# Emerging therapeutics: The imidazo[1,2-b]pyridazine scaffold as a novel drug candidate for eumycetoma, a neglected tropical disease

Lamis Yahia Mohamed Elkheir, Pierre-Olivier Delaye, Mélanie Penichon, Kimberly Eadie, Magdi Awadalla Mohamed, Pierre Besson, Adélaïde Chesnay, Guillaume Desoubeaux, Sébastien Roger, Wendy Wilhelmina Johanna van de Sande, et al.

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Lamis Yahia Mohamed Elkheir, Pierre-Olivier Delaye, Mélanie Penichon, Kimberly Eadie, Magdi Awadalla Mohamed, et al. Emerging therapeutics: The imidazo[1,2-b]pyridazine scaffold as a novel drug candidate for eumycetoma, a neglected tropical disease. European Journal of Medicinal Chemistry, 2024, 277, pp.116720. 10.1016/j.ejmech.2024.116720. hal-04995966

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# Emerging therapeutics: The imidazo[1,2-b]pyridazine scaffold as a novel drug candidate for eumycetoma, a neglected tropical disease☆

Lamis Yahia Mohamed Elkheir<sup>a, b, c</sup>, Pierre-Olivier Delaye<sup>c</sup>, Mélanie Penichon<sup>c</sup>,
Kimberly Eadie<sup>d</sup>, Magdi Awadalla Mohamed<sup>e</sup>, Pierre Besson<sup>f</sup>, Adélaïde Chesnay<sup>g</sup>,
Guillaume Desoubeaux<sup>g</sup>, Sébastien Roger<sup>f</sup>, Wendy Wilhelmina Johanna van de Sande<sup>d</sup>, Ahmed
Hassan Fahal<sup>a</sup>, Cécile Enguehard-Gueiffier<sup>c, ∗</sup>

ARTICLE INFO

Keywords:

imidazo[1,2-b]pyridazine

Antifungal agents

Madurella mycetomatis

Eumycetoma

Novel therapeutics

Drug discovery

Mycetoma

Neglected tropical disease

A B S T R A C T

Mycetoma is a neglected invasive infection endemic in tropical and subtropical regions, presenting as a chronic subcutaneous inflammatory mass that can spread to deeper structures, leading to deformities, disabilities, and potentially mortality. The current treatment of eumycetoma, the fungal form of mycetoma, involves antifungal agents, such as itraconazole, combined with surgical intervention. However, this approach has limited success, with low cure rates and a high risk of recurrence. This study addresses to the urgent need for more effective therapeutics by designing and synthesising 47 diversely pharmacomodulated imidazo [1,2-b]pyridazine derivatives using a simple synthetic pathway with good yields and purity. Of these, 17 showed promising *in vitro* activity against *Madurella mycetomatis*, the prime causative agent of eumycetoma, with IC₅₀ ≤ 5 µM and demonstrated significantly lower cytotoxicity compared to standard treatments in NIH-3T3 fibroblasts. Notably, compound 14d exhibited an excellent activity with an IC₅₀ of 0.9 µM, in the same order then itraconazole (IC₅₀ = 1.1 µM), and achieved a favourable selectivity index of 16 compared to 0.8 for itraconazole. These promising results warrant further research to evaluate the clinical potential of these novel compounds as safer, more effective treatments for eumycetoma, thus addressing a profound gap in current therapeutic strategies.

# 1. Introduction

Mycetoma is a chronic, progressively destructive infectious disease primarily affecting impoverished communities in tropical and subtropical regions such as Sudan, India, and parts of Central America [1,2]. Despite centuries of documented cases, mycetoma was only recognised in 2016 as a neglected tropical disease by the World Health Organization, signalling a renewed focus on combating its spread [3]. The disease can be caused by either bacterial (actinomycetoma) or fungal (eumycetoma) pathogens, which form grains within the infected tissues—a hallmark of mycetoma [4]. Transmission typically occurs through direct

contact with contaminated soil, thorns, or animal dung *via* cuts or abrasions in the skin, making agricultural workers and herdsmen particularly vulnerable [5]. Mycetoma commonly affects the extremities of young adults and children, yet it has also been reported in other body parts, such as the head, neck, chest, and abdominal walls [2]. Regardless of the causative agent, the initial infection typically presents as a small, painless subcutaneous swelling that could occur after years from the inculcation of the causative microorganism into the body. If left untreated, the infection can progressively worsen, leading to severe deformities and disabilities due to extensive damage to skin and bone tissue [6].

<footnote><sup>a</sup> The Mycetoma Research Center, University of Khartoum, Khartoum, Sudan</footnote>

<footnote><sup>b</sup> Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Khartoum, Sudan</footnote>

<footnote><sup>c</sup> EA 7502 Synthèse et Isolement de Molécules Bioactives (SIMBA), University of Tours, France</footnote>

<footnote><sup>d</sup> Department of Microbiology and Infectious Diseases, Erasmus Medical Center, University of Rotterdam, the Netherlands</footnote>

<footnote><sup>e</sup> Department of Pharmaceutical Chemistry, College of Pharmacy, Jouf University, KSA, Saudi Arabia</footnote>

<footnote><sup>f</sup> UMR INSERM U1327 ISCHEMIA "Membrane signalling and inflammation in reperfusion injuries", University of Tours, France</footnote>

<footnote><sup>g</sup> UMR INSERM U 1100 Centre d'Etudes des Pathologies Respiratoires, University of Tours, France</footnote>

<footnote>* This work is dedicated to the memory of Pr Alain Gueiffier.</footnote>

<footnote>* Corresponding author.</footnote>

<footnote>E-mail address: cecile.gueiffier@univ-tours.fr (C. Enguehard-Gueiffier).</footnote>
Treatment for actinomycetoma involves the administration of anti-biotics, which are generally effective [7]. On the other hand, treatment of eumycetoma is quite challenging, requiring long-term antifungal therapy combined with surgery [7]. Despite these efforts, treatment outcomes are frequently suboptimal, with low cure rates and a high incidence of recurrence, largely due to the limited efficacy and potential toxicity of available antifungals [8]. Several classes of antifungals have been employed in managing eumycetoma caused by *Madurella mycetomatis*, the prime causative agent of eumycetoma [2]. Of these classes, azoles remain the most used. For many years, ketoconazole was used until its withdrawal by the FDA in 2013 due to life-threatening hepatic and adrenal toxicity, leaving itraconazole as the primary treatment of choice for eumycetoma [8,9]. Despite prolonged periods of treatment with itraconazole, the outcomes are quite variant and suboptimal, with a high incidence of recurrence [10]. Furthermore, the required extended treatment durations with itraconazole expose eumycetoma patients to its serious adverse effect, namely hepatotoxicity [11]. The use of newer and safer azoles, such as voriconazole and posaconazole, has also been reported in the literature, yet their high cost greatly limits their availability to patients in poor endemic areas [8]. This situation underscores an urgent need for novel, more effective therapeutic options.

Being a disease predominantly affecting the neglected and impoverished, eumycetoma has historically been overlooked in drug discovery efforts, with only minimal innovations in therapeutic approaches. Recent initiatives, such as the Mycetoma Open Source (MycetOS) drug discovery project, have begun to address this gap [12]. The project involved screening of compounds from the Open Access Pathogen, Stasis, and Pandemic Response boxes provided by Medicines for Malaria Venture against *M. mycetomatis* [13,14]. Although this screening

successfully identified azoles, strobilurins, and fenarimol as classes of compounds with promising *in vitro* activity, the limited *in vivo* efficacy and inconsistent clinical effectiveness of azoles, in particular, underscore the urgent need for innovative therapeutic strategies [13–17]. Therefore, there is a critical need to identify novel scaffolds that could lead to more effective therapeutics for eumycetoma [13,14].

In this context, this study aimed to address this gap by designing, synthesising and evaluating the *in vitro* efficacy of a new series of compounds with an imidazo [1,2-b]pyridazine scaffold against *M. mycetomatis*. In this publication, we detail the synthesis methods used, characterise the newly developed compounds, and assess their *in vitro* antifungal activity. Furthermore, we present an analysis of their cytotoxicity profiles compared to currently used antifungals, namely itraconazole and amphotericin B. The goal is to lay the groundwork for developing more effective and safer treatments for this devastating disease, potentially leading to significant advancements in the therapeutic management of eumycetoma caused by *M. mycetomatis*.

## 2. Results and discussion

### 2.1. Chemistry

In this publication, we presented imidazo [1,2-b]pyridazines diversely substituted at positions 2, 6 and 8. Position 2 was functionalised during the heterocyclisation step. In contrast, substitutions at positions 8 and 6 were introduced on the imidazo [1,2-b]pyridazine scaffold during subsequent Suzuki-Miyaura type cross-couplings and nucleophilic aromatic substitution steps, respectively.

The synthesis starts with the bromination of commercially available

<figure><img src="image_4.png" /><figcaption>**Scheme 1.** Synthesis of imidazo [1,2-b]pyridazines 4–8. Reagents and conditions: i) Br₂ (1 eq), NaHCO₃ (2 eq), MeOH, r.t., 24h, 52 % yield. ii) α-brominated ketones (1.3 eq), n-BuOH, 120 °C, overnight. iii) boronic acid (1.4 eq), Cs₂CO₃ (3 eq), PdCl₂(dppf).CH₂Cl₂ (5 mol%), THF/H₂O (10/1), microwave 70–100 °C, 1–4h.</figcaption></figure>
3-amino-6-chloropyridazine **1** in the presence of bromine and NaHCO<sub>3</sub> in methanol for 24h at room temperature (Scheme 1). The brominated product **2** was obtained with a moderate yield of 52%. Compound **2** was then engaged in a cyclisation reaction with various α-brominated ketones to give the imidazo [1,2-b]pyridazines **3a-e**, diversely substituted in position 2 with 4-fluorophenyl (**3a**), 3-fluorophenyl (**3b**), 2-fluorophenyl (**3c**), 2-methoxyphenyl (**3d**) and phenyl (**3e**) in moderate to good yields (59–74%). Suzuki-Miyaura type cross-couplings were then performed on products **3a-e** to functionalise position 8 in the presence of cesium carbonate and PdCl<sub>2</sub>(dppf).CH<sub>2</sub>Cl<sub>2</sub> in THF/water mixture. Heating was performed under microwave irradiation (70–100 °C) for 1–4 h. The coupling products **4–8** were obtained with very variable yields depending on the boronic acid or ester used: from low for thien-2-yl **4i** (30 %) to good for indazol-4-yl **4e** (92 %). Different aromatic and heteroaromatic substituents were introduced via this method, including phenyl **4a**, indole (**4b-d**, **5a**, **6a**, **7a**, **8a**), pyridine (**4f-g**, **5b**, **6b-d**), thiophene (**4h-i**, **5c**, **6e**, **7b**, **8b**), pyrazole (**4j-l**), indazole (**4e**), furan (**4m**), and pyrimidine (**4n**, **5d**, **6f**, **8c**). It should be noted that this coupling reaction was selective of the carbon-bromine bond and that the

products of double couplings at both positions 6 and 8 were not observed (except for thienylboronic acids, which yield a significant amount of di-substituted products, ca. 16–27 %).

The further synthesis consisted of nucleophilic aromatic substitution at position 6, using different nucleophiles (Scheme 2). Using N-methylpiperazine, products **9–13** were obtained in moderate to excellent yields (42–96 %). When this reaction was applied to compound **3a** ($R^8 = Br$), the substitution occurred only at position 6, leading to **9a** ($R^8 = Br$ and $R^6 = N$-methylpiperazinyl). The scope of the nucleophilic aromatic substitution was then extended to various nitrogen nucleophiles: N-(2-hydroxyethyl)piperazine (**14a-f**), S-(2-aminobutan-1-ol (**15a-c**) and morpholine (**16**). Finally, an oxygenated nucleophile, 2-methoxyethanol, was used to form products **17a-c**.

We observed that the substitution applied to substrates bearing 2-methoxypprimidin-5-yl substituent at 8-position **4n** ($R^2 = 4-F$), **5d** ($R^2 = 3-F$), **6f** ($R^2 = 2-F$) and **8c** ($R^2 = H$) did not lead to expected monosubstituted products (Scheme 3). Compounds **9o**, **10d**, **11f**, **13c** and **14g** both substituted at the 6-position of the imidazo [1,2-b]pyridazine and at the 2-position of the 8-pyrimidine were obtained.

<figure><img src="image_5.png" /></figure>

<table><caption>**Scheme 2.** Synthesis of compounds **9–17**. Reagents and conditions: i) N-methylpiperazine (18 eq), 170 °C, overnight. ii) N-(2-hydroxyethyl)piperazine (18 eq), 180 °C, 6h. iii) S-(2-aminobutan-1-ol (18 eq), 180 °C, 24h. iv) morpholine (18 eq), 170 °C, overnight. v) NaH (3.4 eq), 2-methoxyethanol (4 eq), THF, 70 °C, 54h.</caption><tbody><tr><td>9a</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = Br</td><td>72%</td></tr><tr><td>9b</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = Ph</td><td>83%</td></tr><tr><td>9c</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = indol-4-yl</td><td>85%</td></tr><tr><td>9d</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = indol-5-yl</td><td>68%</td></tr><tr><td>9e</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = indol-6-yl</td><td>53%</td></tr><tr><td>9f</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = indazol-4-yl</td><td>84%</td></tr><tr><td>9g</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = pyridin-4-yl</td><td>50%</td></tr><tr><td>9h</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = pyridin-3-yl</td><td>50%</td></tr><tr><td>9i</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = thien-3-yl</td><td>90%</td></tr><tr><td>9j</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = thien-2-yl</td><td>73%</td></tr><tr><td>9k</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = 1-CH<sub>3</sub>pyrazol-4-yl</td><td>88%</td></tr><tr><td>9l</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = 1-CH<sub>3</sub>pyrazol-5-yl</td><td>86%</td></tr><tr><td>9m</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = 1-<i>i</i>-pent-pyrazol-4-yl</td><td>84%</td></tr><tr><td>9n</td><td>R<sup>2</sup> = 4-F-Ph, R<sup>8</sup> = furan-3-yl</td><td>71%</td></tr><tr><td>10a</td><td>R<sup>2</sup> = 3-F-Ph, R<sup>8</sup> = indol-4-yl</td><td>49%</td></tr><tr><td>10b</td><td>R<sup>2</sup> = 3-F-Ph, R<sup>8</sup> = pyridin-3-yl</td><td>80%</td></tr><tr><td>10c</td><td>R<sup>2</sup> = 3-F-Ph, R<sup>8</sup> = thien-3-yl</td><td>83%</td></tr><tr><td>11a</td><td>R<sup>2</sup> = 2-F-Ph, R<sup>8</sup> = indol-4-yl</td><td>69%</td></tr><tr><td>11b</td><td>R<sup>2</sup> = 2-F-Ph, R<sup>8</sup> = pyridin-4-yl</td><td>96%</td></tr><tr><td>11c</td><td>R<sup>2</sup> = 2-F-Ph, R<sup>8</sup> = pyridin-3-yl</td><td>65%</td></tr><tr><td>11d</td><td>R<sup>2</sup> = 2-F-Ph, R<sup>8</sup> = 6-aminopyridin-3-yl</td><td>42%</td></tr><tr><td>11e</td><td>R<sup>2</sup> = 2-F-Ph, R<sup>8</sup> = thien-3-yl</td><td>68%</td></tr><tr><td>12a</td><td>R<sup>2</sup> = 2-CH<sub>3</sub>O-Ph, R<sup>8</sup> = indol-4-yl</td><td>66%</td></tr><tr><td>12b</td><td>R<sup>2</sup> = 2-CH<sub>3</sub>O-Ph, R<sup>8</sup> = thien-3-yl</td><td>73%</td></tr><tr><td>13a</td><td>R<sup>2</sup> = Ph, R<sup>8</sup> = indol-4-yl</td><td>86%</td></tr><tr><td>13b</td><td>R<sup>2</sup> = Ph, R<sup>8</sup> = thien-3-yl</td><td>81%</td></tr></tbody></table>
<figure><img src="image_6.png" /><figcaption>**Scheme 3.** Dual substitution of 8-methoxypyrimidinyl derivatives. Reagents and conditions: i) N-methylpiperazine (18 eq), 170 °C, overnight. ii) N-(2-hydroxyethyl) piperazine (18 eq), 180 °C, 6h.</figcaption></figure>

Finally, some compounds (9a-c, 9g-i, 9k-l, 9n, 10b, 11c, 13a) were transformed into hydrochloride salts in order to improve their aqueous solubility for biological tests (Scheme 4). Indeed, some previous laboratory tests on a series of similar molecules confronted us with recurrent problems of solubility in aqueous media. The 18a-l salts were obtained from the corresponding bases by the addition of HCl in an isopropanol/water mixture at 60 °C for 30 min, followed by acetone and stirring for 1.5 h. The yields obtained ranged from moderate to excellent (45–97 %).

## 2.2. Biological evaluation

In a first attempt, the bioassays on the 18a-l salts were performed *in vitro* on *Madurella mycetomatis* strain MM55 at two concentrations: 100 µM and 25 µM. Compounds that inhibited more than 80 % of fungal growth at these concentrations were then selected, and their activity was assessed against MM55 at concentrations ranging from 16 µM to 0.03 µM to calculate IC₅₀. The results are presented in Table 1.

Many valuable insights can be drawn from these initial results. Firstly, it is clear that a heterocyclic group at position 8 is essential for

<figure><img src="image_7.png" /><figcaption>**Scheme 4.** Synthesis of salts 18a-l. Reagents and conditions: i) HCl (37 %, 2.2 eq), i-PrOH/H₂O, 60 °C, 30 min, then acetone, 60 °C, 1h30.</figcaption></figure>
<caption>**Table 1**</caption>

Biological effect of compounds **18a-1** against *Madurella mycetomatis* (MM55 strain).

<figure><img src="image_8.png" /></figure>

<table><thead><tr><th rowspan="2">Cpd</th><th rowspan="2">R<sup>2</sup></th><th rowspan="2">R<sup>8</sup></th><th colspan="2">MM55<sup>a</sup></th><th rowspan="2">IC<sub>50</sub> (µM)<sup>b</sup></th></tr><tr><th>100 µM</th><th>25 µM</th></tr></thead><tbody><tr><td>18a</td><td>4-F-Ph</td><td>Br</td><td>x</td><td>x</td><td>n.d.<sup>c</sup></td></tr><tr><td>18b</td><td>4-F-Ph</td><td>Ph</td><td>x</td><td>x</td><td>n.d.</td></tr><tr><td>18c</td><td>4-F-Ph</td><td>indol-4-yl</td><td>✓</td><td>✓</td><td>10.6 ± 4.3</td></tr><tr><td>18d</td><td>4-F-Ph</td><td>pyridin-4-yl</td><td>✓</td><td>x</td><td>n.d.</td></tr><tr><td>18e</td><td>4-F-Ph</td><td>pyridin-3-yl</td><td>✓</td><td>✓</td><td>>16</td></tr><tr><td>18f</td><td>4-F-Ph</td><td>thien-3-yl</td><td>✓</td><td>✓</td><td>5.0 ± 0.4</td></tr><tr><td>18g</td><td>4-F-Ph</td><td>1-methylpyrazol-4-yl</td><td>✓</td><td>✓</td><td>9.5 ± 0.2</td></tr><tr><td>18h</td><td>4-F-Ph</td><td>1-methylpyrazol-5-yl</td><td>✓</td><td>x</td><td>n.d.</td></tr><tr><td>18i</td><td>4-F-Ph</td><td>furan-3-yl</td><td>✓</td><td>x</td><td>n.d.</td></tr><tr><td>18j</td><td>3-F-Ph</td><td>pyridin-3-yl</td><td>✓</td><td>✓</td><td>11.7 ± 0.03</td></tr><tr><td>18k</td><td>2-F-Ph</td><td>pyridin-3-yl</td><td>✓</td><td>✓</td><td>11.5 ± 0.5</td></tr><tr><td>18l</td><td>Ph</td><td>indol-4-yl</td><td>✓</td><td>✓</td><td>8.3 ± 2.7</td></tr><tr><td>Itra<sup>d</sup></td><td></td><td></td><td></td><td></td><td>1,1 ± 0,4 [14]</td></tr></tbody></table>

<sup>a</sup> x: Compounds not active, √: Compounds active.

<sup>b</sup> n = 3 independent experiments.

<sup>c</sup> n.d. = not determined.

<sup>d</sup> Itra = itraconazole.

the activity. Indeed, the two compounds **18a** (R<sup>8</sup> = Br) and **18b** (R<sup>8</sup> = Ph) are the only inactive compounds, even at 100 µM. The best IC<sub>50</sub> values are obtained with compounds presenting a thien-3-yl (**18f**, 5.0 µM), indol-4-yl (**18c**, 10.6 µM and **18l**, 8.3 µM), or 1-methylpyrazol-4-yl (**18g**, 9.5 µM) group. In the 4-fluorophenyl series, the presence of 1-methylpyrazol-5-yl (**18h**, >25 µM), pyridine (**18d**, >25 µM and **18e**, >16 µM) or furan (**18i**, >25 µM) at position 8 led to less active compounds. Concerning the influence of the R<sup>2</sup> group, the presence of a fluorine atom on the phenyl ring does not appear to be a determining factor (**18c**, 10.6 µM and **18l**, 8.3 µM), nor the position of the fluorine on the phenyl ring (**18k**, **18j**, **18e**, respectively 11.5, 11.7 and > 16 µM). Nevertheless, substitution in the para position does not seem to be deleterious for the interaction with the target (**18f** and **18g**).

To further understand the structure-activity relationships of this series, another set of molecules was tested (Table 2). First, the base form (**9c**, **9i** and **9k**) of the three best molecules previously obtained in the 4-fluorophenyl series **18c**, **18f** and **18g** respectively, were tested to determine the importance of transforming our molecules into salt. Contrary to what we observed previously in the laboratory, the bases appeared to be sufficiently soluble in aqueous media to be tested. Moreover, the activity seems to be either enhanced or not affected with the base form, depending on the R<sup>8</sup> group. The antifungal activity is improved with the 8-indolyl base **9c** (1.7 µM) and the 8-pyrazolyl base **9k** (3.8 µM) compare to their corresponding salts **18c** (10.6 µM) and **18g** (9.5 µM) respectively, while the 8-thien-3-yl **9i** (5.5 µM) and its salt **18f** (5.0 µM) present the same order of activity. We, therefore, decided to test all the following molecules in base form (see Table 2).

This second set of biological evaluations allows us to draw some new conclusions, notably concerning the influence of the heterocycle nature at position 8. In the 4-fluorophenyl series, comparing the indol-4-yl derivative **9c** (1.7 µM) with its analogues indol-5-yl **9d** (3.4 µM) and indol-6-yl **9e** (7.1 µM), the orientation of the indole ring at position 8 had a slight influence on the anti-mycetomal activity, with the indol-4-yl compound having the best activity. Replacing indol-4-yl (**9c**) with its isostere indazol-4-yl (**9f**) resulted in a total loss of activity. The moderate effect of the heterocycle orientation at position 8 was also observed in the thiophene series, with compound **9i** (R<sup>8</sup> = thien-3-yl, 5.5 µM)

<caption>**Table 2**</caption>

Biological effect of the second set of compounds against *Madurella mycetomatis* (MM55 strain).

<figure><img src="image_9.png" /></figure>

<table><thead><tr><th>Cpd</th><th>R<sup>2</sup></th><th>R<sup>8</sup></th><th>IC<sub>50</sub> (µM)<sup>a</sup></th></tr></thead><tbody><tr><td>9c</td><td>4-F-Ph</td><td>indol-4-yl</td><td>1.7 ± 0.1</td></tr><tr><td>9d</td><td>4-F-Ph</td><td>indol-5-yl</td><td>3.4 ± 0.5</td></tr><tr><td>9e</td><td>4-F-Ph</td><td>indol-6-yl</td><td>7.1 ± 2.7</td></tr><tr><td>9f</td><td>4-F-Ph</td><td>indazol-4-yl</td><td>n.d.<sup>b</sup></td></tr><tr><td>9i</td><td>4-F-Ph</td><td>thien-3-yl</td><td>5.5 ± 0.4</td></tr><tr><td>9j</td><td>4-F-Ph</td><td>thien-2-yl</td><td>8.5 ± 4.5</td></tr><tr><td>9k</td><td>4-F-Ph</td><td>1-methylpyrazol-4-yl</td><td>3.8 ± 1.1</td></tr><tr><td>9m</td><td>4-F-Ph</td><td>1-iso-pentylpyrazol-4-yl</td><td>10.3 ± 4.2</td></tr><tr><td>9o</td><td>4-F-Ph</td><td>2-N-CH<sub>3</sub>piperazinylpyrimidin-5-yl</td><td>2.9 ± 1.6</td></tr><tr><td>10a</td><td>3-F-Ph</td><td>indol-4-yl</td><td>1.8 ± 1.6</td></tr><tr><td>10c</td><td>3-F-Ph</td><td>thien-3-yl</td><td>2.1 ± 0.5</td></tr><tr><td>10d</td><td>3-F-Ph</td><td>2-N-CH<sub>3</sub>piperazinylpyrimidin-5-yl</td><td>4.7 ± 0.2</td></tr><tr><td>11a</td><td>2-F-Ph</td><td>indol-4-yl</td><td>9.2 ± 2.9</td></tr><tr><td>11d</td><td>2-F-Ph</td><td>6-aminopyridin-3-yl</td><td>9.8 ± 3.7</td></tr><tr><td>11e</td><td>2-F-Ph</td><td>thien-3-yl</td><td>1.9 ± 0.2</td></tr><tr><td>11f</td><td>2-F-Ph</td><td>2-N-CH<sub>3</sub>piperazinylpyrimidin-5-yl</td><td>4.0 ± 1.5</td></tr><tr><td>12a</td><td>2-CH<sub>3</sub>O-Ph</td><td>indol-4-yl</td><td>>16</td></tr><tr><td>12b</td><td>2-CH<sub>3</sub>O-Ph</td><td>thien-3-yl</td><td>5.8 ± 0.3</td></tr><tr><td>13a</td><td>Ph</td><td>indol-4-yl</td><td>4.8 ± 0.7</td></tr><tr><td>13b</td><td>Ph</td><td>thien-3-yl</td><td>3.5 ± 2.7</td></tr><tr><td>13c</td><td>Ph</td><td>2-N-CH<sub>3</sub>piperazinylpyrimidin-5-yl</td><td>2.0 ± 0.6</td></tr><tr><td>Itraconazole</td><td></td><td></td><td>1,1 ± 0,4 [14]</td></tr></tbody></table>

<sup>a</sup> n = 3 independent experiments.

<sup>b</sup> n.d. = not determined.

presenting a slightly lower IC<sub>50</sub> than its analogue **9j** (R<sup>8</sup> = thien-2-yl, 8.5 µM). Concerning the 8-pyrazolyl compounds, changing the methyl substituent (**9k**, 3.8 µM) to isopentyl (**9m**, 10.3 µM) did not improve the activity. In the 2-fluorophenyl series, substituting the pyridin-3-yl with an amino group (**11d** R<sup>8</sup> = 6-aminopyridin-3-yl, 9.8 µM) did not improve the activity compared to the salt **18k** (R<sup>8</sup> = pyridin-3-yl, 11.5 µM) but was not detrimental. Finally, the new heterocycle 2-N-methyl-piperazinylpyrimidin-5-yl at position 8 also led to very active compounds whatever the substitution pattern at position 2: **9o** (R<sup>2</sup> = 4-F-Ph, 2.9 µM), **10d** (R<sup>2</sup> = 3-F-Ph, 4.7 µM), **11f** (R<sup>2</sup> = 2-F-Ph, 4.0 µM) and **13c** (R<sup>2</sup> = Ph, 2.0 µM).

