Optimization of an Aminothiazine Ring Formation ... - ACS Publications

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Organic Process Research & Development

Optimization of an Aminothiazine Ring Formation: Integrating Modeling with Experiments to Maximize Yield by Minimizing Impurity Formation Amy C. DeBaillie,* Zhiwei Zhang, ‡

† ‡

†

Paul J. Jasper,

‡

Su Li,

‡

Michael McCulley,† Shankar Vaidyaraman,†

Eli Lilly and Company, Small Molecule Design and Development, Indianapolis, Indiana 46285 RES Group, Inc. 75 Second Avenue, Needham, MA 02494

*CORRESPONDING AUTHOR [email protected].

ACS Paragon Plus Environment

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Table of Contents Graphic:


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ABSTRACT

The aminothiazine formation step is a key transformation in the process to synthesize a potent and selective inhibitor of Beta-Amyloid Cleaving Enzyme (BACE). There are several impurities formed during the telescoped process that impacted the overall yield of the transformation. In order to improve the overall yield and design the impurity control strategy, a mechanistic model was developed to understand the impact of different process parameters on yield and impurity levels. This work describes how mechanistic models were integrated with experiments to determine process conditions to maximize the yield by minimizing impurity formation.

KEYWORDS Modeling, Impurity Formation, Aminothiazine, Impurity control strategy, Telescoped process


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INTRODUCTION LY2886721 (1)1 is a potent and selective inhibitor of Beta-Amyloid Cleaving Enzyme (BACE) and was in Phase 2 clinical trials as a potential treatment for Alzheimer’s disease.

The

development of the process to prepare kilogram quantities of 1 has recently been described (Scheme 1).2

Specifically, for the aminothiazine ring formation (Step 7) it was disclosed that

control of the benzoyl isothiocyanate (BzNCS) and carbonyl diimidazole (CDI) stoichiometry and in situ conversion monitoring were critical to minimize competing impurity formation. Herein, we describe how mechanistic models were integrated with experiments to optimize the process for this key step to synthesize aminothiazine 3 from amino alcohol 2•HCl (Step 7).

Scheme 1. Synthesis of BACE Inhibitor 1


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RESULTS AND DISCUSSION Our prior work 2 demonstrated that the best conditions for forming the aminothiazine ring from the amino alcohol (2•HCl) were through the formation of the thiourea (4) with BzNCS in tetrahydrofuran after treatment with triethylamine (Scheme 2; Step 7a).3 The aminothiazine (3) was then formed via activation of the thiourea with CDI to generate a CDI ester (5) that cyclized upon heating (Step 7b and Step 7c).4,5

Since attempts to isolate 4 via crystallization or

precipitation were unsuccessful, a two-step telescoped conversion process was selected for further development. Due to the complexity of the transformations and sensitivity to BzNCS and CDI stoichiometry, the use of mechanistic modeling6 was implemented to gain a better understanding of the overall reaction mechanism with the goal of maximizing the overall yield of the telescoped process by minimizing impurity formation. The impurities described in the following section were process based impurities. Impurities coming from reagents and 2•HCl were controlled in the specifications of 2•HCl, BzNCS, and CDI and were not included in the model. Additionally, the formation of 3 as the tosylate salt was not in scope of the research and was not included in the model at this stage of development. 7 Step 7a

Step 7b

H O F

H OH

OH 1. Et3 N

NH2 HCl

O F

N H

HO

S NHBz

CDI

NHAc 2

O F

2. BzNCS NHAc 4

Step 7c

O N S

N H

H

N

NHBz

∆

F NHAc

5

S

O N

NHCOPh TsOH NHAc 3

Scheme 2. Preferred Method for Forming Aminothiazine 3

1.

Key Impurities and Proposed Mechanisms of Formation 1.1.

Step 7a: Thiourea Formation

Development of the thiourea formation step identified the impurities shown in Figure 1 as reaction based impurities.


