficacy of a kinetic resolution process (e.g. an enzymatic kinetic resolution process). Therefore, investigations were carried out to identify whether it would be possible to hydrolyse a racemic ester (referred to herein as intermediate C6a) to provide an enantiomerically enriched (R)-C6 product as shown in scheme 2.

Scheme 2 - showing the screening of hydrolytic kinetic resolution conditions.

The results are shown in table 2 below.

^a Conversion calculated using HPLC on Luna C8 stationary phase. ^b Enantiomeric excess calculated on the acetamide of unreacted C6 methyl ester using Daicel Chiralcel AD-H column (eluant heptane/IPA 88/12, retention times were 17.5’ (major) 18.5 (minor). ^c Stereoselectivity factor evaluated using ln[(0-c)(1 -ee)]/ln[(0- c)(1 +ee)]. ^d ALCALASE enzyme, Bacillus licheniformis, Merck-Aldrich 126741. ^e Protease from Bacillus licheniformis > 2.4U/g, Aldrich P 4860. r.t. = room temperature; n.d. = not determined;

Table 2. Screening of enzymes for the kinetic resolution of racemic C6-methyl ester

Despite the presence of the methoxy functional groups on the racemic C6-methyl ester, it was unexpectedly identified that a hydrolytic kinetic resolution of C6a could provide the product (R)-C6 in good yield and with good levels of enantioselectivity using a variety of different enzymes.

On the basis of these data, the use of a protease from Bacillus licheniformis as reported in entries 1 1 and 15 was particularly useful in performing the hydrolysis of methyl ester C6a. The only difference between the two entries is the use of half amount of protease in entry 1 1 with respect to entry 15 (see experimental section for details). Therefore, the reaction was carried out on multigram scale using the condition reported in entry 15 and focusing on an efficient production of p-aminoacid (R)-C6 (39% yield, 97.4 % ee) that the inventors knew to have the required absolute configuration to obtain (3S, 4aS, 1 OaR)- (-)-quinagolide. Methyl 3-amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoate (C6a)

3-amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoic acid (10.0 g, 34.6 mmol) was suspended in MeOH (158.9 mL), a solution of H2SO4 (6.06 mL, 3.3 eq.) in MeOH (37.2 mL) was dropwise added. The obtained solution was refluxed for 3.5h, then MeOH was partially evaporated, water (80 mL) was added, and the mixture was basified to pH >7 using NasCOs. The ester was extracted using AcOEt (3 x 30 mL), the organic phase was washed with brine (2 x 20 mL), dried with NasSO4 and the organic solvent was evaporated affording the desired compound as a yellow oil (9.8 g, 32.1 mmol, yield 93%). ¹H NMR (400 MHz, CDCI₃) 6 7.97 (s, 1 H), 7.33 - 7.27 (m, 2H), 7.05 (s, 1 H), 6.67 (dd, J = 6.1 , 2.5 Hz, 1 H), 3.96 (s, 3H), 3.92 (s, 3H), 3.66 (s, 3H), 3.10 (dd, J = 13.2, 5.9 Hz, 1 H), 3.00 (dd, J = 13.2, 6.5 Hz, 1 H), 2.96 - 2.83 (m, 3H).

¹³C NMR (101 MHz, CDCh) 6 175.6, 156.8, 155.4, 135.1 , 128.0, 126.2, 123.8, 120.4, 119.0 105.0, 102.2, 55.5, 55.4, 51.6, 49.2, 43.5, 31.1.

Methodology for testing of enzymes for kinetic resolution

Entry 1 : Protease from bacillus amiloliquefaciens (see, for example, WO 2005/085462) C6-ester (100mg, 0.33mmol) was suspended in phosphate buffer pH=7 (4mL) at 37°C obtaining a white suspension, protease from bacillus amiloliquefaciens (8.4mL) was added slowly. The mixture was left to react at 37°C and sampled several times for 5 days, the maximum conversion was 10% circa. The reaction was filtered and the filtered solution (showing pH=6.5) was basified to pH=9.3, then was extracted with DCM, dried over Na2SO4, and the solvent was evaporated affording C6 ester (31.4mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (8.2mg).

Entry 2: Protease from bacillus species (see, for example, procedure described in WO 2005/085462)

C6-ester (100mg, 0.33mmol) was suspended in phosphate buffer pH=7 (4mL) at 37°C obtaining a white suspension, protease from bacillus species (0.42mL) was added slowly. The mixture was left to react at 40°C and sampled several times for 21 hours, the maximum conversion was 49% circa. The reaction was filtered and the filtered solution (showing pH=7.15) was basified to pH=9.7, then was extracted with DCM, dried over Na₂SC>4, and the solvent was evaporated affording C6-ester (44.1 mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (38.2mg).