Concerning, the influence of the R<sup>2</sup> group, substitution in ortho position seemed to be deleterious in the 8-(indol-4-yl) series comparing **9c** (R<sup>2</sup> = 4-F-Ph, 1.7 µM), **10a** (R<sup>2</sup> = 3-F-Ph, 1.8 µM), **11a** (R<sup>2</sup> = 2-F-Ph, 9.2 µM) and **12a** (R<sup>2</sup> = 2-OCH<sub>3</sub>-Ph, >16 µM). As already noted in the first set of compounds, the presence of a substituent on the phenyl ring was not essential for the anti-mycetomal activity (**13a**: R<sup>2</sup> = Ph, 4.8 µM). As previsouly observed with the 2-N-methylpiperazinylpyrimidin-5-yl at position 8, the substitution pattern of the 2-phenyl in the 8-thien-3-yl series, had almost no influence with either p-fluoro (**9i**, 5.5 µM), m-fluoro (**10c**, 2.1 µM), o-fluoro (**11e**, 1.9 µM), o-methoxy (**12b**, 5.8 µM) or no substituent (**13b**, 3.5 µM).

In the third and last set of molecules, we decided to evaluate the influence of position 6 by testing compounds **14–17** (Table 3). These results allowed us to conclude that in the 4-fluorophenyl series and in the presence of an indol-4-yl group at position 8, substitution at position 6 by 4-(2-hydroxyethyl)piperazinyl (**14a**), (1-hydroxybut-2-yl)amino (**15a**), morpholino (**16**) or 2-methoxyethoxy (**17a**) group led to a total loss of activity. Similar substitutions at position 6 in the presence of 1-methylpyrazol-4-yl group at position 8 and 4-fluorophenyl at position 2, also resulted in a complete loss of activity as evident by **14c** (4-(2-hydroxyethyl)piperazinyl), **15c** ((1-hydroxybut-2-yl)amino), and **17c** (2-methoxyethoxy). Good anti-mycetomal activity was maintained when 4-(2-hydroxyethyl)piperazinyl group was introduced at both
<figure><img src="image_10.png" /><figcaption>**Table 3**

Biological effect of the third set of compounds against *Madurella mycetomatis* (MM55 strain).</figcaption></figure>

<table><thead><tr><th>Cpd</th><th>R</th><th>R<sup>6</sup></th><th>R<sup>8</sup></th><th>IC<sub>50</sub> (µM)<sup>a</sup></th></tr></thead><tbody><tr><td>14a</td><td>4-F</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>indol-4-yl</td><td>&gt;16</td></tr><tr><td>14b</td><td>4-F</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>thien-3-yl</td><td>5.3 ± 0.7</td></tr><tr><td>14c</td><td>4-F</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>1-methylpyrazol-4-yl</td><td>n.d.<sup>b</sup></td></tr><tr><td>14d</td><td>3-F</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>thien-3-yl</td><td>0.9 ± 0.8</td></tr><tr><td>14e</td><td>2-F</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>thien-3-yl</td><td>1.3 ± 0.1</td></tr><tr><td>14f</td><td>H</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>thien-3-yl</td><td>2.6 ± 0.6</td></tr><tr><td>14g</td><td>2-F</td><td>4-(2-hydroxyethyl)piperazin-1-yl</td><td>2-(4-(2-hydroxyethyl)piperazin-1-yl)pyrimidin-5-yl</td><td>3.8 ± 3.0</td></tr><tr><td>15a</td><td>4-F</td><td>(1-hydroxybut-2-yl)amino</td><td>indol-4-yl</td><td>n.d.</td></tr><tr><td>15b</td><td>4-F</td><td>(1-hydroxybut-2-yl)amino</td><td>thien-3-yl</td><td>n.d.</td></tr><tr><td>15c</td><td>4-F</td><td>(1-hydroxybut-2-yl)amino</td><td>1-methylpyrazol-4-yl</td><td>n.d.</td></tr><tr><td>16</td><td>4-F</td><td>morpholino</td><td>indol-4-yl</td><td>n.d.</td></tr><tr><td>17a</td><td>4-F</td><td>(2-methoxyeth-1-yl)oxy</td><td>indol-4-yl</td><td>n.d.</td></tr><tr><td>17b</td><td>4-F</td><td>(2-methoxyeth-1-yl)oxy</td><td>thien-3-yl</td><td>n.d.</td></tr><tr><td>17c</td><td>4-F</td><td>(2-methoxyeth-1-yl)oxy</td><td>1-methylpyrazol-4-yl</td><td>n.d.</td></tr><tr><td colspan="3">Itraconazole</td><td></td><td>1,1 ± 0,4 [14]</td></tr></tbody></table>

positions 6 and 8 (14g, 3.8 µM). For compounds presenting a thien-3-yl group at the 8 position, only the introduction of the 4-(2-hydroxyethyl) piperazinyl group was tolerated and led to the most active series of compounds with 14b (R² = 4-F-Ph, 5.3 µM), 14d (R² = 3-F-Ph, 0.9 µM), 14e (R² = 2-F-Ph, 1.3 µM) and 14f (R² = Ph, 2.6 µM).

In light of all these results, we can conclude the following SARs for our series of molecules (Fig. 1): 1) At position 6, the N-substitutedpiperazine is the best group introduced so far. 2) At position 2, the presence of a substituent and its ortho/meta/para position on the phenyl ring is not crucial. Only in the 8-indol-4-yl series, the ortho position seems detrimental. 3) At position 8, an heterocycle is required with preferably an indol-4-yl, indol-5-yl, thien-3-yl, 1-methylpyrazol-4-yl or 2-(4-methylpiperazin-1-yl)pyrimidin-5-yl.

Using itraconazole (IC₅₀ = 1.1 ± 0.4 µM) and amphotericin B (IC₅₀ = 9.7 ± 0.7 µM) as reference antifungals for treating eumycetoma caused by *M. mycetomatis*, we found that 25 of our synthesised compounds demonstrated IC₅₀ values within this range. Notably, compound **14d** exhibited an IC₅₀ of 0.9 ± 0.8 µM, in a same order of activity than itraconazole, indicating potent antifungal activity [13,14].

We then selected our best compounds (with IC₅₀ ≤ 5 µM) for testing on a panel of *M. mycetomatis* strains of diverse origins: MM50 and MM55 from Sudan, SO1 from Somalia, l1, l3 and l11 from India, P1 from Mali, Peru72012 from Peru, AL1 from Algeria and CBS247.48 from the USA. The objective was to evaluate if genetic heterogeneity has an impact on the activity. For each strain, we measured the minimum inhibitory concentration (MIC) in µM that inhibits 80 % of the fungal growth. The results are shown in Table 4.

These results showed that most of our molecules have a stable activity against the different strains of *M. mycetomatis*. This is a positive indicator that our compounds would probably have similar activity against the other tested strains of *M. mycetomatis*. Despite itraconazole low *in vitro* IC₅₀ and MIC₅₀ values, it actually demonstrates limited effectiveness when tested in *in vivo* models or used in clinical settings, indicating that its *in vitro* potency does not accurately predict its *in vivo* and clinical efficacy. Contrary to expectations set by its *in vitro* results, itraconazole does not extend survival in *Galleria mellonella* larvae infected with *Madurella mycetomatis*, nor does it prevent grain formation in experimental mycetoma in mice, tasks at which amphotericin B and terbinafine, which show much weaker *in vitro* activity, actually excel [15–18]. Furthermore, the standard treatment regimen at the Mycetoma Research Centre in Khartoum, Sudan, which involves administering 200–400 mg of itraconazole daily for six months, only resulted in encapsulating the infected lesions without eradicating the viable *M. mycetomatis* within, further highlighting itraconazole inadequate clinical efficacy [19]. Therefore, while our compounds exhibit higher MIC₅₀ values compared to itraconazole, this should not be interpreted as an indicator of lesser effectiveness *in vivo* or in clinical settings.

Intriguingly, studies on fenarimols within the MycetOS project have indicated a possible correlation between the polarity of compounds (expressed as logD at pH 7.4) and their ability to prolong survival in the *G. mellonella* larval model infected with *M. mycetomatis* [13]. Compounds with a logD <2.5 (as calculated using Sardrop) or ≤3.6 (as calculated using ChemAxon) demonstrated enhanced *in vivo* performance [13]. Notably, five of our top-acting compounds, namely 90, 10d, 11f, 13c, and 14f, each with a logD ≤3.6 (as calculated using ChemAxon) and an IC₅₀ < 5 µM, show promise based on these criteria. Testing the *in vivo* efficacy of these five compounds could be particularly insightful.

The efficacy of ten of our products was further evaluated *in vitro* against *Candida albicans* and *Aspergillus fumigatus* to determine their broad-spectrum antifungal capabilities. However, the compounds exhibited negligible activity against these fungi (data not shown). This outcome underscores the compounds unique antifungal activity specifically targeted against *M. mycetomatis*.

Finally, we assessed the basal cytotoxicity of our best molecules (with IC₅₀ ≤ 5 µM) towards mammalian cells using NIH-3T3 murine fibroblast as a model [20]. Most of our compounds have a cytotoxicity value of around 10 µM (Table 5), ranging from 9.0 µM (90) to 14.6 µM (11f). In comparison, the reference antifungals itraconazole and amphotericin B had a cytotoxicity of 1.4 µM and 13.7 µM, respectively, against this cell line. All of our best-acting compounds had a selectivity index (SI), which is the ratio between cytotoxicity and biological activity, greater than 2, making them less toxic than both reference antifungals. Compound **14d** exhibited the highest selectivity index of 16. These findings suggest that our molecules possess a very promising toxicity profile.

### 3. Conclusion

This study has successfully demonstrated the potential of imidazo [1,2-b]pyridazine derivatives as potent antifungal agents against *M. mycetomatis*, the major causative agent of eumycetoma. This discovery is particularly crucial given the limited active scaffolds currently available for this neglected tropical disease. Notably, out of 47 compounds, 25 displayed IC₅₀ values that fall between those of the commonly used antifungals itraconazole (IC₅₀ = 1.1 ± 0.4 µM) and amphotericin B (IC₅₀ = 9.7 ± 0.7 µM), which serve as benchmarks in the treatment of eumycetoma. Furthermore, 16 of these compounds exhibited IC₅₀ values under 5 µM, with compound **14d** emerging as the most potent, achieving an IC₅₀ of 0.9 µM. This compound also demonstrated minimal cytotoxic effects on NIH-3T3 murine fibroblasts, with an IC₅₀ of 14.3 µM, resulting in a selectivity index of 16, the highest among the synthesised compounds and reference antifungals.

<footnote><sup>a</sup> n = 3 independent experiments.</footnote>

<footnote><sup>b</sup> n.d. = not determined.</footnote>
<figure><img src="image_11.png" /><figcaption>Fig. 1. Structure-Activity Relationship for imidazo [1,2-b]pyridazines against *Madurella mycetomatis* (MM55).</figcaption></figure>

<table><caption>**Table 4**
MICª for best-acting compounds against a panel of 10 strains of *Madurella mycetomatis*.</caption><thead><tr><th>Cpd</th><th>MM55</th><th>SO1</th><th>l1</th><th>l3</th><th>l11</th><th>P1</th><th>Peru72012</th><th>AL1</th><th>CBS247.48</th><th>MM50</th></tr></thead><tbody><tr><td>9c</td><td>8</td><td>16</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td></tr><tr><td>9d</td><td>16</td><td>8</td><td>4</td><td>8</td><td>4</td><td>8</td><td>4</td><td>8</td><td>4</td><td>4</td></tr><tr><td>9k</td><td>&gt;16</td><td>&gt;16</td><td>16</td><td>&gt;16</td><td>&gt;16</td><td>16</td><td>&gt;16</td><td>&gt;16</td><td>16</td><td>&gt;16</td></tr><tr><td>9o</td><td>8</td><td>4</td><td>4</td><td>8</td><td>8</td><td>4</td><td>4</td><td>4</td><td>4</td><td>4</td></tr><tr><td>10a</td><td>8</td><td>16</td><td>16</td><td>8</td><td>8</td><td>8</td><td>16</td><td>8</td><td>8</td><td>8</td></tr><tr><td>10c</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td></tr><tr><td>10d</td><td>8</td><td>4</td><td>4</td><td>4</td><td>4</td><td>4</td><td>4</td><td>4</td><td>2</td><td>4</td></tr><tr><td>11e</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>16</td><td>8</td><td>8</td><td>8</td></tr><tr><td>11f</td><td>16</td><td>4</td><td>4</td><td>16</td><td>4</td><td>4</td><td>4</td><td>4</td><td>4</td><td>4</td></tr><tr><td>13a</td><td>8</td><td>16</td><td>8</td><td>16</td><td>&gt;16</td><td>8</td><td>8</td><td>16</td><td>16</td><td>16</td></tr><tr><td>13b</td><td>8</td><td>8</td><td>8</td><td>8</td><td>4</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td></tr><tr><td>13c</td><td>2</td><td>16</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td></tr><tr><td>14d</td><td>4</td><td>8</td><td>8</td><td>8</td><td>4</td><td>4</td><td>8</td><td>8</td><td>4</td><td>8</td></tr><tr><td>14e</td><td>8</td><td>8</td><td>8</td><td>8</td><td>4</td><td>8</td><td>8</td><td>8</td><td>8</td><td>4</td></tr><tr><td>14f</td><td>4</td><td>16</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td><td>8</td></tr><tr><td>14g</td><td>8</td><td>&gt;16</td><td>16</td><td>&gt;16</td><td>&gt;16</td><td>&gt;16</td><td>16</td><td>&gt;16</td><td>&gt;16</td><td>&gt;16</td></tr><tr><td>Itra<sup>b</sup></td><td>0.5</td><td>0.25</td><td>8</td><td>1</td><td>0.5</td><td>0.25</td><td>0.5</td><td>0.06</td><td>2</td><td>1</td></tr></tbody></table>

Our findings suggest that imidazo [1,2-b]pyridazine derivatives have substantial potential as candidates for treating eumycetoma, offering potential improvements over current therapeutic options. It is essential to acknowledge the need for further *in vivo* evaluation using *Galleria mellonella* larvae [16,17] and mammalian [18] experimental models, to comprehensively assess the efficacy and safety profiles of these compounds. Addressing these limitations will be crucial for advancing these derivatives towards clinical application.

Future research should focus on exploring the mechanisms underlying the enhanced activity of these compounds and investigating their potential in combination therapies. Additionally, efforts should be directed towards evaluating the pharmacokinetic properties of the most promising candidates.

This study significantly enhances the existing knowledge base, supporting the ongoing development of innovative treatments for neglected tropical diseases, particularly eumycetoma. The findings offer

substantial promise for reducing the burden of this disease, potentially transforming the treatment landscape and markedly improving the quality of life for numerous individuals suffering from this devastating disease.

# 4. Experimental section

## 4.1. General methods of chemistry

All reagents were used directly as obtained commercially. Thin-layer chromatography (TLC) were performed using Merk® silica gel 60F₂₅₄ or Merk® aluminium oxide 60F₂₅₄ neutral plates. Column chromatography were performed manually using Merck Geduran® Si 60 (40–63 µm) silica, Merk® aluminium oxide 90 standardized, or with an Interchim PuriFlash® XS520Plus apparatus. Melting points were determined on a capillary apparatus (Stuart, Staffordshire, United Kingdom) and are

<footnote>ª n = 3 independent experiments.</footnote>

<footnote>ᵇ Itra = itraconazole.</footnote>
<caption>**Table 5**</caption>

Effect of selected compounds on cell viability of NIH-3T3 murine fibroblast and selectivity index.

<table><thead><tr><th rowspan="2">Compound</th><th colspan="2">IC<sub>50</sub> (µM)</th><th rowspan="2">Selectivity index</th></tr><tr><th>Fibroblasts ± SD</th><th>MM55 ± SD</th></tr></thead><tbody><tr><td>9c</td><td>12.9 ± 2.8</td><td>1.7 ± 0.1</td><td>7.5</td></tr><tr><td>9d</td><td>9.0 ± 2.3</td><td>3.4 ± 0.5</td><td>2.6</td></tr><tr><td>9o</td><td>9.0 ± 3.6</td><td>2.9 ± 1.6</td><td>3.1</td></tr><tr><td>10a</td><td>12.3 ± 0.2</td><td>1.8 ± 1.6</td><td>6.8</td></tr><tr><td>10c</td><td>10.1 ± 1.7</td><td>2.1 ± 0.5</td><td>4.8</td></tr><tr><td>10d</td><td>11.9 ± 0.3</td><td>4.7 ± 0.2</td><td>2.5</td></tr><tr><td>11e</td><td>10.5 ± 1.4</td><td>1.9 ± 0.2</td><td>5.5</td></tr><tr><td>11f</td><td>14.6 ± 3.2</td><td>4.0 ± 1.5</td><td>3.7</td></tr><tr><td>13a</td><td>9.6 ± 1.4</td><td>4.8 ± 0.7</td><td>2</td></tr><tr><td>13b</td><td>9.9 ± 1.4</td><td>3.5 ± 2.7</td><td>2.8</td></tr><tr><td>13c</td><td>10.2 ± 3.0</td><td>2.0 ± 0.6</td><td>5.1</td></tr><tr><td>14d</td><td>14.3 ± 1.6</td><td>0.9 ± 0.8</td><td>16</td></tr><tr><td>14e</td><td>10.7 ± 1.1</td><td>1.3 ± 0.1</td><td>8.2</td></tr><tr><td>14f</td><td>10.5 ± 3.3</td><td>2.6 ± 0.6</td><td>4.0</td></tr><tr><td>14g</td><td>13.1 ± 1.0</td><td>3.8 ± 3.0</td><td>3.4</td></tr><tr><td>Itraconazole</td><td>1.4 ± 0.1</td><td>1.13 ± 0.4</td><td>0.8</td></tr><tr><td>Amphotericin B</td><td>13.7 ± 2.7</td><td>9.7 ± 0.7</td><td>1.4</td></tr></tbody></table>

uncorrected. NMR experiments were performed at 300 MHz (¹H) and 75 MHz (¹³C) on a Bruker-Avance 300 MHz spectrometer. The standards used are the residual peaks of deuterated solvents. The abbreviations used to describe the multiplicities are: s (singlet), bs (broad singlet), d (doublet), t (triplet), q (quadruplet), dd (doublet of doublets), ddd (doublet of doublets of doublets), dt (doublet of triplets), td (triplet of doublets), AB (AB system). For the coupling constants TS stands for “Through Space”. The assignment of the carbons/protons noted C*/H* can be interchanged. Product exact mass were check using liquid chromatography-mass spectroscopy (LC-ESI-MS). LC-ESI-MS analyses were performed on an Acquity H-Class with a SQD detector (Waters, Saint Quentin en Yvelines, France). The system was fitted with a BEH C18 (50 mm × 2.1 mm; 1.7 µm particle size). Column oven was set at 40 °C. Mobile phases were A 0.01 % formic acid (aq) and mobile phase B was acetonitrile containing 0.01 % formic acid (aq). Flow rate was 0.28 mL min⁻¹ and gradient was set as follow: initial solvent B content was 50 %, raised to 100 % in 2 min, and maintained for 1 min. The column was then re-equilibrated to initial conditions. Full scan ESI in positive mode was performed between 100 and 600 m/z with cone voltage set at 25, 50 and 75 V and capillary voltage at 3.0 kV. Scan duration was set at 0.05 s. Source temperature was set at 150 °C, and desolvation temperature was set at 400 °C. Nitrogen flow was set at 900 L h⁻¹ for desolvation and 60 L h⁻¹ for cone. Samples were prepared at 50 µg mL⁻¹ in MeOH, 1 µL of the samples were injected in the system. LC-MS purity was >98 % for all final products except for **9f** (97 %), **13b** (95 %), **14f** (87 %), **15c** (96 %) and **17c** (93 %). Compounds **2** [21,22], **3e** [23] were already described in the literature.

### 4.1.1. 4-Bromo-6-chloropyridazin-3-amine (2)

To a solution of 3-amino-6-chloropyridazine (11.7 g, 90.5 mmol, 1 eq) and NaHCO₃ (15.2 g, 181 mmol, 2 eq) in methanol (200 mL) was added bromine (4.64 mL, 90.5 mmol, 1 eq) dropwise. The reaction mixture was stirred at room temperature for 24 h 50 mL of water and 25 mL of a saturated aqueous Na₂S₂O₃ solution were then added. The mixture was extracted with 3 times 100 mL of ethyl acetate. The organic phases were combined, dried over MgSO₄ and evaporated. The crude mixture was then purified by chromatography on alumina column (eluent CH₂Cl₂ 100 % → CH₂Cl₂/AcOEt 90/10) to obtain product **2** (9.74 g, 46.7 mmol, 52 %) as a brown solid. ¹H NMR (300 MHz, DMSO-d₆) δ: 8.01 (s, 1H, H-5), 7.00 (bs, 2H, NH₂). ¹³C NMR (75 MHz, DMSO-d₆) δ: 158.5 (C-6), 143.4 (C-3), 131.3 (C-5), 115.0 (C-4). **m.p.:** 178–182 °C.

### 4.1.2. General procedure for the synthesis of 3a-e

Product **2** (e.g. 600 mg, 2.88 mmol) and brominated alpha ketone (1.3 eq) are weighed into a round-bottom flask. Butanol was added (e.g. 8 mL) and the reaction mixture is stirred at 120 °C overnight. The butanol was then evaporated under vacuum. Acetone (e.g. 2.3 mL) was added to the obtained solid and the mixture was cooled for 15 min in an ice bath. The solid was then filtered and washed three times with diethyl ether. The filtrate was then dissolved in 25 mL of chloroform. 25 mL of a saturated aqueous Na₂CO₃ solution was added. After extraction the phases were separated and the aqueous phase was extracted again twice with 10 mL of chloroform. The organic phases were combined, dried over MgSO₄ and evaporated. The crude mixture was then purified by alumina column chromatography if necessary.

#### 4.1.2.1. 8-Bromo-6-chloro-2-(4-fluorophenyl)imidazo [1,2-b]pyridazine (3a)
Eluent: CH₂Cl₂, yellow solid, 5 g, yield: 65 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.21 (s, 1H, H-3), 7.99–7.94 (m, 2H, 4-F-Ph-2,6), 7.36 (s, 1H, H-7), 7.18–7.12 (m, 2H, 4-F-Ph-3,5). ¹³C NMR (75 MHz, CDCl₃) δ: 163.5 (d, ¹J<sub>C-F</sub> = 248.7 Hz, 4-F-Ph-4), 146.4 (C-2*), 145.6 (C-6*), 137.5 (C-8a), 128.7 (d, ⁴J<sub>C-F</sub> = 3.0 Hz, 4-F-Ph-1), 128.3 (d, ³J<sub>C-F</sub> = 8.3 Hz, 4-F-Ph-2,6), 123.5 (C-8), 121.3 (C-7), 116.1 (d, ²J<sub>C-F</sub> = 21.9 Hz, 4-F-Ph-3,5), 114.4 (C-3). ¹⁹F NMR (282 MHz, CDCl₃) δ: -112.2. **m.p.:** 238–242 °C.

#### 4.1.2.2. 8-Bromo-6-chloro-2-(3-fluorophenyl)imidazo [1,2-b]pyridazine (3b)
Without purification, beige solid, 782 mg, yield: 64 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.24 (s, 1H, H-3), 7.78–7.68 (m, 2H, 3-F-Ph-2,6), 7.49–7.35 (m, 2H, H-7 & 3-F-Ph-5), 7.08 (td, J = 8.4, 1.7 Hz, 1H, 3-F-Ph-4). ¹³C NMR (75 MHz, CDCl₃) δ: 163.4 (d, ¹J<sub>C-F</sub> = 246.0 Hz, 3-F-Ph-3), 145.9 (C-6), 137.5 (C-8a), 134.8 (d, ³J<sub>C-F</sub> = 8.6 Hz, 3-F-Ph-1), 134.2 (C-2), 130.6 (d, ³J<sub>C-F</sub> = 8.2 Hz, 3-F-Ph-5), 123.8 (C-8), 122.1 (d, ⁴J<sub>C-F</sub> = 2.8 Hz, 3-F-Ph-6), 121.6 (C-7), 116.0 (d, ²J<sub>C-F</sub> = 21.4 Hz, 3-F-Ph-4), 115.0 (C-3), 113.4 (d, ²J<sub>C-F</sub> = 23.0 Hz, 3-F-Ph-2). ¹⁹F NMR (282 MHz, CDCl₃) δ: -112.6. **m.p.:** 178–182 °C.

#### 4.1.2.3. 8-Bromo-6-chloro-2-(2-fluorophenyl)imidazo [1,2-b]pyridazine (3c)
Without purification, yellow solid, 1.58 g, yield: 59 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.47–8.36 (m, 2H, H-3 & 2-F-Ph-6), 7.41–7.27 (m, 3H, H-7 & 2-F-Ph-4,5), 7.16 (ddd, J = 9.3, 7.4, 1.2 Hz, 1H, 2-F-Ph-3). ¹³C NMR (75 MHz, CDCl₃) δ: 160.5 (d, ¹J<sub>C-F</sub> = 250.3 Hz, 2-F-Ph-2), 145.7 (C-6), 140.8 (d, ³J<sub>C-F</sub> = 2.0 Hz, C-2), 136.7 (C-8a), 130.3 (d, ³J<sub>C-F</sub> = 8.6 Hz, 2-F-Ph-4), 129.1 (d, ²J<sub>C-F</sub> = 3.4 Hz, 2-F-Ph-6), 124.8 (d, ⁴J<sub>C-F</sub> = 3.3 Hz, 2-F-Ph-5), 123.6 (C-8), 121.5 (C-7), 120.5 (d, ²J<sub>C-F</sub> = 12.1 Hz, 2-F-Ph-1), 118.4 (d, ⁴J<sub>C-F</sub> = 15.4 Hz, C-3), 115.9 (d, ²J<sub>C-F</sub> = 21.9 Hz, 2-F-Ph-3). ¹⁹F NMR (282 MHz, CDCl₃) δ: -113.7. **m.p.:** 178–182 °C.

#### 4.1.2.4. 8-Bromo-6-chloro-2-(2-methoxyphenyl)imidazo [1,2-b]pyridazine (3d)
Eluent: petroleum ether/CH₂Cl₂ (70/30 → 60/40), beige solid, 484 mg, yield: 74 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.60 (s, 1H, H-3), 8.48–8.44 (m, 1H, 2-MeO-Ph-6), 7.39–7.33 (m, 2H, H-7 & 2-MeO-Ph-4), 7.14–7.08 (m, 1H, 2-MeO-Ph-5), 7.01 (d, J = 8.3 Hz, 1H, 2-MeO-Ph-3), 3.99 (s, 3H, OMe). ¹³C NMR (75 MHz, CDCl₃) δ: 157.1 (2-MeO-Ph-2), 145.0 (C-6), 143.0 (C-2), 136.1 (C-8a), 130.0 (2-MeO-Ph-4), 129.1 (2-MeO-Ph-6), 129.0 (2-MeO-Ph-1), 123.2 (C-8), 121.2 (2-MeO-Ph-5), 120.7 (C-7), 119.0 (C-3), 111.0 (2-MeO-Ph-3), 55.6 (CH₃). **m.p.:** 183–187 °C.

#### 4.1.2.5. 8-Bromo-6-chloro-2-phenylimidazo [1,2-b]pyridazine (3e)
Eluent: petroleum ether/Et₂O (80/20), yellow solid, 658 mg, yield: 74 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.26 (s, 1H, H-3), 8.01–7.98 (m, 2H, Ph-2,6), 7.49–7.44 (m, 2H, Ph-3,5), 7.41–7.36 (m, 2H, H-7 & Ph-4). ¹³C NMR (75 MHz, CDCl₃) δ: 147.2 (C-2), 145.5 (C-6), 137.4 (C-8a), 132.4 (Ph-1), 129.2 (Ph-4), 129.0 (Ph-3,5), 126.5 (Ph-2,6), 123.5 (C-8), 121.2 (C-7), 117.4 (C-3). **m.p.:** 194–198 °C.
### 4.1.3. General procedure for Suzuki-Miyaura cross coupling: synthesis of 4a-n, 5a-d, 6a-f, 7a-b and 8a-c

Compound 3a-e (e.g. 200 mg, 1 eq), boronic acid (1.4 eq), cesium carbonate (3 eq) and PdCl2(dppf).CH2Cl2 (5 mol%) were weighed into a microwave tube. Three vacuum/argon cycles were applied and then THF (e.g. 3.3 mL) and water (e.g. 0.37 mL) were added. The reaction mixture was heated under microwave irradiation (ca. 70-100 °C) for 1-4 h 20 mL water and 20 mL CH2Cl2 were then added. After extraction the phases were separated and the aqueous phase was extracted with 5 times 20 mL CH2Cl2. The organic phases were combined, dried over MgSO4 and evaporated. The crude mixture was then purified by alumina or silica column chromatography.