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H

OH

O

O

O Ph

N H

F

H

H O

N

F NHAc

O

O Ph

N H

F

O

NHAc

NHAc

6

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8

7

S H OBz S O

N H

F

H O

NHBz

H

S NHBz

O

N H

F NHAc

9

NHBz

O

O N

F NHAc

10

H

NBz

NHAc 11

Figure 1. Identified Impurities From Step 7a

Impurities 6 and 7 are related to each other in that 7 is the product from the reaction of 6 with CDI. Impurity 6 formed from attack of 2 at the less electrophilic carbon of BzNCS.8,9 Impurity 8 formed during Steps 7b-7c (Scheme 2) from the reaction of unreacted 2 with CDI. Experimental results confirmed that if the freebasing step was slow relative to the reaction time to form the thiourea, then the thiourea (4) was present in a large excess of BzNCS to form impurities 9 and 10 (Scheme 3).10,11 Through the use of in-situ NMR, it was confirmed that impurity 10 converted to the aminooxazine impurity (11), presumably via the intermediacy of 12 (Scheme 3).12


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H OH

H OH NH 2 HCl

O F

BzNCS Et 3N, THF

O

N H

F

NHAc

H OBz S

S NHBz

O excess BzNCS

N H

F

NHAc

NHAc

2

NHBz

4

9

excess BzNCS

S H O

S

NHBz

H OH

S O

N H

F

O

NHBz

H S

N

NHBz

O

O

NHBz

F

N

F

NHAc

NHAc

10

H

NBz

NHAc

12

11

Scheme 3. Proposed Mechanism for the Formation of Impurities from Excess BzNCS

1.2.

Steps 7b And 7c: Aminothiazine Formation

As previously described,2 CDI was selected as the reagent of choice for the cyclization of 4 to the desired aminothiazine ring via the formation of CDI ester 5 (Scheme 2). Two impurities were known to form during this step, impurity 11 and impurity 13 (Figure 2). As noted in section 1.1, impurity 11 is also formed in Step 7a, and for the purpose of the model, it was assumed that the majority of impurity 11 was formed during Step 7a.13 Therefore, the formation and minimization of impurity 13 was the main focus for this step. H

H O

O N H

F

NBz

O NBz

N

F

N N

NHAc 11

NHAc 13

Figure 2. Known Impurities that Form During Steps 7b-7c

Impurity 13 formed when excess CDI was charged after step 7a had been deemed complete by HPLC. For example, when 1.1 equiv of CDI was added, less than 0.5% 13 was observed by HPLC analysis. When 2.0 equiv of CDI was charged, 5.4 area % 13 was formed;


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when 5.0 equiv of CDI was charged, 21 area % of 13 was formed. These results led to the mechanism proposed in Scheme 4.

Scheme 4. Proposed Mechanism for the Formation of Impurity 13

The main reaction (Scheme 2) and the mechanisms of formation of the reaction based impurities discussed in this section have been combined into a schematic representation of the overall system (Figure 3). It should be noted that there were other impurities formed during Steps 7a-7c that are less than 0.2 area % that are described as “unknowns” in the following sections. These impurities were not investigated individually since they did not have any quality implications on the isolated product and were not deemed as significant process impurities. Additionally, the decomposition of BzNCS in the presence of water and triethylamine to form Nbenzoylbenzamide (16) was added to the model to account for residual water that was present in the system.14 The next section discusses the development and calibration of a mechanistic model for this system (Steps 7a-7c).


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2.

Model Development and Calibration

Figure 3: Components, Phases and Reactions in the Mechanistic Model for Steps 7a-7c

As described, this is a complex system due to the high number of impurities formed during the process and the high number of process parameters impacting their formation. A mechanistic model (Figure 3) was needed to understand how the process parameters impacted the overall yield and impurity formation. The calibration of model parameters was done by breaking the system into smaller sub-systems and conducting targeted experiments to calibrate the subsystems. These results were then combined into a model covering all the reactions. The data used for model calibration was species concentration (mol/L, mole fraction) obtained from HPLC data.15 The weighted least squares approach was used to estimate model parameters by minimizing the weighted squares of residuals between experimental and predicted data (equal


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weighting for all data and responses was used). The model calibration (parameter estimation) was performed in the J2 software package by RES Group, Inc.16 2.1.