Entry 3: Protease type XIII aspergillus Saitoi (see, for example, WO 2005/085462) C6-ester (100mg, 0.33mmol) was suspended in phosphate buffer pH=7 (5mL) at 37°C obtaining a white suspension, protease from bacillus species (100mg) was added slowly. The mixture was left to react at 37°C and sampled several times for 48 hours, the maximum conversion was <5%. The reaction was filtered and the filtered solution (showing pH=7.2) was basified to pH=9.3, then was extracted with AcOEt, dried over Na₂SC>4, and the solvent was evaporated affording C6-ester (38.3mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (2.2mg).

Entry 4: Lipase B Candida Antarctica (see, for example, Tetrahedron: Asymmetry 2015, 26, 325-332.

C6-ester (100mg, 0.33mmol) was suspended in hexane (2.06mL), H₂O (0.5 eq, 2.97 pL) was added and then Lipase B Candida antarctica (33mg) was added. The reaction was left to react at 45°C for 12 hours and sampled several times, because of the lack of solubility of the starting material in the solvent, 20% acetone was added to the mixture, the maximum conversion after 48 hours was <5%. The reaction was filtered and the filtered solution was dried, affording C6-ester in a rather complicated crude mixture (98.2mg).

Entry 5: Lipase A Candida Antarctica (see, for example, Tetraedron: Asymmetry 2015, 26, 325-332).

C6-ester (100mg, 0.33mmol) was suspended in CH₃CN (2.0mL), H₂O (0.5 eq, 2.97 pL) was added and then Lipase A Candida antarctica (33mg) was added. The reaction was left to react at 45°C for 5 days and sampled several times, the maximum conversion was <5%. %. The reaction was filtered and the filtered solution was dried, affording a complicated crude mixture (103. 2mg).

Entry 6: Lipase B Candida Antarctica (see, for example, Bioprocess and Biosystems Engineering 2020, 43, 605-613).

C6-ester (67mg, 0.17mmol) was solubilized in acetone (1.4mL), H₂O (10.5mL) was added and then Lipase B Candida antarctica (17.0mg) was added. The mixture was stirred for 24 hours at 37°C and sampled various times, the maximum conversion was 46%. The reaction was filtered and the filtered solution (showing pH=7.4) was basified to pH=10.3, then was extracted with AcOEt, dried over NasSC , and the solvent was evaporated affording C6-ester (42.8mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (39.4mg).

Entry 7: Lipase A Candida Antarctica (see, for example, Bioprocess and Biosystems Engineering 2020, 43, 605-613).

C6-ester (67mg, 0.17mmol) was solubilized in CH3CN (1.4mL), H2O (10.5mL) was added and then Lipase A Candida antarctica (17.0mg) was added. The mixture was stirred for 18 hours at 37°C and sampled various times, the maximum conversion was 58%. The reaction was filtered and the filtered solution (showing pH=7.4) was basified to pH=10.3, then was extracted with AcOEt, dried over NasSO^ and the solvent was evaporated affording C6-ester (41 ,3mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (36.4mg).

Entry 8: Trypsin tpck treated from bovine pancreas (see, for example, DE 3919029) C6-ester (152.7mg, 0.5mmol) was suspended in phosphate buffer pH=7 (3.2mL) at r.t. obtaining a white suspension, Trypsin tpck treated from bovine pancreas (33mg) was added. The mixture was left to react at r.t. and sampled several times for 3 days, the maximum conversion was 14%. The reaction was filtered and the filtered solution (showing pH=8.2) was basified to pH=10.1 , then was extracted with AcOEt, dried over NasSOzi, and the solvent was evaporated affording C6-ester (42.1 mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (31 .2mg).

Entry 9: Protease from bacillus streptomices griseus (see, for example, J. Am. Chem. Soc. 1987, 109, 2845-2846).

C6-ester (100mg, 0.33mmol) was suspended in carbonate buffer (22mL) pH=7 at r.t. obtaining a white suspension, Protease from bacillus streptomices griseus (15mg) was added. The mixture was left to react at r.t. and sampled several times for 52 hours, the maximum conversion was 40%. The reaction was filtered and the filtered solution (showing pH=8.1 ) was basified to pH=10.2, then was extracted with AcOEt, dried over NasSOzi, and the solvent was evaporated affording C6-ester (39.2mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (28.1 mg).

Entry 10: Alcalase from bacillus licheniformis (see, for example, Tetrahedron: Asymmetry 2015, 26, 638-643).

C6-ester (100mg, 0.33mmol) was suspended in phosphate buffer (2mL) pH=7 at 37°C obtaining a white suspension, Alcalase from bacillus licheniformis (111 mg of solution that corresponds to 10 mg of pure enzyme) was added. The mixture was left to react at 37°C and sampled several times for 24 hours, the maximum conversion was 47%. The reaction was filtered and the filtered solution (showing pH=7.4) was basified to pH=9.5, then was extracted with AcOEt, dried over NasSC , and the solvent was evaporated affording C6-ester (36.2mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (29.5mg).

