Synthesis of 2-Cyanobenzothiazoles via Pd-Catalyzed/Cu-Assisted C-H Functionalization/Intramolecular C-S Bond Formation from N-Arylcyanothioformamides

We report herein on a catalytic system involving palladium and copper to achieve the cyclization of N-arylcyanothioformamides and the synthesis of 2-cyanobenzothiazoles. The C-H functionalization/intramolecular C-S bond formation reaction was achieved in the presence of air, using 2.0 equiv of an inorganic additive (KI). In many cases, the reaction led to a sole product regioselectively obtained in good yields, allowing the synthesis of a wide range of substituted 2-cyanobenzothiazole derivatives, providing valuable building blocks for the design of more complex heterocyclic or molecular labeling systems.

All these studies have shown that the biological activities of benzothiazoles are highly dependent on the nature and position of their substituents. The most favorable positions are carbon C2, C5 and C6 of the benzothiazole skeleton, and the number of functional groups can vary from 1 to 3 and range from a simple chemical function to more complex aliphatic or heterocyclic systems [1][2][3][4][5][6][7][8][9][10][11][12][13]. It is important to note that the benzothiazoles showing significant biological activity are mainly substituted at the C2 position of the thiazole ring. In terms of antiproliferative activity, the most remarkable compounds are benzothiazole derivatives substituted by a nitrogen atom (e.g., amine, urea, hydrazone and semicarbazone), sulfur atom (e.g., sulfanyl derivatives), or substituted aromatic group or a hetero-aromatic group (e.g., thiazole, pyridine, imidazole, pyrazole and oxazole). All these efforts led to numerous innovative synthetic routes for preparing such compounds [14][15][16].
For the last 10 years, our group investigated the chemical application of Appel Salt and its 5-N-arylimino-4-chloro-1,2,3-dithiazole derivatives for fusing the 2-cyanobenzothiazole motif on pyrimidine or pyrimidinone systems, and synthesizing bioactive thiazoloquinazolines and quinazolinones, which are able to affect the activity of kinases involved in neurodegenerative diseases (Alzheimer's disease, Down's syndrome) and cancers [58][59][60][61]. Recently a new strategy in our molecular and biological targets incited us to develop more practical and efficient general synthetic protocols. It appeared relevant and useful to allow easy access to diversely substituted and functionalized 2-cyanobenzothiazole derivatives. The present study thoroughly investigates a convenient palladium-catalyzed and copper-assisted method for the synthesis of a large array of these compounds and improves upon the existing literature. It also aims at exploring the regioselectivity of the thiazole ring closure under the steric or electronic influence of substituents present on the starting anilines. For the first time, a reaction mechanism is suggested in adequation with the data obtained ( Figure 1).
The best result was obtained with 20 mol% of PdCl 2 and a starting molar concentration of 0.025 M in the solvent mixture. Under these conditions, the expected benzothiazole 4a was obtained with a 49% yield (entry 4). Expecting improvement, TBAB was replaced by tetrabutylammonium iodide (TBAI) and produced 4a a similar yield (51%).
In the preceding work [53,54], Doi and colleagues discovered that the addition of an inorganic additive such as CsF led to a significant improvement in the C-H functionalization/intramolecular C-S bond formation reaction from thiobenzanilides. We, therefore, replaced TBAB by 2 equiv of CsF, but this provided 4a in only a 15 % yield (entry 1 in Table 2). Based on these preliminary results, inorganic additives in place of TBAB or CsF were screened. Table 2 reports our results with various inorganic salts (2.0 equiv) added to the reaction mixture. The other reactants and solvent proportions remained unchanged. In a first attempt, based on the experimental conditions described by Doi [53,54] and Prescher [55,56]; 3a was solubilized in DMSO/DMF (1:1, v/v, [0.025 or 0. 050 M]) and heated at 120 °C for 4 h in the presence of 10 or 20 mol% of PdCl2, 50 mol% of CuI and 2 equiv of tetrabutylammonium bromide (TBAB), as depicted in Table 1.
The best result was obtained with 20 mol% of PdCl2 and a starting molar concentration of 0.025 M in the solvent mixture. Under these conditions, the expected benzothiazole 4a was obtained with a 49% yield (entry 4). Expecting improvement, TBAB was replaced by tetrabutylammonium iodide (TBAI) and produced 4a a similar yield (51%).