#### 4.1.3.1. 6-Chloro-2-(4-fluorophenyl)-8-phenylimidazo [1,2-b]pyridazine (4a)
Column: alumina. Eluent: petroleum ether/CH2Cl2 (70/30), yellow solid, 484 mg, yield: 74 %. ¹H NMR (300 MHz, DMSO-d6) δ: 8.96 (s, 1H, H-3), 8.47-8.44 (m, 2H, Ph-2,6), 8.14-8.09 (m, 2H, 4-F-Ph-2,6), 7.71 (s, 1H, H-7), 7.66-7.58 (m, 3H, Ph-3,4,5), 7.36-7.30 (m, 2H, 4-F-Ph-3,5). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.3 (d, $^1J_{CF}$= 245.3 Hz, 4-F-Ph-4), 146.4 (C-6), 144.2 (C-2), 137.1 (C-8a), 136.7 (C-8), 132.3 (Ph-1), 130.8 (Ph-4), 129.51 (Ph-2,6), 129.45 (4-F-Ph-1), 128.8 (Ph-3,5), 127.8 (d, $^3J_{CF}$= 8.3 Hz, 4-F-Ph-2,6), 115.8 (d, $^2J_{CF}$= 21.7 Hz, 4-F-Ph-3,5), 115.4 (C-7), 114.2 (C-3). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.3 m.p.: 183-187 °C. LC-MS: retention time = 3.35 min, m/z [M+H]⁺ = 324.05.

#### 4.1.3.2. 6-Chloro-2-(4-fluorophenyl)-8-(1H-indol-4-yl)imidazo [1,2-b] pyridazine (4b)
Column: silica. Eluent: petroleum ether/CH2Cl2 (70/30), yellow solid, 793 mg, yield: 79 %. ¹H NMR (300 MHz, DMSO-d6) δ: 11.50 (bs, 1H, NH), 8.96 (s, 1H, H-3), 8.08-8.03 (m, 2H, 4-F-Ph-2,6), 7.90 (d, J = 6.8 Hz, 1H, indol-5), 7.64 (d, J = 8.1 Hz, 1H, indol-7), 7.54-7.53 (m, 2H, H-7 & indol-2), 7.34-7.27 (m, 3H, 4-F-Ph-3,5 & indol-6), 6.65 (bs, 1H, indol-3). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.3 (d, $^1J_{CF}$= 245.1 Hz, 4-F-Ph-4), 146.2 (C-6), 144.2 (C-2), 138.8 (C-8), 137.0 (C-8a), 136.5 (indol-7a), 129.6 (d, $^4J_{CF}$= 2.8 Hz, 4-F-Ph-1), 127.8 (d, $^3J_{CF}$= 8.2 Hz, 4-F-Ph-2,6), 127.0 (indol-2), 126.0 (indol-3a), 124.0 (indol-4), 122.0 (indol-5), 120.8 (indol-6), 116.6 (C-7), 115.8 (d, $^2J_{CF}$= 21.5 Hz, 4-F-Ph-3,5), 114.2 (C-3), 114.0 (indol-7), 100.6 (indol-3). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.4. m.p.: 246-250 °C. LC-MS: retention time = 3.00 min, m/z [M+H]⁺ = 363.05.

#### 4.1.3.3. 6-Chloro-2-(4-fluorophenyl)-8-(1H-indol-5-yl)imidazo [1,2-b] pyridazine (4c)
Column: alumina. Eluent: CH2Cl2, yellow solid, 181 mg, yield: 90 %. ¹H NMR (300 MHz, DMSO-d6) δ: 11.4 (bs, 1H, NH), 8.92-8.88 (m, 2H, H-3 & indol-4), 8.23 (dd, J = 8.7, 1.7 Hz, 1H, indol-6), 8.17-8.12 (m, 2H, 4-F-Ph-2,6), 7.67 (s, 1H, H-7), 7.59 (d, J = 8.7 Hz, 1H, indol-7), 7.49-7.47 (m, 1H, indol-2), 7.38-7.32 (m, 2H, 4-F-Ph-3,5), 6.64 (bs, indol-3). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.2 (d, $^1J_{CF}$= 245.3 Hz, 4-F-Ph-4), 146.4 (C-6), 143.8 (C-2), 138.8 (C-8), 137.3 (C-8a*), 137.1 (indol-7a*), 129.7 (d, $^4J_{CF}$= 3.0 Hz, 4-F-Ph-1), 127.8 (d, $^3J_{CF}$= 8.2 Hz, 4-F-Ph-2,6), 127.77 (indol-3a), 126.9 (indol-2), 122.9 (indol-5), 122.8 (indol-4), 122.3 (indol-6), 115.8 (d, $^2J_{CF}$= 21.5 Hz, 4-F-Ph-3,5), 113.9 (C-3 & C-7), 111.7 (indol-7), 102.5 (indol-3). ¹⁹F NMR (282 MHz, CDCl3) δ: -112.9. m.p.: 258-262 °C. LC-MS: retention time = 3.07 min, m/z [M+H]⁺ = 363.15.

#### 4.1.3.4. 6-Chloro-2-(4-fluorophenyl)-8-(1H-indol-6-yl)imidazo [1,2-b] pyridazine (4d)
Column: alumina. Eluent: CH2Cl2/petroleum ether (50/50), yellow solid, 132 mg, yield: 59 %. ¹H NMR (300 MHz, acetone-d6) δ: 10.7 (bs, 1H, NH), 9.03 (m, 1H, indol-7), 8.62 (s, 1H, H-3), 8.21-8.16 (m, 2H, 4-F-Ph-2,6), 8.02 (dd, J = 8.5, 1.7 Hz, 1H, indol-5), 7.77 (d, J = 8.4 Hz, 1H, indol-4), 7.58-7.56 (m, 2H, H-7 & indol-2), 7.28-7.22 (m, 2H, 4-F-Ph-3,5), 6.62-6.60 (m, 1H, indol-3). ¹³C NMR (300 MHz, acetone-d6) δ: 163.8 (d, $^1J_{CF}$= 245.6 Hz, 4-F-Ph-4), 147.6 (C-6), 145.4 (C-2), 140.1 (C-8), 138.4 (C-8a), 137.1 (indol-7a), 131.2 (indol-6), 131.1 (d, $^4J_{CF}$= 3.1 Hz, 4-F-Ph-1), 128.83 (indol-2), 128.80 (d, $^3J_{CF}$= 8.3 Hz, 4-F-Ph-2,6), 126.2 (indol-3a), 121.3 (indol-4), 120.7 (indol-5), 116.4 (d, $^2J_{CF}$= 21.9 Hz, 4-F-Ph-3,5), 115.3 (indol-7), 115.0 (C-7), 114.3 (C-3), 102.8 (indol-3). ¹⁹F NMR (282 MHz, acetone-d6) δ: -115.2. m.p.: 245-249 °C. LC-MS: retention time = 3.13 min, m/z [M+H]⁺ = 363.15.

#### 4.1.3.5. 6-Chloro-2-(4-fluorophenyl)-8-(1H-indazol-4-yl)imidazo [1,2-b] pyridazine (4e)
Column: silica. Eluent: CH2Cl2/EtOAc (100/0 to 98/2), yellow solid, 90 mg, yield: 92 %. ¹H NMR (300 MHz, DMSO-d6) δ: 13.39 (bs, 1H, NH), 9.00 (s, 1H, H-3), 8.25 (bs, 1H, indazol-3), 8.07-8.02 (m, 2H, 4-F-Ph-2,6), 7.94 (d, J = 6.8 Hz, 1H, indazol-5), 7.79 (d, J = 8.4 Hz, 1H, indazol-7), 7.63 (s, 1H, H-7), 7.59 (dd, J = 8.3, 7.2 Hz, 1H, indazol-6), 7.34-7.28 (m, 2H, 4-F-Ph-3,5). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.3 (d, $^1J_{CF}$= 245.4 Hz, 4-F-Ph-4), 146.2 (C-6), 144.3 (C-2), 140.3 (C-8), 137.2 (C-8a), 136.7 (indazol-7a), 133.5 (indazol-3), 129.5 (d, $^4J_{CF}$= 2.9 Hz, 4-F-Ph-1), 127.8 (d, $^3J_{CF}$= 8.4 Hz, 4-F-Ph-2,6), 125.9 (indazol-3a), 125.3 (indazol-4), 123.0 (indazol-6), 121.0 (indazol-5), 117.1 (C-7), 115.8 (d, $^2J_{CF}$= 21.7 Hz, 4-F-Ph-3,5), 114.3 (C-3), 112.6 (indazol-7). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.3. m.p.: 301-305 °C. LC-MS: retention time = 2.50 min, m/z [M+H]⁺ = 364.15.

#### 4.1.3.6. 6-Chloro-2-(4-fluorophenyl)-8-(pyridin-4-yl)imidazo [1,2-b] pyridazine (4f)
Column: alumina. Eluent: CH2Cl2, yellow solid, 138 mg, yield: 69 %. ¹H NMR (300 MHz, DMSO-d6) δ: 8.93 (s, 1H, H-3), 8.82-8.80 (m, 2H, pyr-2,6), 8.41-8.38 (m, 2H, Pyr-3,5), 8.11-8.06 (m, 2H, 4-F-Ph-2,6), 7.85 (H-7), 7.34-7.28 (4-F-Ph-3,5). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.4 (d, $^1J_{CF}$= 245.6 Hz, 4-F-Ph-4), 150.2 (pyr-2,6), 146.3 (C-6), 144.6 (C-2), 139.5 (pyr-4), 136.1 (C-8a), 134.1 (C-8), 129.2 (d, $^4J_{CF}$= 3.0 Hz, 4-F-Ph-1), 127.9 (d, $^3J_{CF}$= 8.4 Hz, 4-F-Ph-2,6), 123.2 (pyr-3,5), 116.4 (C-7), 115.7 (d, $^2J_{CF}$= 21.6 Hz, 4-F-Ph-3,5), 114.3 (C-3). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.0. m.p.: 220-224 °C. LC-MS: retention time = 2.02 min, m/z [M+H]⁺ = 325.05.

#### 4.1.3.7. 6-Chloro-2-(4-fluorophenyl)-8-(pyridin-3-yl)imidazo [1,2-b] pyridazine (4g)
Column: alumina. Eluent: CH2Cl2, yellow solid, 163 mg, yield: 40 %. ¹H NMR (300 MHz, DMSO-d6) δ: 9.58 (d, J = 1.9 Hz, 1H, pyr-2), 8.98 (s, 1H, H-3), 8.84 (dt, J = 8.0, 1.7 Hz, 1H, pyr-4), 8.77 (dd, J = 4.8, 1.5 Hz, 1H, pyr-6), 8.14-8.09 (m, 2H, 4-F-Ph-2,6), 7.86 (s, 1H, H-7), 7.66 (dd, J = 8.0, 4.8 Hz, 1H, pyr-5), 7.36-7.30 (m, 2H, 4-F-Ph-3,5). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.3 (d, $^1J_{CF}$= 245.2 Hz, 4-F-Ph-4), 151.2 (pyr-6), 149.9 (pyr-2), 146.3 (C-6), 144.4 (C-2), 136.8 (pyr-4), 136.3 (C-8a), 134.3 (C-8), 129.3 (d, $^4J_{CF}$= 2.9 Hz, 4-F-Ph-1), 128.3 (pyr-3), 127.8 (d, $^3J_{CF}$= 8.4 Hz, 4-F-Ph-2,6), 123.6 (pyr-5), 115.8 (d, $^2J_{CF}$= 21.6 Hz, 4-F-Ph-3,5), 115.7 (C-7), 114.3 (C-3). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.2. m.p.: 199-203 °C. LC-MS: retention time = 2.28 min, m/z [M+H]⁺ = 325.15.

#### 4.1.3.8. 6-Chloro-2-(4-fluorophenyl)-8-(thien-3-yl)imidazo [1,2-b] pyridazine (4h)
Column: silica. Eluent: cyclohexane/CH2Cl2 (71/29), yellow solid, 342 mg, yield: 84 %. ¹H NMR (300 MHz, acetone-d6) δ: 9.40 (dd, J = 3.0, 1.3 Hz, 1H, thien-2), 8.63 (s, 1H, H-3), 8.24-8.18 (m, 3H, 4-F-Ph-2,6 & thien-4), 7.73 (dd, J = 5.2, 3.0 Hz, 1H, thien-5), 7.69 (s, 1H, H-7), 7.30-7.24 (4-F-Ph-3,5). ¹³C NMR (75 MHz, acetone-d6) δ: 163.8 (d, $^1J_{CF}$= 245.7 Hz, 4-F-Ph-4), 147.7 (C-6), 145.5 (C-2), 137.5 (C-8a), 134.3 (C-8), 132.7 (thien-3), 132.4 (thien-2), 130.8 (d, $^4J_{CF}$= 3.2 Hz, 4-F-Ph-1), 128.8 (d, $^3J_{CF}$= 8.2 Hz, 4-F-Ph-2,6), 127.7 (thien-4*), 127.5 (thien-5*), 116.4 (d, $^2J_{CF}$= 21.8 Hz, 4-F-Ph-3,5), 114.5 (C-7*), 114.3 (C-3*). ¹⁹F NMR (282 MHz, acetone-d6) δ: -115.0. m.p.: 182-186 °C. LC-MS: retention time = 3.40 min, m/z [M+H]⁺ = 330.05.

#### 4.1.3.9. 6-Chloro-2-(4-fluorophenyl)-8-(thien-2-yl)imidazo [1,2-b] pyridazine (4i)
Column: alumina. Eluent: CH2Cl2/petroleum ether (50/50), yellow solid, 61 mg, yield: 30 %. ¹H NMR (300 MHz, DMSO-d6) δ: 8.91 (s, 1H, H-3), 8.53 (d, J = 3.7 Hz, 1H, thien-3), 8.17-8.13 (m, 2H, 4-
F-Ph-2,6), 8.04 (d, J = 5.0 Hz, thien-5), 7.83 (s, 1H, H-7), 7.38–7.32 (m, 3H, 4-F-Ph-3,5 & Thien-4). **¹³C NMR** (75 MHz, DMSO-d₆) *δ*: 162.3 (d, *¹J*<sub>C-F</sub> = 245.2 Hz, 4-F-Ph-4), 146.2 (C-6), 143.9 (C-2), 135.2 (C-8a), 133.6 (thiopen-5), 133.5 (C-8*), 131.7 (thien-3), 130.8 (thien-2*), 129.4 (d, *⁴J*<sub>C-F</sub> = 3.1 Hz, 4-F-Ph-1), 128.3 (thien-4), 127.8 (d, *³J*<sub>C-F</sub> = 8.3 Hz, 4-F-Ph-2,6), 115.8 (d, *²J*<sub>C-F</sub> = 21.7 Hz, 4-F-Ph-3,5), 114.3 (C-3), 111.9 (C-7). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –113.2. **m.p.:** 202–206 °C. **LC-MS:** retention time = 3.38 min, m/z [M+H]⁺ = 330.05.

#### 4.1.3.10. 6-Chloro-2-(4-fluorophenyl)-8-(1-methyl-1h-pyrazol-4-yl)imidazo [1,2-b]pyridazine (4j).
Column: alumina. Eluent: CH₂Cl₂/petroleum ether (50/50), yellow solid, 160 mg, yield: 80 %. **¹H NMR** (300 MHz, CDCl₃) *δ*: 8.85 (s, 1H, pyrazol-5), 8.20 (s, 1H, pyrazol-3), 8.12 (s, 1H, H-3), 8.00–7.96 (m, 2H, 4-F-Ph-2,6), 7.19–7.13 (m, 3H, H-7 & 4-F-Ph-3,5), 4.06 (s, 3H, CH₃). **¹³C NMR** (75 MHz, CDCl₃) *δ*: 163.2 (d, *¹J*<sub>C-F</sub> = 247.9 Hz, 4-F-Ph-4), 146.9 (C-6), 144.7 (C-2), 138.8 (pyrazol-3), 136.4 (C-8a), 133.2 (pyrazol-5), 130.9 (C-8), 129.5 (d, *⁴J*<sub>C-F</sub> = 3.3 Hz, 4-F-Ph-1), 127.9 (d, *³J*<sub>C-F</sub> = 8.2 Hz, 4-F-Ph-2,6), 115.9 (d, *²J*<sub>C-F</sub> = 21.8 Hz, 4-F-Ph-3,5), 114-8 (pyrazol-4), 113.1 (C-3), 111.5 (C-7), 39.6 (CH₃). **¹⁹F NMR** (282 MHz, CDCl₃) *δ*: –113.1. **m.p.:** 221–225 °C. **LC-MS:** retention time = 2.55 min, m/z [M+H]⁺ = 328.15.

#### 4.1.3.11. 6-Chloro-2-(4-fluorophenyl)-8-(1-methyl-1h-pyrazol-5-yl)imidazo [1,2-b]pyridazine (4k).
Column: silica. Eluent: CH₂Cl₂, yellow solid, 247 mg, yield: 61 %. **¹H NMR** (300 MHz, DMSO-d₆) *δ*: 8.98 (s, 1H, H-3), 8.12–8.07 (m, 2H, 4-F-Ph-2,6), 7.66 (d, J = 2.0 Hz, 1H, pyrazol-3), 7.55 (s, 1H, H-7), 7.34–7.29 (m, 2H, 4-F-Ph-3,5), 7.03 (d, J = 2.0 Hz, pyrazol-4), 4.06 (s, 3H, CH₃). **¹³C NMR** (75 MHz, DMSO-d₆) *δ*: 162.4 (d, *¹J*<sub>C-F</sub> = 245.6 Hz, 4-F-Ph-4), 145.9 (C-6), 144.4 (C-2), 138.3 (pyrazol-3), 135.8 (C-8a), 134.8 (pyrazol-5), 129.3 (d, *⁴J*<sub>C-F</sub> = 3.0 Hz, 4-F-Ph-1), 128.1 (C-8), 127.9 (d, *³J*<sub>C-F</sub> = 8.4 Hz, 4-F-Ph-2,6), 117.2 (C-7), 115.8 (d, *²J*<sub>C-F</sub> = 21.6 Hz, 4-F-Ph-3,5), 114.4 (C-3); 110.5 (pyrazol-4), 39.1 (CH₃). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –113.1. **m.p.:** 222–226 °C. **LC-MS:** retention time = 2.36 min, m/z [M+H]⁺ = 328.05.

#### 4.1.3.12. 6-Chloro-2-(4-fluorophenyl)-8-(1-isopentyl-1h-pyrazol-4-yl)imidazo [1,2-b]pyridazine (4l).
Column: silica. Eluent: CH₂Cl₂, beige solid, 128 mg, yield: 85 %. **¹H NMR** (300 MHz, DMSO-d₆) *δ*: 9.05 (s, 1H, pyrazol-5), 8.84 (s, 1H, pyrazol-3), 8.61 (s, 1H, H-3), 8.21–8.17 (m, 2H, 4-F-Ph-2,6), 7.69 (s, 1H, H-7), 7.36–7.30 (m, 2H, 4-F-Ph-3,5), 4.30 (t, J = 7.2 Hz, 2H, NCH₂), 1.76 (q, J = 6.9 Hz, 2H, NCH₂CH₂), 1.60–1.49 (m, 1H, CH), 0.93 (d, J = 6.6 Hz, 6H, CH₃). **¹³C NMR** (75 MHz, DMSO-d₆) *δ*: 162.2 (d, *¹J*<sub>C-F</sub> = 245.1 Hz, 4-F-Ph-4), 146.3 (C-6), 143.5 (C-2), 139.4 (pyrazol-3), 135.7 (C-8a), 132.4 (pyrazol-5), 130.6 (C-8), 129.5 (d, *⁴J*<sub>C-F</sub> = 2.9 Hz, 4-F-Ph-1), 127.8 (d, *³J*<sub>C-F</sub> = 8.2 Hz, 4-F-Ph-2,6), 115.7 (d, *²J*<sub>C-F</sub> = 21.6 Hz, 4-F-Ph-3,5), 113.9 (pyrazol-4*), 113.7 (C-3*), 111.3 (C-7), 50.0 (NCH₂), 38.5 (NCH₂CH₂), 25.0 (CH), 22.2 (CH₃). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –113.4. **m.p.:** 169–173 °C. **LC-MS:** retention time = 3.44 min, m/z [M+H]⁺ = 384.15.

#### 4.1.3.13. 6-Chloro-2-(4-fluorophenyl)-8-(furan-3-yl)imidazo [1,2-b]pyridazine (4m).
Column: silica. Eluent: CH₂Cl₂/petroleum ether (50/50 → 80/20), yellow solid, 154 mg, yield: 40 %. **¹H NMR** (300 MHz, acetone-d₆) *δ*: 9.21 (bs, 1H, furan-2), 8.61 (s, 1H, H-3), 8.24–8.19 (m, 2H, 4-F-Ph-2,6), 7.84 (t, J = 1.8 Hz, 1H, furan-5), 7.58 (s, 1H, H-7), 7.42 (dd, J = 2.0, 0.7 Hz, 1H, furan-4), 7.29–7.23 (m, 2H, 4-F-Ph-3,5). **¹³C NMR** (75 MHz, acetone-d₆) *δ*: 163.8 (d, *¹J*<sub>C-F</sub> = 245.7 Hz, 4-F-Ph-4), 148.2 (furan-2), 147.5 (C-6), 145.4 (furan-5), 145.2 (C-2), 137.0 (C-8a), 131.4 (C-8), 130.8 (d, *⁴J*<sub>C-F</sub> = 2.9 Hz, 4-F-Ph-1), 128.8 (d, *³J*<sub>C-F</sub> = 8.2 Hz, 4-F-Ph-2,6), 119.9 (furan-3), 116.4 (d, *²J*<sub>C-F</sub> = 21.8 Hz, 4-F-Ph-3,5), 114.3 (C-3), 114.1 (C-7), 109.2 (furan-4). **¹⁹F NMR** (282 MHz, acetone-d₆) *δ*: –115.0. **m.p.:** 173–177 °C. **LC-MS:** retention time = 3.21 min, m/z [M+H]⁺ = 314.05.

#### 4.1.3.14. 6-Chloro-2-(4-fluorophenyl)-8-(2-methoxypyrimidin-5-yl)imidazo [1,2-b]pyridazine (4n).
Column: silica. Eluent: CH₂Cl₂/cyclohexane (20/80 → 100/0), yellow solid, 161 mg, yield: 74 %. **¹H NMR** (300 MHz, DMSO-d₆) *δ*: 9.69 (s, 2H, pyrimidin-4,6), 8.95 (s, 1H, H-3), 8.10 (dd, J = 8.7, 5.6 Hz, 2H, 4-F-Ph-2,6), 7.90 (s, 1H, H-7), 7.32 (t, J = 8.9 Hz, 2H, 4-F-Ph-3,5), 4.05 (s, 3H, CH₃). **¹³C NMR** (75 MHz, DMSO-d₆) *δ*: 165.6 (pyrimidin-2), 162.4 (d, *¹J*<sub>C-F</sub> = 248.7 Hz, 4-F-Ph-4), 160.0 (pyrimidin-4,6), 146.3 (C-6), 144.3 (C-2), 135.9 (C-8a), 131.4 (C-8), 129.2 (d, *⁴J*<sub>C-F</sub> = 3.6 Hz, 4-F-Ph-1), 127.8 (d, *³J*<sub>C-F</sub> = 8.8 Hz, 4-F-Ph-2,6), 120.3 (pyrimidin-1), 115.8 (d, *²J*<sub>C-F</sub> = 21.8 Hz, 4-F-Ph-3,5), 114.3 (C-3 & C-7), 55.2 (OCH₃). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –113.1. **m.p.:** 335–339 °C. **LC-MS:** retention time = 2.69 min, m/z [M+H]⁺ = 356.15.

#### 4.1.3.15. 6-Chloro-2-(3-fluorophenyl)-8-(indol-4-yl)imidazo [1,2-b]pyridazine (5a).
Column: silica. Eluent: CH₂Cl₂/cyclohexane (20/80 → 100/0), yellow solid, 128 mg, yield: 58 %. **¹H NMR** (300 MHz, DMSO-d₆) *δ*: 11.51 (bs, 1H, NH), 9.04 (s, 1H, H-3), 7.95–7.76 (m, 3H, indol-5, 3-F-Ph-2 & 3-F-Ph-6), 7.64 (d, J = 8.1 Hz, 1H, indol-7), 7.57–7.44 (m, 3H, H-7, indol-2 & 3-F-Ph-5), 7.32 (t, J = 7.8 Hz, 1H, indol-6), 7.19 (td, J = 8.7, 2.0 Hz, 1H, 3-F-Ph-4), 6.66 (s, 1H, indol-3). **¹³C NMR** (75 MHz, DMSO-d₆) *δ*: 162.63 (d, *¹J*<sub>C-F</sub> = 242.9 Hz, 3-F-Ph-3), 146.4 (C-6), 143.7 (d, *⁴J*<sub>C-F</sub> = 2.8 Hz, C-2), 138.9 (C-8), 137.1 (C-8a), 136.5 (indol-7a), 135.6 (d, *³J*<sub>C-F</sub> = 8.4 Hz, 3-F-Ph-1), 130.9 (d, *³J*<sub>C-F</sub> = 8.5 Hz, 3-F-Ph-5), 127.0 (indol-2), 126.0 (indol-3a), 123.9 (indol-4), 122.0 (indol-5), 121.7 (d, *⁴J*<sub>C-F</sub> = 2.3 Hz, 3-F-Ph-6), 120.8 (indol-6), 116.8 (C-7), 115.00 (d, *²J*<sub>C-F</sub> = 21.2 Hz, 3-F-Ph-4), 114.97 (C-3), 114.0 (indol-7), 112.1 (d, *²J*<sub>C-F</sub> = 22.8 Hz, 3-F-Ph-2), 100.6 (indol-3). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –113.0. **m.p.:** 159–163 °C. **LC-MS:** retention time = 3.00 min, m/z [M+H]⁺ = 363.05.

#### 4.1.3.16. 6-Chloro-2-(3-fluorophenyl)-8-(pyridin-3-yl)imidazo [1,2-b]pyridazine (5b).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 178 mg, yield: 60 %. **¹H NMR** (300 MHz, DMSO-d₆) *δ*: 9.57 (dd, J = 2.3, 0.7 Hz, 1H, pyridin-2), 9.04 (s, 1H, H-3), 8.84 (ddd, J = 8.1, 2.2, 1.7 Hz, 1H, pyridin-4), 8.77 (dd, J = 4.8, 1.6 Hz, 1H, pyridin-6), 7.93–7.84 (m, 3H, H-7, 3-F-Ph-2 & 3-F-Ph-6), 7.66 (ddd, J = 8.1, 4.8, 0.8 Hz, 1H, pyridin-5), 7.57–7.49 (m, 1H, 3-F-Ph-5), 7.25–7.19 (m, 1H, 3-F-Ph-4). **¹³C NMR** (75 MHz, DMSO-d₆) *δ*: 162.7 (d, *¹J*<sub>C-F</sub> = 242.8 Hz, 3-F-Ph-3), 151.3 (pyridin-6), 149.8 (pyridin-2), 146.6 (C-6), 143.9 (d, *⁴J*<sub>C-F</sub> = 2.9 Hz, C-2), 136.9 (pyridine-4), 136.4 (C-8a), 135.2 (d, *³J*<sub>C-F</sub> = 8.3 Hz, 3-F-Ph-1), 134.5 (pyridin-C-8), 130.9 (d, *³J*<sub>C-F</sub> = 8.4 Hz, 3-F-Ph-5), 128.2 (pyridin-3), 123.7 (pyridin-5), 121.8 (d, *⁴J*<sub>C-F</sub> = 2.4 Hz, 3-F-Ph-6), 116.0 (C-7), 115.2 (d, *²J*<sub>C-F</sub> = 20.6 Hz, 3-F-Ph-4), 115.1 (C-3), 112.2 (d, *²J*<sub>C-F</sub> = 22.7 Hz, 3-F-Ph-2). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –112.9. **m.p.:** 181–185 °C. **LC-MS:** retention time = 2.33 min, m/z [M+H]⁺ = 325.05.

