Synthesis of 1,4-Disubstituted Pyrazolo[3,4-d]pyrimidines from 4,6-Dichloropyrimidine-5-carboxaldehyde: Insights into Selectivity and Reactivity

Dedicated to Prof. Scott E. Denmark on the occasion of his 60^th birthday

Abstract

Strategies for carrying out the reaction of 4,6-dichloropyrimidine-5-carboxaldehyde with both aromatic and aliphatic hydrazines to generate 1-substituted 4-chloropyrazolo[3,4-d]pyrimidines in a selective, high-yielding, and operationally simple manner are presented. For aromatic hydrazines, the reaction is performed at a high temperature in the absence of an external base. For aliphatic hydrazines, the reaction proceeds at room temperature in the presence of an external base. The observed selectivity and reactivity­ trends are rationalized through consideration of the proposed­ reaction mechanism. The 1-substituted 4-chloropyrazolo[3,4-d]pyrimidine products serve as versatile synthetic intermediates, through further functionalization of the 4-chloride moiety, enabling the rapid generation of a structurally diverse array of 1,4-disubstituted pyrazolo[3,4-d]pyrimidines.

Pyrazolo[3,4-d]pyrimidines have attracted much attention in drug discovery programs. Because of their structural resemblance to purine nucleobases, they are ideally suited to selectively interact with a diverse array of pharmaceutically relevant targets. For example, pyrazolo[3,4-d]pyrimidine substrates have been established as inhibitors of PDE9 (Alzheimer’s disease, diabetes),[1] Src (osteoporosis),[2] p38α (inflammatory and autoimmune diseases),[3] MRP4 (resistance to anticancer drugs),[4] and ADA (ischemia);[5] as well as antagonists of adenosine A_2A (Parkinson’s disease)[6] and adenosine A₃ (inflammation, regulation of cell growth).[7] They have demonstrated both anticonvulsant[8] and antibacterial[9] activities. They have also been extensively exploited as inhibitors of oncogenic kinases, including Bcr,[10] Abl,[10] [11] mTOR,[12] PI3K,[12e–f] ^, [13] Src,[2] [11b] [14] EGF-R,[15] and CDK.[16] Pyrazolo[3,4-d]pyrimidines serve as excellent core structures for drug analogues because they offer multiple options for diversification, through functionalization of the 1-, 2-, 3-, 4-, or 6-position (Figure [1]).

As pyrazolo[3,4-d]pyrimidines are ubiquitous in the pharmaceutical industry, a number of different strategies have been employed for their synthesis. Most of these strategies, however, can be categorized into two disconnections (Scheme [1]). The most commonly cited synthetic route involves generation of the pyrazolo[3,4-d]pyrimidine ring system through the annulation of a carbonyl compound onto a 5-amino-4-cyanopyrazole, which in turn is synthesized from 2-(ethoxymethylene)malononitrile and a hydrazine (route A),[1a] [3] [5] ^, [7] [8] [9] ^, [12a] or some variation thereof.[1c] [2] [11a] ^, [14] [15] [16] The second most common synthetic route reverses the construction order of the two rings by adding a hydrazine onto an existing 4-chloropyrimidine-5-carboxaldehyde through a combined condensation/ nucleophilic­ aromatic substitution reaction (route B).[3] [4] [6] [7] [10] [12] [13] [17] In these cases the pyrimidine starting material is generally purchased or synthesized from another commercially available pyrimidine. Less common synthetic routes include a Diels–Alder reaction between 5-amino-1-phenyl-4-pyrazolecarboxylic acid and 1,3,5-triazines,[18] the cyclization of 4-aminopyrimidine-5-carboxaldehyde oximes,[19] and the reaction of 5-(benzo­yl­-amino)pyrazoles with nitriles.[20]

Recent interest in our laboratories has focused on the generation of a structurally diverse library of 1,4-disubstituted pyrazolo[3,4-d]pyrimidines. The most efficient strategy to obtain these compounds would be to utilize route B (Scheme [1]), performing the direct condensation of commercially available 4,6-dichloropyrimidine-5-carboxaldehyde (1) with various substituted hydrazines (Scheme [2]). An array of 1-substituted 4-chloropyrazolo[3,4-d]pyrimidines (4) would thus be generated, which could undergo subsequent diversification at the 4-position through displacement of the 4-chloro substituent. Whereas application of route A (Scheme [1]) toward the formation of key intermediate 4 would necessitate multiple synthetic steps, usage of route B could potentially generate 4 in a single step.

This proposed synthetic route is advantageous in terms of its simplicity. However, the requisite condensation reaction faces selectivity issues. Desired product 4 is generally observed to be the major product of this reaction,[3] [4] [10] [12] [13] ^, [17a] [b] [c] but several other products are possible, namely hydrazone 3 and 2-substituted pyrazolo[3,4-d]pyrimidine 5 (Scheme [2]). Indeed all three products were observed in our early attempts to condense carboxaldehyde 1 with various arylhydrazines (vide infra). Although product­ mixtures have been previously reported in the literature for condensation reactions involving 1 [17a] or related pyrimidine substrates,[10] [17c] the development of methods for achieving selectivity remains challenging. Our efforts in this area led us to establish strategies to selectively generate 4 in high yield, which we reported in a recent communication.[21] Herein, we present additional insights into both the scope of this reaction and the factors that influence its selectivity. We also describe the further functionalization of 4 to generate a variety of 1,4-disubstituted pyrazolo[3,4-d]pyrimidines.

Reactions Involving Aromatic Hydrazines

Our initial efforts were directed toward the reaction of carboxaldehyde 1 with arylhydrazines 2a–d. We applied the same reaction conditions that had been reported in the literature for the direct condensation of 1 with arylhydrazines.[17a] None of these experiments yielded selective reactions (Table [1], entries 1–4). The major products were pyrazolo[3,4-d]pyrimidines 4 and 5, with the formation of small quantities of hydrazone 3. A clear electronic trend was observed. Electron-rich hydrazines like 2a strongly favored the formation of 4 (entry 1), whereas electron-deficient­ hydrazines like 2d favored the formation of 5 (entry 4). Performing these same reactions in the absence of triethylamine led to dramatically different results: hydrazone 3 was the predominant product in all cases (entries 5–8).

Table 1 Initial Reactions of 1 with Arylhydrazines^a

Entry	R	Et3N (equiv)	1 (%)c	3 (%)c	4 (%)c	5 (%)c
1	4-MeOC6H4 (a)	2.1	ND	ND	91	9
2	Ph (b)	2.1	ND	2	63	35
3	4-ClC6H4 (c)	2.1	2	7	40	51
4	4-F3CC6H4 (d)	2.1	9	9	8	74
5	4-MeOC6H4 (a)	0	3	92	5	ND
6	Ph (b)	0	4	96	ND	ND
7	4-ClC6H4 (c)	0	2	98	ND	ND
8	4-F3CC6H4 (d)	0	21	79	ND	ND

^a Reaction conditions: 0.2 M, 1.05 equiv of 2, 0.3–0.6 mmol scale.

^b HCl salt.

^c Determined from relative ¹H NMR ratios in the crude reaction mixture. ND = not detected.

These observations prompted us to consider the reaction mechanism, suspecting that it would provide insight into developing a strategy to achieve product selectivity. The presence of two different electrophilic sites on 1 and two different nucleophilic sites on 2 results in three possible reaction pathways (Scheme [3]). Initial condensation with the aldehyde moiety of 1 can only proceed with the external nitrogen of 2, reversibly generating tetrahedral intermediate i, which then eliminates water to form hydrazone 3 (path A). Hydrazone 3 can be isolated as such, or it can cyclize to form 4. Initial displacement of the chloro substituent of 1, on the other hand, can occur with either nitrogen of 2, reversibly generating either ii (path B) or iv (path C), which in turn form iii or v, followed by 4 or 5, respectively.

Product 5 can only form if 2 displaces the chloro substituent of 1 prior to the condensation process (Scheme [3], path C), whereas product 4 can be generated through either the initial condensation of 2 with the aldehyde (path A) or initial chloride displacement (path B). We therefore needed to determine which pathway was generating 4 in the examples shown in entries 1–4 of Table [1]. Was 2 initially condensing with the aldehyde or displacing the chloride? To answer this question, hydrazones 3a–d were isolated and resubjected to the original reaction conditions (Table [2]). Within the original reaction time of one hour, cyclization to form 4 was not observed with any of the hydrazone­s (Table [2], entries 1–4). Even with a prolonged reaction time, only small quantities of 4 were observed (entries 5–8). These results indicate that at 65 °C in the presence of an external base, 4 does not readily form via cyclization of hydrazone 3 (Scheme [3], path A). Although 3 itself does form to a small extent under these reaction conditions (Table [1], entries 2–4), it does not cyclize. Product 4 must therefore arise primarily through initial chloride displacement by 2 (Scheme [3], path B).

Table 2 Resubjection of 3 to Original Reaction Conditions^a

Entry	R	Time (h)	3 (%)b	4 (%)b
1	4-MeOC6H4 (a)	1	>99	ND
2	Ph (b)	1	>99	ND
3	4-ClC6H4 (c)	1	>99	ND
4	4-F3CC6H4 (d)	1	>99	ND
5	4-MeOC6H4 (a)	24	88	12
6	Ph (b)	24	95	5
7	4-ClC6H4 (c)	24	96	4
8	4-F3CC6H4 (d)	24	99	1

^a Reaction conditions: 0.2 M, 1.05 equiv of HCl, 1 equiv of H₂O, 0.3 mmol scale.

^b Determined from relative ¹H NMR ratios in the crude reaction mixture. ND = not detected.

The results given in Table [1] suggest that the preferred reaction pathway is dictated by the presence or absence of an external base. We believe that this observation is consistent with the reaction mechanisms illustrated in Scheme [3]. For each possible pathway, product formation is dependent upon the productive collapse of a tetrahedral intermediate, either i, ii, or iv, whose formation is reversible. One would expect the productive collapse of i to be facilitated by acidic conditions, therefore advancing path A. Basic conditions, on the other hand, should instead accelerate the productive collapse of ii and iv, promoting both paths B and C.