Step 7a: Freebasing Step

The parameters in the slurry-to-slurry freebasing reaction are: 1. Solubilities for (2•HCl), (2), and triethylamine hydrochloride (Et3N HCl) 2. pKa difference between Et3N and 2•HCl (pKa_diff) 3. Rate of freebasing reaction 4. Mass transfer between solid and liquid The measured solubilities are shown in Table 1. Since the measured solubility of 2•HCl is very low, and the pKa difference in the THF reaction system could be different from the aqueous pKas used for 2•HCl and triethylamine (Et3N), the solubility of 2•HCl was set to 0.01mg/g, and experimental data was used to calibrate pKa_diff. The experimental data and model prediction with calibrated pKa_diff are shown in Figure 4. The estimated pKa_diff was 0.96 (which was significantly lower than aqueous pKa_diff of 4.43).17 The rate of freebasing reaction was estimated using fifteen minutes as the amount of time to reach equilibrium.18

Table 1. Solubility data for 2•HCl, 2 and Et3N HCl

Figure 4. Concentration of 2 During Step 7a


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2.2.

Step 7a: Thiourea Formation

Experiments were conducted to calibrate the model parameters for Step 7a to specifically include the formation of impurities 9, 10, 11, the decomposition of BzNCS, and to account for minimal amounts of aminothiazine (3) formed during Step 7a.19 (Figure 5).

Figure 5. Sub-mechanism for Step 7a

The experiments to understand and calibrate the impact of Et3N were conducted by reacting one equiv of BzNCS and one equiv of 4 with 0.1, 0.6, and 2 equiv of Et3N at 50 °C with the freebasing occurring at 50 °C. Model calibration results for 4, 3, 9, 10, 11 and BzNCS are shown in Figure 6. A first order dependence of Et3N dependence on the formation of 9 and 10 did not give a good fit to the data. As a result, the fitted order of Et3N dependence for formation of 9 was 1.2 and formation of 10 was 1.3.


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Figure 6: Formation of Step 7a Related Impurities: Comparison of Experimental Data with Model Fitted Data

Two alternative mechanisms (one used Et3N as a catalyst for the BzNCS reaction with water and the other did not use Et3N as a catalyst) were investigated for the reaction of BzNCS with water (Table 2).


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Index

Reaction

Rate Expression

1-1

BzNCS + H2O=> Benzamide

k6[BzNCS][H2O]

1-2

BzNCS + H2O=> Benzamide (Et3N catalyst)

k6[BzNCS][H2O][Et3N]

2

BzNCS + Benzamide => N-benzoylbenzamide (Et3N k7[BzNCS][Benzamide][Et3N] catalyst)

Without With Et3N at Et3N as catalyst catalyst √

√ √

√

√

Table 2: BzNCS Decomposition Pathways

The mechanism with Et3N as a catalyst for the BzNCS reaction with water was chosen since the calibration of BzNCS decomposition using the mechanism with Et3N gave a better fit than a mechanism without Et3N as a catalyst (Figure 7).

Figure 7: Calibration of BzNCS Decomposition Pathways With and Without Using Et3N as a Catalyst

2.3.

Calibration of Remaining Model Parameters

The experiments in Table 3 were used to calibrate the remaining model parameters (parameters that were not part of freebasing reaction, impurity forming reactions from 4, or BzNCS decomposition).


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Experiment Initial Baseline

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Freebasing

Process parameters for experiments Condensation Alcohol Activation

Et3N: 1.1 equiv

BzNCS: 1 equiv over 60min

CDI: 1 equiv over 20min

Hold 30 min

Hold 30 min CDI: 0.04 equiv Hold 30 min

Hold 990 min

CDI: 1 equiv in < 1 min

50 C->reflux

o

o

20 C to 50 C in 30 min

Initial calibration expt 1

Et3N: 1.1 equiv 20 C to 50 C in 15 min

Hold 180 min


Et3N: 1.1 equiv

BzNCS: 1 equiv over 60 min

20 C to 0 C

Hold 240 min


Et3N: 1.1 equiv

BzNCS: 1 equiv over 60min

o

o

o

o

o

Cyclization o o 50 C to 60 C over 30min

BzNCS: 1 equiv over 60 min

o

20 C to 50 C in 206 min Hold 30 min Hold 30 min Hold 7 min Partially Optimized 20oC to 51.7oC in 30 min BzNCS: 1.02 equiv over 84min CDI: 1.02 equiv in 3 shots every 20 min Hold 75 min Hold 74 min Et3N: 1.05 equiv