In the preceding work [53,54], Doi and colleagues discovered that the addition of an inorganic additive such as CsF led to a significant improvement in the C-H functionalization/intramolecular C-S bond formation reaction from thiobenzanilides. We, therefore, replaced TBAB by 2 equiv of CsF, but this provided 4a in only a 15 % yield (entry 1 in Table  2). Based on these preliminary results, inorganic additives in place of TBAB or CsF were screened. Table 2 reports our results with various inorganic salts (2.0 equiv) added to the reaction mixture. The other reactants and solvent proportions remained unchanged.  4 Replacing KI with a base of K 2 CO 3 or LiOtBu did not provide successful results. 5 Adding a ligand (50 mol%) including phenantroline or L-proline gave 0% and 7% yields of 4a, respectively.
Among the salts tested, cesium derivatives (CsF and CsI) were found to be the least effective additives (entries 1 and 2) while sodium, potassium and lithium salts produced good results, producing the desired 6-methyl-2-cyanobenzothiazole 4a in moderate to good yields (53-70%) (entries 3-5, 7 and 9), except in the case of KF and LiCl, which led to yields of 16 and 33%, respectively (entries 6 and 8). In our case, KI gave the best results leading to 4a with a 70% yield (entry 5). Atmospheric oxygen plays a crucial role since an inert atmosphere (argon) gave a lower yield of 23% (see footnote 2 for entry 5). Increasing the quantity of KI (3.0 equiv) also had a negative effect on the yield of the reaction, which fell to 39% (see footnote 3 for entry 5). It is noteworthy that in the optimizing experiments described by Doi et al. [53], LiBr gave the expected product in the same yield as that obtained with TBAB. In our case, LiBr allowed the synthesis of 4a in only a 53% yield (entry 7). Entry 10 confirms that the additive is required to obtain 4a in a good yield.
To complete the optimization of the reaction conditions, various sources of palladium and copper were also tested. Solvent and co-solvent were also investigated, as depicted in Table 3. Table 3. Optimization of the palladium and copper sources as well as the solvent.
fell to 39% (see footnote 3 for entry 5). It is noteworthy that in the optimizing experiments described by Doi et al. [53], LiBr gave the expected product in the same yield as that obtained with TBAB. In our case, LiBr allowed the synthesis of 4a in only a 53% yield (entry 7). Entry 10 confirms that the additive is required to obtain 4a in a good yield.
To complete the optimization of the reaction conditions, various sources of palladium and copper were also tested. Solvent and co-solvent were also investigated, as depicted in Table 3. None of the new conditions tested improved the reaction efficiency except PdBr2, which led to a similar yield to PdCl2 (entry 2). Using Pd(OAc)2 and Pd2dba3 drastically decreased the quantity of 4a obtained (22 and 7%, respectively) (entries 3 and 4). The absence of a palladium source in the reaction mixture gave no reaction (entry 5) while the lack of copper led to a lower yield (41% instead of 70%) (entry 9), confirming the need for these components in the chemical equation. Out of all the solvents tested, the initial None of the new conditions tested improved the reaction efficiency except PdBr 2 , which led to a similar yield to PdCl 2 (entry 2). Using Pd(OAc) 2 and Pd 2 dba 3 drastically decreased the quantity of 4a obtained (22 and 7%, respectively) (entries 3 and 4). The absence of a palladium source in the reaction mixture gave no reaction (entry 5) while the lack of copper led to a lower yield (41% instead of 70%) (entry 9), confirming the need for these components in the chemical equation. Out of all the solvents tested, the initial mixture of DMSO/DMF in equal proportions remained the best for this reaction (entry 1 compared to entries 10, 11 and 12). Table 4 also reports the results obtained when initial amounts of palladium chloride (PdCl 2 ) and copper iodide (CuI) were optimized, as well as the starting molar concentration [c] in the solvent mixture. It confirms that heating the starting cyanothioformamide 3a at 120 • C for 4 h in DMSO/DMF (1:1, v/v; [0. 025 M]), in the presence of 20 mol% PdCl 2 , 50 mol% CuI and 2 equiv of potassium iodide (KI), was the most efficient method for the synthesis of 4a (entry 1). In all cases, changing the initial concentration of PdCl2 or CuI, or the amount of solvent, led to lower yields (entries 2, 3, 4 and 5). mixture of DMSO/DMF in equal proportions remained the best for this reaction (entry 1 compared to entries 10, 11 and 12). Table 4 also reports the results obtained when initial amounts of palladium chloride (PdCl2) and copper iodide (CuI) were optimized, as well as the starting molar concentration [c] in the solvent mixture. It confirms that heating the starting cyanothioformamide 3a at 120 °C for 4 h in DMSO/DMF (1:1, v/v; [0. 025 M]), in the presence of 20 mol% PdCl2, 50 mol% CuI and 2 equiv of potassium iodide (KI), was the most efficient method for the synthesis of 4a (entry 1). In all cases, changing the initial concentration of PdCl2 or CuI, or the amount of solvent, led to lower yields (entries 2, 3, 4 and 5). Considering our experience investigating the role of microwaves in the thermal activation of chemical reactions [63], a series of tests were performed in a microwave reactor operating at atmospheric pressure. Compound 4a was synthesized by applying the same operating parameters (quantities of reagents, solvent, temperature) as those described above. The programmed temperature was controlled by an external infrared pyrometer, which allowed feedback control of the power input in the cavity. A TLC control showed operating at atmospheric pressure. Compound 4a was synthesized by applying the same operating parameters (quantities of reagents, solvent, temperature) as those described above. The programmed temperature was controlled by an external infrared pyrometer, which allowed feedback control of the power input in the cavity. A TLC control showed the disappearance of the reagents after 1 h of irradiation, and no change was observed on prolonged heating. Compound 4a was isolated in a lower yield (57%) than under the standard thermal conditions (70%).