Returning to our goal of developing strategies to selectively synthesize 4, we postulated that our observations regarding the effect of an external base on the preferred reaction pathway could be exploited. Performing the condensation in the presence of an external base inherently leads to product mixtures, as it facilitates reaction through both paths B and C (Scheme [3]). However, carrying out the reaction in the absence of an external base promotes only path A and thus generates a single product, hydrazone 3. Although 3 did not readily cyclize to form 4 in the examples presented in Tables 1 and [2, a] subsequent cyclization reaction, utilizing alternative reaction conditions, might allow conversion of 3 into 4. While this strategy would result in a two-step procedure, selectivity issues would be completely precluded.

Our next task was to optimize the synthesis and isolation of hydrazone 3. Having established reaction conditions to selectively generate 3 from 1 and 2 at 65 °C (Table [1], entries 5–8), we screened additional solvents at room temperature. Nucleophilic solvents such as isopropyl alcohol were not compatible with 1, as they readily displaced the chloro substituents. The conversion of 1 into 3 did proceed cleanly in THF, 1,4-dioxane, acetonitrile, acetic acid, and DMF. The reaction was the most efficient in DMF, exhibiting essentially quantitative conversion within two hours. Either the free base or the HCl salt of 2 was a viable starting material, but for electron-deficient hydrazines such as 2d, reactions involving the HCl salt progressed more slowly. Adding cold aqueous sodium bicarbonate to the crude reaction mixture and filtering the resultant precipitate led to the isolation of 3. Purification via column chromatography was not necessary. Table [3] shows the conversion of 1 into aromatic hydrazones 3a–o using our optimized reaction conditions.[21] The reaction worked efficiently for electron-rich substrates (entries 1–3), electron-deficient substrates (entries 6–10), sterically hindered substrates (entries 1, 6, and 11), and heteroaromatic substrates (entries 12–15).

Table 3 Optimized Conversion of 1 into 3 ^a

Entry	R	Isolated yield (%)
1	2-MeOC6H4 (e)	93
2	4-MeOC6H4 (a)	87
3	4-MeC6H4 (f)	92
4	Ph (b)	97
5	4-FC6H4 (g)	94
6	2-ClC6H4 (h)	92
7	3-ClC6H4 (i)	89
8	4-ClC6H4 (c)	93
9	4-BrC6H4 (j)	96
10c	4-F3CC6H4 (d)	99
11	1-naphthyl (k)	92
12d	2-pyridyl (l)	85
13d	2-quinoxalyl (m)	88
14d	2-benzo[d]oxazolyl (n)	91
15d	4-(5-methylthieno[2,3-d]pyrimidyl) (o)	98

^a Reaction conditions: 0.3–1 M, 1.05 equiv of 1, 0.5–3 mmol scale.

^b HCl salt, unless otherwise noted.

^c Free base of 2 was used.

^d Bis-HCl salt of 2 was generated in situ using 2 equiv of 4 M HCl in 1,4-dioxane.

With hydrazones 3a–o in hand, we needed to identify reaction conditions under which they would efficiently cyclize to form 4a–o. Hydrazones like 3 tend to resist cyclization, presumably because they exist in the E-configuration.[17a] [22] Heating at elevated temperatures can sometimes facilitate the process.[17a] Even prolonged heating of 3 at 65 °C in the presence of triethylamine was insufficient to substantially effect cyclization to form 4 (Table [2], entries 5–8). Performing the cyclization of 3a in the absence of triethylamine, however, led to vastly improved conversion (Table [4], entry 1). Unfortunately we also observed a small amount of side product 6a, which presumably resulted from a reaction between 4a and an equivalent of unreacted 3a. Performing the reaction in acetonitrile led to increased consumption of 3a, but a significantly greater amount of 6a formed as well (entry 2). A similar scenario was observed at 80 °C (entry 3). It was thus clear that, although this cyclization occurred at temperatures as low as 65 °C, higher temperatures were necessary in order to increase the cyclization rate such that all of 3a would cyclize before reacting with 4a. At 140 °C, 3a was completely consumed within 20 minutes, and only a small amount of 6a formed (entry 4). Formation of 6a could be further suppressed by either reducing the concentration (entry 5) or further increasing the temperature (entries 6, 7).[23]

Table 4 Optimization of the Cyclization of 3 to Form 4 ^a

Entry	R	Temp (°C)b	Time (h)	3 (%)c	4 (%)c	6 (%)c
1d	4-MeOC6H4 (a)	65	24	47	48	5
2	4-MeOC6H4 (a)	65	24	16	16	68
3	4-MeOC6H4 (a)	80	16	ND	33	67
4	4-MeOC6H4 (a)	140	0.3	ND	95	5
5e	4-MeOC6H4 (a)	140	0.3	ND	>99	ND
6	4-MeOC6H4 (a)	160	0.3	ND	98	2
7	4-MeOC6H4 (a)	180	0.3	ND	>99	ND
8	Ph (b)	140	0.3	19	76	5
9	Ph (b)	160	0.3	ND	97	3
10	Ph (b)	180	0.3	ND	>99	ND
11	4-ClC6H4 (c)	140	0.3	52	47	1
12	4-ClC6H4 (c)	160	0.3	7	91	2
13	4-ClC6H4 (c)	180	0.3	ND	>99	ND
14	4-F3CC6H4 (d)	140	0.3	86	10	4
15	4-F3CC6H4 (d)	180	0.3	10	90	ND
16	4-F3CC6H4 (d)	200	0.3	ND	>99	ND

^a Reaction conditions: 0.2 M in MeCN unless otherwise noted, 0.1–0.9 mmol scale.

^b Microwave heating was used in all cases except for entries 1–3.

^c Determined from relative ¹H NMR ratios in the crude reaction mixture. ND = not detected.

^d 0.2 M in THF.

^e 0.02 M in MeCN.

The reactions of hydrazones 3b–d were evaluated next. The cyclization proceeded more slowly as the phenyl substituent of 3 became less electron-rich. For example, after 20 minutes at 140 °C, all of 3a had been consumed (Table [4], entry 4), 19% of 3b remained (entry 8), 52% of 3c remained (entry 11), and 86% of 3d remained (entry 14). Complete conversion of 3 into 4, without formation of 6, was again achieved by increasing the temperature (entries 8–16). A temperature of 180 °C was sufficient to effect the quantitative conversion of 3 into 4 within 20 minutes for every substrate except 3d, which required a temperature of 200 °C (entries 15, 16).

After establishing the optimal conditions to convert 3a–d into 4a–d, this cyclization reaction was performed with a variety of other aromatic hydrazones (Table [5]).[21] For the sake of simplicity, these reactions were performed at 200 °C, since most substrates cyclized within 20 minutes at this temperature, regardless of their electronic properties. Acceptable solvents for this process included THF, 1,4-dioxane, and acetonitrile. Both microwave and conventional heating (sealed tube) were evaluated, and no difference was observed between the two methods in terms of the reaction efficiency. 1,4-Dioxane was utilized for reactions involving conventional heating, due to its higher boiling point. Acetonitrile, on the other hand, was the preferred solvent for the microwave reactions, because it reached 200 °C more readily in our microwave reactor compared to THF or 1,4-dioxane. Pyrazolo[3,4-d]pyrimidine products 4a–o were isolated by adding cold aqueous sodium bicarbonate and filtering the resultant precipitate. Purification via column chromatography was not necessary.

Table 5 Optimized Cyclization of 3 to Form 4 ^a

Entry	R	Isolated yield (%)
1	2-MeOC6H4 (e)	85
2b	4-MeOC6H4 (a)	97
3	4-MeC6H4 (f)	98
4b	Ph (b)	91
5	4-FC6H4 (g)	99
6c	2-ClC6H4 (h)	94
7	3-ClC6H4 (i)	92
8b	4-ClC6H4 (c)	97
9	4-BrC6H4 (j)	86
10b	4-F3CC6H4 (d)	96
11	1-naphthyl (k)	92
12d	2-pyridyl (l)	90
13	2-quinoxalyl (m)	92
14	2-benzo[d]oxazolyl (n)	92
15	4-(5-methylthieno[2,3-d]pyrimidyl) (o)	90

^a Reaction conditions: 0.5 M in 1,4-dioxane using conventional heating (sealed tube) unless otherwise noted, 0.5 mmol scale.

^b Reaction conditions: 0.5 M in MeCN using microwave heating, 0.5 mmol scale.

^c Reaction time: 40 min.

^d Product 4 was isolated as the HCl salt.

Reactions Involving Aliphatic Hydrazines

Having developed an efficient two-step procedure to selectively synthesize 4 from 1 and various aromatic hydrazines, the reaction of 1 with aliphatic hydrazines was investigated. Unlike the aromatic hydrazines (Table [1], entries 1–4), aliphatic substrates selectively generated 4 in high yield in the presence of an external base (Table [6]).[21] [24] Essentially quantitative conversion to 4 was observed within one hour at room temperature, and isomeric product 5 was not detected. Products 4p–t were cleanly isolated by removing the solvent in vacuo, following an aqueous workup, without the use of column chromatography. We observed significantly higher yields than those reported in the literature for similar reactions.[17a] This improvement in yield may exist because our reactions were carried out at room temperature instead of 65 °C, as some product decomposition could occur at elevated temperatures.[25]

Table 6 Reactions of 1 with Aliphatic Hydrazines^a

Entry	R	Et3N (equiv)	Isolated yield (%)
1	H (p)	1	74
2b	Me (q)	1	92
3	CH2CH2OH (r)	1	81
4c	cyclohexyl (s)	2	84
5d	Bn (t)	3	95

^a Reaction conditions: 0.3–0.8 M, 1.05 equiv of 2, 0.5–0.8 mmol scale.

^b i-Pr₂NEt was used instead of Et₃N.

^c HCl salt of 2 was used.

^d Bis-HCl salt of 2 was used.