o

Hold 427 min o

o

51.7 C to 68.1 C over 30 min Hold 410 min

Hold 246 min

Experiment Initial Baseline

2

Species Measured Experimentally 4 5 3 6 10 9 8 13

7

X

X

X

X


X

X

X

X

X


X

X

X

X

X

X

X


X

X

X

X

X

X

X

X

X

X

X

X

X

Partially Optimized X

X

X

X

Table 3: Experiments Used for Model Calibration

The calibration results for the four selected experiments are shown in Figures 8 through 11.


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Figure 8: Model Fit vs Experimental Data (Initial Baseline)

Figure 9: Model Fit vs Experimental Data (initial calibration expt 3)

Figure 10: Model Fit vs Experimental Data (initial calibration experiment 1)


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Figure 11: Model Fit vs Experimental Data (initial calibration experiment 2)

2.4.

Model Testing with a Partially Optimized Experiment

A partially optimized experiment with conditions shown in Table 4 was conducted to use as a test data set to check the model prediction. The model parameters were further tuned to fit the data from this experiment.

Table 4: Partially Optimized Experiment

Figure 12 shows the calibration results of the main products in the partially optimized experiment. The model shows a reasonable match to the experimental data. In Step 7a, the model slightly underpredicted the formation of 4, which is possibly due to experimental error in sampling the reaction since it is a slurry (Et3N HCl is not in solution).


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Figure 12: Calibration Results of the Main Products in the Partially Optimized Experiment. * Means in the Condensation Step, Unfiltered Data is Used.

Figure 13 shows the calibration results for the impurities in the partially optimized experiment. A reasonable match to the experimental data for impurity formation was also acquired. The decrease of 10 in Step 7b and Step 7c is captured in the model with 10 converting to 11.


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Figure 13: Calibration Results of the Impurities in the Partially Optimized Experiment

With the partially optimized experiment, the full mechanism is confirmed to be consistent with the previous work. The main products of Steps 7a-7c and most of the impurities fit the data well. In Step 7b and Step 7c, “unknown” species are considered to be impurities from 5. This is considered to be a conservative handling for impurity amounts. The calibrated parameters are listed in Table 5. The temperature dependence of reaction rates were estimated only for certain reactions (where activation energy shown in Table 5 is greater than zero) where there were more opportunities to get a better impurity profile by changing temperature. The next section describes how the model was used to optimize the process by determining conditions to maximize the yield by minimizing impurity formation. Since the role of the model in this work was to find a better operating point and not test boundaries of failure, the current model was considered adequate to use for determining an operating point to meet the


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overall goal. The search for an optimal point took into consideration the parameter space covered by experiments. For example, since most of the experiments focused on higher temperatures (50 °C for Step 7a, 60-68 °C for Step 7c), the search for an optimum point was restricted to higher

temperatures. The next phase in model development would have been to conduct more experiments to investigate a broader space of parameters to refine the model so that it could be used for determining the critical process parameters and design space.

Index

Reaction

k@50°C*

E** [kJ/mole]

Solubility Model 21

2•HCl _solid 2•HCl

kla21*V*(C2.HCl_LIMIT-[2•HCl])

1.00E+00

0

22

2•HCl 2 + HCl

kr22(Ka2.HCl[2•HCl]-[2][HCl])

5.00E+08

0

23

2 2_solid

kla23*V*([2]- C2_LIMIT)

1.00E+00

0

24

Et3N HCl Et3N + HCl

kr24(KaEt3NH[Et3N HCl][Et3N][HCl])

1.00E+08

0

25

Et3N HCl Et3N HCl_solid

kla25*V*([Et3N HCl]- CEt3N HCl_LIMIT)