Entry 11 : Protease from bacillus licheniformis (see, for example, WO 2005/085462) C6-ester (1 OOmg, 0.33mmol) was suspended in phosphate buffer (3.9mL) pH=7 at 37°C obtaining a white suspension, Protease from bacillus licheniformis (1 .4 mL) was added. The mixture was left to react at 37°C and sampled several times for 24 hours, the maximum conversion was 49%. The reaction was filtered and the filtered solution (showing pH=7.4) was basified to pH=10.1 , then was extracted with AcOEt, dried over NasSOzi, and the solvent was evaporated affording C6-ester (37.6mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (28.7mg).

Entry 12: Protease from bacillus amiloliquefaciens (see, for example, WO 2005/085462 and J. Basic Microbiol. 2016, 56, 138-152).

C6-ester (100mg, 0.33mmol) was suspended in phosphate buffer pH=9 (4mL) at 60°C obtaining a white suspension, protease from bovine pancreas type I (8.4mL) was added in portions. The mixture was left to react at 60°C and sampled after 6h, the maximum conversion was 50% circa. The reaction was filtered and the filtered solution (showing pH=7.3) was basified to pH=9.4, then was extracted with AcOEt, dried over NasSO^ and the solvent was evaporated affording C6-ester (36.2mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (39.8mg).

Entry 13: Protease from bovine pancreas Type I (see, for example, WO 2005/085462). C6-ester (100mg, 0.33mmol) was suspended in phosphate buffer pH=7 (5mL) at 37°C obtaining a white suspension, protease from bovine pancreas type I (1 OOmg) was added in portions. The mixture was left to react at 37°C and sampled after 6h, the maximum conversion was 50% circa. The reaction was filtered and the filtered solution (showing pH=7.3) was basified to pH=9.4, then was extracted with AcOEt, dried over NasSO^ and the solvent was evaporated affording C6 ester (36.2mg). From the filtration a white solid was recovered from the filter, NMR analysis showed to be C6 (39.8mg).

Entry 14: C6-ester (150mg, 0.5mmol) was suspended in acetone (0.2mL), then phosphate buffer pH=7 (1.1 mL) was added and then Lipase from Candida Rugosa (19mg) was added. The reaction was left to react at room temperature for 72 hours and conversions determined by HPLC was <10%.

Entry 15: (Details in Part C below)

Part C - Preparation of intermediates

An outline of the synthesis is illustrated in Figures 7a and 7b (the compound numbers in the procedures below correspond to the compound numbering shown in Figures 7a and 7b).

Methyl 3-amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoate (8a). 3- amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoic acid (10.0 g, 34.6 mmol) was suspended in MeOH (158.9 mL), a solution of H2SO4 (6.06 mL, 3.3 eq.) in MeOH (37.2 mL) was dropwise added. The obtained solution was refluxed for 3.5h, then MeOH was partially evaporated, water (80 mL) was added, and the mixture was basified to pH>7 using NasCOs. The ester was extracted using AcOEt (3 x 30 mL), the organic phase was washed with brine (2 x 20 mL), dried with NasSO4 and the organic solvent was evaporated affording the title compound as a yellow oil (9.8 g, 32.1 mmol, yield 93%). ¹H NMR (400 MHz, CDCI₃) 6 7.97 (s, 1 H), 7.33 - 7.27 (m, 2H), 7.05 (s, 1 H), 6.67 (dd, J = 6.1 , 2.5 Hz, 1 H), 3.96 (s, 3H), 3.92 (s, 3H), 3.66 (s, 3H), 3.10 (dd, J = 13.2, 5.9 Hz, 1 H), 3.00 (dd, J = 13.2, 6.5 Hz, 1 H), 2.96 - 2.83 (m, 3H). ¹³C NMR (101 MHz, CDCI3) 6 175.6, 156.8, 155.4, 135.1 , 128.0, 126.2, 123.8, 120.4, 1 19.0 105.0, 102.2, 55.5, 55.4, 51.6, 49.2, 43.5, 31.1.

(R)-3-amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoic acid (R)-7. (Also conditions for entry 15 in part B).