The reactions described in Table [6] could proceed via either path A or path B (Scheme [3]). To determine whether or not path A was operative in these cases, isolated hydrazones 3s and 3t were stirred with two equivalents of triethylamine in THF at room temperature, and subsequent examination of the crude reaction mixture by ¹H NMR analysis revealed no formation of 4s or 4t. These observations suggest that the reactions shown in Table [6] must proceed through path B, analogous to the reactions shown in entries 1–4 of Table [1]. This interpretation is consistent with our earlier postulation that reactions between 1 and 2 initially undergo chloride displacement by 2 when an external base is present.

Discussion of the Reaction Mechanism

The difference in product selectivity between aliphatic and aromatic hydrazines in the presence of an external base is striking. Aliphatic substrates selectively generate 4 (Table [6]), proceeding exclusively through path B (Scheme [3]). Aromatic substrates, on the other hand, generate mixtures of 3, 4, and 5 (Table [1], entries 1–4); with 3 resulting from path A, 4 resulting from path B, and 5 resulting from path C. The high selectivity with which aliphatic hydrazines 2q–t react with 1 could be attributed to the reactivity difference between the two nitrogen atoms of 2q–t. It has been previously observed that the internal nitrogen of methylhydrazine (2q) is more nucleophilic than the external nitrogen.[26] A similar situation likely exists for hydrazines 2r–t. If the internal nitrogen atom of 2 serves as the predominant nucleophile in the reaction with 1, then intermediate ii (Scheme [3]) will form preferentially over iv. Assuming that productive collapse of ii is rapid in the presence of an external base, then path B will be the sole reaction pathway, ensuring the selective formation of 4. With aromatic hydrazines the situation becomes more complicated, as it appears that all reaction pathways shown in Scheme [3] are operative. We suggest that this reduced selectivity exists because the two nitrogen atoms of aromatic hydrazines are competitive nucleophiles. Initial reaction at the internal nitrogen of 2 initiates path B, while initial reaction at the external nitrogen leads to both paths A and C.

Despite the inherent lack of selectivity that exists for aromatic hydrazines in the presence of an external base, a clear electronic trend is evident in entries 1–4 of Table [1] and warrants an explanation. Electron-rich hydrazines like 2a favor reaction through path B (Scheme [3]). Electron-deficient hydrazines like 2d show the opposite selectivity, with path C being preferred. Path B and path C proceed through tetrahedral intermediates ii and iv, respectively, which presumably exist as an equilibrium mixture. In this scenario, the product distribution would be dictated by the position of the equilibrium. Considering the structures of ii and iv, one would expect ii to be favored if R is electron-donating and disfavored if R is electron-withdrawing, whereas iv should not be significantly affected by the electronic nature of the R group. This interpretation supports the results presented in entries 1–4 of Table [1]. With electron-rich 2a, ii is favored and thus 4 forms selectively (Table [1], entry 1). With electron-deficient 2d, ii is disfavored, and thus 5 is the major product (Table [1], entry 4). The above discussions provide a rationale to explain our observations, which can serve as a predictive model for other systems. However, further mechanistic studies are required in order to more fully determine the operative reaction pathways.

Some additional studies were performed to determine if the selectivity that was inherent in reactions involving aliphatic hydrazines would hold if the internal hydrazine nitrogen atom was less reactive than those of hydrazines 2q–t. The internal nitrogen atom of aliphatic hydrazine 2u is sterically hindered with a bulky tert-butyl group. At 65 °C, the reaction of 2u with 1 resulted in a nearly 1:1 mixture of 4u and 5u (Table [7], entry 1). Selectivity toward 4u improved somewhat at lower temperatures (entries 2, 3), but 5u still formed. In the case of hydrazine 2v, the internal nitrogen is sterically unencumbered, but it is rendered electron-deficient through an electron-withdrawing trifluoroethyl substituent. At 65 °C, the reaction of 2v with 1 led to a mixture of 4v and 5v, with 5v being the major product (entry 4). This result is similar to that obtained with electron-deficient arylhydrazines 2c and 2d (Table [1], entries 3, 4) and is consistent with our hypothesis regarding intermediate ii being disfavored by an electron-withdrawing R group (vide supra). Lowering the reaction temperature did not significantly alter the product selectivity in this case (Table [7], entry 5).[27] As with the aromatic hydrazines, product selectivity toward 4 could be achieved with hydrazines 2u and 2v by instead performing their reactions with 1 as two-step procedures carried out in the absence of an external base (Scheme [4]).

Table 7 Reactions of 1 with 2u and 2v ^a

Entry	R	Temp (°C)	Time (h)	1 (%)b	4 (%)b	5 (%)b
1	t-Bu (u)	65	1	ND	54	46
2	t-Bu (u)	r.t.	1	2	70	28
3	t-Bu (u)	0	7	4	85	11
4	CH2CF3 (v)	65	1	ND	36	64
5	CH2CF3 (v)	r.t.	1	4	33	63

^a Reaction conditions: 0.2 M, 1.05 equiv of 2, 2.1 equiv of Et₃N, 0.2 mmol scale. HCl salt of 2u was used. 2v was used as a 70 wt% solution in H₂O.

^b Determined from relative ¹H NMR ratios in the crude reaction mixture. ND = not detected.

One-Pot Reactions Involving Arylhydrazines

Thus far we had established efficient procedures to selectively react 1 with 2 to obtain 1-substituted 4-chloropyrazolo[3,4-d]pyrimidines (4), using either aromatic or aliphatic hydrazines. Reactions with most aliphatic hydrazines could achieve selectivity through a single-step reaction. Aromatic hydrazines, however, required a two-step procedure. We wondered if this transformation could instead be carried out as a one-pot reaction. Several literature procedures describe single-step, high temperature reactions of other heteroaromatic chlorocarboxaldehydes with substituted hydrazines, performed in the absence of an external base. These procedures selectively generated the desired pyrazolo-quinoline,[28] -naphthyridine,[29] or -pyrazole[28a] isomers, analogous to 4, in a single step. However, none of these products contained a sensitive substituent comparable to the 4-chloride of 4.

Arylhydrazines 2a–d were used to evaluate the efficiency of a one-pot, high temperature reaction with 1 in the absence of an external base (Table [8]). As expected, these reactions demonstrated high selectivity toward pyrazolo[3,4-d]pyrimidine 4 over 5. A slight excess of 1 was employed in these reactions, as excess 2 could displace the 4-chloro substituent of 4 and thus generate by-products. The primary issue for these reactions was that the in situ generated water partially hydrolyzed the 4-chloro substituent of 4, generating by-product 7. When 1 and 2a were subjected to the previously optimized cyclization conditions (Table [5]), 35% conversion to by-product 7a was observed (Table [8], entry 1). Production of 7a could be significantly reduced by decreasing the concentration (entry 2). Further suppression of 7a was accomplished by shortening the reaction time (entries 3–5). By employing a reaction time of only one minute, chloride hydrolysis could be reduced to <5%, while maintaining quantitative cyclization of 3, with both 2a and 2b (entries 5, 6).

Table 8 Optimization of One-Pot Reactions of 1 with 2a–d ^a

Entry	R	Time (min)	Conc. (M)	3 (%)c	4 (%)c	7 (%)c
1	4-MeOC6H4 (a)	20	0.5	ND	65	35
2	4-MeOC6H4 (a)	20	0.2	ND	83	17
3	4-MeOC6H4 (a)	10	0.2	ND	92	8
4	4-MeOC6H4 (a)	5	0.2	ND	94	6
5	4-MeOC6H4 (a)	1	0.2	ND	97	3
6	Ph (b)	1	0.2	ND	98	2
7	4-ClC6H4 (c)	1	0.2	9	77	14
8	4-ClC6H4 (c)	3	0.2	ND	66	34
9	4-ClC6H4 (c)	3	0.1	ND	76	24
10	4-ClC6H4 (c)	3	0.01	ND	98	2
11	4-F3CC6H4 (d)	3	0.2	25	65	10
12	4-F3CC6H4 (d)	15	0.2	ND	53	47
13	4-F3CC6H4 (d)	15	0.01	ND	97	3
14d	4-F3CC6H4 (d)	15	0.2	ND	96	4
15d	4-F3CC6H4 (d)	15	0.1	ND	98	2
16e	4-ClC6H4 (c)	3	0.1	ND	95	5
17e,f	4-ClC6H4 (c)	5	0.1	ND	97	3

^a Reactions were performed in MeCN unless otherwise noted; heating was performed using a microwave reactor; 1.05 equiv of 1, 0.2 mmol scale.

^b HCl salt unless otherwise noted.

^c Determined from relative ¹H NMR ratios in the crude reaction mixture. ND = not detected.

^d Free base of 2 was used.

^e MgSO₄ (3 equiv) was used.

^f Reaction was performed in THF with 1 equiv of i-Pr₂NEt.

Electron-deficient products 4c and 4d were both slower to form and more sensitive to hydrolysis. These reactions needed more time to reach completion and required a higher level of dilution in order to suppress chloride hydrolysis (Table [8], entries 7–13). For these more challenging substrates, we investigated the options of minimizing the presence of HCl or water. For example, using 2d in its free base form, as opposed to the HCl salt, significantly suppressed the hydrolysis process (entries 14, 15). These results suggest that chloride hydrolysis is promoted by acid. Alternatively, it was found that performing the reaction of 1 with 2c in the presence of magnesium sulfate could minimize the formation of 7c (entries 16, 17).[30] The optimized one-pot reactions of 1 with 2a–d were performed on a preparative scale as well (Table [9]). Products 4a–d were isolated in good yield after adding cold aqueous sodium bicarbonate to the crude reaction mixture and filtering the resultant precipitate. This method of purification was adequate to remove the small quantities of by-product 7 from the reactions involving 2a and 2b, but not from those of 2c and 2d.

Comparing the one-pot procedure (Table [9]) to the two-step procedure (Tables 3 and 5) for reactions involving aromatic­ hydrazines, there are advantages and disadvantages to each. The one-pot procedure is operationally simpler, but it suffers from substrate-dependent sensitivity to both concentration and reaction time. These issues would become especially important if larger scale reactions were desired. The two-step procedure requires an additional isolation step, but it is clearly more robust and is still quite facile. We suggest that for small scale reactions, in which 4 will undergo a subsequent transformation, the one-pot procedure is the method of choice. If large quantities of 4 are desired in highly pure form, then we recommend the usage of the two-step synthesis.