1.00E+00

0

Product Formation 2

2 + BzNCS => 4

k2[2][BzNCS]

4.50E+00

3

4 + CDI =>5 + Imidazole

k3[4][CDI]

3.14E+00

4

5 => 3 + Imidazole + CO2

k4[5]

2.03E+10

35

4 + BzNCS + Et3N => Inter+ Et3N HCl

K35[4][BzNCS][Et3N]

2.30E-02

36

Inter=>3

K36[Inter]

2.50E-02

20 0 80 0 0

BzNCS Decomposition 26

BzNCS + H2O=> Benzamide (Et3N catalyst)

K26[BzNCS][H2O][Et3N]

2.60E-02

0

37

BzNCS + H2O=> Benzamide

K37[BzNCS][H2O]

8.76E-04

0

27

BzNCS + Benzamide => N-benzoylbenzamide (Et3N catalyst)

K27[BzNCS][Benzamide][Et3N] 2.00E+00

0

k9[CDI][H2O]

0

CDI Decomposition 9

CDI + H2O => 2 Imidazole + CO2

7.93E-02


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Impurities from 2 – 6, 7 & 8 7 2 + BzNCS => 6 + HNCS

k [2][BzNCS]

3.50E-02

22

8

6 + CDI => CDI ester of 6

k [6][CDI]

2.13E+00

0

16

CDI ester of 6=> 7 + 2 Imidazole + CO2

k [CDI ester of 6]

4.02E-03

0

14 2 + CDI => 8 Impurities from 4 – 9, 10 & 13

k [2][CDI]

40.0E+01

0

17

K [4][BzNCS][Et3N]

1.20E-01

29

K [4][BzNCS][Et3N]

6.25E-02

12

28 10 + Et3N => 11 + Et3N HCl Impurities from 5 – 14/15 13 & Unknown 5 5 + CDI => 14/15 + 2 Imidazole + CO2

K [10][Et3N]

1.40E-01

0

k [5][CDI]

3.90E-03

0

6 13

k [14/15][Imidazole]

1.14E+02 2.20E-04

0 0

7 8

16 14

1.3

4 + BzNCS + Et3N => 10 + Et3N HCl

17

1.2

19

4 + BzNCS + Et3N => 9 + Et3N HCl

14/15 + Imi => 13 + CSO + Imidazole 5 => Unknown

19 28

5 6

k13[5]

* rate constants all used base unit of mole, L and min,**Activation energy of zero does not imply that the reaction rate is not dependent on temperature but that it was not calibrated.

Table 5: Calibrated Model Parameters

3.

Process Optimization

The mechanistic model enabled simulation of different scenarios of process parameters to determine conditions to optimize the overall yield by minimizing impurities. Table 6 compares the initial baseline conditions prior to developing the model, the partially optimized conditions used for final model calibration and the model suggested optimum conditions. This shows that there is potential to have a 5% increase in the in-situ yield at 50 ºC. The model suggested conditions deviate from the original conditions by decreasing Et3N equiv, increasing BzNCS equiv, increasing BzNCS addition time, increasing Step 7a reaction time , increasing Step 7c reaction temperature, and reducing Step 7c reaction time. The optimized conditions were determined by a two stage optimization procedure: 1. Determine conditions that maximize the overall yield of Step 7a at 50 ºC. a. Optimize Et3N and BzNCS equiv b. Optimize BzNCS addition time and reaction time with fixed Et3N and BzNCS equiv from step (a). 2. Determine conditions that maximize product yield from Step 7b-Step 7c


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a. With parameters fixed from step (1), optimize CDI equiv, Step 7b reaction time and Step 7c reaction temperature b. Optimize THF volume with other parameters fixed from previous steps.

Table 6: Comparison of Initial Baseline, Partially Optimized and Optimized Conditions

3.1.