#### 4.1.3.17. 6-Chloro-2-(3-fluorophenyl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (5c).
Column: silica. Eluent: CH₂Cl₂/cyclohexane (2/98 → 80/20), yellow solid, 152 mg, yield: 75 %. **¹H NMR** (300 MHz, CDCl₃) *δ*: 9.15 (dd, J = 3.0, 1.3 Hz, 1H, thien-2), 8.18 (s, 1H, H-3), 7.81 (dd, J = 5.2, 1.3 Hz, 1H, thien-4), 7.79–7.73 (m, 2H, 3-F-Ph-2 & 3-F-Ph-6), 7.50 (dd, J = 5.1, 3.1 Hz, 1H, thien-5), 7.47–7.37 (m, 1H, 3-F-Ph-5), 7.25 (s, 1H, H-7), 7.06 (tdd, J = 8.5, 2.3, 1.2 Hz, 1H, 3-F-Ph-4). **¹³C NMR** (75 MHz, CDCl₃) *δ*: 163.5 (d, *¹J*<sub>C-F</sub> = 245.3 Hz, 3-F-Ph-3), 147.2 (C-6), 144.7 (d, *⁴J*<sub>C-F</sub> = 2.8 Hz, C-2), 136.8 (C-8a), 135.5 (d, *³J*<sub>C-F</sub> = 8.4 Hz, 3-F-Ph-1), 133.2 (C-8*), 132.4 (thien-3*), 131.7 (thien-2), 130.5 (d, *³J*<sub>C-F</sub> = 8.5 Hz, 3-F-Ph-5), 126.7 (thien-5), 126.2 (thien-4), 121.8 (d, *⁴J*<sub>C-F</sub> = 2.8 Hz, 3-F-Ph-6), 115.4 (d, *²J*<sub>C-F</sub> = 21.4 Hz, 3-F-Ph-4), 113.8 (C-3*), 113.7 (C-7*), 113.1 (d, *²J*<sub>C-F</sub> = 22.9 Hz, 3-F-Ph-2). **¹⁹F NMR** (282 MHz, DMSO-d₆) *δ*: –112.8. **m.p.:** 198–202 °C. **LC-MS:** retention time = 3.39 min, m/z [M+H]⁺ = 330.05.

#### 4.1.3.18. 6-Chloro-2-(3-fluorophenyl)-8-(2-methoxypyrimidin-5-yl)imidazo [1,2-b]pyridazine (5d).
Column: silica. Eluent: CH₂Cl₂/cyclohexane (20/80 → 100/0), yellow solid, 109 mg, yield: 50 %. **¹H NMR
(300 MHz, CDCl₃) δ: 9.54 (s, 2H, pyrimidin-4,6), 8.28 (s, 1H, H-3), 7.80-7.70 (m, 2H, 3-F-Ph-2 & 3-F-Ph-6), 7.48-7.37 (m, 1H, 3-F-Ph-5), 7.25 (s, 1H, H-7), 7.08 (tdd, J = 8.4, 2.6, 0.9 Hz, 1H, 3-F-Ph-4), 4.16 (s, 3H, OMe). ¹³C NMR (75 MHz, CDCl₃) δ: despite all our efforts (testing all the deuterated solvents available in the laboratory, increasing the number of scans, increasing the relaxation delay) we have never succeeded in obtaining a useable carbon spectrum. ¹⁹F NMR (282 MHz, CDCl₃) δ: -112.9. m.p.: 257-261 °C. LC-MS: retention time = 2.72 min, m/z [M+H]⁺ = 356.15.

### 4.1.3.19. 6-Chloro-2-(2-fluorophenyl)-8-(1H-indol-4-yl)imidazo [1,2-b] pyridazine (6a).
Column: silica. Eluent: CH₂Cl₂/petroleum ether (50/50 → 60/40), yellow solid, 161 mg, yield: 80 %. ¹H NMR (300 MHz, acetone-d6) δ: 10.64 (bs, 1H, NH), 8.50 (d, TSJ = 4.2 Hz, 1H, H-3), 8.34 (td, J = 7.8, 2.0 Hz, 1H, 2-F-Ph-6), 8.07 (dd, J = 7.5, 0.7, 1H, indol-5), 7.69 (d, J = 8.2 Hz, 1H, indol-7), 7.56 (s, 1H, H-7), 7.54-7.53 (m, 1H, indol-2), 7.46-7.24 (m, 4H, 2-F-Ph-3,4,5 & indol-6), 7.81-7.80 (m, 1H, indol-3). ¹³C NMR (75 MHz, acetone-d6) δ: 161.2 (d, $^{1}J_{C-F}$ = 248.5 Hz, 2-F-Ph-2), 147.7 (C-6), 140.5 (C-8), 138.0 (C-8a), 130.7 (d, $^{3}J_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 129.7 (d, $^{3}J_{C-F}$ = 3.7 Hz, 2-F-Ph-6), 127.45 (indol-3a), 127.38 (indol-2), 125.5 (d, $^{4}J_{C-F}$ = 3.3 Hz, 2-F-Ph-5), 125.4 (indol-4), 123.6 (indol-5), 122.0 (indol-6), 118.0 (C-7), 117.6 (d, $^{4}TSJ_{C-F}$ = 14.6 Hz, C-3), 116.6 (d, $^{2}J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 114.8 (indol-7), 102.0 (indol-3). (Note 1: 2-F-Ph-1, C-2 and indol-7a are missing. Note 2: For the whole series with a 2-fluoro-phenyl group at position 2 we can note that the proton and the carbon at position 3 present abnormally high H-F and C-F coupling constants. This is due to the proximity of fluorine to this position in space, which generates a through space spin-spin coupling). ¹⁹F NMR (282 MHz, acetone-d6) δ: -115.2. m.p.: 237-241 °C. LC-MS: retention time = 3.21 min, m/z [M+H]⁺ = 363.15.

### 4.1.3.20. 6-Chloro-2-(2-fluorophenyl)-8-(pyridin-4-yl)imidazo [1,2-b] pyridazine (6b).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 115 mg, yield: 58 %. ¹H NMR (300 MHz, DMSO-d6) δ: 8.85-8.83 (m, 2H, pyridin-2,6), 8.67 (d, TSJ = 4.0 Hz, 1H, H-3), 8.45-8.42 (m, 2H, pyridin-3,5), 8.33-8.27 (m, 1H, 2-F-Ph-6), 7.95 (s, 1H, H-7), 7.51-7.33 (m, 3H, 2-F-Ph-3,4,5). ¹³C NMR (75 MHz, DMSO-d6) δ: 159.6 (d, $^{1}J_{C-F}$ = 249.1 Hz, 2-F-Ph-2), 150.3 (pyridin-2,6), 146.7 (C-6), 139.3 (pyridin-4), 138.9 (d, $^{3}J_{C-F}$ = 1.8 Hz, C-2), 135.6 (C-8a), 134.3 (C-8), 130.4 (d, $^{3}J_{C-F}$ = 8.4 Hz, 2-F-Ph-4), 128.5 (d, $^{3}J_{C-F}$ = 3.3 Hz, 2-F-Ph-6), 124.9 (d, $^{4}J_{C-F}$ = 3.1 Hz, 2-F-Ph-5), 123.3 (pyridin-3,5), 120.3 (d, $^{2}J_{C-F}$ = 12.3 Hz, 2-F-Ph-1), 117.1 (C-7), 116.9 (d, $^{4TS}J_{C-F}$ = 13.8 Hz, C-3), 116.1 (d, $^{2}J_{C-F}$ = 21.4 Hz, 2-F-Ph-3). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.4. m.p.: 192-196 °C. LC-MS: retention time = 2.33 min, m/z [M+H]⁺ = 325.05.

### 4.1.3.21. 6-Chloro-2-(2-fluorophenyl)-8-(pyridin-3-yl)imidazo [1,2-b] pyridazine (6c).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 102 mg, yield: 34 %. ¹H NMR (300 MHz, DMSO-d6) δ: 9.58 (d, J = 1.8 Hz, 1H, pyridin-2), 8.83 (dt, J = 8.1, 2.1 Hz, 1H, pyridin-4), 8.77 (dd, J = 4.7, 1.4 Hz, 1H, pyridin-6), 8.62 (d, TSJ = 4.0 Hz, 1H, H-3), 8.26 (td, J = 7.8, 2.1 Hz, 1H, 2-F-Ph-6), 7.88 (s, 1H, H-7), 7.65 (dd, J = 7.7, 4.8 Hz, 1H, pyridin-5), 7.49-7.32 (m, 3H, 2-F-Ph-3,4,5). ¹³C NMR (75 MHz, DMSO-d6) δ: 159.6 (d, $^{1}J_{C-F}$ = 249.0 Hz, 2-F-Ph-2), 151.3 (pyridin-6), 149.9 (pyridin-2), 146.7 (C-6), 138.8 (d, $^{3}J_{C-F}$ = 1.6 Hz, C-2), 136.9 (pyridin-4), 135.9 (C-8a), 134.5 (C-8), 130.3 (d, $^{3}J_{C-F}$ = 8.5 Hz, 2-F-Ph-4), 128.4 (d, $^{3}J_{C-F}$ = 3.2 Hz, 2-F-Ph-6), 128.2 (pyridin-3), 124.9 (d, $^{4}J_{C-F}$ = 3.2 Hz, 2-F-Ph-5), 123.7 (pyridin-5), 120.3 (d, $^{2}J_{C-F}$ = 12.2 Hz, 2-F-Ph-1), 116.9 (d, $^{4TS}J_{C-F}$ = 13.7 Hz, C-3), 116.3 (C-7), 116.0 (d, $^{2}J_{C-F}$ = 21.3 Hz, 2-F-Ph-3). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -113.5. m.p.: 179-183 °C. LC-MS: retention time = 2.54 min, m/z [M+H]⁺ = 325.05.

### 4.1.3.22. 6-Chloro-2-(2-fluorophenyl)-8-(6-aminopyridin-3-yl)imidazo [1,2-b]pyridazine (6d).
Column: silica. Eluent: CH₂Cl₂/EtOAc (50/50 → 0/100), yellow solid, 201 mg, yield: 64 %. ¹H NMR (300 MHz, acetone-d6) δ: 9.37 (d, J = 2.3 Hz, 1H, pyridin-2), 8.62 (dd, J = 8.8, 2.5 Hz, 1H, pyridin-4, 8.48-8.37 (m, 2H, H-3 & 2-F-Ph-6), 7.51 (s, 1H, H-7), 7.48-7.22 (m, 3H, 2-F-Ph-3,4,5), 6.76 (d, J = 8.8 Hz, 1H, pyridin-5), 6.16 (bs, 2H, NH₂). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.2 (pyridin-6), 161.2 (d, $^{1}J_{C-F}$ = 248.5 Hz, 2-F-Ph-2), 151.6 (pyridin-2), 147.9 (C-6), 139.6 (d, $^{3}J_{C-F}$ = 1.9 Hz, C-2), 138.6 (pyridin-4), 137.2 (C-8*), 136.9 (C-8a*), 130.7 (d, $^{3}J_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 129.5 (d, $^{3}J_{C-F}$ = 3.7 Hz, 2-F-Ph-6), 125.6 (d, $^{4}J_{C-F}$ = 3.3 Hz, 2-F-Ph-5), 122.0 (d, $^{2}J_{C-F}$ = 12.5 Hz, 2-F-Ph-1), 117.9 (pyridin-3), 117.4 (d, $^{4TS}J_{C-F}$ = 14.6 Hz, C-3), 116.6 (d, $^{2}J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 113.1 (C-7), 108.5 (pyridin-5). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -115.1. m.p.: 207-211 °C. LC-MS: retention time = 0.96 min, m/z [M+H]⁺ = 340.05.

### 4.1.3.23. 6-Chloro-2-(2-fluorophenyl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (6e).
Column: silica. Eluent: CH₂Cl₂/cyclohexane (5/95 → 40/60), yellow solid, 113 mg, yield: 56 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.17 (dd, J = 3.0, 1.2 Hz, 1H, thien-2), 8.44 (td, J = 7.6, 2.2 Hz, 1H, 2-F-Ph-6), 8.37 (d, TSJ = 4.1 Hz, 1H, H-3), 7.82 (dd, J = 5.2, 1.3 Hz, 1H, thien-4), 7.50 (dd, J = 5.1, 3.1 Hz, 1H, thien-5), 7.40-7.27 (m, 2H, 2-F-Ph-4,5), 7.25 (s, 1H, H-7), 7.17 (ddd, J = 11.3, 7.7, 1.5 Hz, 1H, 2-F-Ph-3). ¹³C NMR (75 MHz, CDCl₃) δ: 160.5 (d, $^{1}J_{C-F}$ = 249.9 Hz, 2-F-Ph-2), 147.0 (C-6), 139.5 (d, $^{3}J_{C-F}$ = 2.0 Hz, C-2), 136.0 (C-8a), 133.3 (C-8*), 132.1 (thien-3*), 131.5 (thien-2), 129.7 (d, $^{3}J_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 128.7 (d, $^{3}J_{C-F}$ = 3.6 Hz, 2-F-Ph-6), 126.6 (thien-5), 126.2 (thien-4), 124.6 (d, $^{4}J_{C-F}$ = 3.3 Hz, 2-F-Ph-5), 121.1 (d, $^{2}J_{C-F}$ = 12.2 Hz, 2-F-Ph-1), 117.1 (d, $^{4TS}J_{C-F}$ = 15.3 Hz, C-3), 115.9 (d, $^{2}J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 113.7 (C-7). ¹⁹F NMR (282 MHz, CDCl₃) δ: -113.5. m.p.: 160-164 °C. LC-MS: retention time = 3.56 min, m/z [M+H]⁺ = 329.95.

### 4.1.3.24. 6-Chloro-2-(2-fluorophenyl)-8-(2-methoxypyrimidin-5-yl)imidazo [1,2-b]pyridazine (6f).
Column: alumina. Eluent: CH₂Cl₂/petroleum ether (80/20), yellow solid, 145 mg, yield: 66 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.55 (bs, 2H, pyrimidin-4,6), 8.45 (d, TSJ = 4.1 Hz, 1H, H-3), 8.41 (td, J = 7.7, 1.9 Hz, 1H, 2-F-Ph-6), 7.40-7.13 (m, 4H, H-7 & 2-F-Ph-3,4,5), 4.16 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 166.6 (pyrimidin-2), 160.6 (d, $^{1}J_{C-F}$ = 249.9 Hz, 2-F-Ph-2), 159.9 (pyrimidin-4,6), 146.9 (C-6), 140.4 (d, $^{3}J_{C-F}$ = 2.1 Hz, C-2), 135.6 (C-8a), 132.2 (C-8), 130.1 (d, $^{3}J_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 128.8 (d, $^{3}J_{C-F}$ = 3.3 Hz, 2-F-Ph-6), 124.7 (d, $^{4}J_{C-F}$ = 3.2 Hz, 2-F-Ph-5), 120.72 (pyrimidin-5), 120.71 (d, $^{2}J_{C-F}$ = 11.9 Hz, 2-F-Ph-1), 117.6 (d, $^{4TS}J_{C-F}$ = 15.3 Hz, C-3), 116.0 (d, $^{2}J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 113.8 (C-7), 55.7 (CH₃). ¹⁹F NMR (282 MHz, CDCl₃) δ: -113.5. m.p.: 235-239 °C. LC-MS: retention time = 2.92 min, m/z [M+H]⁺ = 356.05.

### 4.1.3.25. 6-Chloro-8-(1H-indol-4-yl)-2-(2-methoxyphenyl)imidazo [1,2-b]pyridazine (7a).
Column: silica. Eluent: CH₂Cl₂/petroleum ether (70/30), yellow solid, 84 mg, yield: 76 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.63 (s, 1H, H-3), 8.48 (bs, 1H, NH), 8.42 (dd, J = 7.7, 1.8 Hz, 1H, 2-MeO-Ph-6), 8.07 (dd, J = 7.4, 0.9 Hz, 1H, indol-5), 7.51 (d, J = 8.2 Hz, 1H, indol-7), 7.40 (s, 1H, H-7), 7.38-7.27 (m, 3H, indol-2,6 & 2-MeO-Ph-4), 7.05 (td, J = 7.6, 1.1 Hz, 1H, 2-MeO-Ph-5), 7.01 (d, J = 8.3 Hz, 1H, 2-MeO-Ph-3), 6.78 (ddd, J = 3.1, 2.0, 0.9 Hz, 1H, indol-3), 4.01 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 157.1 (2-MeO-Ph-2), 146.4 (C-6), 142.0 (C-2), 139.0 (C-8), 136.7 (C-8a*), 136.5 (indol-7a*), 129.4 (2-MeO-Ph-4), 129.0 (2-MeO-Ph-6), 126.5 (indol-4), 125.5 (indol-2), 125.3 (2-MeO-Ph-1), 123.1 (indol-5), 122.1 (indol-6), 121.1 (2-MeO-Ph-5), 117.8 (C-3), 116.7 (C-7), 113.5 (indol-7), 111.0 (2-MeO-Ph-3), 102.3 (indol-3), 55.6 (CH₃). (Note: indol-3a is missing). m.p.: 203-207 °C. LC-MS: retention time = 3.05 min, m/z [M+H]⁺ = 375.15.

### 4.1.3.26. 6-Chloro-2-(2-methoxyphenyl)-8-(thien-3-yl)imidazo [1,2-b] pyridazine (7b).
Column: silica. Eluent: CH₂Cl₂/petroleum ether (20/80 → 30/70), yellow solid, 81 mg, yield: 54 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.19 (d, J = 2.5 Hz, 1H, thien-2), 8.54 (s, 1H, H-3), 8.51 (d, J = 7.7 Hz, 1H, 2-MeO-Ph-6), 7.83 (d, J = 5.1 Hz, 1H, thien-4), 7.48 (dd, J = 4.8, 3.2 Hz, 1H, thien-5), 7.35 (t, J = 7.7 Hz, 1H, 2-MeO-Ph-4), 7.22 (s, pyridin-4), 8.48-8.37 (m, 2H, H-3 & 2-F-Ph-6), 7.51 (s, 1H, H-7), 7.48-7.22 (m, 3H, 2-F-Ph-3,4,5), 6.76 (d, J = 8.8 Hz, 1H, pyridin-5), 6.16 (bs, 2H, NH₂). ¹³C NMR (75 MHz, DMSO-d6) δ: 162.2 (pyridin-6), 161.2 (d, $^{1}J_{C-F}$ = 248.5 Hz, 2-F-Ph-2), 151.6 (pyridin-2), 147.9 (C-6), 139.6 (d, $^{3}J_{C-F}$ = 1.9 Hz, C-2), 138.6 (pyridin-4), 137.2 (C-8*), 136.9 (C-8a*), 130.7 (d, $^{3}J_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 129.5 (d, $^{3}J_{C-F}$ = 3.7 Hz, 2-F-Ph-6), 125.6 (d, $^{4}J_{C-F}$ = 3.3 Hz, 2-F-Ph-5), 122.0 (d, $^{2}J_{C-F}$ = 12.5 Hz, 2-F-Ph-1), 117.9 (pyridin-3), 117.4 (d, $^{4TS}J_{C-F}$ = 14.6 Hz, C-3), 116.6 (d, $^{2}J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 113.1 (C-7), 108.5 (pyridin-5). ¹⁹F NMR (282 MHz, DMSO-d6) δ: -115.1. m.p.: 207-211 °C. LC-MS: retention time = 0.96 min, m/z [M+H]⁺ = 340.05.
1H, H-7), 7.13 (t, J = 7.4 Hz, 1H, 2-MeO-Ph-5), 7.01 (d, J = 8.2 Hz, 1H, 2-MeO-Ph-3), 4.00 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 157.2 (2-MeO-Ph-2), 146.5 (C-6), 141.7 (C-2), 135.5 (C-8a), 133.6 (C-8), 131.9 (thien-3), 131.4 (thien-2), 129.5 (2-MeO-Ph-4), 128.8 (2-MeO-Ph-6), 126.5 (thien-4*), 126.3 (thien-5*), 121.9 (2-MeO-Ph-1), 121.1 (2-MeO-Ph-5), 117.8 (C-3), 113.2 (C-7), 111.1 (2-MeO-Ph-3), 55.6 (CH₃). m.p.: 194–198 °C. LC-MS: retention time = 3.45 min, m/z [M+H]⁺ = 342.05.

#### 4.1.3.27. 6-Chloro-8-(1H-indol-4-yl)-2-phenylimidazo [1,2-b]pyridazine [8a].
Column: alumina. Eluent: CH₂Cl₂/petroleum ether (20/80 → 40/60), yellow solid, 330 mg, yield: 60 %. ¹H NMR (300 MHz, acetone-d6) δ: 10.65 (bs, 1H, NH), 8.68 (s, 1H, H-3), 8.13–8.04 (m, 3H, indol-5 & Ph-2,6), 7.68 (d, J = 8.1 Hz, 1H, indol-7), 7.56–7.51 (m, 2H, H-7 & indol-2), 7.49–7.41 (m, 2H, Ph-3,5), 7.39–7.30 (m, 2H, indol-6 & Ph-4), 6.84–6.76 (m, 1H, indol-3). ¹³C NMR (75 MHz, acetone-d6) δ: 147.3 (C-6*), 146.8 (C-2*), 140.3 (C-8), 138.4 (C-8a), 137.9 (indol-7a), 134.6 (Ph-1), 129.5 (Ph-3,5), 129.2 (Ph-4), 127.4 (indol-4), 127.3 (indol-2), 126.8 (Ph-2,6), 125.5 (indol-3a), 123.5 (indol-5), 121.9 (indol-6), 117.4 (C-7), 114.7 (C-3*), 114.6 (indol-7*), 101.9 (indol-3). m.p.: 212–216 °C. LC-MS: retention time = 3.00 min, m/z [M+H]⁺ = 345.15.

#### 4.1.3.28. 6-Chloro-2-phenyl-8-(thien-3-yl)imidazo [1,2-b]pyridazine (8b).
Column: silica. Eluent: CH₂Cl₂/cyclohexane (2/98 → 40/60), yellow solid, 175 mg, yield: 87 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.19 (dd, *J* = 3.0, 1.2 Hz, 1H, thien-2), 8.21 (s, 1H, H-3), 8.08–7.99 (m, 2H, Ph-2,6), 7.83 (dd, *J* = 5.2, 1.2 Hz, 1H, thien-4), 7.53–7.44 (m, 3H, thien-5 & Ph-3,5), 7.42–7.34 (m, 1H, Ph-4), 7.24 (s, 1H, H-7). ¹³C NMR (75 MHz, CDCl₃) δ: 146.9 (C-6), 146.0 (C-2), 136.8 (C-8a), 133.3 (C-8*), 133.2 (thien-3*), 132.2 (Ph-1), 131.6 (thien-2), 129.0 (Ph-3,5), 128.7 (Ph-4), 126.6 (thien-5), 126.25 (thien-4), 126.22 (Ph-2,6), 113.5 (C-7), 113.3 (C-3). m.p.: 186–200 °C. LC-MS: retention time = 3.40 min, m/z [M+H]⁺ = 312.04.

#### 4.1.3.29. 6-Chloro-8-(2-methoxypyrimidin-5-yl)-2-phenylimidazo [1,2-b]pyridazine (8c).
Column: silica. Eluent: ethyl acetate/cyclohexane (5/95 → 30/70), yellow solid, 113 mg, yield: 52 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.55 (s, 2H, pyrimidin-4,6), 8.27 (s, 1H, H-3), 8.06–7.97 (m, 2H, Ph-2,6), 7.50–7.43 (m, 2H, Ph-3,5), 7.42–7.35 (m, 1H, Ph-4), 7.22 (s, 1H, H-7), 4.15 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 166.4 (pyrimidin-2), 159.8 (pyrimidin-4,6), 146.7 (C-6*), 146.6 (C-2*), 136.1 (C-8a), 132.6 (Ph-1), 132.0 (C-8), 128.9 (Ph-3,4,5), 126.1 (Ph-2,6), 120.6 (pyrimidin-5), 113.6 (C-3), 113.3 (C-7), 55.6 (CH₃). m.p.: 205–209 °C. LC-MS: retention time = 2.66 min, m/z [M+H]⁺ = 338.15.

### 4.1.4. General procedure for nucleophilic aromatic substitution using nitrogen nucleophiles, synthesis of 9a-o, 10a-d, 11a-f, 12a-b, 13a-c, 14a-g, 15a-c and 16

Compounds 4a-n, 5a-d, 6a-f, 7a-b and 8a-c were weighed into a glass tube with a screw cap (ca. 100–150 mg). The nitrogen nucleophile was added (18 eq). The tube was sealed, and the medium was heated for 6–24 h, depending on the base, at 170–180 °C. After cooling, the mixture was evaporated under vacuum, and the residue was purified by alumina or silica column chromatography.

#### 4.1.4.1. 8-Bromo-2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (9a).
Column: alumina. Eluent: CH₂Cl₂, white solid, 88 mg, yield: 72 %. ¹H NMR (300 MHz, CD₂Cl₂) δ: 7.97 (s, 1H, H-3), 7.93–7.87 (m, 2H, 4-F-Ph-2,6), 7.18–7.06 (m, 2H, 4-F-Ph-3,5), 6.09 (s, 1H, H-7), 4.16–4.07 (m, 4H, 2 CH₂ piperazine), 2.64–2.55 (m, 4H, 2 CH₂ piperazine), 2.33 (s, 3H, CH₃). ¹³C NMR (75 MHz, CD₂Cl₂) δ: 163.2 (d, ¹J<sub>C-F</sub> = 246.3 Hz, 4-F-Ph-4), 148.2 (C-6), 145.0 (C-2), 141.6 (C-8a), 134.3 (C-8), 130.4 (d, ⁴J<sub>C-F</sub> = 3.3 Hz, 4-F-Ph-1), 127.9 (d, ³J<sub>C-F</sub> = 8.1 Hz, 4-F-Ph-2,6), 116.07 (d, ²J<sub>C-F</sub> = 21.7 Hz, 4-F-Ph-3,5), 113.3 (C-3), 96.6 (C-7), 55.2 (2 CH₂ piperazine), 48.3 (2 CH₂ piperazine), 46.3 (CH₃). ¹⁹F NMR (282 MHz, CD₂Cl₂) δ: –114.1. LC-MS: retention time = 0.64 min, m/z [M + H - C₂H₆N]⁺ = 346.15 et [M - C₅H₁₂N₂]⁺ = 289.14.

#### 4.1.4.2. 2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-phenylimidazo [1,2-b]pyridazine (9b).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 290 mg, yield: 83 %. ¹H NMR (300 MHz, acetone-d6) δ: 8.42–8.36 (m, 2H, Ph-2,6), 8.31 (s, 1H, H-3), 8.12–8.02 (m, 2H, 4-F-Ph-2,6), 7.62–7.51 (m, 3H, Ph-3,4,5), 7.32 (s, 1H, H-7), 7.25–7.14 (m, 2H, 4-F-Ph-3,5), 3.67–3.55 (m, 4H, 2 CH₂ piperazine), 2.58–2.46 (m, 4H, 2 CH₂ piperazine), 2.28 (s, 3H, CH₃). ¹³C NMR (75 MHz, acetone-d6) δ: 163.2 (d, ¹J<sub>C-F</sub> = 244.1 Hz, 4-F-Ph-4), 156.6 (C-6), 143.3 (C-2), 137.5 (C-8), 136.8 (C-8a), 135.4 (Ph-1), 132.0 (d, ⁴J<sub>C-F</sub> = 3.0 Hz, 4-F-Ph-1), 130.6 (Ph-4), 130.2 (Ph-2,6), 129.2 (Ph-3,5), 128.1 (d, ³J<sub>C-F</sub> = 8.1 Hz, 4-F-Ph-2,6), 116.1 (d, ²J<sub>C-F</sub> = 21.7 Hz, 4-F-Ph-3,5), 113.8 (C-3), 108.1 (C-7), 55.5 (2 CH₂ piperazine), 47.0 (2 CH₂ piperazine), 46.4 (CH₃). ¹⁹F NMR (282 MHz, acetone-d6) δ: –116.6. m.p.: 192–196 °C. LC-MS: retention time = 0.89 min, m/z [M+H]⁺ = 388.15.