Racemic compound 8a (5.0 g, 16.38 mmol) was suspended in phosphate buffer pH=7 (195 mL) at 37°C and stirred for 10 minutes. Protease from Bacillus licheniformis (>2.4 U/g, 140 mL) was portion-wise added, and the mixture was left to react for 48h under vigorous stirring. The obtained suspension was filtered under vacuum affording a filtered dark yellow solution and a solid cake. The filtered solution was basified using NasCOs to pH=9 and extracted with AcOEt (3 x 25 mL). The organic phase was washed with brine (2 x 20 mL) and dried with Na2SO4. The organic solvent was evaporated affording (S)-8 (2.2 g, 7.2 mmol, yield 44%, ee > 99%, Daicel AD-H column, eluent heptane/IPA 88:12). The solid cake was suspended in a mixture of EtOH/H₂O (1 :1 ) (300 mL), LiOH*H₂O was added to pH=1 1 and the mixture was stirred overnight at 60°C. The suspension was filtered under vacuum, obtaining a brown slime and a filtered solution. The filtered solution was heated to 60°C and a solution of AcOH (1 :1 ) in EtOH/H₂O (1 :1 ) was dropwise added to pH=9-9.5. The mixture was stirred for 30 minutes, then AcOH in EtOH/H₂O was added to pH=8-8.5, and the mixture was cooled to room temperature. The pH was adjusted to pH=8-8.5, then the suspension was filtered under vacuum affording (R)-7 as a white solid [1.97g, 6.80 mmol, total yield 39%, single enantiomer (73% yield), ee= 97.4%, [a]²⁶= -38.4], ¹H NMR (400 MHz, MeOD) 5 8.03 (s, 1 H), 7.33 - 7.25 (m, 2H), 7.18 (s, 1 H), 6.73 (dd, J = 6.1 , 2.5 Hz, 1 H), 3.97 (s, 6H), 3.10 (dd, J = 13.2, 5.9 Hz, 1 H), 3.02-2.94 (m, 1 H), 2.93 - 2.81 (m, 3H). ¹³C NMR (101 MHz, MeOD) 5 169.2,

156.6, 155.2, 135.4, 135.2, 127.8, 125.9, 123.5, 120.3, 118.6, 104.8, 101.8, 54.4, 44.9,

40.6, 31.4.

(S)-3-amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoic acid [(S)-7J. Methyl (S)-3-amino-2-((3,8-dimethoxynaphthalen-2-yl)methyl)propanoate [(S)-8a] (1.53 g, 5.0 mmol) was dissolved in a mixture of EtOH/H₂O (1 :1 ) (28.0 mL) and LiOH*H₂O was added (4.3 eq, 902.15 mg), the reaction mixture was refluxed for 3h. The mixture was cooled to 60°C, and AcOH (1 :1 ) in EtOH/H₂O (1 :1 ) was dropwise added to pH=9-9.5. The mixture was stirred for 30 minutes, then AcOH in EtOH/H₂O was added to pH=8- 8.5, and the mixture was cooled to room temperature. The pH was adjusted to pH=8- 8.5, then the suspension was filtered under vacuum affording title compound as a white solid (1.17 g, 4.0 mmol, yield 80%). ¹H NMR (400 MHz, MeOD) 5 8.03 (s, 1 H), 7.33 - 7.25 (m, 2H), 7.18 (s, 1 H), 6.73 (dd, J = 6.1 , 2.5 Hz, 1 H), 3.97 (s, 6H), 3.10 (dd, J = 13.2, 5.9 Hz, 1 H), 3.02-2.94 (m, 1 H), 2.93 - 2.81 (m, 3H). ¹³C NMR (101 MHz, MeOD) 5 169.2,

40.6, 31.4.

(3R, 4aS,/?)-3-carboxy-6-methoxy-2,3,4,4a,5,10-hexahydrobenzo[g]quinolin-1-ium chloride (3H, 4aS,R)-(2). (R)-3-amino-2-((3,8-dimethoxynaphthalen-2- yl)methyl)propanoic acid [( R)-7] (1 .83 g, 6.34 mmol) was suspended in dry THE (18,334 mL, 10 Vol.) in argon atmosphere using mechanical stirring; dry t-BuOH (2.4 eq, 13.2 mmol, 1 .45 mL) was added and the mixture was cooled to -70°C. NH3 (9.167 mL, 5 Vol.) was condensed in a trap at -50°C then added to the reaction mixture. Lithium wires (5.2 eq, 32.97 mmol, 227.48 mg) in oil were dried and portion-wise added, after the addition, the mixture turned from white to dark green and after 2-3 minutes it turned dark blue. The reaction mixture is left to react at -70°C for 2.5h (if during this time there is a loss of color, extra lithium has to be added). NH₃ was evaporated overnight at room temperature, then the reaction was quenched with water (13.0 mL). The organic solvents were evaporated at 50°C and 100mbar, and the aqueous phase was dropwise added to a solution of concentrated HCI (4.6 mL) at 0°C. The obtained suspension was left to stir for 4. Oh, then it was filtered under vacuum, and washed with HCI 2N (2 mL) and water (2 mL x 2) affording title compound as a beige solid after drying for 48h at 50°C under vacuum (780 mg, 2.64 mmol, yield 42%). ¹H NMR (400 MHz, MeOD) 5 7.25 (t, J = 7.9 Hz, 1 H), 6.90 (dd, J = 8.3, 4.9 Hz, 1 H), 6.81 (t, J = 7.8 Hz, 1 H), 4.00 - 3.90 (m, 2H), 3.86 (s, 3H), 3.55 (t, J = 16.0 Hz, 1 H), 3.22 - 3.08 (m, 1 H), 2.67 (d, J = 16.3 Hz, OH), 2.61 - 2.51 (m, 1 H), 2.45 - 2.35 (m, 1 H), 2.08 (dd, J = 14.0, 4.2 Hz, 1 H), 1 .82 (dd, J = 13.8,

11.9 Hz, 1 H), 1.29 (s, 1 H). ¹³C NMR (101 MHz, MeOD) 5 193.8, 174.6, 158.0, 132.3,

129.4, 123.8, 121.1 , 1 10.0, 56.1 , 46.8, 46.8, 37.4, 35.9, 29.9, 28.2, 26.7.