Optimizing Et3N and BzNCS Equiv and BzNCS Addition Time

The molar fraction of 4 was simulated for different Et3N and BzNCS equiv at different BzNCS addition times. The Step 7a reaction time was set as 30 minutes for freebasing and 75 minutes for condensation time for these simulations. The overlay of regions with greater than 97% yield for different BzNCS addition times is shown in Figure 14. The optimized conditions reduced Et3N equiv and increased BzNCS equiv compared to the original conditions. The optimized conditions were set as: 1.05 equiv of Et3N and 1.15 equiv of BzNCS.


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Figure 14: Overlay of Regions of Greater Than 97% Yield as a Function of Et3N Equiv, BzNCS Equiv and BzNCS Addition Time

As BzNCS addition time increases, the Step 7a yield increases (Figure 15) and impurities 9 and 10 decrease (Figure 16). After 120 minutes of BzNCS addition time, there are no significant differences in the Step 7a yield. As a result, the BzNCS addition time was set to 120 minutes.

Figure 15: Effect of the BzNCS Addition Time on Step 7a Yield


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Figure 16: Effect of BzNCS Addition Time on the Formation of Impurities 9 and 10

The impact of Et3N equiv and BzNCS equiv on the formation of impurities 6, 9, 10, and 11 and the consumption of 2 is shown in Figure 17. This shows that an increase in Et3N and BzNCS equiv could increase the formation of impurities 9, 10, and 11 and lead to the consumption of 2. However, reducing Et3N equiv while increasing BzNCS equiv ensures that the impurity levels for 6, 9, and 11 will stay the same or decrease, while impurity 10 will slightly increase and the amount of unreacted 2 will increase.


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Figure 17: Impact of Et3N Equiv and BzNCS Equiv on Impurities 6, 9, 10 and 11

3.2.

Optimizing Step 7a Reaction Time

The change in molar fraction of Step 7a yield at different reaction times is shown in Figure 18. At approximately forty to sixty minutes of reaction time, the Step 7a yield reaches a maximum. Over sixty minutes, the Step 7a yield starts to decrease as 4 converts to impurities 9, 10, and 11. As a result, the Step 7a reaction time was set to sixty minutes.

Figure 18: Effect of Step 7a Reaction Time on Yield

The species molar fraction profile after Step 7a is shown in Figure 19 with 1.05 equiv of Et3N and 1.15 equiv of BzNCS. This figure represents thirty minutes of freebasing time, 120 minutes of BzNCS addition time, and sixty minutes of reaction time. The yield for Step 7a is predicted to be 97%.


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Figure 19: Species Molar Fraction Profile After Step 7a

3.3.

Optimizing Step 7b Reaction Time and Step 7c Reaction Temperature

Figure 20 shows the impact of CDI equiv on the formation of impurity 13. This suggests operating at a lower excess of CDI equiv (where formation of 13 is low and is not sensitive to time).

Figure 20: Sensitivity of Impurity 13 Formation to CDI Equiv

Hence, the CDI equiv were set to 1.02 equiv relative to the amount 4 so that the process operated in a region that reduces the formation of 13. The effect of Step 7b reaction time was also examined (Figure 21). It was concluded that a longer Step 7b reaction time will lead to a decrease in the overall yield. This conclusion is also supported by the graphs in Figure 22.


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Figure 21: Effect of Step 7b Reaction Time on the Overall Yield

An increase in Step 7b reaction time could increase the formation of impurity 13 and unknowns. This is a result of the fact that the formation of 3, impurity 13, and unknowns are from 5. These reactions are assumed to be in competition, and as the reaction activation energy of 5 to 3 is much higher than the other reactions to form impurities, an increase in temperature as soon as CDI is charged could favor the formation of 3.20 Hence, the optimized Step 7b reaction time was set to zero minutes and the Step 7c reaction temperature was set close to reflux (68ºC).


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Figure 22: Effect of Step 7b Reaction Time on Impurity Formation

3.4.

Optimizing THF Volume

Figure 23 shows a summary of the THF volume effect on the Step 7a-7c reactions. A significant decrease or increase in THF volume could decrease the overall yield (Figure 24).