#### 4.1.4.3. 2-(4-fluorophenyl)-8-(1H-indol-4-yl)-6-(4-methylpiperazin-1-yl) imidazo [1,2-b]pyridazine (9c).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 150 mg, yield: 85 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.53 (bs, 1H, NH), 8.03 (s, 1H, H-3), 7.96–7.82 (m, 3H, 4-F-Ph-2,6 & indol-5), 7.46 (d, *J* = 8.1 Hz, 1H, indol-7), 7.33 (t<sub>AB</sub>, *J* = 7.6 Hz, 1H, indol-6), 7.24–7.21 (m, 1H, indol-2), 7.13 (s, 1H, H-7), 7.09–7.04 (m, 2H, 4-F-Ph-3,5), 6.70–6.68 (m, 1H, indol-3), 3.67–3.53 (m, 4H, 2 CH₂ piperazine), 2.66–2.53 (m, 4H, 2 CH₂ piperazine), 2.38 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.6 (d, ¹J<sub>C-F</sub> = 246.0 Hz, 4-F-Ph-4), 155.5 (C-6), 143.0 (C-2), 138.1 (C-8*), 136.7 (C-8a*), 136.6 (indol-7a*), 130.7 (d, ⁴J<sub>C-F</sub> = 3.2 Hz, 4-F-Ph-1), 127.5 (d, ³J<sub>C-F</sub> = 8.0 Hz, 4-F-Ph-2,6), 126.7 (indol-4*), 126.5 (indol-3a*), 125.2 (indol-2), 122.1 (indol-5), 121.9 (indol-6), 115.6 (d, ²J<sub>C-F</sub> = 21.6 Hz, 4-F-Ph-3,5), 112.9 (C-3*), 112.8 (indol-7*), 109.0 (C-7), 102.1 (indol-3), 54.8 (2 CH₂ piperazine), 46.5 (2 CH₂ piperazine), 46.3 (CH₃). ¹⁹F NMR (282 MHz, CDCl₃) δ: –115.0. m.p.: 242–246 °C. LC-MS: retention time = 0.59 min, m/z [M+H]⁺ = 427.26.

#### 4.1.4.4. 2-(4-fluorophenyl)-8-(1H-indol-5-yl)-6-(4-methylpiperazin-1-yl) imidazo [1,2-b]pyridazine (9d).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 113 mg, yield: 68 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.33 (s, 1H, NH), 8.71 (s, 1H, indol-4), 8.51 (s, 1H, H-3), 8.15 (d, *J* = 8.6 Hz, 1H, indol-6), 8.03 (dd, *J* = 8.3, 5.7 Hz, 2H, 4-F-Ph-2,6), 7.55 (d, *J* = 8.6 Hz, 1H, indol-7), 7.45 (s, 1H, indol-2), 7.34 (s, 1H, H-7), 7.27 (t, *J* = 8.8 Hz, 2H, 4-F-Ph-3,5), 6.60 (s, 1H, indol-3), 3.56 (bs, 4H, 2 CH₂ piperazine), 2.25 (s, 3H, CH₃). (Note: two of the four CH₂ are overlapped with DMSO). ¹³C NMR (75 MHz, DMSO-d₆) δ: 161.6 (d, ¹J<sub>C-F</sub> = 243.7 Hz, 4-F-Ph-4), 155.4 (C-6), 141.3 (C-2), 137.5 (C-8), 136.7 (indol-7a), 135.9 (C-8a), 130.7 (d, ⁴J<sub>C-F</sub> = 2.9 Hz, 4-F-Ph-1), 127.6 (indol-3a), 127.0 (d, ³J<sub>C-F</sub> = 8.1 Hz, 4-F-Ph-2,6), 126.3 (indol-5), 124.5 (indol-2), 122.2 (indol-4*), 121.7 (indol-6*), 115.4 (d, ²J<sub>C-F</sub> = 21.6 Hz, 4-F-Ph-3,5), 113.1 (C-3), 111.2 (indol-7), 106.0 (C-7), 102.1 (indol-3), 54.2 (2 CH₂ piperazine), 45.7 (2 CH₂ piperazine & CH₃). ¹⁹F NMR (282 MHz, DMSO-d₆) δ: –115.0. m.p.: 245–249 °C. LC-MS: retention time = 0.67 min, m/z [M+H]⁺ = 427.26.

#### 4.1.4.5. 2-(4-fluorophenyl)-8-(1H-indol-6-yl)-6-(4-methylpiperazin-1-yl) imidazo [1,2-b]pyridazine (9e).
Column: silica. Eluent: acetone/MeOH (98/2), yellow solid, 74 mg, yield: 53 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.72 (bs, 1H, NH), 8.66 (s, 1H, indol-7), 8.01 (s, 1H, H-3), 7.96–7.91 (m, 2H, 4-F-Ph-2,6), 7.74 (d<sub>AB</sub>, *J* = 8.3 Hz, 1H, indol-4*), 7.68 (d<sub>AB</sub>, *J* = 8.4 Hz, 1H, indol-5*), 7.08 (m, 2H, 4-F-Ph-3,5), 7.00 (s, 1H, H-7), 6.57 (bs, 1H, indol-3), 3.68–3.49 (m, 4H, 2 CH₂ piperazine), 2.68–2.53 (m, 4H, 2 CH₂ piperazine), 2.39 (s, 3H, CH₃). (Note: indol-2 is overlapped with CHCl₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.6 (d, ¹J<sub>C-F</sub> = 246.1 Hz, 4-F-Ph-4), 155.6 (C-6), 142.9 (C-2), 138.5 (C-8a), 136.9 (indol-7a), 135.9 (C-8), 130.7 (d, ⁴J<sub>C-F</sub> = 2.9 Hz, 4-F-Ph-1), 129.4 (indol-6), 127.8 (indol-3a), 127.4 (d, ³J<sub>C-F</sub> = 8.0 Hz, 4-F-Ph-2,6), 126.3 (indol-2), 121.0 (indol-4), 127.4 (d, ²J<sub>C-F</sub> = 8.0 Hz, 4-F-Ph-3,5), 113.3 (C-3), 96.6 (C-7), 55.2 (2 CH₂ piperazine), 48.3 (2 CH₂ piperazine), 46.3 (CH₃). ¹⁹F NMR (282 MHz, CD₂Cl₂) δ: –114.1. LC-MS: retention time = 0.64 min, m/z [M + H - C₂H₆N]⁺ = 346.15 et [M - C₅H₁₂N₂]⁺ = 289.14.
120.0 (indol-5), 115.6 (d, $^2J_{CF}$ = 21.5 Hz, 4-F-Ph-3,5), 113.4 (indol-7), 112.9 (C-3), 106.8 (C-7), 102.8 (indol-3), 54.8 (2 CH$_2$ piperazine), 46.5 (2 CH$_2$ piperazine), 46.3 (CH$_3$). $^{19}$F NMR (282 MHz, CDCl$_3$) δ: −114.9. m.p.: 231–235 °C. **LC-MS**: retention time = 0.76 min, m/z [M+H]$^+$ = 427.26.

### 4.1.4.6. 2-(4-fluorophenyl)-8-(1H-indazol-4-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (9f).
Column: silica. Eluent: CH$_2$Cl$_2$/MeOH (90/10), yellow solid, 87 mg, yield: 84 %. $^1$H NMR (300 MHz, CDCl$_3$) δ: 8.24 (s, 1H, indazol-3), 8.06 (s, 1H, H-3), 7.94-7.84 (m, 3H, 4-F-Ph-2,6 & indazol-5), 7.55 (d$_{AB}$, J = 8.3 Hz, 1H, indazol-7), 7.49 (dd$_{AB}$, J = 8.4, 6.8 Hz, 1H, indazol-6), 7.10-7.04 (m, 3H, 4-F-Ph-3,5 & H-7), 3.68-3.55 (m, 4H, 2 CH$_2$ piperazine), 2.65-2.56 (m, 4H, 2 CH$_2$ piperazine), 2.39 (s, 3H, CH$_3$). $^{13}$C NMR (75 MHz, CDCl$_3$) δ: 162.7 (d, $^1J_{CF}$ = 246.5 Hz, 4-F-Ph-4), 155.4 (C-6), 143.4 (C-2), 140.8 (C-8), 136.6 (indazol-7a), 136.1 (C-8a), 135.0 (indazol-3), 130.4 (d, $^4J_{CF}$ = 3.1 Hz, 4-F-Ph-1), 127.9 (indazol-4), 127.5 (d, $^3J_{CF}$ = 8.1 Hz, 4-F-Ph-2,6), 126.9 (indazol-6), 123.2 (indazol-5), 122.0 (indazol-3a), 115.7 (d, $^2J_{CF}$ = 21.6 Hz, 4-F-Ph-3,5), 113.1 (C-3), 111.3 (indazol-7), 108.9 (C-7), 54.7 (2 CH$_2$ piperazine), 46.5 (2 CH$_2$ piperazine), 46.3 (CH$_3$). $^{19}$F NMR (282 MHz, CDCl$_3$) δ: −114.9. m.p.: 190–194 °C. **LC-MS**: retention time = 0.56 min, m/z [M+H]$^+$ = 428.26.

### 4.1.4.7. 2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(pyridin-4-yl) imidazo [1,2-b]pyridazine (9g).
Column: alumina. Eluent: CH$_2$Cl$_2$/MeOH (98/2), yellow solid, 97 mg, yield: 50 %. $^1$H NMR (300 MHz, DMSO-d$_6$) δ: 8.85-8.74 (m, 2H, pyridin-2,6), 8.57 (s, 1H, H-3), 8.45-8.34 (m, 2H, pyridin-3,5), 8.12-7.90 (m, 2H, 4-F-Ph-2,6), 7.56 (s, 1H, H-7), 7.40-7.12 (m, 2H, 4-F-Ph-3,5), 3.68-3.47 (m, 4H, 2 CH$_2$ piperazine), 2.24 (s, 3H, CH$_3$). (Note: two of the four CH$_2$ are overlapped with DMSO). $^{13}$C NMR (75 MHz, DMSO-d$_6$) δ: 161.8 (d, $^1J_{CF}$ = 244.2 Hz, 4-F-Ph-4), 155.1 (C-6), 150.0 (pyridin-2,6), 141.9 (C-2), 141.0 (pyridin-4), 134.6 (C-8*), 133.0 (C-8a*), 130.3 (d, $^4J_{CF}$ = 2.8 Hz, 4-F-Ph-1), 127.1 (d, $^3J_{CF}$ = 8.2 Hz, 4-F-Ph-2,6), 123.2 (pyridin-3,5), 115.5 (d, $^2J_{CF}$ = 21.5 Hz, 4-F-Ph-3,5), 113.6 (C-3), 108.4 (C-7), 54.1 (2 CH$_2$ piperazine), 45.7 (2 CH$_2$ piperazine), 45.6 (CH$_3$). $^{19}$F NMR (282 MHz, DMSO-d$_6$) δ: −114.6. m.p.: 213–217 °C. **LC-MS**: retention time = 0.53 min, m/z [M+H]$^+$ = 389.25.

### 4.1.4.8. 2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(pyridin-3-yl) imidazo [1,2-b]pyridazine (9h).
Column: alumina. Eluent: CH$_2$Cl$_2$/MeOH (99/1), yellow solid, 152 mg, yield: 50 %. $^1$H NMR (300 MHz, DMSO-d$_6$) δ: 9.52 (d, J = 1.7 Hz, 1H, pyridin-2), 8.80-8.75 (m, 1H, pyridin-4), 8.72 (dd, J = 4.8, 1.4 Hz, 1H, pyridin-6), 8.56 (s, 1H, H-3), 8.01 (dd, J = 8.7, 5.6 Hz, 2H, 4-F-Ph-3,5), 7.61 (dd, J = 7.9, 4.9 Hz, 1H, pyridin-5), 7.51 (s, 1H, H-7), 7.26 (t, J = 8.9 Hz, 2H, 4-F-Ph-2,6), 3.57 (bs, 4H, 2 CH$_2$ piperazine), 2.25 (s, 3H, CH$_3$). (Note: two of the four CH$_2$ are overlapped with DMSO). $^{13}$C NMR (75 MHz, CD$_2$Cl$_2$) δ: 163.1 (d, $^1J_{CF}$ = 246.1 Hz, 4-F-Ph-4), 150.8 (pyridin-6), 148.7 (pyridin-2), 145.0 (C-6), 141.8 (C-2), 135.0 (pyridin-4*), 134.7 (C-8*), 133.3 (pyridin-3), 130.7 (d, $^4J_{CF}$ = 3.1 Hz, 4-F-Ph-1), 127.8 (d, $^3J_{CF}$ = 8.1 Hz, 4-F-Ph-2,6), 124.0 (pyridin-5), 116.0 (d, $^2J_{CF}$ = 21.8 Hz, 4-F-Ph-3,5), 113.2 (C-7), 94.7 (C-3), 55.3 (2 CH$_2$ piperazine), 48.4 (2 CH$_2$ piperazine), 46.4 (CH$_3$). (Note: C-8a is missing). $^{19}$F NMR (282 MHz, DMSO-d$_6$) δ: −116.6. m.p.: 201–205 °C. **LC-MS**: retention time = 0.54 min, m/z [M+H]$^+$ = 389.25.

### 4.1.4.9. 2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (9i).
Column: alumina. Eluent: CH$_2$Cl$_2$/MeOH (100/0 → 98/2), yellow solid, 135 mg, yield: 90 %. $^1$H NMR (300 MHz, acetone-d$_6$) δ: 9.27 (dd, J = 3.0, 1.2 Hz, 1H, thien-2), 8.28 (s, 1H, H-3), 8.22–8.00 (m, 3H, thien-4 & 4-F-Ph-2,6), 7.64 (dd, J = 5.2, 3.1 Hz, 1H, thien-5), 7.48 (s, 1H, H-7), 7.27–7.12 (m, 2H, 4-F-Ph-3,5), 3.70–3.49 (m, 4H, 2 CH$_2$ piperazine), 2.56–2.45 (m, 4H, 2 CH$_2$ piperazine), 2.27 (s, 3H, CH$_3$). $^{13}$C NMR (75 MHz, acetone-d$_6$) δ: 163.2 (d, $^1J_{CF}$ = 244.1 Hz, 4-F-Ph-4), 156.7 (C-6), 143.0 (C-2), 136.3 (C-8a*), 135.9 (C-8*), 132.0 (d, $^4J_{CF}$ = 3.2 Hz, 4-F-Ph-1), 131.5 (thien-3), 130.3 (thien-2), 128.1 (d, $^3J_{CF}$ = 8.1 Hz, 4-F-Ph-2,6), 127.7 (thien-4), 126.7 (thien-5), 116.1 (d, $^2J_{CF}$ = 21.7 Hz, 4-F-Ph-3,5), 113.7 (C-3), 106.4 (C-7), 55.5 (2 CH$_2$ piperazine), 47.1 (2 CH$_2$ piperazine), 46.4 (CH$_3$). $^{19}$F NMR (282 MHz, acetone-d$_6$) δ: −116.6. m.p.: 182–186 °C. **LC-MS**: retention time = 1.06 min, m/z [M+H]$^+$ = 394.15.

### 4.1.4.10. 2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(thien-2-yl) imidazo [1,2-b]pyridazine (9j).
Column: alumina. Eluent: CH$_2$Cl$_2$/petroleum ether (60/40), yellow solid, 87 mg, yield: 73 %. $^1$H NMR (300 MHz, CDCl$_3$) δ: 8.50 (dd, J = 3.7, 1.0 Hz, 1H, thien-3), 7.99–7.94 (m, 3H, H-3 & 4-F-Ph-2,6), 7.54 (dd, J = 5.1, 1.0 Hz, 1H, thien-5), 7.24 (dd, J = 5.1, 3.8 Hz, 1H, thien-4), 7.17–7.08 (m, 2H, 4-F-Ph-3,5), 6.99 (s, 1H, H-7), 3.67–3.49 (m, 4H, 2 CH$_2$ piperazine), 2.70–2.54 (m, 4H, 2 CH$_2$ piperazine), 2.40 (s, 3H, CH$_3$). $^{13}$C NMR (75 MHz, CDCl$_3$) δ: 163.0 (d, $^1J_{CF}$ = 242.1 Hz, 4-F-Ph-4), 155.4 (C-6), 143.0 (C-2), 136.5 (C-8a), 135.1 (C-8), 131.0 (thien-5), 130.8 (thien-2), 130.5 (d, $^4J_{CF}$ = 3.2 Hz, 4-F-Ph-1), 128.9 (thien-3), 128.4 (thien-4), 127.4 (d, $^3J_{CF}$ = 8.0 Hz, 4-F-Ph-2,6), 115.7 (d, $^2J_{CF}$ = 21.6 Hz, 4-F-Ph-3,5), 112.9 (C-3), 104.0 (C-7), 54.8 (2 CH$_2$ piperazine), 46.5 (2 CH$_2$ piperazine), 46.2 (CH$_3$). $^{19}$F NMR (282 MHz, CDCl$_3$) δ: −114.7. m.p.: 165–169 °C. **LC-MS**: retention time = 1.01 min, m/z [M+H]$^+$ = 394.25.

### 4.1.4.11. 2-(4-fluorophenyl)-8-(1-methyl-1H-pyrazol-4-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (9k).
Column: alumina. Eluent: CH$_2$Cl$_2$, yellow solid, 362 mg, yield: 88 %. $^1$H NMR (300 MHz, CDCl$_3$) δ: 8.84 (s, 1H, pyrazol-5), 8.12 (s, 1H, pyrazol-3), 8.01–7.86 (m, 3H, H-3 & 4-F-Ph-2,6), 7.19–7.06 (m, 2H, 4-F-Ph-3,5), 6.92 (s, 1H, H-7), 4.04 (s, 3H, CH$_3$ pyrazole), 3.66–3.49 (m, 4H, 2 CH$_2$ piperazine), 2.68–2.54 (m, 4H, 2 CH$_2$ piperazine), 2.38 (s, 3H, CH$_3$ piperazine). $^{13}$C NMR (75 MHz, CDCl$_3$) δ: 162.6 (d, $^1J_{CF}$ = 246.3 Hz, 4-F-Ph-4), 155.6 (C-6), 142.5 (C-2), 138.0 (pyrazol-3), 135.6 (C-8a), 132.7 (pyrazol-5), 130.6 (d, $^4J_{CF}$ = 3.1 Hz, 4-F-Ph-1), 129.5 (C-8), 127.3 (d, $^3J_{CF}$ = 8.1 Hz, 4-F-Ph-2,6), 116.0 (pyrazol-4), 115.7 (d, $^2J_{CF}$ = 21.7 Hz, 4-F-Ph-3,5), 112.8 (C-3), 102.9 (C-7), 54.8 (2 CH$_2$ piperazine), 46.6 (2 CH$_2$ piperazine), 46.3 (CH$_3$ piperazine), 39.5 (3H pyrazole). $^{19}$F NMR (282 MHz, CDCl$_3$) δ: −114.7. m.p.: 246–250 °C. **LC-MS**: retention time = 0.59 min, m/z [M+H]$^+$ = 392.15.

### 4.1.4.12. 2-(4-fluorophenyl)-8-(1-methyl-1H-pyrazol-5-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (9l).
Column: alumina. Eluent: CH$_2$Cl$_2$/EtOAc (80/20), yellow solid, 230 mg, yield: 86 %. $^1$H NMR (300 MHz, CDCl$_3$) δ: 8.33 (s, 1H, H-3), 8.10–7.98 (m, 2H, 4-F-Ph-2,6), 7.54 (d, J = 1.9 Hz, 1H, pyrazol-3), 7.25–7.11 (m, 3H, H-7 & 4-F-Ph-3,5), 6.80 (d, J = 1.9 Hz, 1H, pyrazol-4), 4.07 (s, 3H, CH$_3$ pyrazole), 3.64–3.54 (m, 4H, 2 CH$_2$ piperazine), 2.60–2.43 (m, 4H, 2 CH$_2$ piperazine), 2.28 (s, 3H, CH$_3$ piperazine). $^{13}$C NMR (75 MHz, CDCl$_3$) δ: 163.3 (d, $^1J_{CF}$ = 244.5 Hz, 4-F-Ph-4), 156.0 (C-6), 143.6 (C-2), 138.9 (pyrazol-3), 137.6 (pyrazol-5), 135.7 (C-8a), 131.8 (d, $^4J_{CF}$ = 3.0 Hz, 4-F-Ph-1), 128.7 (C-8), 128.1 (d, $^3J_{CF}$ = 8.1 Hz, 4-F-Ph-3,5), 116.2 (d, $^2J_{CF}$ = 21.7 Hz, 4-F-Ph-3,5), 114.0 (C-3), 110.5 (C-7), 109.9 (pyrazol-4), 55.4 (2 CH$_2$ piperazine), 46.9 (2 CH$_2$ piperazine), 46.4 (CH$_3$ piperazine), 39.4 (CH$_3$ pyrazole). $^{19}$F NMR (282 MHz, CDCl$_3$) δ: −116.4. m.p.: 196–200 °C. **LC-MS**: retention time = 0.55 min, m/z [M+H]$^+$ = 392.25.

### 4.1.4.13. 2-(4-fluorophenyl)-8-(1-isopentyl-1H-pyrazol-4-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (9m).
Column: silica. Eluent: CH$_2$Cl$_2$/MeOH (98/2 → 96/4), yellow solid, 117 mg, yield: 84 %. $^1$H NMR (300 MHz, CDCl$_3$) δ: 8.83 (s, 1H, pyrazol-5), 8.15 (d, J = 0.5 Hz, 1H, pyrazol-3), 8.01–7.87 (m, 3H, H-3 & 4-F-Ph-2,6), 7.19–7.07 (m, 2H, 4-F-Ph-3,5), 6.92 (s, 1H, H-7), 4.26 (t, J = 7.5 Hz, 2H, NCH$_2$), 3.65–3.49 (m, 4H, 2 CH$_2$ piperazine), 2.69–2.55 (m, 4H, 2 CH$_2$ piperazine), 2.39 (s, 3H, CH$_3$ piperazine), 1.94–1.80 (m, 2H, NCH$_2$CH$_2$), 1.74–1.57 (m, 1H, CH), 0.99 (d, J = 6.6 Hz, 6H, 2 CH$_3$). $^{13}$C NMR (75 MHz, CDCl$_3$) δ: 162.6 (d, $^1J_{CF}$ = 246.2 Hz, 4-F-Ph-4), 155.6 (C-6), 142.5 (C-2), 137.8 (pyrazol-
3), 135.6 (C-8a), 131.7 (pyrazol-5), 130.6 (d, $ ^{4}J_{C-F} = 3.0 $ Hz, 4-F-Ph-1), 129.6 (C-8), 127.3 (d, $ ^{3}J_{C-F} = 8.0 $ Hz, 4-F-Ph-2,6), 115.7 (d, $ ^{4}J_{C-F} = 21.7 $ Hz, 4-F-Ph-3,5), 115.5 (pyrazol-4), 112.8 (C-3), 102.8 (C-7), 54.8 (2 CH₂ piperazine), 51.1 (NCH₂), 46.5 (2 CH₂ piperazine), 46.2 (CH₃ piperazine), 39.2 (NCH₂CH₂), 25.8 (CH), 22.5 (2 CH₃). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -114.7. **m.p.:** 215–219 °C. **LC-MS:** retention time = 1.44 min, m/z [M+H]⁺ = 448.26.

#### 4.1.4.14. 2-(4-fluorophenyl)-8-(furan-3-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (9n).
Column: alumina. Eluent: CH₂Cl₂, yellow solid, 110 mg, yield: 71 %. $ ^{1}H$ NMR (300 MHz, CDCl₃) $\delta$: 9.14 (dd, *J* = 1.5, 0.9 Hz, 1H, furan-2), 8.26 (s, 1H, H-3), 8.17–8.07 (m, 2H, 4-F-Ph-2,6), 7.76 (t, *J* = 1.8 Hz, 1H, furan-5), 7.39 (s, 1H, H-7), 7.37 (dd, *J* = 1.9, 0.7 Hz, 1H, furan-4), 7.25–7.16 (m, 2H, 4-F-Ph-3,5), 3.64–3.53 (m, 4H, 2 CH₂ piperazine), 2.56–2.46 (m, 4H, 2 CH₂ piperazine), 2.28 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, CDCl₃) $\delta$: 163.2 (d, $ ^{1}J_{C-F} = 244.2 $ Hz, 4-F-Ph-4), 156.6 (C-6), 147.0 (furan-2), 144.7 (furan-5), 142.8 (C-2), 136.0 (C-8a), 132.0 (d, $ ^{4}J_{C-F} = 2.9 $ Hz, 4-F-Ph-1), 129.8 (C-8), 128.1 (d, $ ^{3}J_{C-F} = 8.1 $ Hz, 4-F-Ph-2,6), 120.9 (furan-3), 116.1 (d, $ ^{2}J_{C-F} = 21.6 $ Hz, 4-F-Ph-3,5), 113.6 (C-3), 109.2 (furan-4), 105.9 (C-7), 55.4 (2 CH₂ piperazine), 47.0 (2 CH₂ piperazine), 46.4 (CH₃). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -116.6. **m.p.:** 188–192 °C. **LC-MS:** retention time = 0.89 min, m/z [M+H]⁺ = 378.15.

#### 4.1.4.15. 2-(4-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(2-(4-methylpiperazin-1-yl)pyrimidin-5-yl)imidazo [1,2-b]pyridazine (9o).
Column: silica. Eluent: CH₂Cl₂/MeOH (98/2 → 80/20), yellow solid, 78 mg, yield: 57 %. $ ^{1}H$ NMR (300 MHz, CDCl₃) $\delta$: 9.25 (s, 2H, pyrimidin-4,6), 7.98 (s, 1H, H-3), 7.97–7.88 (m, 2H, 4-F-Ph-2,6), 7.09 (t, *J* = 8.7 Hz, 2H, 4-F-Ph-3,5), 6.82 (s, 1H, H-7), 4.04–3.93 (m, 4H, 2 CH₂ piperazine), 3.60–3.51 (m, 4H, 2 CH₂ piperazine), 2.63–2.55 (m, 4H, 2 CH₂ piperazine), 2.55–2.47 (m, 4H, 2 CH₂ piperazine), 2.38 (s, 3H, CH₃), 2.36 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, CDCl₃) $\delta$: 162.7 (d, $ ^{1}J_{C-F} = 246.3 $ Hz, 4-F-Ph-4), 161.5 (pyrimidin-2), 158.1 (pyrimidin-4,6), 155.5 (C-6), 143.0 (C-2), 135.7 (C-8a), 132.5 (C-8), 130.4 (d, $ ^{4}J_{C-F} = 3.2 $ Hz, 4-F-Ph-1), 127.3 (d, $ ^{3}J_{C-F} = 8.1 $ Hz, 4-F-Ph-2,6), 116.5 (pyrimidin-5), 115.6 (d, $ ^{2}J_{C-F} = 21.6 $ Hz, 4-F-Ph-3,5), 112.8 (C-3), 103.6 (C-7), 55.0 (2 CH₂ piperazine), 54.8 (2 CH₂ piperazine), 46.6 (2 CH₂ piperazine), 46.32 (CH₃), 46.30 (CH₃), 44.0 (2 CH₂ piperazine). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -114.6. **m.p.:** 210–214 °C. **LC-MS:** retention time = 0.38 min, m/z [M+H]⁺ = 488.36.