Table 9 Optimized One-Pot Reactions of 1 with 2a–d ^a

Entry	R	Conditions	Isolated yield (%)
1	4-MeOC6H4 (a)	MeCN (0.2 M), 1 min	87
2	Ph (b)	MeCN (0.2 M), 1 min	95
3c	4-ClC6H4 (c)	THF (0.1 M), 5 min	86d
4e	4-F3CC6H4 (d)	MeCN (0.1 M), 15 min	95d

^a Heating was performed using a microwave reactor, 1.05 equiv of 1, 0.5 mmol scale.

^b HCl salt unless otherwise noted.

^c MgSO₄ (3 equiv), i-Pr₂NEt (1 equiv).

^{d 1}H NMR analysis of 4 shows the presence of 3 mol% of 7.

^e Free base of 2 was used.

Further Functionalization of the 4-Position

Finally, we demonstrated the utility of 4-chloropyrazolo[3,4-d]pyrimidine 4 as a key intermediate to generate a variety of 1,4-disubstituted pyrazolo[3,4-d]pyrimidines. Due to the highly reactive nature of the 4-chloro substituent, it is possible to further functionalize the 4-position of 4 through nucleophilic aromatic substitution reactions. The one-pot procedures presented in Table [9] were combined with the subsequent addition of a nucleophile, generating pyrazolo[3,4-d]pyrimidines 8 and 9 using two-step, one-pot reactions (Scheme [5]).[31] This procedure was not only successful with amines (equation 1), but unactivated 1-methylindole was also able to displace the chloride (equation 2).

Additional examples of functionalization at the 4-position of 4 can be readily demonstrated using isolated 4 as the starting material. As illustrated in Scheme [6] [31] the 4-chloro substituent of 4c was directly displaced with alkoxides (equations 1 and 2) or Grignard reagents (equation 3). 4c also underwent Suzuki cross-coupling reactions (equation 4). In all of these cases, the 4-chloride of 4c was selectively displaced over the chloro substituent on the 1-phenyl moiety, providing an additional opportunity for selective functionalization of these molecules. Many other examples of further functionalization at the 4-position of 4 can be found in the literature. The 4-chloride of 4b has been transformed into various sulfur moieties.[32] The 4-chloride of 4t has been phosphonated.[33] Compounds 4b and 4q have undergone both cyanation (with potassium cyanide)[34] and aroylation (with aromatic aldehydes)[35] at the 4-position. Despite the highly electrophilic nature of the 4-chloride of 4, this position can even be rendered nucleophilic. For example, the 4-position of 4b has been lithiated and subsequently added to aldehydes and ketones.[36]

In summary, we have developed efficient procedures to selectively convert 4,6-dichloropyrimidine-5-carboxaldehyde (1) and various hydrazines into 1-substituted 4-chloropyrazolo[3,4-d]pyrimidines (4), using either aromatic or aliphatic hydrazines. Each hydrazine class has a distinct set of requisite reaction conditions to achieve selectivity. For aromatic substrates, the key is to carry out the reaction in the absence of an external base. This reaction can be performed as either a two-step procedure or a one-pot process. Aliphatic hydrazines, on the other hand, generally exhibit high selectivity toward 4 in the presence of an external base. The observations that we have documented have been rationalized within the proposed reaction mechanism, establishing a model that can be utilized to predict the optimal conditions needed to effect selective reactions between other combinations of 4-chloropyrimidine-5-carboxaldehydes and substituted hydrazines. Finally, all of the protocols reported herein are operationally simple, high-yielding, and involve reaction conditions that are mild enough to preserve the 4-chloro substituent of 4. The highly reactive nature of this 4-chloride can be exploited through further functionalization at the 4-position. Thus 4 serves as a highly versatile synthetic intermediate, capable of rapidly generating a structurally diverse array of 1,4-disubstituted pyrazolo[3,4-d]pyrimidines.

¹H and ¹³C NMR spectra were recorded on a Bruker 500 MHz NMR spectrometer. Flash column chromatography was performed on a CombiFlash Companion system (Isco, Inc.) with Silicycle prepacked silica gel cartridges. HRMS (ESI positive) analyses were performed on an LTQ Orbitrap Discovery mass spectrometer. Anhydrous solvents were purchased from Acros and used without further purification. All commercially available reagents (Aldrich, Acros, Matrix Scientific) were purchased and used without further purification unless otherwise noted.

Microwave Irradiation Experiments: Microwave irradiation experiments were performed in sealed vessels using a Biotage Initiator 60 microwave reactor, which utilized a continuous focused microwave power delivery system with operator-selectable power output from 0 to 400 W. The reaction temperature was continuously monitored by measuring the outer temperature of the reaction vessel with a calibrated infrared temperature control mounted on the side of the reaction vessel. Reactions generally reached their target temperature within 2 min upon initiation of the irradiation process, and they remained within five degrees of that temperature throughout the designated reaction time. All reactions were performed using a stirring option, which was accomplished through use of a Teflon-coated magnetic stir bar located inside of the reaction vessel and a rotating magnetic plate located below the floor of the microwave cavity.

Purification of 1 : 4,6-Dichloropyrimidine-5-carboxaldehyde (1) was not stable at room temperature for extended periods of time. Carboxaldehyde 1 obtained from commercial sources was a yellow-orange solid that was contaminated with its monohydrolyzed derivative, 4-chloro-6-hydroxypyrimidine-5-carboxaldehyde (5–10 mol% by ¹H NMR, DMSO-d ₆), along with other minor impurities. Within days, the amount of this impurity had increased, and over time the bis-hydrolyzed derivative, 4,6-dihydroxypyrimidine-5-carboxaldehyde, was also detected. For the studies described herein, commercially purchased 1 (Aldrich or Matrix Scientific) was initially purified by silica gel chromatography (0 to 10% EtOAc gradient in hexanes) to obtain a white solid (which usually contained faint yellow portions) whose sole impurity was 4-chloro-6-hydroxypyrimidine-5-carboxaldehyde in <5 mol% by ¹H NMR (DMSO­-d ₆). This material was stored in the refrigerator when not in use and maintained its integrity for several months.

Conversion of 1 into 3a–o and 3u–v; 4,6-Dichloro-5-{[2-(4-methoxy­phenyl)hydrazono]methyl}pyrimidine (3a); Typical Procedure A

To a 50 mL round-bottomed flask was added 1 (97%, 500 mg, 2.74 mmol), 4-methoxyphenylhydrazine hydrochloride (2a; 456 mg, 2.61 mmol), and DMF (10 mL). The reaction was allowed to stir under a N₂ atmosphere at r.t. for 1 h. Ice (5 g) was added with stirring, followed by a solution of NaHCO₃ (330 mg, 3.9 mmol) in H₂O (5 mL). The resultant precipitate was filtered, washed with H₂O, and dried at 40–50 °C under vacuum overnight to obtain 3a; yield: 678 mg (87%); yellow powder; mp 114–115 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.01 (s, 1 H), 8.70 (s, 1 H), 7.96 (s, 1 H), 7.01–7.10 (m, 2 H), 6.84–6.94 (m, 2 H), 3.70 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.5, 153.9, 153.7, 137.9, 126.5, 125.4, 114.7, 113.6, 55.2.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₁₀Cl₂N₄O: 297.0304; found: 297.0301 (100), 301.0240 (10), 299.0270 (64), 261 (60).

4,6-Dichloro-5-[(2-phenylhydrazono)methyl]pyrimidine (3b)

Yield (Procedure A): 678 mg (97%); yellow powder; mp 152–153 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.15 (s, 1 H), 8.75 (s, 1 H), 8.04 (s, 1 H), 7.23–7.32 (m, 2 H), 7.11 (d, J = 7.57 Hz, 2 H), 6.86 (t, J = 7.25 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.9, 154.5, 144.1, 129.3, 127.0, 126.4, 120.4, 112.5.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₈Cl₂N₄: 267.0199; found: 267.0194 (100), 271.0137 (10), 269.0165 (60), 231 (98).

4,6-Dichloro-5-{[2-(4-chlorophenyl)hydrazono]methyl}pyrimidine (3c)

Yield (Procedure A): 734 mg (93%); yellow powder; mp 167–168 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.24 (s, 1 H), 8.76 (s, 1 H), 8.04 (s, 1 H), 7.27–7.35 (m, 2 H), 7.04–7.15 (m, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.1, 154.7, 143.1, 129.2, 128.0, 126.2, 123.7, 114.0.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₇Cl₃N₄: 300.9809; found: 300.9804 (100), 306.9709 (3), 304.9746 (31), 302.9775 (92), 265 (64).

4,6-Dichloro-5-({2-[4-(trifluoromethyl)phenyl]hydrazono}methyl)pyrimidine (3d)

Yield (Procedure A): 862 mg (99%); yellow powder; mp 159–160 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.48 (s, 1 H), 8.80 (s, 1 H), 8.12 (s, 1 H), 7.61 (d, J = 8.51 Hz, 2 H), 7.23 (d, J = 8.51 Hz, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.4, 155.1, 147.2, 129.8, 126.6 (q, J = 3.78 Hz), 126.0, 124.8 (q, J = 271.90 Hz), 120.1 (q, J = 32.34 Hz), 112.4.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₇Cl₂F₃N₄: 335.0073; found: 335.0066 (100), 338.9996 (10), 337.0036 (68), 299 (40).

4,6-Dichloro-5-{[2-(2-methoxyphenyl)hydrazono]methyl}pyrimidine (3e)

Yield (Procedure A): 413 mg (93%); yellow powder; mp 113–114 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 10.67 (s, 1 H), 8.74 (s, 1 H), 8.41 (d, J = 1.26 Hz, 1 H), 7.39 (dd, J = 1.58, 7.88 Hz, 1 H), 6.99 (dd, J = 1.42, 8.04 Hz, 1 H), 6.91 (dt, J = 1.10, 7.65 Hz, 1 H), 6.81–6.88 (m, 1 H), 3.86 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.9, 154.4, 145.6, 133.3, 128.7, 126.5, 121.3, 120.4, 112.4, 111.2, 55.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₁₀Cl₂N₄O: 297.0304; found: 297.0296 (98), 301.0239 (13), 299.0267 (60), 291 (100).