An

explanation for a reduction of the overall yield at lower and higher THF volumes is as follows: a lower THF volume increases the concentration of 4, BzNCS, and Et3N, which leads to an increase in the amounts of impurities 9 and 10 that will form. A higher THF volume decreases the BzNCS concentration and decreases the reaction rate of Step 7a, which leads to an increase in the amount of unreacted 2 and subsequently an increase in the amount of impurity 8.


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Figure 23: THF Volume Effect

Figure 24: Illustration of Cause of Yield Loss at Low or High THF Volume

The model predicted that if the reaction volume was increased from the original conditions (9.5 volumes) to 1.5 to 2 times that this would lead to an increase in yield of approx. 2%. Since the predicted increase in yield did not warrant such an increase in the reaction volume and


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subsequent process mass intensity, it was decided to slightly increase the reaction volume to 10 volumes for the optimized conditions. 3.5.

Further Optimization Opportunities

Since more experiments were targeted at higher temperatures, the search for optimum conditions was restricted to 50 ºC for Step 7a. The thermodynamic model suggested that some operations could be optimized at lower temperatures (approx. 0 or 20 ºC). The model was used to explore these areas and limited experiments were performed as confirmation, but kinetic data at multiple temperatures was not obtained for most parameters. However, this would have been part of next phase of process optimization and was not investigated further in this work due to the discontinuation of this project. CONCLUSION In summary, a model to gain insight into the mechanism of formation of the aminothiazine and identified impurities was developed. The model was used to provide direction in the experimental program for achieving the desired yield of the aminothiazine by minimizing impurity formation. It was concluded that all of the sub-steps within Step 7 impact the overall yield and impurity profile with the freebasing of 2•HCl to 2 having the highest impact on the overall yield. This step is also equilibrium controlled and depends on the species (2•HCl, 2, triethylamine HCl) solubility and the pKa difference between triethylamine and 2•HCl. For the Step 7a reaction the optimal equivalents of Et3N and BzNCS have been identified for 50 °C. Increasing the addition time of BzNCS will increase the yield, but the benefit decreases after 120 min at 50 °C. For Step 7b and 7c, the CDI equiv and timing were optimized. An immediate temperature increase for cyclization is recommended after the addition of CDI. Lastly, the


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model suggests that diluting the overall reaction by 1.5 to 2 times the current volume will provide a slight increase in the yield of 3. The model predicts a 5% increase in the in-situ yield of 3 with the optimized conditions at 50 °C. AUTHOR INFORMATION PRESENT ADDRESS † Michael McCulley, Baxalta US Inc., 505 Baxter Parkway, Social Circle, GA 30025.21 AUTHOR CONTRIBUTIONS The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. SUPPORTING INFORMATION HPLC and data processing method and the Model calibration method.


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REFERENCES

1

Mergott, D. J.; Vaught, G. M., BACE Inhibitors, US 2012/8278441.

2

(a) Kolis, S. P.; Hansen, M. M.; Arslantas, E.; Brandli, L.; Buser, J.; DeBaillie, A. C.; Frederick, A. L.; Hoard, D. W.; Hollister, A.; Huber, D.; Kull, T.; Linder, R. L.; Martin, T. J.; Richey, R. N.; Stultz, A.; Waibel, M.; Ward, J. A.; Zamfir, A.; Zweifel, T. Org. Process Res. Dev. 2015, manuscript submitted. (b) Hansen, M. M.; Jarmer, D. J.; Arslantas, E.; DeBaillie, A. C.; Frederick, A. L.; Harding, M.; Hoard, D. W.; Hollister, A.; Huber, D.: Kolis, S. P.; KuehneWillmore, J. E.; Kull, T.; Laurila, M. E.; Linder, R. J.; Martin, T.; Martinelli, J. R.; McCulley, M. J.; Richey, R. N.; Ward, J. A.; Zaborenko, N.; Zweifel, T. Org. Process Res. Dev. 2015, manuscript submitted. 3

For literature conditions that would circumvent use of the benzoyl protecting group see: a) Herr, R. J.; Kuhler, J. L.; Meckler, H.; Opalka, C. J. Synthesis 2000, 1569. b) Gauthier, J.; Duceppe, J. S. J. Heterocyclic Chem. 1984, 21, 1081. 4

For the use of CDI to form an aminothiazine ring see: Bernacki, A. L.; Zhu, L.; Hennings, D. D. Org. Lett. 2010, 12, 5526.