Methyl (3/?,4a/?,10a/?)-6-methoxy-1 ,2,3,4,4a,5,10,10a-octahydrobenzo[g]quinoline- 3-carboxylate [(3/?,4a/?,10a/?-3a)].

Iminium chloride ([3R, 4aS,R)-(2) (700 mg, 2.36 mmol) was solubilized in dry MeOH (14 mL) in argon atmosphere and cooled to -70°C, and LiBH₄ (77.8 mg, 3.57 mmol) was portion-wise added. After 2.0h the mixture was heated to -20°C and a solution of H2SO4 in MeOH (2.64 M, 2.4 mL) was dropwise added. The reaction mixture was refluxed for 3.5h. Once cooled to room temperature, MeOH was partially evaporated and water (14 mL) was added. NasCOs was added to pH>7, and the biphasic mixture was extracted with AcOEt (15 mL x 3). The organic phase was dried with Na2SO₄ and the solvents were evaporated to afford the title compound as dark yellow oil (403 mg, 1.47 mmol, yield 62%, dr = 5.7). ¹H NMR (400 MHz, CDCI₃) 5 7.10 (t, J = 7.8 Hz, 1 H), 6.69 (dd, J = 17.2,

7.9 Hz, 2H), 3.81 (s, 3H), 3.69 (s, 3H), 3.40 - 3.34 (m, 1 H), 3.01 - 2.78 (m, 3H), 2.67 - 2.54 (m, 2H), 2.29 - 2.23 (m, 1 H), 2.22 - 2.12 (m, 1 H), 1 .45 - 1 .36 (m, 1 H). ¹³C NMR (101 MHz, CDCI₃) 5 174.1 , 155.4, 143.0, 135.1 , 125.8, 122.3, 121.7, 1 19.2, 118.1 , 107.0,

100.4, 55.4, 52.1 , 43.8, 38.6, 30.5.

(3H,4a/?,10afi)-6-methoxy-3-(rnethoxycarbonyl)-1 ,2, 3, 4, 4a, 5,10,10a- octahydrobenzo[g]quinolin-1-ium 4-methylbenzenesulfonate [(3H,4a/?,10aR)-9)].

(3R,4aR,10a/^:?)-(3a) (550 mg, 2.0 mmol) was solubilized in AcOEt (3.0 mL) at 70°C, and a solution of p-toluene sulphonic acid (378 mg, 2.19 mmol) in AcOEt (3.30 mL) was dropwise added. The obtained suspension was cooled to 0°C for 3. Oh and filtered under vacuum. The afforded purple solid was washed using cold AcOEt (3 mL) affording a light pink powder that was dried under vacuum (695 mg, 1.55 mmol, yield 78%, dr >10). ¹H NMR (400 MHz, CDCI3) 5 9.59 (d, J = 11 .9 Hz, 1 H), 8.90 (d, J = 1 1 .6 Hz, 1 H), 7.73 (d, J = 8.2 Hz, 2H), 7.15 (d, J = 7.9 Hz, 2H), 7.10 (t, J = 7.9 Hz, 1 H), 6.69 - 6.63 (m, 2H), 3.85 (d, J = 12.6 Hz, 1 H), 3.80 (s, 3H), 3.71 (s, 3H), 3.31 (dd, J = 15.7, 5.0 Hz, 1 H), 3.20 - 2.95 (m, 4H), 2.42 - 2.35 (m, 1 H), 2.33 (s, 3H), 2.20 (dd, J = 17.4, 11 .4 Hz, 1 H), 1 .44 - 1.33 (m, 1 H). ¹³C NMR (101 MHz, CDCI₃) 172.0, 156.8, 141.8, 140.5, 133.7, 129.2, 127.2, 125.8, 123.3, 121.4, 107.8, 58.8, 55.2, 52.4, 49.4, 45.3, 38.5, 32.7, 29.6, 21.5, 19.1.

Methyl (3R,4aR,10aR)-6-methoxy-1 -propyl-1 , 2, 3, 4, 4a, 5,10,10a- octahydrobenzo[g]quinoline-3-carboxylate [(3R,4aR,10aR)-10]. (3H,4aH,10aH)-(9)

(675 mg, 1.5 mmol) was suspended in DMF (6.5 mL), K2CO3 (384 mg, 3.0 mmol) was added in argon atmosphere. 1 -lodopropane (364 mL, 637.5 mg, 3.75 mmol) was dropwise added at 50°C, and the reaction mixture was left to stir for 4.0h. Toluene (14.5 mL) was added, then water (12.9 mL) was added to the mixture at 0°C. The biphasic mixture was extracted with toluene (15 mL x 3), dried with NasSC , and the organic solvents were evaporated affording title compound as a brown sticky oil (457 mg, 1.44 mmol, yield 96%, dr >10).