With the optimized conditions identified, the scope of N-arylcyanothioformamides 3 was explored to generate a valuable array of variously substituted 2-cyanobenzothiazoles. As described above for 3a, all N-arylcyanothioformanilides 3 were obtained using a twostep procedure in which starting anilines 1 were stirred with Appel salt (1.1 equiv) and pyridine (2.0 equiv) in dichloromethane (DCM), at r.t. for 1 h, to give the corresponding imino-1,2,3-dithiazoles 2, which were then treated by 3 equiv Considering our experience investigating the role of microwaves in the therm vation of chemical reactions [63], a series of tests were performed in a microwave operating at atmospheric pressure. Compound 4a was synthesized by applying th operating parameters (quantities of reagents, solvent, temperature) as those de above. The programmed temperature was controlled by an external infrared pyr which allowed feedback control of the power input in the cavity. A TLC control the disappearance of the reagents after 1 h of irradiation, and no change was obse prolonged heating. Compound 4a was isolated in a lower yield (57%) than un standard thermal conditions (70%).
thiazole-2-carbonitriles 4l and 4i produced a more significant difference of 30 a respectively.
To increase the range of 2-cyanobenzothiazoles, derivatives di-substitute 5,6-, 4,5-and 4,6-positions (compounds 4q-4x) were prepared from the corres N-arylcyanothioformamides 3q-3x, difunctionalized in the 3,4-, 2,3-and 2,4 -p (Scheme 3). Di-substituted benzothiazoles in positions 5 and 6 were obtained in good (67 to excellent yields (e.g., 96 and 94% for 4q and 4r, respectively). In these cases, the uents were mainly activating groups while a bromide in p-position for the nitrog led to a decrease in the yield (71% for 4s compared with 96% for 4q). This res accordance with those described in Scheme 1 for 4a and 4e with yields of 70 a respectively. Microwave-assisted synthesis of 4q and 4s was also tested and co the results previously obtained for 4a.
The synthesis of 4,5-di-substituted 2-cyanobenzothiazoles (4v, 4w and 4x) p be more difficult and yields were lower than those obtained with the 5,6-disub compounds. However, these results are close to those obtained in the synthesis o thiazoles 4n-4p from cyanothioformanilides 3n-3p, which are substituted in po Di-substituted benzothiazoles in positions 5 and 6 were obtained in good (67% for 4t) to excellent yields (e.g., 96 and 94% for 4q and 4r, respectively). In these cases, the substituents were mainly activating groups while a bromide in p-position for the nitrogen atom led to a decrease in the yield (71% for 4s compared with 96% for 4q). This result is in accordance with those described in Scheme 1 for 4a and 4e with yields of 70 and 54%, respectively. Microwave-assisted synthesis of 4q and 4s was also tested and confirmed the results previously obtained for 4a.
The synthesis of 4,5-di-substituted 2-cyanobenzothiazoles (4v, 4w and 4x) proved to be more difficult and yields were lower than those obtained with the 5,6-disubstituted compounds. However, these results are close to those obtained in the synthesis of benzothiazoles 4n-4p from cyanothioformanilides 3n-3p, which are substituted in position 2. These results suggest a significant steric effect when substituents are in the C2 position of the reagent. This effect is apparently counterbalanced by the electron-donor effect of the substituents in the C4 position, as shown by the data obtained for the synthesis of 2-cynobenzothiazoles 4z and 4aa.