4,6-Dichloro-5-[(2-p-tolylhydrazono)methyl]pyrimidine (3f)

Yield (Procedure A): 387 mg (92%); yellow powder; mp 145–146 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.05 (s, 1 H), 8.72 (s, 1 H), 7.99 (s, 1 H), 7.05–7.12 (m, 2 H), 6.98–7.04 (m, 2 H), 2.23 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.7, 154.2, 141.8, 129.7, 129.1, 126.4, 126.1, 112.6, 20.3.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₁₀Cl₂N₄: 281.0355; found: 281.0348 (93), 285.0288 (9), 283.0317 (69), 245 (100).

4,6-Dichloro-5-{[2-(4-fluorophenyl)hydrazono]methyl}pyrimidine (3g)

Yield (Procedure A): 400 mg (94%); yellow powder; mp 150–151 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.12 (s, 1 H), 8.73 (s, 1 H), 8.01 (d, J = 0.95 Hz, 1 H), 7.06–7.16 (m, 4 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.9, 156.7 (d, J = 235.62 Hz), 154.4, 140.7 (d, J = 1.26 Hz), 126.9, 126.3, 115.9 (d, J = 23.94 Hz), 113.6 (d, J = 7.56 Hz).

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₇Cl₂FN₄: 285.0105; found: 285.0097 (100), 289.0042 (11), 287.0068 (68), 249 (76).

4,6-Dichloro-5-{[2-(2-chlorophenyl)hydrazono]methyl}pyrimidine (3h)

Yield (Procedure A): 415 mg (92%); tan powder; mp 161–163 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 10.76 (s, 1 H), 8.79 (s, 1 H), 8.56 (d, J = 0.95 Hz, 1 H), 7.55 (dd, J = 1.26, 8.20 Hz, 1 H), 7.38 (dd, J = 1.26, 7.88 Hz, 1 H), 7.26–7.33 (m, 1 H), 6.89 (ddd, J = 1.42, 7.25, 8.04 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.5, 155.1, 140.5, 131.2, 129.6, 128.2, 126.2, 121.2, 116.8, 114.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₇Cl₃N₄: 300.9809; found: 300.9803 (92), 306.9713 (3), 304.9746 (27), 302.9775 (100), 265 (79).

4,6-Dichloro-5-{[2-(3-chlorophenyl)hydrazono]methyl}pyrimidine (3i)

Yield (Procedure A): 400 mg (89%); bright yellow powder; mp 172–174 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.28 (s, 1 H), 8.78 (s, 1 H), 8.05 (d, J = 0.95 Hz, 1 H), 7.29 (t, J = 8.04 Hz, 1 H), 7.10 (t, J = 2.05 Hz, 1 H), 6.98–7.06 (m, 1 H), 6.88 (ddd, J = 0.79, 2.05, 7.88 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.2, 154.9, 145.6, 133.9, 131.0, 128.7, 126.1, 119.8, 111.8, 111.2.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₇Cl₃N₄: 300.9809; found: 300.9803 (100), 306.9716 (3), 304.9744 (31), 302.9774 (94), 265 (73).

5-{[2-(4-Bromophenyl)hydrazono]methyl}-4,6-dichloropyrimidine (3j)

Yield (Procedure A): 495 (96%); yellow powder; mp 161–162 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.23 (s, 1 H), 8.76 (s, 1 H), 8.05 (d, J = 0.95 Hz, 1 H), 7.40–7.48 (m, 2 H), 7.01–7.09 (m, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.1, 154.7, 143.4, 132.0, 128.1, 126.2, 114.5, 111.4.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₇BrCl₂N₄: 344.9304; found: 344.9301 (60), 350.9214 (4), 348.9250 (47), 346.9275 (100), 311 (45).

4,6-Dichloro-5-{[2-(naphthalen-1-yl)hydrazono]methyl}pyrimidine (3k)

Yield (Procedure A): 290 mg (92%); brown powder; mp 159–160 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.33 (s, 1 H), 8.79 (s, 1 H), 8.53 (d, J = 0.63 Hz, 1 H), 8.27–8.37 (m, 1 H), 7.85–7.94 (m, 1 H), 7.50–7.59 (m, 3 H), 7.41–7.49 (m, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.2, 154.8, 139.3, 133.9, 129.9, 128.2, 126.6, 126.3, 126.0, 125.0, 121.5, 121.2, 120.2, 107.7.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₅H₁₀Cl₂N₄: 317.0355; found: 317.0351 (100), 321.0296 (10), 319.0323 (60), 281 (28).

4,6-Dichloro-5-{[2-(pyridin-2-yl)hydrazono]methyl}pyrimidine (3l)

Yield (Procedure A): 455 mg (85%); tan powder; mp 187–188 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.54 (br s, 1 H), 8.79 (s, 1 H), 8.24 (s, 1 H), 8.14–8.20 (m, 1 H), 7.70 (ddd, J = 1.73, 7.09, 8.51 Hz, 1 H), 7.26 (d, J = 8.20 Hz, 1 H), 6.87 (ddd, J = 0.95, 5.04, 7.25 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.5, 156.2, 155.2, 147.7, 138.4, 129.8, 126.2, 116.3, 106.8.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₀H₇Cl₂N₅: 268.0151; found: 268.0150 (100), 272.0090 (10), 270.0120 (64), 232 (2).

2-{2-[(4,6-Dichloropyrimidin-5-yl)methylene]hydrazinyl}quinoxaline (3m)

Yield (Procedure A): 282 mg (88%); orange powder; mp 222–223 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 12.27 (s, 1 H), 9.07 (s, 1 H), 8.86 (s, 1 H), 8.32–8.37 (m, 1 H), 7.96 (d, J = 8.20 Hz, 1 H), 7.69–7.80 (m, 2 H), 7.53–7.63 (m, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 159.0, 155.9, 149.9, 140.5, 138.3, 136.0, 132.9, 130.6, 128.8, 126.5, 126.0, 125.8.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₃H₈Cl₂N₆: 319.0260; found: 319.0259 (100), 323.0198 (10), 321.0230 (64), 315 (2).

2-{2-[(4,6-Dichloropyrimidin-5-yl)methylene]hydrazinyl}benzo[d]oxazole (3n)

Yield (Procedure A): 279 mg (91%); yellow powder; mp 207–208 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 12.63 (br s, 1 H), 8.90 (s, 1 H), 8.37 (s, 1 H), 7.55 (d, J = 7.57 Hz, 1 H), 7.45 (br m, 1 H), 7.24 (t, J = 7.57 Hz, 1 H), 7.15 (t, J = 7.57 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 159.5, 159.2, 156.7, 147.5, 137.4, 126.1, 124.3, 122.0, 116.0, 109.6. Note that not all ¹³C peaks are visible, and several are broad (see spectrum in the Supporting Information). Product 3n likely exists as a mixture of rotamers.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₇Cl₂N₅O: 308.0100; found: 308.0099 (100), 312.0038 (10), 310.0069 (66), 304 (8).

4-{2-[(4,6-Dichloropyrimidin-5-yl)methylene]hydrazinyl}-5-methylthieno[2,3-d]pyrimidine (3o)

Yield (Procedure A): 330 mg (98%); yellow powder; mp 182–184 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 11.69 (br s, 1 H), 8.91 (s, 1 H), 8.48 (s, 1 H), 7.90 (d, J = 3.47 Hz, 1 H), 7.21 (s, 1 H), 2.58 (s, 3 H).

¹³C NMR spectrum for 3o could not be obtained due to the highly insoluble nature of this compound in DMSO-d ₆ and all other common NMR solvents.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₈Cl₂N₆S: 338.9981; found: 338.9997 (100), 342.99 (13), 340.997 (34), 165 (24).

5-[(2-tert-Butylhydrazono)methyl]-4,6-dichloropyrimidine (3u)

Yield (Procedure A): 590 mg (91%); white powder; mp 114–115 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.65 (s, 1 H), 8.25 (br s, 1 H), 7.71 (s, 1 H), 1.20 (s, 9 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.3, 153.6, 127.1, 123.4, 53.6, 28.4.

HRMS (ESI): m/z [M + H]⁺ calcd for C₉H₁₂Cl₂N₄: 247.0512; found: 247.0509 (100), 251.0454 (12), 249.0481 (72), 204 (1).

4,6-Dichloro-5-{[2-(2,2,2-trifluoroethyl)hydrazono]methyl}pyrimidine (3v)

Compound 3v was synthesized according to Procedure A, from 1 (97%, 400 mg, 2.19 mmol) and 2,2,2-trifluoroethylhydrazine (2v; 70 wt% in H₂O, 535 mg, 3.28 mmol) in DMF (2.2 mL) to obtain 396 mg of 3v as a yellow powder, which contained 1 wt% DMF by ¹H NMR (66% yield); mp 113–114 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.79 (br s, 1 H), 8.73 (s, 1 H), 7.80 (s, 1 H), 4.04 (q, J = 9.46 Hz, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.5, 155.2, 126.9, 126.8, 125.0 (q, J = 281.82 Hz), 50.0 (q, J = 31.50 Hz).

HRMS (ESI): m/z [M + H]⁺ calcd for C₇H₅Cl₂F₃N₄: 272.9916; found: 272.9924 (100), 105 (22).

Conversion of 3a–o into 4a–o; 4-Chloro-1-(4-methoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (4a); Typical Procedure B

To a microwave vial was added 3a (149 mg, 0.5 mmol) and MeCN (1 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 20 min; then cooled to r.t. A cold solution of NaHCO₃ (63 mg, 0.75 mmol) in H₂O (4 mL) was added with stirring. The resultant precipitate was filtered, washed with H₂O, and dried at r.t. under vacuum overnight to obtain 4a (Note: Isolation of equally pure 4 could alternatively be accomplished by simply concentrating the crude reaction mixture in vacuo. The in situ generated HCl, which was present in the crude reaction mixture, was generally not observed to form a salt with 4 upon concentration and thus presumably evaporated); yield: 126 mg (97%); tan powder; mp 122–123 °C.