Methyl (3S,4aR,10aR)-6-methoxy-1 -propyl-1 , 2, 3, 4, 4a, 5,10,10a- octahydrobenzo[g]quinoline-3-carboxylate [(3S,4aR,10aR)-11)]. In a Schlenk at - 50°C, LDA (0.85 mL, 1 .7 mmol, 2M in THF) was added in argon atmosphere, a solution of (3F?,4aF?,10aF?)-10 (250 mg, 0.79 mmol) in THF (1.95 mL, 0.4M) was dropwise added, and the mixture was left to react for 40 minutes. Freshly distilled TMSCI (0.2 mL, 170 mg, 1.56 mmol) was dropwise added and the mixture was left to react for 1.0 h. The mixture was then slowly dropwise added to a solution of HCI 15% (2.44 mL) at -30°C and left to stir for 15 minutes. A saturated solution of NasCOs was added at -10°C and then the mixture was left to stir at room temperature for 1 ,0h. The biphasic mixture was extracted with toluene (10 mL x 3), the organic phase was dried with NasSC , and the solvents were evaporated affording crude [(3S,4aF?,10aF?)-11)] as a brown sticky oil (210 mg). The solid was purified by flash chromatography (Hex/AcOEt:8/2) affording a light yellow solid (105 mg, 0.32 mmol, yield 41%). A simple crystallization from MeOH afforded the title compound as single stereoisomer. [a]²⁰ _D= -137 (c = 0.5, CHCI3). The comparison of the sign of the optical rotatory power reported in the literature (S. P. Chavan, A. L. Kadam, R. G. Gonnade, Org. Let. 2019, 21, 9089-9094) established the absolute configuration of the final product. ¹H NMR (400 MHz, CDCI3) 6 7.08 (t, J = 7.9 Hz, 1 H), 6.71 (d, J = 7.7 Hz, 1 H), 6.64 (d, J = 8.1 Hz, 1 H), 3.81 (s, 3H), 3.69 (s, 3H), 3.45 (dt, J = 1 1.4, 2.4 Hz, 1 H), 3.11 (dd, J = 16.1 , 5.0 Hz, 1 H), 2.96 (dd, J= 17.3, 5.0 Hz, 1 H), 2.78 - 2.57 (m, 3H), 2.41 - 2.29 (m, 3H), 2.20 - 2.06 (m, 2H), 1 .92 - 1 .80 (m, 1 H), 1 .48 (q, J = 7.4 Hz, 2H), 1 .32 - 1 .27 (m, 1 H), 0.88 (t, J = 7.4 Hz, 3H). ¹³C NMR (101 MHz, CDCI3) 6 174.2, 156.5, 136.6, 125.9, 125.4, 121.1 , 106.6, 61.3, 55.0, 54.3, 53.4, 51.4, 39.8, 34.6, 34.0, 32.2, 30.5, 18.0, 11 .7. Although the present invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, the descriptions and examples should not be construed as limiting the scope of the invention. The disclosures of all patent and scientific literature cited herein are expressly incorporated herein in their entirety by reference.

Claims

1. A method for preparing a compound of formula (II’) in an enantiomerically enriched form:

R², R³ and R⁴ are each independently selected from H, optionally substituted Ci-Ce alkyl, optionally substituted CO(Ci-Ce alkyl), optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; and the stereochemistry at the starred position is (R).

2. The method of claim 1 , comprising preparing and/or obtaining a compound of formula (II’) from a compound of formula (I’):

wherein R¹, R², R³ and R⁴ are as defined for formula (II’); and

R⁵ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; wherein the stereochemistry at starred (*) position is (R) or (S).

3. The method according to claim 1 or 2, wherein the compound of formula (II’) comprises a structure according to formula (II) or formula (H’a):

(i) wherein formula (II) is:

wherein R¹

R², R³ and R⁴ are as defined in claim 1 ; and the stereochemistry at the starred position is (R); and (ii) wherein formula (H’a) is:

wherein R¹, R², R³, R⁴ and * are as defined in claim 1 ; and the stereochemistry at the starred position is (R).