#### 4.1.4.16. 2-(3-fluorophenyl)-8-(1H-indol-4-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (10a).
Column: silica. Eluent: CH₂Cl₂/ EtOH (90/10 → 10/90), beige solid, 58 mg, yield: 49 %. $ ^{1}H$ NMR (300 MHz, acetone-d6) $\delta$: 10.59 (bs, 1H, NH), 8.40 (s, 1H, H-3), 7.92 (dd, *J* = 7.4, 0.6 Hz, 1H, indol-5), 7.85–7.71 (m, 2H, 3-F-Ph-2 & 3-F-Ph-6), 7.61 (d, *J* = 8.2 Hz, 1H, indol-7), 7.48–7.44 (m, 1H, indol-2), 7.44–7.37 (m, 1H, 3-F-Ph-5), 7.35–7.26 (m, 2H, H-7 & indol-6), 7.02 (td, *J* = 8.2, 2.3 Hz, 1H, 3-F-Ph-4), 6.75–6.68 (m, 1H, indol-3), 3.66–3.57 (m, 4H, 2 CH₂ piperazine), 2.59–2.49 (m, 4H, 2 CH₂ piperazine), 2.29 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, acetone-d6) $\delta$: 164.1 (d, $ ^{1}J_{C-F} = 242.3 $ Hz, 3-F-Ph-3), 156.5 (C-6), 142.8 (d, $ ^{4}J_{C-F} = 3.2 $ Hz, C-2), 138.9 (C-8), 138.3 (d, $ ^{3}J_{C-F} = 8.5 $ Hz, 3-F-Ph-1), 137.8 (C-8a), 137.3 (Indol-7a), 131.2 (d, $ ^{3}J_{C-F} = 8.4 $ Hz, 3-F-Ph-5), 127.5 (indol-4), 127.2 (indol-3a*), 126.7 (indol-2*), 122.8 (indol-5), 122.1 (d, $ ^{4}J_{C-F} = 3.0 $ Hz, 3-F-Ph-6), 121.8 (indol-6), 114.50 (d, $ ^{2}J_{C-F} = 21.4 $ Hz, 3-F-Ph-4), 114.49 (C-3), 113.7 (indol-7), 112.6 (d, $ ^{2}J_{C-F} = 24.9 $ Hz, 3-F-Ph-2), 109.9 (C-7), 102.1 (indol-3), 55.4 (2 CH₂ piperazine), 47.0 (2 CH₂ piperazine), 46.38 (CH₃). $ ^{19}F$ NMR (282 MHz, acetone-d6) $\delta$: -115.1. **m.p.:** 165–169 °C. **LC-MS:** retention time = 0.64 min, m/z [M+H]⁺ = 427.16.

#### 4.1.4.17. 2-(3-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(pyridin-3-yl)imidazo [1,2-b]pyridazine (10b).
Column: alumina. Eluent: CH₂Cl₂/ MeOH (98/2), yellow solid, 191 mg, yield: 80 %. $ ^{1}H$ NMR (300 MHz, CDCl₃) $\delta$: 9.35 (dd, *J* = 2.1, 0.6 Hz, 1H, pyridin-2), 8.69–8.59 (m, 2H, pyridin-4 & pyridin-6), 8.18 (s, 1H, H-3), 7.70–7.56 (m, 3H, 3-F-Ph-2, 3-F-Ph-6 & pyridin-5), 7.42–7.32 (m, 1H, 3-F-Ph-5), 7.21 (s, 1H, H-7), 7.05–6.94 (m, 1H, 3-F-Ph-4), 3.63–3.53 (m, 4H, 2 CH₂ piperazine), 2.64–2.54 (m, 4H, 2 CH₂ piperazine), 2.36 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, CDCl₃) $\delta$: 164.7 (d, $ ^{1}J_{C-F} = 243.4 $ Hz, 3-F-Ph-3), 156.6 (C-6), 150.9 (pyridin-6), 150.3 (pyridin-2), 143.8 (d, $ ^{4}J_{C-F} = 2.8 $ Hz, C-2), 138.6 (C-8a*), 137.6 (d, $ ^{3}J_{C-F} = 8.3 $ Hz, 3-F-Ph-1), 136.8 (pyridin-4), 134.7 (C-8*), 132.1 (pyridin-3), 131.4 (d, $ ^{3}J_{C-F} = 8.3 $ Hz, 3-F-Ph-5), 125.1 (pyridin-5), 122.4 (d, $ ^{4}J_{C-F} = 2.7 $ Hz, 3-F-Ph-6), 115.3 (C-3), 115.2 (d, $ ^{2}J_{C-F} = 21.5 $ Hz, 3-F-Ph-4), 113.1 (d, $ ^{2}J_{C-F} = 23.1 $ Hz, 3-F-Ph-2), 109.5 (C-7), 55.5 (2 CH₂ piperazine), 46.8 (2 CH₂ piperazine), 46.2 (CH₃). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -115.3. **m.p.:** 157–161 °C. **LC-MS:** retention time = 0.54 min, m/z [M+H]⁺ = 389.15.

#### 4.1.4.18. 2-(3-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (10c).
Column: silica. Eluent: EtOAc/EtOH (95/5 → 70/30), yellow solid, 69 mg, yield: 83 %. $ ^{1}H$ NMR (300 MHz, CDCl₃) $\delta$: 9.01 (dd, *J* = 2.9, 1.1 Hz, 1H, thien-2), 8.02 (s, 1H, H-3), 7.79 (dd, *J* = 5.1, 1.0 Hz, 1H, thien-4), 7.76–7.68 (m, 2H, 3-F-Ph-2,6), 7.46 (dd, *J* = 5.1, 3.1 Hz, 1H, thien-5), 7.43–7.32 (m, 1H, 3-F-Ph-5), 7.05–6.94 (m, 2H, H-7 & 3-F-Ph-4), 3.62–3.52 (m, 4H, 2 CH₂ piperazine), 2.64–2.55 (m, 4H, 2 CH₂ piperazine), 2.38 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, CDCl₃) $\delta$: 163.5 (d, $ ^{1}J_{C-F} = 246.2 $ Hz, 3-F-Ph-3), 155.6 (C-6), 142.4 (C-2), 136.7 (d, $ ^{3}J_{C-F} = 7.8 $ Hz, 3-F-Ph-1), 135.9 (C-8a), 134.7 (C-8), 131.5 (thien-3), 130.2 (d, $ ^{3}J_{C-F} = 8.2 $ Hz, 3-F-Ph-5), 129.6 (thien-2), 126.1 (thien-4,5), 121.2 (3-F-Ph-6), 114.3 (d, $ ^{2}J_{C-F} = 21.3 $ Hz, 3-F-Ph-4), 113.4 (C-3), 112.5 (d, $ ^{2}J_{C-F} = 22.8 $ Hz, 3-F-Ph-2), 105.3 (C-7), 54.8 (2 CH₂ piperazine), 46.5 (2 CH₂ piperazine), 46.3 (CH₃). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -113.4. **m.p.:** 140–144 °C. **LC-MS:** retention time = 1.11 min, m/z [M+H]⁺ = 394.15.

#### 4.1.4.19. 2-(3-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(2-(4-methylpiperazin-1-yl)pyrimidin-5-yl)imidazo [1,2-b]pyridazine (10d).
Column: silica. Eluent: CH₂Cl₂/MeOH (98/2 → 80/20), beige solid, 73 mg, yield: 53 %. $ ^{1}H$ NMR (300 MHz, CDCl₃) $\delta$: 9.25 (s, 2H, pyrimidin-4,6), 8.02 (s, 1H, H-3), 7.75–7.63 (m, 2H, 3-F-Ph-2,6), 7.41–7.29 (m, 1H, 3-F-Ph-5), 6.97 (td, *J* = 7.9, 1.7 Hz, 1H, 3-F-Ph-4), 6.83 (s, 1H, H-7), 4.04–3.93 (m, 4H, 2 CH₂ piperazine), 3.63–3.47 (m, 4H, 2 CH₂ piperazine), 2.63–2.55 (m, 4H, 2 CH₂ piperazine), 2.55–2.46 (m, 4H, 2 CH₂ piperazine), 2.37 (s, 3H, CH₃), 2.36 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, CDCl₃) $\delta$: 163.4 (d, $ ^{1}J_{C-F} = 244.7 $ Hz, 3-F-Ph-3), 161.5 (pyrimidin-2), 158.1 (pyrimidin-4,6), 155.5 (C-6), 142.6 (d, $ ^{4}J_{C-F} = 2.7 $ Hz, C-2), 136.5 (d, $ ^{3}J_{C-F} = 8.2 $ Hz, 3-F-Ph-1), 135.7 (C-8a), 132.6 (C-8), 130.2 (d, $ ^{3}J_{C-F} = 8.4 $ Hz, 3-F-Ph-5), 121.2 (d, $ ^{4}J_{C-F} = 2.5 $ Hz, 3-F-Ph-6), 116.4 (pyrimidin-5), 114.4 (d, $ ^{2}J_{C-F} = 21.4 $ Hz, 3-F-Ph-4), 113.5 (C-3), 112.5 (d, $ ^{2}J_{C-F} = 22.7 $ Hz, 3-F-Ph-2), 103.8 (C-7), 55.0 (2 CH₂ piperazine), 54.8 (2 CH₂ piperazine), 46.5 (2 CH₂ piperazine), 46.34 (CH₃), 46.30 (CH₃), 44.0 (2 CH₂ piperazine). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -113.4. **m.p.:** 200–204 °C. **LC-MS:** retention time = 0.39 min, m/z [M+H]⁺ = 488.36.

#### 4.1.4.20. 2-(2-fluorophenyl)-8-(1H-indol-4-yl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (11a).
Column: silica. Eluent: CH₂Cl₂/ MeOH (100/0 → 98/2), white solid, 65 mg, yield: 69 %. $ ^{1}H$ NMR (300 MHz, DMSO-d₆) $\delta$: 11.40 (bs, 1H, NH), 8.23 (d, *J* = 4.3 Hz, 1H, H-3), 8.17–8.04 (m, 1H, 2-F-Ph-6), 7.76 (d, *J* = 7.2 Hz, 1H, indol-5), 7.58 (d, *J* = 8.1 Hz, 1H, indol-7), 7.48 (t, *J* = 2.6 Hz, 1H, indol-2), 7.39–7.20 (m, 5H, H-7, indol-6, 2-F-Ph-3,4,5), 6.57 (s, 1H, indol-3), 3.56 (s, 4H, 2 CH₂ piperazine), 2.26 (s, 3H,CH₃). (Note 1: two of the four CH₂ are overlapped with DMSO). $ ^{13}C$ NMR (75 MHz, DMSO-d₆) $\delta$: 159.2 (d, $ ^{1}J_{C-F} = 247.6 $ Hz, 2-F-Ph-2), 155.1 (C-6), 137.3 (C-8a), 136.3 (C-2*), 135.9 (indol-7a*), 135.3 (C-8*), 128.8 (d, $ ^{3}J_{C-F} = 8.2 $ Hz, 2-F-Ph-4), 127.8 (d, $ ^{3}J_{C-F} = 2.8 $ Hz, 2-F-Ph-6), 126.3 (indol-2), 126.0 (indol-4), 125.6 (indol-3a*), 124.5 (d, $ ^{4}J_{C-F} = 2.1 $ Hz, 2-F-Ph-5), 121.5 (d, $ ^{2}J_{C-F} = 12.5 $ Hz, 2-F-Ph-1), 121.1 (indol-5), 120.6 (indol-6), 115.9 (d, $ ^{4}J_{C-F} = 13.8 $ Hz, C-3), 115.7 (d, CDCl₃) $\delta$: 9.35 (dd, *J* = 2.1, 0.6 Hz, 1H, pyridin-2), 8.69–8.59 (m, 2H, pyridin-4 & pyridin-6), 8.18 (s, 1H, H-3), 7.70–7.56 (m, 3H, 3-F-Ph-2, 3-F-Ph-6 & pyridin-5), 7.42–7.32 (m, 1H, 3-F-Ph-5), 7.21 (s, 1H, H-7), 7.05–6.94 (m, 1H, 3-F-Ph-4), 3.63–3.53 (m, 4H, 2 CH₂ piperazine), 2.64–2.54 (m, 4H, 2 CH₂ piperazine), 2.36 (s, 3H, CH₃). $ ^{13}C$ NMR (75 MHz, CDCl₃) $\delta$: 164.7 (d, $ ^{1}J_{C-F} = 243.4 $ Hz, 3-F-Ph-3), 156.6 (C-6), 150.9 (pyridin-6), 150.3 (pyridin-2), 143.8 (d, $ ^{4}J_{C-F} = 2.8 $ Hz, C-2), 138.6 (C-8a*), 137.6 (d, $ ^{3}J_{C-F} = 8.3 $ Hz, 3-F-Ph-1), 136.8 (pyridin-4), 134.7 (C-8*), 132.1 (pyridin-3), 131.4 (d, $ ^{3}J_{C-F} = 8.3 $ Hz, 3-F-Ph-5), 125.1 (pyridin-5), 122.4 (d, $ ^{4}J_{C-F} = 2.7 $ Hz, 3-F-Ph-6), 115.3 (C-3), 115.2 (d, $ ^{2}J_{C-F} = 21.5 $ Hz, 3-F-Ph-4), 113.1 (d, $ ^{2}J_{C-F} = 23.1 $ Hz, 3-F-Ph-2), 109.5 (C-7), 55.5 (2 CH₂ piperazine), 46.8 (2 CH₂ piperazine), 46.2 (CH₃). $ ^{19}F$ NMR (282 MHz, CDCl₃) $\delta$: -115.3. **m.p.:** 157–161 °C. **LC-MS:** retention time = 0.54 min, m/z [M+H]⁺ = 389.15.
$^2J_{C-F} = 21.5$ Hz, 2-F-Ph-3), 112.8 (indol-7), 109.1 (C-7), 100.6 (indol-3), 54.0 (2 CH₂ piperazine), 45.5 (2 CH₂ piperazine), 45.4 (CH₃). (Note 2: For the whole series with a 2-fluoro-phenyl group at position 2 we can note that the proton and the carbon at position 3 present abnormally high H–F and C–F coupling constants. This is due to the proximity of fluorine to this position in space, which generates a through space spin-spin coupling). **¹⁹F NMR** (282 MHz, DMSO-d₆) δ: –114.5. **m.p.:** 222–226 °C. **LC-MS:** retention time = 0.65 min, m/z [M+H]⁺ = 427.16.

#### 4.1.4.21. 2-(2-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(pyridin-4-yl) imidazo [1,2-b]pyridazine (11b).
Column: alumina. Eluent: CH₂Cl₂/MeOH (99/1), yellow solid, 87 mg, yield: 96 %. **¹H NMR** (300 MHz, DMSO-d₆) δ: 8.86–8.75 (m, 2H, pyridin-2,6), 8.45–8.36 (m, 2H, pyridin-3,5), 8.30–8.19 (m, 3H, H-3 & 2-F-Ph-6), 7.61 (s, 1H, H-7), 7.44–7.26 (m, 3H, 2-F-Ph-3,4,5), 3.63–3.52 (m, 4H, 2 CH₂ piperazine), 2.24 (s, 3H, CH₃) (Note: two of the four CH₂ are overlapped with DMSO). **¹³C NMR** (75 MHz, DMSO-d₆) δ: 159.4 (d, $^1J_{C-F}$ = 247.8 Hz, 2-F-Ph-2), 155.2 (C-6), 150.0 (pyridin-2,6), 140.9 (pyridin-4), 136.2 (d, $^3J_{C-F}$ = 1.2 Hz, C-2), 134.2 (C-8*), 133.1 (C-8a*), 129.2 (d, $^3J_{C-F}$ = 8.5 Hz, 2-F-Ph-4), 128.0 (d, $^3J_{C-F}$ = 3.6 Hz, 2-F-Ph-6), 124.8 (d, $^4J_{C-F}$ = 3.1 Hz, 2-F-Ph-5), 123.3 (pyridin-3,5), 121.2 (d, $^2J_{C-F}$ = 12.7 Hz, 2-F-Ph-1), 116.3 (d, $^{4TSJ_{C-F}}$ = 13.9 Hz, C-3), 115.9 (d, $^2J_{C-F}$ = 21.6 Hz, 2-F-Ph-3), 109.1 (C-7), 54.1 (2 CH₂ piperazine), 45.7 (CH₃), 45.6 (2 CH₂ piperazine). **¹⁹F NMR** (282 MHz, CDCl₃) δ: –114.2. **m.p.:** 202–206 °C. **LC-MS:** retention time = 0.54 min, m/z [M+H]⁺ = 389.25.

#### 4.1.4.22. 2-(2-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(pyridin-3-yl) imidazo [1,2-b]pyridazine (11c).
Column: alumina. Eluent: CH₂Cl₂/MeOH (99/1), yellow solid, 68 mg, yield: 65 %. **¹H NMR** (300 MHz, CD₃OD) δ: 9.41 (dd, *J* = 2.1, 0.9 Hz, 1H, pyridin-2), 8.72–8.63 (m, 2H, pyridin-4 & pyridin-6), 8.20 (td, *J* = 7.7, 1.9 Hz, 1H, 2-F-Ph-6), 8.14 (d, $^TSJ$ = 4.4 Hz, 1H, H-3), 7.61 (ddd, *J* = 8.1, 4.8, 0.9 Hz, 1H, pyridin-5), 7.36–7.12 (m, 4H, H-7 & 2-F-Ph-3,4,5), 3.69–3.56 (m, 4H, 2 CH₂ piperazine), 2.69–2.57 (m, 4H, 2 CH₂ piperazine), 2.38 (s, 3H, CH₃). **¹³C NMR** (75 MHz, CD₃OD) δ: 161.4 (d, $^1J_{C-F}$ = 248.0 Hz, 2-F-Ph-2), 156.6 (C-6), 150.9 (pyridin-6), 150.3 (pyridin-2), 138.6 (C-8a*), 138.5 (d, $^3J_{C-F}$ = 2.0 Hz, C-2), 136.1 (pyridin-4), 134.6 (C-8*), 132.1 (pyridin-3), 130.1 (d, $^3J_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 129.3 (d, $^3J_{C-F}$ = 3.7 Hz, 2-F-Ph-6), 125.5 (d, $^4J_{C-F}$ = 3.1 Hz, 2-F-Ph-5), 125.1 (pyridin-5), 122.7 (d, $^2J_{C-F}$ = 12.5 Hz, 2-F-Ph-1), 118.0 (d, $^{4TSJ_{C-F}}$ = 14.6 Hz, C-3), 116.7 (d, $^2J_{C-F}$ = 22.1 Hz, 2-F-Ph-3), 109.6 (C-7), 55.5 (2 CH₂ piperazine), 46.8 (2 CH₂ piperazine), 46.1 (CH₃). **¹⁹F NMR** (282 MHz, CD₃OD) δ: –116.1. **m.p.:** 185–189 °C. **LC-MS:** retention time = 0.54 min, m/z [M+H]⁺ = 389.25.

#### 4.1.4.23. 2-(2-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(6-amino-pyridin-3-yl)imidazo [1,2-b]pyridazine (11d).
Column: alumina. Eluent: CH₂Cl₂/MeOH (99/1), yellow solid, 73 mg, yield: 42 %. **¹H NMR** (300 MHz, CDCl₃) δ: 8.98 (d, *J* = 2.1 Hz, 1H, pyridin-2), 8.45 (dd, *J* = 8.6, 2.3 Hz, 1H, pyridin-4), 8.42–8.34 (m, 1H, 2-F-Ph-6), 8.21 (d, $^TSJ$ = 4.3 Hz, 1H, H-3), 7.25–7.17 (m, 2H, 2-F-Ph-4,5), 7.17–7.06 (m, 1H, 2-F-Ph-3), 6.87 (s, 1H, H-7), 6.64 (d, *J* = 8.6 Hz, 1H, pyridin-5), 4.77 (bs, 2H, NH₂), 3.62–3.52 (m, 4H, 2 CH₂ piperazine), 2.62–2.53 (m, 4H, 2 CH₂ piperazine), 2.37 (s, 3H, CH₃). **¹³C NMR** (75 MHz, CDCl₃) δ: 160.3 (d, $^1J_{C-F}$ = 249.0 Hz, 2-F-Ph-2), 159.3 (pyridin-6), 155.5 (C-6), 149.0 (pyridin-2), 138.5 (pyridin-4), 137.2 (d, $^3J_{C-F}$ = 1.8 Hz, C-2), 135.4 (C-8*), 134.6 (C-8a*), 128.6 (d, $^3J_{C-F}$ = 8.5 Hz, 2-F-Ph-4), 128.4 (d, $^3J_{C-F}$ = 3.9 Hz, 2-F-Ph-6), 122.1 (d, $^2J_{C-F}$ = 12.3 Hz, 2-F-Ph-1), 120.7 (pyridin-3), 117.0 (d, $^{4TSJ_{C-F}}$ = 14.6 Hz, C-3), 115.7 (d, $^2J_{C-F}$ = 22.0 Hz, 2-F-Ph-3), 108.1 (pyridin-5), 104.9 (C-7), 54.8 (2 CH₂ piperazine), 46.6 (2 CH₂ piperazine), 46.3 (CH₃). **¹⁹F NMR** (282 MHz, CDCl₃) δ: –114.4. **m.p.:** 233–237 °C. **LC-MS:** retention time = 0.39 min, m/z [M+H]⁺ = 404.15.

#### 4.1.4.24. 2-(2-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(thien-3-yl) imidazo [1,2-b]pyridazine (11e).
Column: silica. Eluent: EtOAc/EtOH (95/5 → 70/30), yellow solid, 57 mg, yield: 68 %. **¹H NMR** (300 MHz, CDCl₃) δ: 9.02 (dd, *J* = 3.0, 1.1 Hz, 1H, thien-2), 8.48–8.36 (m, 1H, 2-F-Ph-6), 8.19 (d, $^TSJ$ = 4.4 Hz, 1H, H-3), 7.77 (dd, *J* = 5.1, 1.1 Hz, 1H, thien-4), 7.44 (dd, *J* = 5.1, 3.1 Hz, 1H, thien-5), 7.30–7.21 (m, 2H, 2-F-Ph-4,5), 7.18–7.08 (m, 1H, 2-F-Ph-3), 6.97 (s, 1H, H-7), 3.62–3.47 (m, 4H, 2 CH₂ piperazine), 2.69–2.49 (m, 4H, 2 CH₂ piperazine), 2.36 (s, 3H, CH₃). **¹³C NMR** (75 MHz, CDCl₃) δ: 160.3 (d, $^1J_{C-F}$ = 249.1 Hz, 2-F-Ph-2), 155.5 (C-6), 137.1 (d, $^3J_{C-F}$ = 1.6 Hz, C-2), 135.1 (C-8a), 134.7 (C-8), 131.2 (thien-3*), 129.5 (d, $^3J_{C-F}$ = 8.4 Hz, 2-F-Ph-4), 128.3 (d, $^3J_{C-F}$ = 3.9 Hz, 2-F-Ph-6), 126.2 (thien-4*), 126.0 (thien-5*), 124.4 (d, $^4J_{C-F}$ = 3.1 Hz, 2-F-Ph-5), 122.1 (d, $^2J_{C-F}$ = 12.4 Hz, 2-F-Ph-1), 116.9 (d, $^{4TSJ_{C-F}}$ = 14.7 Hz, C-3), 115.7 (d, $^2J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 105.3 (C-7), 54.7 (2 CH₂ piperazine), 46.5 (2 CH₂ piperazine), 46.3 (CH₃). **¹⁹F NMR** (282 MHz, CDCl₃) δ: –114.2. **m.p.:** 167–171 °C. **LC-MS:** retention time = 1.13 min, m/z [M+H]⁺ = 394.15.

#### 4.1.4.25. 2-(2-fluorophenyl)-6-(4-methylpiperazin-1-yl)-8-(2-(4-methyl-piperazin-1-yl)pyrimidin-5-yl)imidazo [1,2-b]pyridazine (11f).
Column: silica. Eluent: CH₂Cl₂/MeOH (90/10), beige solid, 72 mg, yield: 52 %. **¹H NMR** (300 MHz, CDCl₃) δ: 9.28 (s, 2H, pyrimidin-4,6), 8.40 (td, *J* = 7.6, 2.4 Hz, 1H, 2-F-Ph-6), 8.21 (d, $^TSJ$ = 4.4 Hz, 1H, H-3), 7.29–7.21 (m, 2H, 2-F-Ph-4,5), 7.17–7.06 (m, 1H, 2-F-Ph-3), 6.84 (s, 1H, H-7), 4.06–3.96 (m, 4H, 2 CH₂ piperazine), 3.63–3.53 (m, 4H, 2 CH₂ piperazine), 2.66–2.49 (m, 8H, 4 CH₂ piperazine), 2.39 (s, 6H, 2 CH₃). **¹³C NMR** (75 MHz, CDCl₃) δ: 161.5 (pyrimidin-2), 160.3 (d, $^1J_{C-F}$ = 248.8 Hz, 2-F-Ph-2), 158.1 (pyrimidin-4,6), 155.4 (C-6), 137.4 (d, $^3J_{C-F}$ = 1.8 Hz, C-2), 135.0 (C-8a), 132.4 (C-8), 128.7 (d, $^3J_{C-F}$ = 8.4 Hz, 2-F-Ph-4), 128.5 (d, $^3J_{C-F}$ = 3.9 Hz, 2-F-Ph-6), 124.4 (d, $^4J_{C-F}$ = 3.2 Hz, 2-F-Ph-5), 121.9 (d, $^2J_{C-F}$ = 12.3 Hz, 2-F-Ph-1), 117.1 (d, $^{4TSJ_{C-F}}$ = 14.9 Hz, C-3), 116.6 (pyrimidin-5), 115.7 (d, $^2J_{C-F}$ = 21.9 Hz, 2-F-Ph-3), 103.8 (C-7), 55.0 (2 CH₂ piperazine), 54.7 (2 CH₂ piperazine), 46.5 (2 CH₂ piperazine), 46.2 (2 CH₃), 43.9 (2 CH₂ piperazine). **¹⁹F NMR** (282 MHz, CDCl₃) δ: –114.3. **m.p.:** 178–182 °C. **LC-MS:** retention time = 0.39 min, m/z [M+H]⁺ = 488.36.

#### 4.1.4.26. 8-(1H-indol-4-yl)-2-(2-methoxyphenyl)-6-(4-methylpiperazin-1-yl)imidazo [1,2-b]pyridazine (12a).
Column: silica. Eluent: CH₂Cl₂/MeOH (99/1), white solid, 54 mg, yield: 66 %. **¹H NMR** (300 MHz, DMSO-d₆) δ: 11.39 (bs, 1H, NH), 8.34 (s, 1H, H-3), 8.14 (dd, *J* = 7.7, 1.6 Hz, 1H, 2-MeO-Ph-6), 7.79 (d, *J* = 7.1 Hz, 1H, indol-5), 7.58 (d, *J* = 8.1 Hz, 1H, indol-7), 7.48 (t, *J* = 2.7 Hz, 1H, indol-2), 7.33–7.23 (m, 3H, H-7, 2-MeO-Ph-4, indol-6), 7.10 (d, *J* = 8.0 Hz, 1H, 2-MeO-Ph-3*), 6.99 (t, *J* = 7.4 Hz, 1H, 2-MeO-Ph-5*), 6.57 (s, 1H, indol-3), 3.97 (s, 3H, OCH₃), 3.54 (s, 4H, 2 CH₂ piperazine), 2.25 (s, 3H, NCH₃). (Note: two of the four CH₂ are overlapped with DMSO). **¹³C NMR** (75 MHz, DMSO-d₆) δ: 156.1 (C-6), 154.9 (2-MeO-Ph-2), 138.2 (C-2), 137.1 (C-8), 136.3 (C-8a), 134.5 (indol-7a), 128.2 (2-MeO-Ph-4), 127.2 (2-MeO-Ph-6), 126.2 (indol-4*), 126.1 (indol-3a*), 125.9 (indol-2), 122.2 (2-MeO-Ph-1), 121.2 (2-MeO-Ph-5), 120.6 (indol-5), 120.3 (indol-6), 116.5 (C-3), 112.8 (indol-7), 111.3 (2-MeO-Ph-3), 108.5 (C-7), 100.7 (indol-3), 55.3 (2 CH₂ piperazine), 54.1 (2 CH₂ piperazine & OCH₃), 45.6 (NCH₃). **m.p.:** 214–218 °C. **LC-MS:** retention time = 0.44 min, m/z [M+H]⁺ = 439.26.