Alternative One-Pot Synthesis of 4a : To a microwave vial was added 1 (97%, 95 mg, 0.52 mmol), 4-methoxyphenylhydrazine hydrochloride (2a; 87 mg, 0.5 mmol), and MeCN (2.5 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 1 min; then cooled to r.t. The crude reaction mixture was cooled in an ice bath. A cold solution of NaHCO₃ (130 mg, 1.5 mmol) in H₂O (8 mL) was added, with stirring. The resultant precipitate was filtered, washed with H₂O, and dried at r.t. under vacuum overnight to obtain 113 mg (87%) of 4a as a tan powder.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.95 (s, 1 H), 8.72 (s, 1 H), 7.96–8.02 (m, 2 H), 7.13–7.20 (m, 2 H), 3.84 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 158.4, 155.2, 154.1, 152.2, 133.4, 130.9, 123.3, 114.5, 114.3, 55.5.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₉ClN₄O: 261.0538; found: 261.0541 (100), 263.0511 (33), 234 (4).

4-Chloro-1-phenyl-1H-pyrazolo[3,4-d]pyrimidine (4b)

Yield (Procedure B): 105 mg (91%); tan powder; mp 111–112 °C.

Alternative One-Pot Synthesis of 4b : To a microwave vial was added 1 (97%, 95 mg, 0.52 mmol), phenylhydrazine hydrochloride (2b; 72 mg, 0.5 mmol), and MeCN (2.5 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 1 min; then cooled to r.t. The crude mixture was cooled in an ice bath. A cold solution of NaHCO₃ (130 mg, 1.5 mmol) in H₂O (8 mL) was added, with stirring. The resultant precipitate was filtered, washed with H₂O, and dried at r.t. under a N₂ flow to obtain 110 mg (95%) of 4b as a yellow powder.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.99 (s, 1 H), 8.77 (s, 1 H), 8.11–8.18 (m, 2 H), 7.60–7.66 (m, 2 H), 7.42–7.48 (m, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.4, 154.2, 152.6, 137.8, 134.0, 129.4, 127.4, 121.4, 114.7.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₇ClN₄: 231.0432; found: 231.0432 (100), 233.0402 (33), 204 (3).

4-Chloro-1-(4-chlorophenyl)-1H-pyrazolo[3,4-d]pyrimidine (4c)

Yield (Procedure B): 129 mg (97%); tan powder; mp 130–131 °C.

Alternative One-Pot Synthesis of 4c : To a microwave vial was added 4-chlorophenylhydrazine hydrochloride (2c; 90 mg, 0.5 mmol), i-Pr₂NEt (87 μL, 0.5 mmol), MgSO₄ (181 mg, 1.5 mmol), and THF (5 mL). The reaction mixture was stirred at r.t. for 10 min, and 1 (97%, 95 mg, 0.52 mmol) was then added. The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 5 min; then cooled to r.t. The crude mixture was filtered through filter paper, eluting with CH₂Cl₂. The solvent was removed in vacuo. MeCN (2 mL) was added, and the solution was cooled in an ice bath. A cold solution of NaHCO₃ (130 mg, 1.5 mmol) in H₂O (8 mL) was added with stirring. The resultant precipitate was filtered, washed with H₂O, and dried at r.t. under a N₂ flow to obtain 117 mg (86%) of 4c as a yellow powder, which also contained 3 mol% 7c by ¹H NMR analysis.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.99 (s, 1 H), 8.76 (s, 1 H), 8.19 (d, J = 8.51 Hz, 2 H), 7.67 (d, J = 8.83 Hz, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.5, 154.2, 152.6, 136.6, 134.3, 131.4, 129.4, 122.5, 114.8.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₆Cl₂N₄: 265.0042; found: 265.0041 (100), 268.9982 (11), 267.0013 (67), 197 (4).

4-Chloro-1-(4-(trifluoromethyl)phenyl)-1H-pyrazolo[3,4-d]pyrimidine (4d)

Yield (Procedure B): 129 mg (96%); orange powder; mp 88–89 °C.

Alternative One-Pot Synthesis of 4d : To a microwave vial was added 1 (97%, 95 mg, 0.52 mmol), 4-(trifluoromethyl)phenylhydrazine (2d; 88 mg, 0.5 mmol), and MeCN (5 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 15 min; then cooled to r.t. The crude mixture was cooled in an ice bath. A cold solution of NaHCO₃ (130 mg, 1.5 mmol) in H₂O (8 mL) was added, with stirring. The resultant precipitate was filtered, washed with H₂O, and dried at r.t. under a N₂ flow to obtain 146 mg (95%) of 4d as a yellow powder, which also contained 3 mol% 7d by ¹H NMR analysis.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.02 (s, 1 H), 8.80 (s, 1 H), 8.43 (d, J = 8.51 Hz, 2 H), 7.96 (d, J = 8.51 Hz, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.7, 154.3, 153.1, 140.9, 134.9, 127.0 (q, J = 32.13 Hz), 126.7 (q, J = 3.78 Hz), 123.9 (q, J = 272.16 Hz), 120.9, 115.1.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₆ClF₃N₄: 299.0306; found: 299.0304 (100), 301.0275 (30), 189 (6).

4-Chloro-1-(2-methoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidine (4e)

Yield (Procedure B): 110 mg (85%); tan powder; mp 85–87 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.85 (s, 1 H), 8.70 (s, 1 H), 7.56–7.66 (m, 1 H), 7.49 (dd, J = 1.58, 7.57 Hz, 1 H), 7.32 (dd, J = 0.95, 8.20 Hz, 1 H), 7.16 (dt, J = 1.26, 7.57 Hz, 1 H), 3.72 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.2, 154.7, 153.9, 153.8, 133.4, 131.4, 128.9, 125.3, 120.6, 113.2, 112.9, 55.9.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₉ClN₄O: 261.0538; found: 261.0537 (100), 263.0508 (34), 246 (3).

4-Chloro-1-p-tolyl-1H-pyrazolo[3,4-d]pyrimidine (4f)

Yield (Procedure B): 120 mg (98%); yellow powder; mp 122–124 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.98 (s, 1 H), 8.75 (s, 1 H), 7.98–8.07 (m, 2 H), 7.43 (d, J = 7.88 Hz, 2 H), 2.40 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.4, 154.1, 152.4, 136.9, 135.5, 133.8, 129.8, 121.3, 114.5, 20.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₉ClN₄: 245.0589; found: 245.0588 (100), 247.0558 (33), 218 (4).

4-Chloro-1-(4-fluorophenyl)-1H-pyrazolo[3,4-d]pyrimidine (4g)

Yield (Procedure B): 123 mg (99%); tan powder; mp 156–158 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.99 (s, 1 H), 8.78 (s, 1 H), 8.17 (dd, J = 4.89, 8.67 Hz, 2 H), 7.48 (t, J = 8.83 Hz, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 160.7 (d, J = 245.7 Hz), 155.4, 154.2, 152.5, 134.2 (d, J = 2.52 Hz), 134.0, 123.6 (d, J = 8.82 Hz), 116.3 (d, J = 23.94 Hz), 114.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₆ClFN₄: 249.0338; found: 249.0336 (100), 251.0307 (34), 197 (3).

4-Chloro-1-(2-chlorophenyl)-1H-pyrazolo[3,4-d]pyrimidine (4h)

Yield (Procedure B): 125 mg (94%); tan powder; mp 107–110 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.91 (s, 1 H), 8.81 (s, 1 H), 7.80 (dd, J = 1.42, 8.04 Hz, 1 H), 7.73 (dd, J = 1.73, 7.72 Hz, 1 H), 7.68 (dt, J = 1.73, 7.80 Hz, 1 H), 7.59–7.64 (m, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.6, 154.1, 154.0, 134.2, 134.1, 131.9, 131.0, 130.4, 130.2, 128.4, 113.4.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₆Cl₂N₄: 265.0042; found: 265.0040 (100), 268.9983 (10), 267.0013 (65), 238 (3).

4-Chloro-1-(3-chlorophenyl)-1H-pyrazolo[3,4-d]pyrimidine (4i)

Yield (Procedure B): 121 mg (92%); tan powder; mp 112–113 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.05 (s, 1 H), 8.82 (s, 1 H), 8.32 (t, J = 2.05 Hz, 1 H), 8.18 (ddd, J = 0.79, 1.97, 8.28 Hz, 1 H), 7.67 (t, J = 8.20 Hz, 1 H), 7.53 (ddd, J = 0.95, 2.05, 8.04 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.7, 154.3, 152.9, 139.0, 134.6, 133.7, 131.3, 127.1, 120.6, 119.5, 115.0.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₆Cl₂N₄: 265.0042; found: 265.0040 (100), 268.9982 (10), 267.0011 (65), 198 (4).

1-(4-Bromophenyl)-4-chloro-1H-pyrazolo[3,4-d]pyrimidine (4j)

Yield (Procedure B): 210 mg (86%); tan powder; mp 166–168 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.02 (s, 1 H), 8.81 (s, 1 H), 8.15–8.19 (m, 2 H), 7.82–7.86 (m, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.6, 154.3, 152.7, 137.1, 134.5, 132.4, 122.9, 119.8, 114.9.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₆BrClN₄: 308.9537; found: 308.9533 (77), 312.9483 (24), 310.9510 (100), 197 (4).

4-Chloro-1-(naphthalen-1-yl)-1H-pyrazolo[3,4-d]pyrimidine (4k)

Yield (Procedure B): 128 mg (92%); brown powder; mp 97–100 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.86 (s, 1 H), 8.85 (s, 1 H), 8.21 (d, J = 8.20 Hz, 1 H), 8.13 (d, J = 8.20 Hz, 1 H), 7.76–7.81 (m, 1 H), 7.70–7.75 (m, 1 H), 7.63 (ddd, J = 0.95, 6.94, 8.20 Hz, 1 H), 7.52 (ddd, J = 1.10, 6.94, 8.35 Hz, 1 H), 7.41 (dd, J = 0.63, 8.51 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.4, 154.5, 154.1, 134.0, 133.8, 132.9, 130.1, 129.1, 128.3, 127.5, 126.9, 125.9, 125.4, 122.6, 113.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₅H₉ClN₄: 281.0589; found: 281.0586 (100), 283.0558 (31), 218 (2).

