Cinnoline Scaffold—A Molecular Heart of Medicinal Chemistry?

The cinnoline nucleus is a very important bicyclic heterocycle that is used as the structural subunit of many compounds with interesting pharmaceutical properties. Cinnoline derivatives exhibit broad spectrum of pharmacological activities such as antibacterial, antifungal, antimalarial, anti-inflammatory, analgesic, anxiolytic and antitumor activities. Some of them are under evaluation in clinical trials. In the present review, we have compiled studies focused on the biological properties of cinnoline derivatives conducted by many research groups worldwide between 2005 and 2019. Comprehensive and target oriented information clearly indicate that the development of cinnoline based molecules constitute a significant contribution to the identification of lead compounds with optimized pharmacodynamic and pharmacokinetic properties.


Antimicrobial Activity
Infectious diseases constitute a growing therapeutic challenge worldwide due to the developing resistance of pathogens to known drugs [17,18]. As a consequence, there is an urgent need to design new compounds with improved activity against drug-sensitive as well as drug-resistant pathogens. Cinnoline derivatives were widely studied as antimicrobial agents [1]. Cinoxacin 3 ( Figure 3) is a common drug used in urinary tract infections [19]. Since it has a high phototoxicity index, Vargas et al. synthesized the naphthyl ester of cinoxacin 4 ( Figure 3) in order to evaluate its possible application in antibacterial phototherapy. The ester derivative of cinoxacin 4 exhibited comparable photostability and antibacterial activity against E. coli to the parent drug but enhanced antibacterial activity upon irradiation [20]. Depicted in Figure 4, 6-hydroxycinnolines were synthesized and tested for in vitro antifungal activity against Candida and Aspergillus species. It was elucidated that most of the obtained compounds exhibited potent antifungal activity against C. krusei, C. neoformans, and A. niger, with the highest activity towards C. neoformans [21].   Cinnoline, together with other bicyclic scaffolds, is the subject of our special interest as the terminal moiety of symmetrical compounds designed in agreement with the bisintercalators' structural requirements [15,16]. Our previous review describing the biological properties of cinnoline derivatives included papers and patents published until 2004 [1]. Herein, we aimed to review documents published from 2005 to 2019, focusing on the compounds bearing a cinnoline nucleus, in particular with respect to their biological activity and potential therapeutic use.

Antimicrobial Activity
Infectious diseases constitute a growing therapeutic challenge worldwide due to the developing resistance of pathogens to known drugs [17,18]. As a consequence, there is an urgent need to design new compounds with improved activity against drug-sensitive as well as drug-resistant pathogens. Cinnoline derivatives were widely studied as antimicrobial agents [1]. Cinoxacin 3 ( Figure 3) is a common drug used in urinary tract infections [19]. Since it has a high phototoxicity index, Vargas et al. synthesized the naphthyl ester of cinoxacin 4 ( Figure 3) in order to evaluate its possible application in antibacterial phototherapy. The ester derivative of cinoxacin 4 exhibited comparable photostability and antibacterial activity against E. coli to the parent drug but enhanced antibacterial activity upon irradiation [20]. Cinnoline, together with other bicyclic scaffolds, is the subject of our special interest as the terminal moiety of symmetrical compounds designed in agreement with the bisintercalators' structural requirements [15,16]. Our previous review describing the biological properties of cinnoline derivatives included papers and patents published until 2004 [1]. Herein, we aimed to review documents published from 2005 to 2019, focusing on the compounds bearing a cinnoline nucleus, in particular with respect to their biological activity and potential therapeutic use.