4. The method of claim 3, comprising:

(i) preparing and/or obtaining a compound of formula (II) from a compound of formula (I):

wherein R¹, R², R³ and R⁴ are as defined for formula (II); and

R⁵ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; and the stereochemistry at the starred (*) position is (R) or (S), optionally wherein the compound of formula (I) is present in a racemic mixture; or

(ii) preparing and/or obtaining a compound of formula (H’a) from a compound of formula (I’a):

wherein R¹, R², R³, R⁴ and R⁵ are as defined in claim 2; and the stereochemistry at the starred (*) position is (R) or (S), optionally wherein the compound of formula (I’a) is present in a racemic mixture

5. The method of any one of the preceding claims, comprising contacting and/or reacting the compound of formula (I’), with one or more reagent(s) and/or under conditions that facilitate or promote the hydrolysis of a compound of formula (I’) to provide a compound of formula (II’), optionally comprising contacting and/or reacting the compound of formula (I) or (I’a), with one or more reagent(s) and/or under conditions that facilitate or promote the hydrolysis of a compound of formula (I) or (I’a) to provide a compound of formula (II) or (H’a).

6. The method of any one of the preceding claims, comprising a step of contacting and/or reacting the compound of formula (I’) with an enzyme, such as a hydrolytic enzyme to provide the compound of formula (II’), optionally comprising a step of contacting and/or reacting the compound of formula (I) or (I’a) with an enzyme, such as a hydrolytic enzyme to provide the compound of formula (II) or (H’a).

7. The method of any one of the preceding claims, wherein the method is a kinetic resolution process, optionally an enzymatic kinetic resolution.

8. The method according to any one of the preceding claims, wherein the compound of formula (II) is obtained or provided in an enantiomerically enriched form.

9. The method according to any one of the preceding claims, wherein the compound of formula (II’), and optionally formula (II) or (H’a), is provided or obtained in an enantiomeric excess (ee) of at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, or at least about 97%.

10. The method according to any one of claims 6 to 9, wherein the enzyme is selected from the group consisting of lipases and proteases.

11 . The method according to claim 10, wherein the enzyme is a protease.

12. The method according to claim 10 or 1 1 , wherein the enzyme is derived from Bacillus licheniformis, optionally wherein the enzyme is a protease derived from Bacillus licheniformis.

13. The method according to any one of the preceding claims, wherein the method is carried out in a solvent, wherein the solvent is or comprises an aqueous solution.

14. The method according to any one of the preceding claims, wherein the method is carried out in solvent that is or comprises a buffered solution.

15. The method according to any one of the preceding claims, wherein the method takes place at a temperature between about 10 °C and 100 °C, between about 20 °C and 50 °C, or between about 25 °C and 45 °C.

16. The method according to any one of the preceding claims, wherein the method takes place at a pH range between about pH 4 and pH 10, between about pH 5 and pH 8, or at about pH 7.

17. The method according to any one of the preceding claims, wherein, in compounds of formulae (I’), (I), (II’) and (II):

R¹ is absent or is selected from Ci-Ce alkyl, Ci-Ce alkoxy and halo;

R² is selected from Ci-Ce alkyl;

R³ and R⁴ are each H; and/or wherein, in compounds of formula (I’) or (I):

R⁵ is Ci-Ce alkyl.

18. The method according to any one of the preceding claims, wherein the compound of formula (II’) is:

wherein the stereochemistry at the starred position is (R).

19. A method of providing a compound comprising an octahydrobenzoquinoline moiety, optionally an octahydrobenzo[g]quinoline moiety (e.g. quinagolide), in an enantiomerically enriched form, the method comprising a step of preparing a compound of formula (II’), optionally of formula (II), according to any one of the preceding claims.

20. The method according to claim 19, further comprising a step of preparing a compound of formula (III) from a compound of formula (II), wherein a compound of formula (III) is:

wherein R¹ is as defined for formulae (I’), (I), (II’) and (II); and

X- is a counter ion; optionally wherein the method comprises reducing and then acidifying a compound of formula (II) to provide an iminium ion according to formula (III).

21. The method according to claim 20, further comprising a step of preparing a compound of formula (IV) from a compound of formula (III), wherein the compound of formula (IV) is:

wherein R¹ is as is defined for formulae (I’), (I), (II’) and (II); and

R⁶ is selected from H, optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; optionally wherein the method comprises reducing and then esterifying a compound of formula (III) to provide the compound of formula (IV).

22. A method for preparing a compound of formula (IV) in a diastereomerically enriched form:

the method comprising contacting a compound of formula (III) with a reducing agent selected from lithium borohydride and sodium borohydride, wherein when sodium borohydride is used as the reducing agent an additional metal salt is present;

wherein R¹ is as is defined for formula (I’), (I), (II’) and (II); and

R⁶ is selected from H, optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; and

X- is a counter ion.

23. The method according to claim 22, wherein the amount of reducing agent may be between about 1 and 10 equivalents, between about 1 and 8 equivalents, between about 1 and 5 equivalents or about 1 .5 equivalents based on the molar amount of the starting material e.g. the iminium compound of formula (III).

24. The method according to claim 22 or 23, wherein the reducing agent is lithium borohydride.

25. The method according to claim 22 or 23, wherein the reducing agent is sodium borohydride and the additional metal salt is selected from magnesium salts, optionally magnesium halides and magnesium carbonate, and lithium salts, optionally lithium halide.