These results suggest a significant steric effect when substituents are in the C2 po the reagent. This effect is apparently counterbalanced by the electron-donor effe substituents in the C4 position, as shown by the data obtained for the synthesis of benzothiazoles 4z and 4aa.
Compounds 4ab and 4ac were obtained in good yields of 59 and 65%, resp In contrast, when the reaction was produced using dissymmetric cyanothioform on the C3 and C5 positions (3ad and 3ae), a mixture of regioisomers was obtain about 30% yield in both cases. The benzothiazoles 4ae' and 4ae'' were separated column chromatography and isolated in 11 and 24% yields. However, compoun and 4ad'' could not be separated regardless of the techniques used. The intramole S bond formation sequence predominantly occurred on the side of the electron-do stituent with a ratio of 2:1 to the other partner. Unfortunately, the developed failed to cyclize the cyanothioformamides disubstituted in C2 and C5 (3af and Scheme 4).
This result suggests that depending on the reagents, the size of the inorganic may influence the yield of this regiospecific reaction. In the case of 4a (Table 2), constraints were present and KI was more efficient than LiBr. In contrast, for the c oformamide 3af, steric hindrance prevented KI from playing its role in the reactio ertheless, this yield was still lower than the one previously obtained by our grou when compound 2af was subjected to microwave-assisted thermolysis at 150 ° methylpyrrolidinone (NMP) [51]. Notably, Prescher also observed the same d and finally heated 2,5-disubstituted cyanothioformamides at 170 °C in sulfolane t the attempted 4-bromo-7-methyl-benzothiazole-2-carbonitrile derivatives, also yields (10-20%) [32]. Compounds 4ab and 4ac were obtained in good yields of 59 and 65%, respectively. In contrast, when the reaction was produced using dissymmetric cyanothioformamides on the C3 and C5 positions (3ad and 3ae), a mixture of regioisomers was obtained in an about 30% yield in both cases. The benzothiazoles 4ae' and 4ae" were separated by flash column chromatography and isolated in 11 and 24% yields. However, compounds 4ad' and 4ad" could not be separated regardless of the techniques used. The intramolecular C-S bond formation sequence predominantly occurred on the side of the electron-donor substituent with a ratio of 2:1 to the other partner. Unfortunately, the developed method failed to cyclize the cyanothioformamides disubstituted in C2 and C5 (3af and 3ag in Scheme 4).
Cyanobenzothiazole-2-carbonitrile 4af was obtained in only a 22% yield when KI was replaced by 2.0 equiv of LiBr.
This result suggests that depending on the reagents, the size of the inorganic additive may influence the yield of this regiospecific reaction. In the case of 4a (Table 2), no steric constraints were present and KI was more efficient than LiBr. In contrast, for the cyanothioformamide 3af, steric hindrance prevented KI from playing its role in the reaction. Nevertheless, this yield was still lower than the one previously obtained by our group (58%) when compound 2af was subjected to microwave-assisted thermolysis at 150 • C in Nmethylpyrrolidinone (NMP) [51]. Notably, Prescher also observed the same drawback and finally heated 2,5-disubstituted cyanothioformamides at 170 • C in sulfolane to obtain the attempted 4-bromo-7-methyl-benzothiazole-2-carbonitrile derivatives, also in low yields (10-20%) [32].
Scheme 5 depicts the suggested mechanism, based on these results and the literature data [63,68]. Scheme 5 depicts the suggested mechanism, based on these results and the literature data [63,68].

Scheme 5. Suggested mechanism for synthesis of 2-cyanobenzothiazoles (4) from N-arylcyanothioformamides (3).
The reaction is most likely initiated by the base-assisted formation of the Cu (I) thioamidate (I), favoring the coordination of Pd (II) with the sulfur atom to form the intermediate (II). Then, a deprotonative metalation step occurs, forming a sterically hindered transition state driving the regioselectivity (III), to obtain the palladacycle (IV), which undergoes reductive elimination, leading to the desired 2-cyanobenzothiazole 4 and Pd (0) , which can be reoxidized by atmospheric oxygen.