4-Chloro-1-(pyridin-2-yl)-1H-pyrazolo[3,4-d]pyrimidine (4l)

Yield (Procedure B): 121 mg (90%); tan powder; mp 175–177 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.02 (s, 1 H), 8.81 (s, 1 H), 8.68 (d, J = 3.78 Hz, 1 H), 8.11–8.18 (m, 1 H), 8.06–8.11 (m, 1 H), 7.56 (dd, J = 6.78, 5.20 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 155.7, 154.1, 153.1, 149.8, 148.9, 139.2, 134.5, 123.4, 117.2, 114.7.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₀H₆ClN₅: 232.0384; found: 232.0381 (100), 234.0351 (33), 469 (1).

2-(4-Chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)quinoxaline (4m)

Yield (Procedure B): 141 mg (92%); rose-colored powder; mp 220–223 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.79 (s, 1 H), 9.15 (s, 1 H), 8.98 (s, 1 H), 8.23 (dd, J = 1.42, 8.35 Hz, 1 H), 8.16–8.21 (m, 1 H), 7.92–8.02 (m, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 156.3, 155.7, 154.3, 144.7, 141.6, 140.5, 139.5, 135.9, 131.3, 130.2, 129.32, 129.25. Note that not all ¹³C peaks are visible (see spectrum in the Supporting Information). Product 4m was highly insoluble in DMSO-d ₆ and all other common NMR solvents; thus the ¹³C spectrum could not be recorded at a concentration adequate to visualize all ¹³C peaks.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₃H₇ClN₆: 283.0493; found: 283.0490 (100), 285.0461 (32), 267 (5).

2-(4-Chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)benzo[d]oxazole (4n)

Yield (Procedure B): 124 mg (92%); yellow powder; mp 236–239 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.18 (s, 1 H), 9.03 (s, 1 H), 7.85–7.94 (m, 2 H), 7.47–7.53 (m, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.0, 154.61, 154.58, 151.6, 148.7, 140.1, 137.9, 125.61, 125.58, 119.9, 115.4, 111.1.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₆ClN₅O: 272.0334; found: 272.0332 (100), 274.0300 (33), 244 (5).

4-(4-Chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-5-methylthieno[2,3-d]pyrimidine (4o)

Yield (Procedure B): 137 mg (90%); yellow powder; mp 185–190 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.24 (s, 1 H), 8.96 (s, 1 H), 8.95 (s, 1 H), 7.84 (d, J = 1.26 Hz, 1 H), 1.84 (d, J = 1.26 Hz, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 172.1, 156.2, 155.0, 154.6, 152.7, 150.0, 135.2, 128.7, 126.7, 125.6, 113.9, 15.2.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₇ClN₆S: 303.0214; found: 303.0224 (100), 305.019 (34), 105 (26).

Conversion of 1 into 4p–t; 4-Chloro-1H-pyrazolo[3,4-d]pyrimidine (4p); Typical Procedure C

To a 20 mL vial was added 1 (97%, 88 mg, 0.5 mmol), Et₃N (0.070 mL, 0.5 mmol), and THF (1 mL). The reaction mixture was allowed to stir at 0 °C for 10 min under a N₂ atmosphere. To the solution was added hydrazine monohydrate (2p; 0.026 mL, 0.525 mmol) in THF (1 mL) dropwise. The reaction mixture was allowed to warm to r.t. and stirred for 1 h. The solvent was removed in vacuo, and the residue was partitioned between CH₂Cl₂ (6 mL) and H₂O (10 mL). The layers were separated, and the aqueous layer was extracted with CH₂Cl₂ (2 × 6 mL). The combined organic layers were dried (MgSO₄) and concentrated in vacuo to obtain 4p; yield: 56 mg (74%); dark yellow powder; mp 156–158 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 14.51 (br s, 1 H), 8.83 (s, 1 H), 8.45 (d, J = 0.95 Hz, 1 H).

¹³C NMR (126 MHz, CDCl₃): δ = 155.4, 155.1, 154.7, 133.8, 113.5.

HRMS (ESI): m/z [M + H]⁺ calcd for C₅H₃ClN₄: 155.0119; found: 155.0117 (100), 157.0088 (34), 149 (4).

4-Chloro-1-methyl-1H-pyrazolo[3,4-d]pyrimidine (4q)

Yield (Procedure C): 78 mg (92%); yellow powder; mp 94–96 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.88 (s, 1 H), 8.48 (s, 1 H), 4.08 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 154.6, 153.5, 152.7, 131.8, 112.9, 34.3.

HRMS (ESI): m/z [M + H]⁺ calcd for C₆H₅ClN₄: 169.0276; found: 169.0274 (100), 171.0244 (32), 133 (13).

2-(4-Chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethanol (4r)

Yield (Procedure C): 120 mg (81%); yellow powder; mp 91–93 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.86 (s, 1 H), 8.49 (s, 1 H), 4.82 (br s, 1 H), 4.51 (t, J = 5.67 Hz, 2 H), 3.86 (t, J = 5.52 Hz, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 154.5, 153.5, 153.3, 132.0, 113.0, 59.1, 50.1.

HRMS (ESI): m/z [M + H]⁺ calcd for C₇H₇ClN₄O: 199.0381; found: 199.0380 (100), 201.0351 (34), 181 (13).

4-Chloro-1-cyclohexyl-1H-pyrazolo[3,4-d]pyrimidine (4s)

Yield (Procedure C): 100 mg (84%); tan powder; mp 56–57 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.85 (s, 1 H), 8.47 (s, 1 H), 4.72–4.88 (m, 1 H), 1.91–2.00 (m, 4 H), 1.87 (d, J = 13.24 Hz, 2 H), 1.64–1.75 (m, 1 H), 1.40–1.56 (m, 2 H), 1.27 (d, J = 12.93 Hz, 1 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 154.3, 153.5, 151.9, 131.7, 113.0, 56.6, 31.8, 24.82, 24.80.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₁H₁₃ClN₄: 237.0902; found: 237.0902 (100), 239.0873 (33), 155 (33).

1-Benzyl-4-chloro-1H-pyrazolo[3,4-d]pyrimidine (4t)

Yield (Procedure C): 175 mg (95%); tan powder; mp 72–74 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.91 (s, 1 H), 8.53 (s, 1 H), 7.23–7.36 (m, 5 H), 5.70 (s, 2 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 154.9, 153.8, 152.8, 136.2, 132.6, 128.6, 127.8, 127.7, 113.1, 50.7.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₉ClN₄: 245.0589; found: 245.0588 (100), 247.0559 (31), 197 (2).

1-tert-Butyl-4-chloro-1H-pyrazolo[3,4-d]pyrimidine (4u)

To a microwave vial was added 3u (124 mg, 0.5 mmol) and THF (3 mL). The reaction mixture was heated with stirring in a microwave reactor at 100 °C for 1 h; then cooled to r.t. The solvent was removed in vacuo to obtain 103 mg (98%) of 4u as an orange oil, which solidified upon storage in the refrigerator; mp 36–37 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.84 (s, 1 H), 8.40 (s, 1 H), 1.75 (s, 9 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 153.6, 153.4, 152.3, 130.5, 114.1, 61.0, 28.7.

HRMS (ESI): m/z [M + H]⁺ calcd for C₉H₁₁ClN₄: 211.0745; found: 211.0746 (55), 213.0717 (20), 155 (100).

4-Chloro-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-d]pyrimidine (4v)

To a microwave vial was added 3v (110 mg, 0.4 mmol) and MeCN (4 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 5 min; then cooled to r.t. The solvent was removed in vacuo to obtain 85 mg (90%) of 4v as an orange oil, which solidified upon storage in the refrigerator; mp 57–58 °C.

¹H NMR (500 MHz, CDCl₃): δ = 8.85 (s, 1 H), 8.28 (s, 1 H), 5.10 (q, J = 8.20 Hz, 2 H).

¹³C NMR (126 MHz, CDCl₃): δ = 155.4, 155.3, 154.6, 134.0, 122.7 (q, J = 280.56 Hz), 114.0, 48.3 (q, J = 36.54 Hz).

HRMS (ESI): m/z [M + H]⁺ calcd for C₇H₄ClF₃N₄: 237.0149; found: 237.0156 (43), 239.0126 (14), 102 (48).

4-[1-(4-Methoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]morpholine (8)

To a microwave vial was added 1 (97%, 95 mg, 0.52 mmol), 4-methoxyphenylhydrazine hydrochloride (2a; 87 mg, 0.5 mmol), and MeCN (2.5 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 1 min; then cooled to r.t. Morpholine (53 μL, 0.61 mmol) and i-Pr₂NEt (280 μL, 1.6 mmol) were added, and the mixture was allowed to stir, open to air, at r.t. for 30 min. H₂O (6 mL) was added with stirring. The resultant precipitate was filtered, washed with H₂O, and dried at r.t. under vacuum overnight to obtain 122 mg (78%) of 8 as a tan powder; mp 147–148 °C.

¹H NMR (500 MHz, CDCl₃): δ = 8.47 (s, 1 H), 8.11 (s, 1 H), 7.92–7.98 (m, 2 H), 7.02–7.09 (m, 2 H), 4.05 (t, J = 4.57 Hz, 4 H), 3.89–3.93 (m, 4 H), 3.87 (s, 3 H).

¹³C NMR (126 MHz, CDCl₃): δ = 158.4, 157.2, 155.3, 153.9, 133.1, 131.9, 123.8, 114.3, 101.4, 66.5, 55.5, 45.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₆H₁₇N₅O₂: 312.1455; found: 312.1465 (100), 102 (6).