5

For other methods to convert thioureas to aminothiazenes see: a) Xu, X.; Qian, X.; Li, Z.; Song, G.; Chen, W. J. Fluorine Chem. 2005, 126, 297. b) Klayman, D. L.; Woods, T. S. J. Org. Chem. 1975,40, 2000. c) Kim, T. H.; Cha, M.-H. Tetrahedron Lett. 1999, 40, 3125. d) Long, K.; Boyce, M.; Lin, H.; Yuan, J.; Ma, D. Bioorg. Med. Chem.Lett. 2005, 15, 3849. 6

The authors chose to use a mechanistic approach to understand this reaction instead of an empirical model due to the ability of the mechanistic approach to have a more sound extrapolation. The mechanistic model is ideal when there are multiple connected variables. See also: Hallow, D. M; Mudryk, B. M.; Bream, A. D.; Tabora, J. E.; Lyngberg, O. K.; Bergum, J. S.; Rossano, L. T.; Tummala, S. J Pharm Innov 2010, 5, 193.

7

The reactive crystallization to form the tosylate salt of 3 was not included in the model because impurity rejection was excellent and other development efforts were underway to modify the crystallization to improve isolation performance at scale. 8

For an example of non-regioselective attack of BzNCS see: Caille, S.; Boni, J.; Cox, G. B.; Faul, M. M.; Franco, P.; Khattabi, S.; Klingensmith, L. M.; Larrow, J. F.; Lee, J. K.; Martinelli, M. J.; Miller, L. M.; Moniz, G. A.; Sakai, K.; Tedrow, J. S.; Hansen, K. B. Org. Process Res. Dev. 2010, 14, 133.


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9

Additionally, another possible route for the formation of 6 is from benzoyl chloride present in the benzoyl isothiocyanate. Analysis of the lots of benzoyl isothiocyanate utilized for development and manufacture showed that benzoyl chloride was not present. 10

Impurity 10 could not be isolated, but was observed by in situ NMR.

11

The freebasing step of 2•HCl can not be a separate step. Step 7a is a slurry-to-slurry process. 2•HCl, 2, and triethylamine hydrochloride have incomplete solubility in 10 L/kg THF at 50 °C (the reaction conditions).

12

Interestingly, NMR showed that the imine double bond was exocyclic in oxazine 11, and endocyclic in the thiazine analogs. 13

The oxazine impurity (11) likely forms in Step 1c partitioning between alcohol activation and thiourea activation, similar to the pathway proposed from activation by excess BzNCS. Investigation of Step 7b and 7c with purified 4 led to undectable levels by HPLC of impurity 11 formed with CDI as the activating reagent (see ref 2b). 14

Typical reaction Karl Fischer measurements ranged from 0.021%w/w to 0.043%w/w with a specification of ≤ 0.1% w/w.

15

The conversion from HPLC area fraction to mole fraction using response factors and the overall model calibration workflow is described in the supplemental section.

16

J2, A Dynamic Modeling and Optimization Software, RES Group Inc. Needham, MA 2013. Website: http://www.resgroupinc.com 17

The aqueous pKa of 2 was obtained experimentally. The aqueous pKa for triethylamine was obtained from the Evans pKa table: http://evans.rc.fas.harvard.edu/pdf/evans_pKa_table.pdf.

18

It could have been shorter than fifteen minutes, but this was difficult experimentally.

to measure

19

The exact mechanism to explain the formation of 3 from 4 without the use of CDI is not known. Heating the Step 7a reaction mixture does not lead to the productive formation of 3.

20

It is possible to charge CDI at a higher temperature as a solution. For this process, it was decided to charge CDI as a solid charge. For safety reasons, it was preferred to charge CDI and then increase the temperature of the batch. 21

While currently affiliated with Baxalta US Inc., the research relates to prior affiliation at Eli Lilly and Company.


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Optimization of an Aminothiazine Ring Formation ... - ACS Publications

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