#### 4.1.4.27. 2-(2-methoxyphenyl)-6-(4-methylpiperazin-1-yl)-8-(thien-3-yl) imidazo [1,2-b]pyridazine (12b).
Column: silica. Eluent: CH₂Cl₂/MeOH (99/1 → 98/2), white solid, 51 mg, yield: 73 %. **¹H NMR** (300 MHz, DMSO-d₆) δ: 9.24 (d, *J* = 2.2 Hz, 1H, thien-2), 8.40 (d, *J* = 7.6 Hz, 1H, 2-MeO-Ph-6), 8.29 (s, 1H, H-3), 8.26 (d, *J* = 5.0 Hz, 1H, thien-4), 7.76 (dd, *J* = 4.9, 3.0 Hz, 1H, thien-5), 7.53 (s, 1H, H-7), 7.32 (t, *J* = 7.7 Hz, 1H, 2-MeO-Ph-4), 7.18–7.03 (m, 2H, 2-MeO-Ph-3,5), 3.98 (s, 3H, OCH₃), 3.57 (s, 4H, 2 CH₂ piperazine), 2.27 (s, 3H, NCH₃). (Note: two of the four CH₂ are overlapped with DMSO). **¹³C NMR** (75 MHz, DMSO-d₆) δ: 156.2 (2-MeO-Ph-2), 155.3 (C-6), 138.0 (C-2), 134.6 (C-8a*), 133.7 (C-8*), 129.8 (thien-3), 129.5 (thien-2), 128.4 (2-MeO-Ph-4), 127.4 (2-MeO-Ph-6), 127.2 (thien-4), 126.5 (thien-5), 122.0 (2-MeO-Ph-1), 120.5 (2-MeO-Ph-5), 116.5 (C-3), 111.4 (2-MeO-Ph-3), 105.6 (C-7), 55.4 (OCH₃), 54.1 (2
CH₂ piperazine), 45.65 (2 CH₂ piperazine), 45.58 (NCH₃). **m.p.:** 157–161 °C. **LC-MS:** retention time = 1.02 min, *m/z* [M+H]⁺ = 406.15.

### 4.1.4.28. 8-(1H-indol-4-yl)-6-(4-methylpiperazin-1-yl)-2-phenylimidazo [1,2-b]pyridazine (13a).
Column: alumina. Eluent: CH₂Cl₂/EtOAc (80/20), white solid, 274 mg, yield: 86 %. ¹H NMR (300 MHz, acetone-d6) δ: 10.54 (bs, 1H, NH), 8.33 (s, 1H, H-3), 8.04-7.97 (m, 2H, Ph-2,6), 7.93 (dd, *J* = 7.4, 0.9 Hz, 1H, indol-5), 7.60 (d, *J* = 8.1 Hz, 1H, indol-7), 7.48-7.44 (m, 1H, indol-2), 7.42-7.35 (m, 2H, Ph-3,5), 7.34-7.22 (m, 3H, H-7, indol-6 & Ph-4), 6.74-6.68 (m, 1H, indol-3), 3.67-3.54 (m, 4H, 2 CH₂ piperazine), 2.59-2.49 (m, 4H, 2 CH₂ piperazine), 2.29 (s, 3H, CH₃). ¹³C NMR (75 MHz, acetone-d6) δ: 156.4 (C-6), 144.2 (C-2), 138.8 (C-8), 137.8 (C-8a*), 137.2 (indol-7a*), 135.7 (Ph-1), 129.3 (Ph-3,5), 128.1 (Ph-4), 127.5 (indol-4*), 127.4 (indol-3a*), 126.6 (indol-2), 126.2 (Ph-2,6), 122.8 (indol-5), 121.8 (indol-6), 113.8 (C-3), 113.6 (indol-7), 109.5 (C-7), 102.1 (indol-3), 55.5 (2 CH₂ piperazine), 47.1 (2 CH₂ piperazine), 46.4 (CH₃). **m.p.:** 205–209 °C. **LC-MS:** retention time = 0.43 min, *m/z* [M+H]⁺ = 409.25.

### 4.1.4.29. 6-(4-Methylpiperazin-1-yl)-2-phenyl-8-(thien-3-yl)imidazo [1,2-b]pyridazine (13b).
Column: silica. Eluent: EtOAc/EtOH (95/5 → 70/30), yellow solid, 97 mg, yield: 81 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.05 (dd, *J* = 3.0, 1.2 Hz, 1H, thien-2), 8.04 (s, 1H, H-3), 8.03-7.96 (m, 2H, Ph-2,6), 7.80 (dd, *J* = 5.1, 1.2 Hz, 1H, thien-4), 7.50-7.38 (m, 3H, Ph-3,5 & thien-5), 7.34-7.27 (m, 1H, Ph-4), 6.99 (s, 1H, H-7), 3.62-3.53 (m, 4H, 2 CH₂ piperazine), 2.65-2.53 (m, 4H, 2 CH₂ piperazine), 2.38 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 155.5 (C-6), 143.7 (C-2), 135.9 (C-8a), 134.8 (C-8*), 134.3 (Ph-1*), 131.4 thien-3), 129.6 (thien-2), 128.8 (Ph-3,5), 127.7 (Ph-4), 126.2 (thien-4), 126.0 (thien-5), 125.7 (Ph-2,6), 113.1 (C-3), 105.1 (C-7), 54.8 (2 CH₂ piperazine), 46.6 (2 CH₂ piperazine), 46.3 (CH₃). **m.p.:** 139–143 °C. **LC-MS:** retention time = 0.96 min, *m/z* [M+H]⁺ = 376.15.

### 4.1.4.30. 6-(4-Methylpiperazin-1-yl)-8-(2-(4-methylpiperazin-1-yl)pyrimidin-5-yl)-2-phenylimidazo [1,2-b]pyridazine (13c).
Column: silica. Eluent: cyclohexane/EtOH (100/0 → 60/40), yellow solid, 100 mg, yield: 83 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.27 (s, 2H, pyrimidin-4,6), 8.04 (s, 1H, H-3), 8.01-7.92 (m, 2H, Ph-2,6), 7.47-7.35 (m, 2H, Ph-3,5), 7.34-7.27 (m, 1H, Ph-4), 6.82 (s, 1H, H-7), 3.98 (bs, 4H, 2 CH₂ piperazine), 3.55 (bs, 4H, 2 CH₂ piperazine), 2.58 (bs, 4H, 2 CH₂ piperazine), 2.51 (bs, 4H, 2 CH₂ piperazine), 2.37 (s, 3H, CH₃), 2.36 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 161.5 (pyrimidin-2), 158.1 (pyrimidin-4,6), 155.4 (C-6), 143.9 (C-2), 135.6 (C-8a), 134.2 (Ph-1), 132.5 (C-8), 128.7 (Ph-3,5), 127.7 (Ph-4), 125.7 (Ph-2,6), 116.6 (pyrimidin-5), 113.2 (C-3), 103.5 (C-7), 55.0 (2 CH₂ piperazine), 54.8 (2 CH₂ piperazine), 46.6 (2 CH₂ piperazine), 46.32 (CH₃), 46.30 (CH₃), 44.0 (2 CH₂ piperazine). **m.p.:** 151–155 °C. **LC-MS:** retention time = 0.38 min, *m/z* [M+H]⁺ = 470.36.

### 4.1.4.31. 2-(4-fluorophenyl)-8-(1H-indol-4-yl)-6-(4-(2-hydroxyethyl) piperazin-1-yl)imidazo [1,2-b]pyridazine (14a).
Column: alumina. Eluent: CH₂Cl₂/MeOH (100/0 → 99/1), white solid, 94 mg, yield: 67 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.37 (bs, 1H, NH), 8.55 (s, 1H, H-3), 8.01-7.89 (m, 2H, 4-F-Ph-2,6), 7.75 (d, *J* = 7.3 Hz, 1H, indol-5), 7.57 (d, *J* = 8.1 Hz, 1H, indol-7), 7.47 (t, *J* = 2.8 Hz, 1H, indol-2), 7.33-7.15 (m, 4H, H-7, indol-6 & 4-F-Ph-3,5), 6.56 (bs, 1H, indol-3), 4.45 (t, *J* = 5.4 Hz, 1H, OH), 3.61-3.48 (m, 6H, 2 CH₂ piperazine & NCH₂CH₂OH), 2.64-2.54 (m, 4H, 2 CH₂ piperazine), 2.46 (t, *J* = 6.2 Hz, 2H, NCH₂CH₂OH). ¹³C NMR (75 MHz, DMSO-d₆) δ: 161.6 (d, $^{1}_{J_{C-F}}$ = 243.7 Hz, 4-F-Ph-4), 155.1 (C-6), 141.6 (C-2), 137.3 (C-8*), 136.3 (C-8a*), 135.7 (indol-7a*), 130.7 (d, $^{4}_{J_{C-F}}$ = 3.0 Hz, 4-F-Ph-1), 127.0 (d, $^{3}_{J_{C-F}}$ = 8.1 Hz, 4-F-Ph-2,6), 126.3 (indol-2), 126.1 (Indol-4), 125.8 (indol-3a), 121.1 (indol-5), 120.6 (indol-6), 115.4 (d, $^{2}_{J_{C-F}}$ = 21.6 Hz, 4-F-Ph-3,5), 113.2 (C-3), 112.8 (indol-7), 108.6 (C-7), 100.6 (indol-3), 60.2 (NCH₂CH₂OH), 58.5 (NCH₂CH₂OH), 52.7 (2 CH₂ piperazine), 45.8 (2 CH₂ piperazine). ¹⁹F NMR (282 MHz, CDCl₃) δ: -113.4. **m.p.:** 185–189 °C. **LC-MS:** retention time = 1.02 min, *m/z* [M+H]⁺ = 424.25.

### 4.1.4.32. 2-(4-fluorophenyl)-6-(4-(2-hydroxyethyl)piperazin-1-yl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (14b).
Column: silica. Eluent: CH₂Cl₂/MeOH (100/0 → 98/2), white solid, 59 mg, yield: 46 %. ¹H NMR (300 MHz, CD₃OD) δ: 8.95 (dd, *J* = 3.0, 1.2 Hz, 1H, thien-2), 8.16 (s, 1H, H-3), 8.04-7.94 (m, 3H, thien-4 & 4-F-Ph-2,6), 7.58 (dd, *J* = 5.1, 3.0 Hz, 1H, thien-5), 7.30 (s, 1H, H-7), 7.20-7.10 (m, 2H, 4-F-Ph-3,5), 3.77 (t, *J* = 5.8 Hz, 2H, NCH₂CH₂OH), 3.69-3.60 (m, 4H, 2 CH₂ piperazine), 2.85-2.75 (d, 4H, 2 CH₂ piperazine), 2.70 (t, *J* = 5.8 Hz, 2H, NCH₂CH₂OH). ¹³C NMR (75 MHz, CD₃OD) δ: 163.9 (d, $^{1}_{J_{C-F}}$ = 245.1 Hz, 4-F-Ph-4), 157.0 (C-6), 143.9 (C-2), 137.1 (C-8a*), 136.0 (C-8*), 132.6 (thien-3), 131.8 (d, $^{4}_{J_{C-F}}$ = 3.3 Hz, 4-F-Ph-1), 130.1 (thien-2), 128.5 (d, $^{3}_{J_{C-F}}$ = 8.1 Hz, 4-F-Ph-2,6), 127.9 (thien-4), 127.0 (thien-5), 116.4 (d, $^{3}_{J_{C-F}}$ = 21.8 Hz, 4-F-Ph-3,6), 114.2 (C-3), 107.0 (C-7), 61.2 (NCH₂CH₂OH), 59.5 (NCH₂CH₂OH), 54.1 (2 CH₂ piperazine), 46.9 (2 CH₂ piperazine). ¹⁹F NMR (282 MHz, CD₃OD) δ: -116.9. **m.p.:** 164–168 °C. **LC-MS:** retention time = 0.98 min, *m/z* [M+H]⁺ = 424.15.

### 4.1.4.33. 2-(4-fluorophenyl)-6-(4-(2-hydroxyethyl)piperazin-1-yl)-8-(1-methyl-1H-pyrazol-4-yl)imidazo [1,2-b]pyridazine (14c).
Column: silica. Eluent: CH₂Cl₂/MeOH (100/0 → 99/1), white solid, 58 mg, yield: 45 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.85 (s, 1H, pyrazol-5), 8.12 (s, 1H, pyrazol-3), 8.01-7.89 (m, 3H, 4-F-Ph-2,6 & H-3), 7.13 (t, *J* = 8.7 Hz, 2H, 4-F-Ph-3,5), 6.92 (s, 1H, H-7), 4.04 (s, 3H, CH₃), 3.69 (t, *J* = 5.3 Hz, 2H, NCH₂CH₂OH), 3.63-3.50 (m, 4H, 2 CH₂ piperazine), 2.81-2.57 (m, 6H, 2 CH₂ piperazine & NCH₂CH₂OH). ¹³C NMR (75 MHz, CDCl₃) δ: 162.7 (d, $^{1}_{J_{C-F}}$ = 246.4 Hz, 4-F-Ph-4), 155.6 (C-6), 142.6 (C-2), 138.0 (pyrazol-3), 135.6 (C-8a), 132.8 (pyrazol-5), 130.6 (d, $^{4}_{J_{C-F}}$ = 3.1 Hz, 4-F-Ph-1), 129.5 (C-8), 127.3 (d, $^{3}_{J_{C-F}}$ = 8.0 Hz, 4-F-Ph-2,6), 115.9 (pyrazol-4), 115.7 (d, $^{2}_{J_{C-F}}$ = 21.6 Hz, 4-F-Ph-3,5), 112.8 (C-3), 102.9 (C-7), 59.5 (NCH₂CH₂OH), 58.0 (NCH₂CH₂OH), 52.7 (2 CH₂ piperazine), 46.8 (2 CH₂ piperazine), 39.5 (CH₃). ¹⁹F NMR (282 MHz, CDCl₃) δ: -117.7. **m.p.:** 257–261 °C. **LC-MS:** retention time = 0.55 min, *m/z* [M+H]⁺ = 422.25.

### 4.1.4.34. 2-(3-fluorophenyl)-6-(4-(2-hydroxyethyl)piperazin-1-yl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (14d).
Column: silica. Eluent: CH₂Cl₂/EtOH (100/0 → 90/10), yellow solid, 120 mg, yield: 94 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.02 (dd, *J* = 3.0, 1.2 Hz, 1H, thien-2), 8.02 (s, 1H, H-3), 7.79 (dd, *J* = 5.1, 1.1 Hz, 1H, thien-4), 7.77-7.68 (m, 2H, 3-F-Ph-2,6), 7.46 (dd, *J* = 5.1, 3.1 Hz, 1H, thien-5), 7.43-7.33 (m, 1H, 3-F-Ph-5), 7.05-6.94 (m, 2H, H-7 & 3-F-Ph-4), 3.69 (t, *J* = 5.3 Hz, 2H, NCH₂CH₂OH), 3.61-3.53 (m, 4H, 2 CH₂ piperazine), 2.72-2.67 (m, 4H, 2 CH₂ piperazine), 2.64 (t, *J* = 5.1 Hz, 2H, NCH₂CH₂OH). ¹³C NMR (75 MHz, CDCl₃) δ: 163.5 (d, $^{1}_{J_{C-F}}$ = 244.4 Hz, 3-F-Ph-3), 155.6 (C-6), 142.4 (d, $^{4}_{J_{C-F}}$ = 2.7 Hz, C-2), 136.7 (d, $^{3}_{J_{C-F}}$ = 8.2 Hz, 3-F-Ph-1), 135.9 (C-8a), 134.6 (C-8), 131.5 (thien-3), 130.2 (d, $^{3}_{J_{C-F}}$ = 8.5 Hz, 3-F-Ph-5), 129.7 (thien-2), 126.19 (thien-4*), 126.16 (thien-5*), 121.2 (d, $^{4}_{J_{C-F}}$ = 2.7 Hz, 3-F-Ph-6), 114.4 (d, $^{2}_{J_{C-F}}$ = 21.4 Hz, 3-F-Ph-4), 113.4 (C-3), 112.5 (d, $^{2}_{J_{C-F}}$ = 22.8 Hz, 3-F-Ph-2), 105.3 (C-7), 59.5 (NCH₂CH₂OH), 58.0 (NCH₂CH₂OH), 52.6 (2 CH₂ piperazine), 46.7 (2 CH₂ piperazine). ¹⁹F NMR (282 MHz, CDCl₃) δ: -113.4. **m.p.:** 185–189 °C. **LC-MS:** retention time = 1.02 min, *m/z* [M+H]⁺ = 424.25.

### 4.1.4.35. 2-(2-fluorophenyl)-6-(4-(2-hydroxyethyl)piperazin-1-yl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (14e).
Column: silica. Eluent: acetone/EtOH (100/0 → 30/70), yellow solid, 46 mg, yield: 39 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.04 (d, *J* = 2.9 Hz, 1H, thien-2), 8.48-8.39 (m, 1H, 2-F-Ph-6), 8.21 (d, $^{TS}_{J}$ = 4.3 Hz, 1H, H-3), 7.80 (d, *J* = 5.1 Hz, 1H, thien-4), 7.46 (dd, *J* = 4.5, 3.1 Hz, 1H, thien-5), 7.31-7.22 (m, 2H, 2-F-Ph-4,5), 7.20-7.09 (m, 1H, 2-F-Ph-3), 7.01 (s, 1H, H-7), 3.69 (t, *J* = 5.3 Hz, 2H, NCH₂CH₂OH), 3.63-3.53 (m, 4H, 2 CH₂ piperazine), 2.74-2.67 (m, 4H, 2 CH₂ piperazine), 2.64 (t, *J* = 5.1 Hz, 2H,
NCH₂CH₂CH₂OH). ¹³C NMR (75 MHz, CDCl₃) δ: 160.3 (d, $^{1}$J$_{C-F}$ = 249.6 Hz, 2-F-Ph-2), 155.5 (C-6), 137.2 (C-2), 135.2 (C-8a), 134.8 (C-8), 131.3 (thien-3), 129.6 (thien-2), 128.6 (d, $^{3}$J$_{C-F}$ = 8.6 Hz, 2-F-Ph-4), 128.4 (2-F-Ph-6), 126.2 (thien-4*), 126.1 (thien-5*), 124.4 (2-F-Ph-5), 122.1 (d, $^{2}$J$_{C-F}$ = 13.3 Hz, 2-F-Ph-1), 117.0 (d, ⁴TSJ$_{C-F}$ = 14.9 Hz, C-3), 115.8 (d, $^{2}$J$_{C-F}$ = 20.0 Hz, 2-F-Ph-3), 105.3 (C-7), 59.5 (NCH₂CH₂CH₂OH), 57.9 (NCH₂CH₂CH₂OH), 52.6 (2 CH₂ piperazine), 46.7 (2 CH₂ piperazine). ¹⁹F NMR (282 MHz, CDCl₃) δ: −114.2. m.p.: 128–132 °C. **LC-MS:** retention time = 1.02 min, m/z [M+H]⁺ = 424.15.

### 4.1.4.36. 2-Phenyl-6-(4-(2-hydroxyethyl)piperazin-1-yl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (14f).
Column: silica. Eluent: cyclohexane/EtOH (93/7 → 50/50), yellow solid, 81 mg, yield: 62 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.05 (dd, *J* = 3.0, 1.1 Hz, 1H, thien-2), 8.04 (s, 1H, H-3), 8.03–7.97 (m, 2H, Ph-2,6), 7.80 (dd, *J* = 5.1, 1.1 Hz, 1H, thien-4), 7.49–7.39 (m, 3H, thien-5 & ph-3,5), 7.36–7.28 (m, 1H, Ph-4), 6.99 (s, 1H, H-7), 3.69 (t, *J* = 5.1 Hz, 3H, NCH₂CH₂CH₂OH), 3.61–3.53 (m, 4, 2 CH₂ piperazine), 2.74–2.67 (m, 4H, 2 CH₂ piperazine), 2.64 (t, *J* = 5.1 Hz, 3H, NCH₂CH₂CH₂OH). ¹³C NMR (75 MHz, CDCl₃) δ: 155.5 (C-6), 143.7 (C-2), 135.9 (C-8a), 134.8 (C-8), 134.3 (thien-3), 131.5 (Ph-1), 129.6 (thien-2), 128.8 (Ph-3,5), 127.7 (Ph-4), 126.2 (thien-4*), 126.1 (thien-5*), 125.7 (Ph-2,6), 113.1 (C-3), 105.0 (C-7), 59.5 (NCH₂CH₂CH₂OH), 58.0 (NCH₂CH₂CH₂OH), 52.6 (2 CH₂ piperazine), 46.8 (2 CH₂ piperazine). m.p.: 131–135 °C. **LC-MS:** retention time = 0.88 min, m/z [M+H]⁺ = 406.25.

### 4.1.4.37. 2-(2-fluorophenyl)-6-(4-(2-hydroxyethyl)piperazin-1-yl)-8-(2-(4-(2-hydroxyethyl)piperazin-1-yl)-pyrimidin-5-yl)imidazo [1,2-b]pyridazine (14g).
Column: silica. Eluent: CH₂Cl₂/MeOH (100/0 → 99/1), beige solid, 35 mg, yield: 23 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.28 (s, 1H, pyrimidin-4,6), 8.45–8.34 (m, 1H, 2-F-Ph-6), 8.21 (d, TSJ = 4.2 Hz, 1H, H-3), 7.37–7.02 (m, 3H, 2-F-Ph-3,4,5), 6.84 (s, 1H, H-7), 3.99 (bs, 4H, 2 CH₂ piperazine), 3.81–3.64 (m, 4H, 2 NCH₂CH₂CH₂OH), 3.57 (bs, 4H, 2 CH₂ piperazine), 2.83–2.54 (m, 14H, 4 CH₂ piperazine, 2 NCH₂CH₂CH₂OH & 2 OH). ¹³C NMR (75 MHz, CDCl₃) δ: 161.5 (pyrimidin-2), 160.3 (d, $^{1}$J$_{C-F}$ = 248.8 Hz, 2-F-Ph-2), 158.1 (pyrimidin-4,6), 155.4 (C-6), 137.5 (C-2), 135.0 (C-8a), 132.4 (C-8), 128.7 (d, $^{3}$J$_{C-F}$ = 8.8 Hz, 2-F-Ph-4), 128.5 (d, $^{3}$J$_{C-F}$ = 3.9 Hz, 2-F-Ph-6), 124.4 (d, $^{4}$J$_{C-F}$ = 3.2 Hz, 2-F-Ph-5), 121.9 (d, $^{2}$J$_{C-F}$ = 12.5 Hz, 2-F-Ph-1), 117.2 (d, ⁴TSJ$_{C-F}$ = 14.8 Hz, C-3), 116.6 (pyrimidin-5), 115.7 (d, $^{2}$J$_{C-F}$ = 22.4 Hz, 2-F-Ph-3), 103.8 (C-7), 59.62 (NCH₂CH₂CH₂OH), 59.57 (NCH₂CH₂CH₂OH), 58.0 (2 NCH₂CH₂CH₂OH), 53.0 (2 CH₂ piperazine), 52.6 (2 CH₂ piperazine), 46.7 (2 CH₂ piperazine), 44.1 (2 CH₂ piperazine). ¹⁹F NMR (282 MHz, CDCl₃) δ: −114.3. m.p.: 158–162 °C. **LC-MS:** retention time = 0.39 min, m/z [M+H]⁺ = 548.37.

### 4.1.4.38. 6-((S)-1-hydroxybut-2-ylamino)-2-(4-fluorophenyl)-8-(1H-indol-4-yl)imidazo [1,2-b]pyridazine (15a).
Column: alumina. Eluent: CH₂Cl₂/MeOH (100/0 → 99/1), white solid, 109 mg, yield: 86 %. ¹H NMR (300 MHz, CD₃OD) δ: 8.09 (s, 1H, H-3), 7.93–7.77 (m, 2H, 4-F-Ph-2,6), 7.64 (d, *J* = 7.3 Hz, 1H, indol-5), 7.54 (d, *J* = 8.0 Hz, 1H, indol-7), 7.33 (d, *J* = 2.8 Hz, 1H, indol-2), 7.27 (t, *J* = 7.7 Hz, 1H, indol-6), 7.14–7.03 (m, 2H, 4-F-Ph-3,5), 6.94 (s, 1H, H-7), 6.61 (d, *J* = 2.8 Hz, 1H, indol-3), 4.01–3.86 (m, 1H, CHNH), 3.70 (d, *J* = 5.0 Hz, 2H, CH₂OH), 1.90–1.72 (m, 1H, CH₃H₃CH₃), 1.70–1.53 (m, 1H, CH₄H₃CH₃), 1.06 (t, *J* = 7.4 Hz, 3H, CH₂CH₃). ¹³C NMR (75 MHz, CD₃OD) δ: 163.8 (d, $^{1}$J$_{C-F}$ = 244.8 Hz, 4-F-Ph-4), 155.8 (C-6), 142.9 (C-2), 139.0 (C-8*), 138.3 (C-8a*), 138.0 (indol-7a*), 131.9 (d, $^{4}$J$_{C-F}$ = 3.2 Hz, 4-F-Ph-1), 128.6 (d, $^{3}$J$_{C-F}$ = 8.1 Hz, 4-F-Ph-2,6), 127.8 (indol-4*), 127.2 (indol-3a*), 126.5 (indol-2), 122.2 (indol-5*), 122.1 (indol-6*), 116.2 (d, $^{2}$J$_{C-F}$ = 21.8 Hz, 4-F-Ph-3,5), 114.3 (C-3), 113.7 (indol-7), 113.1 (C-7), 102.0 (indol-3), 64.23 (CH₂OH), 55.5 (CHNH), 25.2 (CH₂CH₃), 11.0 (CH₂CH₃). ¹⁹F NMR (282 MHz, CD₃OD) δ: −117.4. m.p.: 227–231 °C. **LC-MS:** retention time = 0.90 min, m/z [M+H]⁺ = 416.05.

### 4.1.4.39. 6-((S)-1-hydroxybut-2-ylamino)-2-(4-fluorophenyl)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (15b).
Column: silica. Eluent: CH₂Cl₂, yellow solid, 51 mg, yield: 45 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.84 (s, 1H, thien-2), 7.97–7.88 (m, 2H, 4-F-Ph-2,6), 7.86 (s, 1H, H-3), 7.63 (d, *J* = 4.3 Hz, 1H, thien-4), 7.36 (s, 1H, thien-5), 7.17–7.08 (m, 2H, 4-F-Ph-3,5), 6.76 (s, 1H, H-7), 4.79 (bs, 1H, NH*), 4.04–3.83 (m, 2H, CHNH & CH₃H₃OH), 3.73 (ddAB, *J* = 10.9, 5.7 Hz, 1H, CH₃H₃OH), 2.83 (bs, 1H, OH*), 1.81–1.55 (m, 2H, CH₂CH₃), 1.04 (t, *J* = 7.4 Hz, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.8 (d, $^{1}$J$_{C-F}$ = 246.8 Hz, 4-F-Ph-4), 154.4 (C-6), 141.3 (C-2), 135.3 (C-8a), 133.8 (C-8), 131.0 (thien-3), 129.6 (thien-2), 127.5 (d, $^{3}$J$_{C-F}$ = 8.1 Hz, 4-F-Ph-2,6), 126.15 (thien-4*), 126.11 (thien-5*), 115.8 (d, $^{3}$J$_{C-F}$ = 21.8 Hz, 4-F-Ph-3,5), 112.8 (C-3), 108.0 (C-7), 64.8 (CH₂OH), 55.3 (CHNH), 24.7 (CH₂CH₃), 10.8 (CH₂CH₃). (Note: 4-F-Ph-1 is missing, probably overlapped with thien-2). ¹⁹F NMR (282 MHz, CDCl₃) δ: −114.0. m.p.: 147–151 °C. **LC-MS:** retention time = 2.51 min, m/z [M+H]⁺ = 383.15.