4-(1-Methyl-1H-indol-3-yl)-1-[4-(trifluoromethyl)phenyl]-1H-pyrazolo[3,4-d]pyrimidine (9)

To a microwave vial was added 1 (97%, 95 mg, 0.52 mmol), 4-(trifluoromethyl)phenylhydrazine (2d; 88 mg, 0.5 mmol), and MeCN (5 mL). The reaction mixture was heated with stirring in a microwave reactor at 200 °C for 15 min; then cooled to r.t. 1-Methylindole (187 μL, 1.5 mmol) was added, and the mixture was heated with stirring in a microwave reactor at 200 °C for 2 h. The crude product mixture was dry-loaded directly onto silica gel and purified twice by silica gel chromatography (12 g then 4 g SiO₂, 0 to 30% EtOAc gradient in hexanes) to obtain 81 mg (40%) of 9 as a yellow powder; mp 198–199 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.09 (s, 1 H), 9.06 (s, 1 H), 8.82 (s, 1 H), 8.73–8.78 (m, 1 H), 8.56 (d, J = 8.51 Hz, 2 H), 7.95 (d, J = 8.83 Hz, 2 H), 7.55–7.61 (m, 1 H), 7.26–7.36 (m, 2 H), 3.96 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 157.4, 156.0, 153.2, 141.6, 137.4, 136.01, 135.97, 126.5 (q, J = 3.78 Hz), 126.2 (q, J = 32.76 Hz), 126.1, 124.1 (q, J = 272.16 Hz), 123.0, 122.9, 121.8, 120.6, 111.1, 110.6, 110.3, 33.3.

HRMS (ESI): m/z [M + H]⁺ calcd for C₂₁H₁₄F₃N₅: 394.1274; found: 394.1285 (100), 102 (2).

1-(4-Chlorophenyl)-4-isopropoxy-1H-pyrazolo[3,4-d]pyrimidine (10)

To a conical vial was added 4c (40 mg, 0.15 mmol), i-PrOH (23 μL, 0.3 mmol), and THF (0.75 mL). The reaction mixture was allowed to stir, open to air, in an ice bath for 5 min, after which t-AmONa (2.5 M in THF, 120 μL, 0.3 mmol) was added. The reaction was allowed to reach r.t. and stirred for 16 h. The crude reaction mixture was transferred to a separatory funnel, and CH₂Cl₂ (10 mL), sat. aq NH₄Cl (10 mL) and brine (5 mL) were added. The layers were separated, and the aqueous layer was extracted with CH₂Cl₂ (2 × 10 mL). The combined organic layers were dried (Na₂SO₄), and the crude material was purified by silica gel chromatography (4 g SiO₂, 0 to 10% EtOAc gradient in hexanes) to obtain 36 mg (83%) of 10 as a white powder; mp 145–146 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.71 (s, 1 H), 8.49 (s, 1 H), 8.21–8.27 (m, 2 H), 7.62–7.68 (m, 2 H), 5.63 (sept, J = 6.20 Hz, 1 H), 1.43 (d, J = 6.31 Hz, 6 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 163.0, 156.1, 154.3, 137.3, 133.5, 130.7, 129.3, 122.2, 103.9, 70.6, 21.6.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₄H₁₃ClN₄O: 289.0851; found: 289.0861 (10), 291.083 (3), 247 (100).

1-(4-Chlorophenyl)-4-phenoxy-1H-pyrazolo[3,4-d]pyrimidine (11)

A procedure analogous to that employed for 10 using sodium phenoxide as the nucleophile was carried out. The crude material was purified by silica gel chromatography (4 g SiO₂, 0 to 10% EtOAc gradient in hexanes) to obtain 42 mg (87%) of 11 as a pale yellow powder; mp 188–189 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.69 (s, 1 H), 8.50 (s, 1 H), 8.23–8.29 (m, 2 H), 7.65–7.71 (m, 2 H), 7.53 (t, J = 7.88 Hz, 2 H), 7.34–7.41 (m, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 163.3, 156.0, 154.8, 151.8, 137.1, 133.6, 131.0, 129.9, 129.4, 126.3, 122.5, 122.0, 103.8.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₇H₁₁ClN₄O: 323.0694; found: 323.0706 (100), 325.067 (32), 102 (13).

1-(4-Chlorophenyl)-4-methyl-1H-pyrazolo[3,4-d]pyrimidine (12)

To a conical vial was added 4c (40 mg, 0.15 mmol) and THF (0.75 mL). The solution was allowed to stir, open to air, in an ice bath for 5 min, after which MeMgBr (1 M in THF, 300 μL, 0.3 mmol) was added. The reaction mixture was allowed to reach r.t. and stirred for 17 h. The crude reaction mixture was transferred to a separatory funnel, and CH₂Cl₂ (10 mL), sat. aq NH₄Cl (10 mL), and brine (5 mL) were added. The layers were separated, and the aqueous layer was extracted with CH₂Cl₂ (2 × 10 mL). The combined organic layers were dried (Na₂SO₄), and the crude material was purified by silica gel chromatography (4 g SiO₂, 0 to 30% EtOAc gradient in hexanes) to obtain 30 mg (82%) of 12 as a yellow powder; mp 145–146 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 8.98 (s, 1 H), 8.77 (s, 1 H), 8.23–8.29 (m, 2 H), 7.61–7.68 (m, 2 H), 2.82 (s, 3 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 164.1, 155.5, 151.8, 137.2, 135.4, 130.6, 129.3, 122.0, 115.2, 21.8.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₂H₉ClN₄: 245.0589; found: 245.0600 (100), 247.057 (32), 218 (2).

1-(4-Chlorophenyl)-4-phenyl-1H-pyrazolo[3,4-d]pyrimidine (13)

To a microwave vial was added 4c (40 mg, 0.15 mmol), phenylboronic acid (20 mg, 0.16 mmol), K₂CO₃ (62 mg, 0.45 mmol), PdCl₂(dppf) (10 mg, 0.014 mmol), THF (2.2 mL), and H₂O (10 μL). The reaction mixture was heated with stirring in a microwave reactor at 170 °C for 10 min. The crude reaction mixture was passed through a small silica gel plug eluting with EtOAc, and the crude material was purified by silica gel chromatography (4 g SiO₂, 0 to 10% EtOAc gradient in hexanes) to obtain 29 mg (63%) of 13 as a white powder; mp 165–166 °C.

¹H NMR (500 MHz, DMSO-d ₆): δ = 9.21 (s, 1 H), 9.03 (s, 1 H), 8.35 (dd, J = 1.73, 7.72 Hz, 2 H), 8.31 (d, J = 9.14 Hz, 2 H), 7.63–7.73 (m, 5 H).

¹³C NMR (126 MHz, DMSO-d ₆): δ = 160.1, 155.7, 153.2, 137.1, 135.8, 135.6, 131.9, 130.9, 129.4, 129.3, 129.1, 122.5, 112.3.

HRMS (ESI): m/z [M + H]⁺ calcd for C₁₇H₁₁ClN₄: 307.0745; found: 307.0758 (100), 309.073 (31), 102 (4).

Acknowledgment

The authors wish to thank Dr. Ramil Baiazitov (PTC Therapeutics, Inc.) and Prof. Scott E. Denmark (University of Illinois, Urbana-Champaign) for helpful discussions and Jane Yang (PTC Therapeutics, Inc.) for analytical support.

• References

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• 24 We also investigated the corresponding reactions of 2q–t in the absence of an external base. In cases where 2 was an HCl salt (2s and 2t), exclusive formation of hydrazone 3 was observed. When 2 was used in its free base form (2q and 2r), a mixture of 1 and 4 (ca. 1:2) was observed, along with several unidentified minor byproducts. When 2.1 equiv of 2q and 2r were used instead of 1.05 equiv, 4 was formed quantitatively. In the latter case 2q and 2r presumably functioned as the external base, thus leading to similar results as those obtained in Table 6.
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• 19 Evans LE, Cheeseman MD, Jones K. Org. Lett. 2012; 14: 3546
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• 20 Madroñero R, Vega S. Synthesis 1987; 628
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• 23 We considered the possibility of 6 serving as an intermediate capable of converting to 4 at elevated temperatures. To test this hypothesis, isolated 6a was heated at 180 °C in MeCN for 20 min, in both the presence and the absence of 1 equiv of HCl (4 M in 1,4-dioxane). In both cases, formation of 4a was not observed (¹H NMR of the crude reaction mixture). In both cases, 6a did, however, undergo 70–80% conversion to a new product, identified by both UPLC/MS and ¹H NMR (taken of the crude reaction mixture) as 4-chloro-6-{(4-methoxyphenyl)[1-(4-methoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]amino}pyrimidine-5-carbonitrile.
• 24 We also investigated the corresponding reactions of 2q–t in the absence of an external base. In cases where 2 was an HCl salt (2s and 2t), exclusive formation of hydrazone 3 was observed. When 2 was used in its free base form (2q and 2r), a mixture of 1 and 4 (ca. 1:2) was observed, along with several unidentified minor byproducts. When 2.1 equiv of 2q and 2r were used instead of 1.05 equiv, 4 was formed quantitatively. In the latter case 2q and 2r presumably functioned as the external base, thus leading to similar results as those obtained in Table 6.
• 25 Another possible explanation is that the isolated yields reported in reference 17a were obtained after silica gel chromatography, whereas those reported in Table 6 were not. We observed that the products shown in Table 6 were partially unstable to silica gel chromatography, with an approximately 25% loss of material when this purification method was attempted.
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• 27 We also evaluated the corresponding reaction of 1 with 2v at 0 °C. We observed a mixture of 4v and 5v, again favoring 5v. The majority of the material, however, formed an unidentified intermediate(s), which did not significantly react further until the reaction warmed to r.t.
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• 30 An equivalent of i-Pr₂NEt was employed in entry 17 (Table 8) because THF was unable to reach 200 °C in our microwave reactor unless this additive was present. It is thus unclear whether or not the presence of i-Pr₂NEt is necessary to minimize chloride hydrolysis, as we were unable to perform the corresponding reaction carried out in the absence of i-Pr₂NEt. MeCN, on the other hand, could be heated to 200 °C in the absence of i-Pr₂NEt. The corresponding reactions to entries 8, 9, and 16 (Table 8), performed with one equivalent of i-Pr₂NEt, exhibited no change in product distribution relative to that shown in Table 8.
• 31 The reactions illustrated in this scheme represent unoptimized procedures carried out on a small scale (0.2–0.5 mmol).
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• Supplementary Material