General Information
All reagents were purchased from commercial suppliers and used without further purification. All reactions were monitored by thin-layer chromatography with aluminum plates (0.25 mm) precoated with silica gel 60 F254 (Merck KGaA, Darmstadt, Germany). Visualization was performed with UV light at a wavelength of 254 nm. Purifications were conducted with a flash column chromatography system (PuriFlash, Interchim, Montluçon, France) using stepwise gradients of petroleum ether (also called light petroleum) (PE) and dichloromethane (DCM) as the eluent. Melting points were measured with an SMP3 Melting Point instrument (STUART, Bibby Scientific Ltd., Roissy, France) with a precision of 1.5 °C. IR spectra were recorded with a Spectrum 100 Series FTIR spectrometer (Perki-nElmer, Villebon S/Yvette, France). Liquids and solids were investigated with a singlereflection attenuated total reflectance (ATR) accessory; the absorption bands are given in cm −1 . NMR spectra ( 1 H, 13 C and 19 F) were acquired at 295 K using an AVANCE 300 MHz spectrometer (Bruker, Wissembourg, France) at 300, 75 and 282 MHz. Coupling constant J was in Hz and chemical shifts were given in ppm. Mass (ESI, EI and field desorption (FD) were recorded with an LCP 1er XR spectrometer (WATERS, Guyancourt, France). Mass spectrometry was performed by the Mass Spectrometry Laboratory of the University of Rouen.
The reaction is most likely initiated by the base-assisted formation of the Cu (I) thioamidate (I), favoring the coordination of Pd (II) with the sulfur atom to form the intermediate (II). Then, a deprotonative metalation step occurs, forming a sterically hindered transition state driving the regioselectivity (III), to obtain the palladacycle (IV), which undergoes reductive elimination, leading to the desired 2-cyanobenzothiazole 4 and Pd (0) , which can be reoxidized by atmospheric oxygen.

General Information
All reagents were purchased from commercial suppliers and used without further purification. All reactions were monitored by thin-layer chromatography with aluminum plates (0.25 mm) precoated with silica gel 60 F254 (Merck KGaA, Darmstadt, Germany). Visualization was performed with UV light at a wavelength of 254 nm. Purifications were conducted with a flash column chromatography system (PuriFlash, Interchim, Montluçon, France) using stepwise gradients of petroleum ether (also called light petroleum) (PE) and dichloromethane (DCM) as the eluent. Melting points were measured with an SMP3 Melting Point instrument (STUART, Bibby Scientific Ltd., Roissy, France) with a precision of 1.5 • C. IR spectra were recorded with a Spectrum 100 Series FTIR spectrometer (PerkinElmer, Villebon S/Yvette, France). Liquids and solids were investigated with a single-reflection attenuated total reflectance (ATR) accessory; the absorption bands are given in cm −1 . NMR spectra ( 1 H, 13 C and 19 F) were acquired at 295 K using an AVANCE 300 MHz spectrometer (Bruker, Wissembourg, France) at 300, 75 and 282 MHz. Coupling constant J was in Hz and chemical shifts were given in ppm. Mass (ESI, EI and field desorption (FD) were recorded with an LCP 1er XR spectrometer (WATERS, Guyancourt, France). Mass spectrometry was performed by the Mass Spectrometry Laboratory of the University of Rouen.
Some compounds of the 4 series (4a, 4b, 4f, 4o, 4i, 4k, 4l, 4n, 4p, 4r, 4u and 4af) were randomly described in studies cited in this paper [44,47,48,51,57,69]. To complete data sometimes uneasy to find in the literature, all compounds 4 were fully characterized. The general procedure of their synthesis and physicochemical characterization are described below. 1 H NMR and 13 C NMR spectra of these products are available in the Supplementary Materials (Sections S12-S47

Conclusions
We have investigated reaction conditions involving palladium and copper to achieve the successful cyclization of cyanothioformamides (3), leading to benzothiazoles 4 substituted in various positions and bearing in position C2 the versatile carbonitrile function. In this process, the presence of 2.0 equiv of an inorganic additive such as KI proved to be essential for a better conversion. The presence of air was also found to be crucial to the reaction, allowing reoxidation of Pd 0 at the end of the process. In many cases, the selective C-H functionalization/C-S bond formation reactions were performed in good to very good yields, allowing a wide range of benzothiazole derivatives. In comparison with previous work, this synthetic route produced only one regioisomer, except in the case of unsymmetrical 3,5-disubstituted thioformanilides wherein steric effects due to substituents may influence the reaction outcome. Moreover, this work allowed the formation of an array of polyfunctionalized 2-cyanobenthiazoles, as building blocks for the construction of more complex heterocyclic systems or potent applications in molecular labeling.

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