26. The method according to any one of claims 22 to 25, wherein the method is carried out in an alcohol solvent (e.g. an anhydrous alcohol).

27. The method according to any one of claims 22 to 26, wherein the compound of formula (III) is obtained and/or provided in an enantiomerically enriched form (e.g. is prepared by a method as defined in any one of claims 1 to 20).

28. The method according to claim 19, further comprising a step of preparing a compound of formula (I I I’a) from a compound of formula (H’a), wherein a compound of formula (II I’a) is:

wherein R¹ is as defined in any preceding claim: and

X- is a counter ion; optionally wherein the method comprises reducing and then acidifying a compound of formula (H’a) to provide an iminium ion according to formula (lll’a), wherein formula (H’a) is:

wherein R¹, R², R³, R⁴ and * are as defined any preceding claim.

29. The method according to claim 28, further comprising a step of preparing a

wherein R¹ and R⁶ are as defined for formula (IV); optionally comprising reacting and/or converting a compound of formula (I H’a), further optionally by reducing the iminium ion of formula (I H’a) and acidifying and/or esterifying the reduced iminium ion to provide the compound of formula (IV’a).

30. The method according to any one of claims 21 to 27, further comprising a step of preparing a compound of formula (V) from a compound of formula (IV), wherein a compound of formula (V) is:

wherein R¹ is as defined in formulae (I’), (I), (II’) and (II);

R⁶ is as defined for formula (IV); and

Y is a counterion; optionally wherein the method comprises forming an acid addition salt of the compound of formula (IV) to provide the compound of formula (V).

31. The method according to claim 30, further comprising a step of preparing a compound of formula (VI) from a compound of formula (V), wherein the compound of formula (VI) is:

wherein R¹ is as defined in formulae (I) and (II);

R⁶ is as defined in formula (VI); and

R⁷ is selected from optionally substituted Ci-Ce alkyl; optionally wherein the method comprises alkylating (e.g. N-alkylating) the compound of formula (V) to provide the compound of formula (VI).

32. The method of claim 31 , further comprising a step of preparing a compound of formula (VII) from a compound of formula (VI), wherein the compound of formula (VII) is:

wherein R¹ is as is defined in formulae (I) and (II); and

R⁷ is as defined in formula (VI); optionally wherein the method comprises epimerizing a compound of formula (VI) to provide the compound of formula (VII).

33. The method of claim 32, wherein the method is for the synthesis of quinagolide and the method comprises preparing and/or obtaining a compound according to formula (Vila) in an enantiomerically enriched form:

34. The method of claim 33, the method further comprising reacting and/or converting the compound according to formula (Vila) to provide an enantiomerically enriched form of (3S,4aS,10aR)-quinagolide using the following steps and/or transformations:

(ii) converting and/or reacting the hydrazide of formula (IXa) to form a quaternary ammonium salt of formula (Xa)

35. The method of claim 34, further comprising converting the enantiomerically enriched (3S,4aS,1 OaR)-quinagolide into a hydrochloride salt.

36. Use of the method of any one of the preceding claims in the synthesis of an enantiomerically enriched form of (3S,4aS,10aR)-quinagolide.

37. A method for preparing a compound of formula (I) in an enantiomerically enriched form:

wherein R¹ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci- Ce alkoxy, optionally substituted Ci-Ce haloalkyl, halo, optionally substituted aryl, and optionally substituted heteroaryl, or R¹ is absent;

R², R³ and R⁴ are each independently selected from H, optionally substituted Ci-Ce alkyl, optionally substituted CO(Ci-Ce alkyl), optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; and

R⁵ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; wherein the stereochemistry at the starred position is (S).

38. A compound of formula (II) in an enantiomerically enriched form:

R², R³ and R⁴ are each independently selected from H, optionally substituted Ci-Ce alkyl, optionally substituted CO(Ci-Ce alkyl), optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; and the stereochemistry at the starred position is (R); optionally wherein the compound of formula (II) has the following structure:

39. A compound of formula (I) in an enantiomerically enriched form:

R⁵ is selected from optionally substituted Ci-Ce alkyl, optionally substituted Ci-Ce haloalkyl, optionally substituted aryl and optionally substituted heteroaryl; wherein the stereochemistry at the starred position is (S); optionally wherein the compound of formula (I) has the following structure:

40. A method for screening for suitable enzymes for use in preparing a compound of formula (II’) or formula (I’) in an enantiomerically enriched form, the method comprising: contacting a compound of formula (I’) with a candidate enzyme; determining if the candidate enzyme promotes, facilitates and/or catalyses the conversion (e.g. hydrolysis) of a compound of formula (I’) to a compound of formula (II’); and determining if the candidate enzyme promotes, facilitates and/or catalyses the conversion in an enantioselective manner, optionally wherein the compound of formula (II’) is a compound of formula (II) or (H’a); and/or the compound of formula (I’) is a compound of formula (I) or (I’a).