### 4.1.4.40. 6-((S)-1-hydroxybut-2-ylamino)-2-(4-fluorophenyl)-8-(1-methyl-1h-pyrazol-4-yl)imidazo [1,2-b]pyridazine (15c).
Column: silica. Eluent: CH₂Cl₂/MeOH (100/0 → 99/1), yellow solid, 51 mg, yield: 44 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.75 (s, 1H, pyrazol-5), 7.99 (s, 1H, pyrazol-3), 7.96–7.87 (m, 2H, 4-F-Ph-2,6), 7.84 (s, 1H, H-3), 7.12 (t, *J* = 8.6 Hz, 2H, 4-F-Ph-3,5), 6.62 (s, 1H, H-7), 4.62 (bs, 1H, OH), 4.07–3.83 (m, 5H, NCH₃, CHNH & CH₃H₃OH), 3.71 (dd, *J* = 10.9, 5.6 Hz, 1H, CH₃H₃OH), 2.85 (bs, 1H, NH), 1.81–1.54 (m, 2H, CH₂CH₃), 1.04 (t, *J* = 7.4 Hz, 3H, CH₂CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.7 (d, $^{1}$J$_{C-F}$ = 246.6 Hz, 4-F-Ph-4), 154.3 (C-6), 141.4 (C-2), 137.9 (pyrazol-3), 135.2 (C-8a), 132.7 (pyrazol-5), 130.0 (4-F-Ph-1), 129.4 (C-8), 127.4 (d, $^{3}$J$_{C-F}$ = 8.1 Hz, 4-F-Ph-2,6), 115.8 (d, $^{2}$J$_{C-F}$ = 21.6 Hz, 4-F-Ph-3,5), 115.2 (pyrazol-4), 112.9 (C-3), 105.1 (C-7), 65.0 (CH₂OH), 55.3 (CHNH), 39.4 (NCH₃), 24.8 (CH₂CH₃), 10.9 (CH₂CH₃). m.p.: 200–204 °C. ¹⁹F NMR (282 MHz, CDCl₃) δ: −113.8. **LC-MS:** **LC-MS:** retention time = 1.17 min, m/z [M+H]⁺ = 381.25.

### 4.1.4.41. 2-(4-fluorophenyl)-8-(1H-indol-4-yl)-6-(N-morpholinyl)imidazo [1,2-b]pyridazine (16).
Column: alumina. Eluent: CH₂Cl₂, brown solid, 136 mg, yield: 94 %. ¹H NMR (300 MHz, acetone-d6) δ: 8.34 (s, 1H, H-3), 8.07–7.98 (m, 2H, 4-F-Ph-2,6), 7.92 (dd, *J* = 7.4, 0.8 Hz, 1H, indol-5), 7.61 (d, *J* = 8.2 Hz, 1H, indol-7), 7.46 (d, *J* = 3.2 Hz, 1H, indol-2), 7.35–7.26 (m, 2H, H-7 & indol-6), 7.21–7.11 (m, 2H, 4-F-Ph-3,5), 6.71 (dd, *J* = 3.2, 0.8 Hz, 1H, indol-3), 3.92–3.74 (m, 4H, 2 CH₂O morpholine), 3.65–3.45 (m, 4H, 2 CH₂N morpholine). ¹³C NMR (75 MHz, acetone-d6) δ: 163.2 (d, $^{1}$J$_{C-F}$ = 244.0 Hz, 4-F-Ph-4), 156.6 (C-6), 143.4 (C-2), 138.9 (C-8a), 137.8 (C-8*), 137.4 (indol-7a*), 132.1 (d, $^{4}$J$_{C-F}$ = 3.0 Hz, 4-F-Ph-1), 128.1 (d, $^{3}$J$_{C-F}$ = 8.0 Hz, 4-F-Ph-2,6), 127.5 (indol-4*), 127.3 (indol-3a*), 126.7 (indol-2), 122.8 (indol-5), 121.8 (indol-6), 116.1 (d, $^{2}$J$_{C-F}$ = 21.7 Hz, 4-F-Ph-3,5), 113.71 (C-3*), 113.68 (indol-7*), 109.4 (C-7), 102.1 (indol-3), 67.1 (2 CH₂O morpholine), 47.5 (2 CH₂N morpholine). ¹⁹F NMR (282 MHz, acetone-d6) δ: −116.7. m.p.: 236–240 °C. **LC-MS:** retention time = 1.78 min, m/z [M+H]⁺ = 414.25.

### 4.1.5. General procedure for nucleophilic aromatic substitution using 2-methoxyethanol, synthesis of 17a-c

3.4 equivalents of NaH were weighed into a round-bottom flask. Anhydrous THF was added (ca. 5 mL) followed by 2-methoxyethanol (4 eq). The reaction mixture was stirred at room temperature for 15 min then compound 4b/4h/4j (ca. 0.3 mmol) was added. The reaction mixture was stirred at 70 °C for 54 h 25 mL of a saturated aqueous NaCl solution were added and the reaction mixture was extracted with three times 10 mL of ethyl acetate. The organic phases were combined, dried over MgSO₄ and evaporated. The crude mixture was then purified by alumina or silica column chromatography.

#### 4.1.5.1. 2-(4-fluorophenyl)-8-(1H-indol-4-yl)-6-(2-methoxyethoxy)imidazo [1,2-b]pyridazine (17a).
Column: silica. Eluent: CH₂Cl₂, yellow
solid, 75 mg, yield: 61 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.61 (bs, 1H, NH), 8.06 (s, 1H, H-3), 7.97-7.87 (m, 3H, 4-F-Ph-2,6 & indol-5), 7.43 (d, J = 8.2 Hz, 1H, indol-7) 7.31 (t<sub>AB</sub>, J = 7.8 Hz, 1H, indol-6), 7.22-7.16 (m, 1H, indol-2), 7.14-7.02 (m, 3H, 4-F-Ph-3,5 & H-7), 6.76-6.70 (m, 1H, indol-3), 4.59-4.52 (m, 2H, OCH₂CH₂CH₂OMe), 3.88-3.79 (m, 2H, OCH₂CH₂CH₂OMe), 3.49 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.8 (d, ¹J<sub>C-F</sub> = 246.7 Hz, 4-F-Ph-4), 160.2 (C-6), 143.0 (C-2), 139.6 (C-8a), 137.3 (C-8*), 136.6 (indol-7a*), 130.0 (d, ⁴J<sub>C-F</sub> = 2.9 Hz, 4-F-Ph-1), 127.8 (d, ³J<sub>C-F</sub> = 8.1 Hz, 4-F-Ph-2,6), 126.5 (indol-2), 125.6 (indol-3a), 125.3 (indol-4), 122.4 (indol-5), 121.8 (indol-6), 115.7 (d, ²J<sub>C-F</sub> = 21.7 Hz, 4-F-Ph-3,5), 113.2 (C-3*), 113.1 (indol-7*), 110.6 (C-7), 102.1 (indol-3). 70.7 (OCH₂CH₂CH₂OMe), 66.6 (OCH₂CH₂CH₂OMe), 59.3 (OCH₃). ¹⁹F NMR (282 MHz, CDCl₃) δ: -114.5. m.p.: 214-218 °C. LC-MS: retention time = 2.51 min, m/z [M+H]⁺ = 403.15.

### 4.1.5.2. 2-(4-fluorophenyl)-6-(2-methoxyethoxy)-8-(thien-3-yl)imidazo [1,2-b]pyridazine (17b).
Column: silica. Eluent: CH₂Cl₂/EtOAc (70/30), white solid, 108 mg, yield: 96 %. ¹H NMR (300 MHz, CDCl₃) δ: 9.09 (dd, J = 3.0, 1.1 Hz, 1H, thien-2), 8.03-7.93 (m, 3H, H-3 & 4-F-Ph-2,6), 7.77 (dd, J = 5.1, 1.1 Hz, 1H, thien-4), 7.46 (dd, J = 5.1, 3.0 Hz, 1H, thien-5), 7.18-7.09 (m, 2H, 4-F-Ph-3,5), 6.98 (s, 1H, H-7), 4.55-4.48 (m, 2H, OCH₂CH₂OMe), 3.85-3.77 (m, 2H, OCH₂CH₂OMe), 3.48 (s, 3H, CH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.8 (d, ¹J<sub>C-F</sub> = 246.7 Hz, 4-F-Ph-4), 160.2 (C-6), 142.9 (C-2), 136.5 (C-8a), 134.0 (C-8), 132.7 (thien-3), 130.2 (thien-2), 130.0 (d, ⁴J<sub>C-F</sub> = 3.1 Hz, 4-F-Ph-1), 127.6 (d, ³J<sub>C-F</sub> = 8.1 Hz, 4-F-Ph-3), 126.3 (thien-4 & thien-7), 115.8 (d, ²J<sub>C-F</sub> = 21.7 Hz, 4-F-Ph-3,5), 113.0 (C-3), 106.8 (C-7), 70.7 (OCH₂CH₂CH₂OMe), 66.6 (OCH₂CH₂CH₂OMe), 59.3 (OCH₃). ¹⁹F NMR (282 MHz, CDCl₃) δ: -114.2. m.p.: 157-161 °C. LC-MS: retention time = 3.19 min, m/z [M+H]⁺ = 370.05.

### 4.1.5.3. 2-(4-fluorophenyl)-6-(2-methoxyethoxy)-8-(1-methyl-1H-pyrazol-4-yl)imidazo [1,2-b]pyridazine (17c).
Column: silica. Eluent: CH₂Cl₂/MeOH (100/0 → 99/1), white solid, 86 mg, yield: 77 %. ¹H NMR (300 MHz, CDCl₃) δ: 8.85 (s, 1H, pyrazol-5), 8.13 (d, J = 0.5 Hz, 1H, pyrazol-3), 8.01-7.91 (m, 3H, H-3 & 4-F-Ph-2,6), 7.20-7.08 (m, 2H, 4-F-Ph-3,5), 6.90 (s, 1H, H-7), 4.53-4.46 (m, 2H, OCH₂CH₂OMe), 4.04 (s, 3H, NCH₃), 3.83-3.76 (m, 2H, OCH₂CH₂CH₂OMe), 3.48 (s, 3H, OCH₃). ¹³C NMR (75 MHz, CDCl₃) δ: 162.8 (d, ¹J<sub>C-F</sub> = 246.8 Hz, 4-F-Ph-4), 160.2 (C-6), 142.7 (C-2), 138.5 (pyrazol-3), 136.2 (C-8a), 132.7 (pyrazol-5), 131.0 (C-8), 130.1 (d, ⁴J<sub>C-F</sub> = 3.1 Hz, 4-F-Ph-1), 127.5 (d, ³J<sub>C-F</sub> = 8.1 Hz, 4-F-Ph-2,6), 115.8 (d, ²J<sub>C-F</sub> = 21.7 Hz, 4-F-Ph-3,5), 115.4 (pyrazol-4), 113.1 (C-3), 104.6 (C-7), 70.7 (OCH₂CH₂CH₂OMe), 66.5 (OCH₂CH₂CH₂OMe), 59.3 (OCH₃), 39.5 (NCH₃). ¹⁹F NMR (282 MHz, CDCl₃) δ: -114.2. m.p.: 153-157 °C. LC-MS: retention time = 2.06 min, m/z [M+H]⁺ = 367.95.

## 4.1.6. General procedure for salt formation, synthesis of 18a-l

To a solution of compounds 9/10/11/13 (ca. 0.47 mmol) in an isopropanol/water mixture (ca. 850 µL/150 µL) was added HCl (37 %, 2.2 eq). The reaction mixture was then stirred for 30 min at 60 °C. Acetone (ca. 1.2 mL) was then added and the reaction mixture was heated to 60 °C for another 1.5 h. After cooling in an ice bath, the solid was filtered and dried under vacuum to obtain the salts 18a-l.

### 4.1.6.1. 4-(8-Bromo-2-(4-fluorophenyl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18a).
White solid. 92 mg, yield: 76 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 10.97 (bs, 1H, N⁺H), 8.68 (s, 1H, H-3), 8.15-7.95 (m, 2H, 4-F-Ph-2,6), 7.41-7.20 (m, 2H, 4-F-Ph-3,5), 6.64 (s, 1H, H-7), 5.26 (bs, 2H, 2 CH piperazine), 3.76-3.46 (m, 4H, 4 CH piperazine), 3.24 (bs, 2H, 2 CH piperazine), 2.80 (s, 3H, CH₃). (Note: For the entire series of salts, the piperazine protons have become diastereotopic due to quaternization of the ammonium. The signals then appear as separate CH and not as CH₂). m.p.: 304-308 °C. LC-MS: retention time = 0.63 min, m/z [M + H-C₂H₆N]⁺ = 346.15 et [M-C₅H₁₂N₂]⁺ = 289.14.

### 4.1.6.2. 4-(2-(4-fluorophenyl)-8-phenylimidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18b).
White solid. 301 mg, yield: 90 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.27 (bs, 1H, N⁺H), 8.64 (s, 1H, H-3), 8.46-8.26 (m, 2H, Ph-2,6), 8.12-7.97 (m, 2H, 4-F-Ph-2,6), 7.66-7.55 (m, 3H, Ph-3,4,5), 7.38-7.22 (m, 2H, 4-F-Ph-3,5), 4.41 (d, J = 13.7 Hz, 2H, 2 CH piperazine), 3.62-3.34 (m, 4H, 4 CH piperazine), 3.29-3.06 (m, 2H, 2 CH piperazine), 2.80 (d, J = 4.6 Hz, 3H, CH₃). m.p.: 287-291 °C. LC-MS: retention time = 0.88 min, m/z [M+H]⁺ = 388.25.

### 4.1.6.3. 4-(2-(4-fluorophenyl)-8-(1H-indol-4-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18c).
Yellow solid. 50 mg, yield: 77 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.61-11.35 (m, 2H, NH & N⁺H), 8.75 (s, 1H, H-3), 8.05-7.92 (m, 2H, 4-F-Ph-2,6), 7.70-7.58 (m, 2H, H-7 & indol-5), 7.58-7.46 (m, 2H, indol-7 & indol-2), 7.40-7.21 (m, 3H, indol-6 & 4-F-Ph-3,5), 6.54 (bs, 1H, indol-3), 4.37 (d, J = 13.4 Hz, 2H, 2 CH piperazine), 3.59-3.41 (m, 4H, 4 CH piperazine), 3.29-3.10 (m, 2H, 2 CH piperazine), 2.80 (d, J = 4.3 Hz, 3H, CH₃). m.p.: 243-247 °C. LC-MS: retention time = 0.59 min, m/z [M+H]⁺ = 427.26.

### 4.1.6.4. 4-(2-(4-fluorophenyl)-8-(pyridin-4-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18d).
Orange solid. 72 mg, yield: 86 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.06 (bs, 1H, N⁺H), 8.98 (d, J = 6.2 Hz, 2H, pyridin-2,6), 8.75 (d, J = 5.8 Hz, 2H, pyridin-3,5), 8.71 (s, 1H, H-3), 8.15-8.02 (m, 2H, 4-F-Ph-2,6), 7.78 (s, 1H, H-7), 7.39-7.25 (m, 2H, 4-F-Ph-3,5), 4.43 (d, J = 13.8 Hz, 2H, 2 CH piperazine), 3.64-3.33 (m, 4H, 4 CH piperazine), 3.30-3.07 (m, 2H, 2 CH piperazine), 2.82 (d, J = 3.7 Hz, 3H, CH₃). m.p.: 231-235 °C. LC-MS: retention time = 0.54 min, m/z [M+H]⁺ = 389.25.

### 4.1.6.5. 4-(2-(4-fluorophenyl)-8-(pyridin-3-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18e).
Yellow solid. 189 mg, yield: 95 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.07 (bs, 1H, N⁺H), 9.66 (d, J = 1.8 Hz, 1H, pyridin-2), 9.03 (d, J = 7.7 Hz, 1H, pyridin-4), 8.84 (dd, J = 5.0, 1.3 Hz, 1H, pyridin-6), 8.67 (s, 1H, H-3), 8.12-7.99 (m, 2H, 4-F-Ph-2,6), 7.82 (dd, J = 7.9, 5.1 Hz, 1H, pyridin-5), 7.69 (s, 1H, H-7), 7.37-7.22 (m, 2H, 4-F-Ph-3,5), 4.43 (d, J = 14.0 Hz, 2H, 2 CH piperazine), 3.62-3.35 (m, 4H, 4 CH piperazine), 3.29-3.08 (m, 2H, 2 CH piperazine), 2.81 (d, J = 4.3 Hz, 3H, CH₃). m.p.: 306-310 °C. LC-MS: retention time = 0.55 min, m/z [M+H]⁺ = 389.25.

### 4.1.6.6. 4-(2-(4-fluorophenyl)-8-(thien-3-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18f).
Yellow solid. 209 mg, yield: 97 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.12 (bs, 1H, N⁺H), 9.23 (d, J = 1.9 Hz, 1H, thien-2), 8.59 (s, 1H, H-3), 8.24 (d, J = 4.4 Hz, 1H, thien-4), 8.17-8.06 (m, 2H, 4-F-Ph-2,6), 7.79 (dd, J = 5.0, 3.0 Hz, 1H, thien-5), 7.61 (s, 1H, H-7), 7.37-7.20 (m, 2H, 4-F-Ph-3,5), 4.40 (d, J = 13.7 Hz, 2H, 2 CH piperazine), 3.59-3.33 (m, 4H, 4 CH piperazine), 3.26-3.09 (m, 2H, 2 CH piperazine), 2.81 (d, J = 4.4 Hz, 3H, CH₃). m.p.: 302-306 °C. LC-MS: retention time = 1.06 min, m/z [M+H]⁺ = 394.15.

### 4.1.6.7. 4-(2-(4-fluorophenyl)-8-(1-methyl-1H-pyrazol-4-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18g).
Yellow solid. 82 mg, yield: 94 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.20 (bs, 1H, N⁺H), 8.98 (s, 1H, pyrazol-5), 8.55 (s, 1H, pyrazol-3), 8.54 (s, 1H, H-3), 8.17-8.08 (m, 2H, 4-F-Ph-2,6), 7.50 (s, 1H, H-7), 7.34-7.23 (m, 2H, 4-F-Ph-3,5), 4.00 (s, 3H, CH₃ pyrazole), 3.58-3.33 (m, 4H, 4 CH piperazine), 3.25-3.07 (m, 2H, 2 CH piperazine), 2.80 (d, J = 4.6 Hz, 3H, CH₃ piperazine). (Note: the two missing piperazine CH signals are overlapped with water). m.p.: 312-316 °C. LC-MS: retention time = 0.58 min, m/z [M+H]⁺ = 392.15.

### 4.1.6.8. 4-(2-(4-fluorophenyl)-8-(1-methyl-1H-pyrazol-5-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18h).
Yellow solid. 210 mg, yield: 93 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.29 (bs, 1H, N⁺H), 8.66 (s, 1H, H-3), 8.10-7.94 (m, 2H, 4-F-Ph-2,6), 7.64 (d, J = 1.9 Hz, N⁺H).
Hz, 1H, pyrazol-3), 7.36 (s, 1H, H-7), 7.33–7.18 (m, 2H, 4-F-Ph-3,5), 6.87 (d, *J* = 1.9 Hz, 1H, pyrazol-4), 4.01 (s, 3H, CH₃ pyrazole), 3.59–3.37 (m, 4H, 4 CH piperazine), 3.27–3.08 (m, 2H, 2 CH piperazine), 2.80 (d, *J* = 4.6 Hz, 3H, CH₃). (Note: *the two missing piperazine CH signals are overlapped with water*). **m.p.:** 305–309 °C. **LC-MS:** retention time = 0.55 min, *m/z* [*M+H]⁺ = 392.15.

4.1.6.9. **4-(2-(4-fluorophenyl)-8-(furan-3-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18i).** Yellow solid. 62 mg, yield: 63 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 10.66 (bs, 1H, N⁺H), 9.11 (d, *J* = 0.9 Hz, 1H, furan-2), 8.56 (s, 1H, H-3), 8.19–8.05 (m, 2H, 4-F-Ph-2,6), 7.94 (t, *J* = 1.7 Hz, 1H, furan-5), 7.56–7.44 (m, 2H, H-7 & furan-4), 7.34–7.21 (m, 2H, 4-F-Ph-3,5), 4.39 (d, *J* = 13.6 Hz, 2H, 2 CH piperazine), 3.55 (d, *J* = 11.4 Hz, 2H, 2 CH piperazine), 3.27–3.10 (m, 2H, 2 CH piperazine), 2.83 (d, *J* = 3.8 Hz, 3H, CH₃). (Note: *the two missing piperazine CH signals are overlapped with water*). **m.p.:** 313–317 °C. **LC-MS:** retention time = 0.90 min, *m/z* [*M+H]⁺ = 378.15.

4.1.6.10. **4-(2-(3-fluorophenyl)-8-(pyridin-3-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18j).** Yellow solid. 173 mg, yield: 92 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 10.87 (bs, 1H, N⁺H), 9.62 (d, *J* = 1.7 Hz, 1H, pyridin-2), 8.97 (d, *J* = 8.1 Hz, 1H, pyridin-4), 8.82 (dd, *J* = 4.9, 1.5 Hz, 1H, pyridin-6), 8.74 (s, 1H, H-3), 7.92–7.73 (m, 3H, 3-F-Ph-2,6 & pyridin-5), 7.68 (s, 1H, H-7), 7.56–7.42 (m, 1H, 3-F-Ph-5), 7.18–7.08 (m, 1H, 3-F-Ph-4), 4.43 (d, *J* = 13.8 Hz, 2H, 2 CH piperazine), 3.55 (d, *J* = 11.5 Hz, 2H, 2 CH piperazine), 3.49–3.34 (m, 2H, 2 CH piperazine), 3.27–3.10 (m, 2H, 2 CH piperazine), 2.82 (d, *J* = 4.2 Hz, 3H, CH₃). **m.p.:** 308–312 °C. **LC-MS:** retention time = 0.55 min, *m/z* [*M+H]⁺ = 389.15.

4.1.6.11. **4-(2-(2-fluorophenyl)-8-(pyridin-3-yl)imidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18k).** Yellow solid. 22 mg, yield: 45 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.15 (bs, 1H, N⁺H), 9.70 (d, *J* = 1.7 Hz, 1H, pyridin-2), 9.08 (d, *J* = 8.1 Hz, 1H, pyridin-4), 8.86 (dd, *J* = 5.0, 1.5 Hz, 1H, pyridin-6), 8.32 (d, *J* = 4.2 Hz, 1H, H-3), 8.30–8.20 (m, 1H, 2-F-Ph-6), 7.86 (dd, *J* = 8.1, 5.1 Hz, 1H, pyridin-5), 7.75 (s, 1H, H-7), 7.46–7.29 (m, 3H, 2-F-Ph-3,4,5), 4.45 (d, *J* = 13.5 Hz, 3H, 2 CH piperazine), 3.61–3.37 (m, 4H, 4 CH piperazine), 3.27–3.10 (m, 2H, 2 CH piperazine), 2.81 (d, *J* = 4.3 Hz, 3H, CH₃). (Note: *The proton at position 3 present an abnormally high H–F coupling constants. This is due to the proximity of fluorine to this position in space, which generates a through space spin-spin coupling*). **m.p.:** 300–304 °C. **LC-MS:** retention time = 0.54 min, *m/z* [*M+H]⁺ = 389.25.

4.1.6.12. **4-(8-(1H-indol-4-yl)-2-phenylimidazo [1,2-b]pyridazin-6-yl)-1-methylpiperazin-1-ium chloride (18l).** Yellow solid. 205 mg, yield: 92 %. ¹H NMR (300 MHz, DMSO-d₆) δ: 11.54 (bs, 1H, N⁺H*), 11.39 (bs, 1H, NH*), 8.79 (s, 1H, H-3), 8.00–7.87 (m, 2H, Ph-2,6), 7.70–7.55 (m, 3H, H-7 & indol-5,7), 7.55–7.42 (m, 3H, indol-2 & Ph-3,5), 7.42–7.27 (m, 2H, indol-6 & Ph-4), 6.54 (s, 1H, indol-3), 4.39 (d, *J* = 14.2 Hz, 2H, 2 CH piperazine), 3.63–3.41 (m, 4H, 4 CH piperazine), 3.29–3.11 (m, 2H, 2 CH piperazine), 2.80 (d, *J* = 4.6 Hz, 3H, CH₃). **m.p.:** 237–241 °C. **LC-MS:** retention time = 0.54 min, *m/z* [*M+H]⁺ = 409.25.

## 4.2. *In vitro* activity against *M. mycetomatis*

Testing the *in vitro* susceptibility of the synthesised imidazo [1,2-b] pyridazine analogues was done via a Viability-Based 2,3-Bis(2-Methoxy-4-Nitro-5-Sulfophenyl)-5-[(Phenylamino)Carbonyl]-2H-Tetrazolium Hydroxide (XTT) Assay first described by Van de Sande et al. [24] In this optimised protocol, *M. mycetomatis* genome strain MM55 was cultured in RPMI 1640 medium supplemented with 0.3 g/L L-glutamine and 20 mM morpholinepropanesulfonic acid (MOPS), then incubated at 37 °C for ten days. The mycelia were harvested through a 5-min centrifugation and washed with sterile saline, followed by sonication for 20 s at 28 µm

(Soniprep, Beun de Ronde, The Netherlands) to achieve a homogeneous suspension. This suspension is then mixed with RPMI medium to obtain a final inoculum suspension with a transmission of 70 % at 660 nm (Novaspec II; Pharmacia Biotech). Antifungal screening was done using 96-well microplates, and end-point reading was done using XTT as a viability indicator; all experiments were done in triplicates. The preliminary screening of the compounds was done at 25 µM or 10 µM and 100 µM concentrations. Only compounds that caused >80 % inhibition of fungal growth at both concentrations were selected for further testing of their IC₅₀ values at concentrations ranging from 16 µM to 0.03 µM [24]. The MIC values against MM55 and other *M. mycetomatis* strains (SO1, l1, l3, l11, P1, Peru72012, AL1, CBS247.48, and MM50) were then determined for compounds with an IC₅₀ ≤ 5 µM [18].

## 4.3. Cytotoxicity against NIH-3T3 murine fibroblast

The investigation of the basal *in vitro* cytotoxicity of the synthesised imidazo [1,2-b]pyridazine analogues involved the utilisation of the NIH-3T3 murine fibroblast cell line and the (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The NIH-3T3 cells were cultured in Dulbecco's Modified Eagle Medium (DMEM), supplemented with 4.5 g/L glucose and 10 % fetal calf serum in 75-cm² culture flasks. The flasks were incubated at 37 °C in a 5 % CO₂ humidified atmosphere until the cells reached a 70–90 % confluence range. Cells were then trypsinised and seeded into 24-well plates at a density of 50,000 cells per well in 0.5 ml of culture medium, after which a 24-h incubation period was instituted. The medium in the wells was then replaced with 0.5 ml of DMEM containing 50 µM, 30 µM, 15 µM, 10 µM, 3 µM, 1 µM, and 0.1 µM concentrations of each compound and plates were incubated for five days at 37 °C. Wells were then washed, and 0.4 ml MTT working solution (0.05 mg/ml of MTT in complete media) was dispensed into each well, followed by a 70-min incubation period. 0.4 ml of DMSO was then added into each well to dissolve the formed formazan crystals. Absorbance readings were conducted at a wavelength of 570 nm using a microplate reader. Experiments were conducted in triplicate, and IC₅₀ values were calculated using GraphPad Prism version 8.

## Author information

The authors declare no competing financial interest.

### CRediT authorship contribution statement

**Lamis Yahia Mohamed Elkheir:** Writing – original draft, Project administration, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. **Pierre-Olivier Delaye:** Writing – original draft, Supervision, Project administration, Methodology, Formal analysis, Data curation, Conceptualization. **Mélanie Penichon:** Methodology, Investigation, Data curation. **Kimberly Eadie:** Investigation. **Magdi Awadalla Mohamed:** Supervision, Project administration. **Pierre Besson:** Validation, Supervision, Conceptualization. **Adélaïde Chesnay:** Investigation. **Guillaume Desoubeaux:** Methodology, Investigation. **Sébastien Roger:** Validation, Supervision, Conceptualization. **Wendy Wilhelmina Johanna van de Sande:** Writing – original draft, Validation, Methodology, Investigation, Formal analysis. **Ahmed Hassan Fahal:** Validation, Supervision, Project administration, Investigation, Funding acquisition, Formal analysis, Data curation, Conceptualization. **Cécile Enguehard-Gueiffier:** Writing – review & editing, Writing – original draft, Supervision, Project administration, Conceptualization.

### Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
## Data availability

No data was used for the research described in the article.

## Acknowledgements

Lamis Yahia Mohamed Elkheir was supported by a grant from the Sudanese government and French embassy of Sudan.

## Appendix A. Supplementary data

Supplementary data to this article can be found online at https://doi.org/10.1016/j.ejmech.2024.116720.

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