Antimicrobial Activity
Infectious diseases constitute a growing therapeutic challenge worldwide due to the developing resistance of pathogens to known drugs [17,18]. As a consequence, there is an urgent need to design new compounds with improved activity against drug-sensitive as well as drug-resistant pathogens. Cinnoline derivatives were widely studied as antimicrobial agents [1]. Cinoxacin 3 ( Figure 3) is a common drug used in urinary tract infections [19]. Since it has a high phototoxicity index, Vargas et al. synthesized the naphthyl ester of cinoxacin 4 ( Figure 3) in order to evaluate its possible application in antibacterial phototherapy. The ester derivative of cinoxacin 4 exhibited comparable photostability and antibacterial activity against E. coli to the parent drug but enhanced antibacterial activity upon irradiation [20]. Depicted in Figure 4, 6-hydroxycinnolines were synthesized and tested for in vitro antifungal activity against Candida and Aspergillus species. It was elucidated that most of the obtained compounds exhibited potent antifungal activity against C. krusei, C. neoformans, and A. niger, with the highest activity towards C. neoformans [21].   Depicted in Figure 4, 6-hydroxycinnolines were synthesized and tested for in vitro antifungal activity against Candida and Aspergillus species. It was elucidated that most of the obtained compounds exhibited potent antifungal activity against C. krusei, C. neoformans, and A. niger, with the highest activity towards C. neoformans [21]. Cinnoline, together with other bicyclic scaffolds, is the subject of our special interest as the terminal moiety of symmetrical compounds designed in agreement with the bisintercalators' structural requirements [15,16]. Our previous review describing the biological properties of cinnoline derivatives included papers and patents published until 2004 [1]. Herein, we aimed to review documents published from 2005 to 2019, focusing on the compounds bearing a cinnoline nucleus, in particular with respect to their biological activity and potential therapeutic use.

Antimicrobial Activity
Infectious diseases constitute a growing therapeutic challenge worldwide due to the developing resistance of pathogens to known drugs [17,18]. As a consequence, there is an urgent need to design new compounds with improved activity against drug-sensitive as well as drug-resistant pathogens. Cinnoline derivatives were widely studied as antimicrobial agents [1]. Cinoxacin 3 ( Figure 3) is a common drug used in urinary tract infections [19]. Since it has a high phototoxicity index, Vargas et al. synthesized the naphthyl ester of cinoxacin 4 ( Figure 3) in order to evaluate its possible application in antibacterial phototherapy. The ester derivative of cinoxacin 4 exhibited comparable photostability and antibacterial activity against E. coli to the parent drug but enhanced antibacterial activity upon irradiation [20]. Depicted in Figure 4, 6-hydroxycinnolines were synthesized and tested for in vitro antifungal activity against Candida and Aspergillus species. It was elucidated that most of the obtained compounds exhibited potent antifungal activity against C. krusei, C. neoformans, and A. niger, with the highest activity towards C.   Cinnoline derivatives bearing sulphonamide moiety 6 ( Figure 5) were synthesized as potential antimicrobial and antifungal agents. Evaluation of their activity against a panel of bacteria strains including P. aeruginosa, E. coli, B. subtilis, S. aureus and fungi C. albicans and A. niger revealed that the combination of two active moieties in one molecule resulted in significant activity improvement. Halogen substituted derivatives showed potent activity at lesser concentrations with approximately the same zone of inhibition as the reference drug [22]. Cinnoline derivatives bearing sulphonamide moiety 6 ( Figure 5) were synthesized as potential antimicrobial and antifungal agents. Evaluation of their activity against a panel of bacteria strains including P. aeruginosa, E. coli, B. subtilis, S. aureus and fungi C. albicans and A. niger revealed that the combination of two active moieties in one molecule resulted in significant activity improvement. Halogen substituted derivatives showed potent activity at lesser concentrations with approximately the same zone of inhibition as the reference drug [22]. Some new cinnoline based chalcones 7 and cinnoline based pyrazoline derivatives 8 ( Figure 6) were evaluated for their antibacterial activity against B. subtilis, E. coli, S. aureus and K. pneumoniae, antifungal activity against A. flavus, F. oxysporum, A. niger and T. viridae and insecticidal activity against Periplaneta americana. The most potent tested compounds were 4-Cl-, 2-NO2-, 4-NO2substituted cinnoline based chalcones as well as 3-Cl-, 2-NO2-and 4-OH-substituted cinnoline based pyrazolines. In addition, all chloro-substituted derivatives of series 7 and hydroxy-substituted derivatives of series 8 exhibited better insecticidal activity in comparison to the standard drug [23]. In the search for potent antibacterial and antimalarial drugs, Unnissa and co-workers synthesized pyrazole based cinnoline derivatives 9 ( Figure 7). All compounds demonstrated significant antitubercular and antifungal activity. Compound 10 4-methyl-3-[5-(4-hydroxy-3methoxyphenyl)-4,5-dihydro-1H-pyrazol-3-yl]cinnoline-6-sulphonamide ( Figure 7) was found to be the most potent with promising activity against resistant strains of M. tuberculosis and various pathogenic fungi [24], as well as against protozoan parasite P. falciparum [25]. Some new cinnoline based chalcones 7 and cinnoline based pyrazoline derivatives 8 ( Figure 6) were evaluated for their antibacterial activity against B. subtilis, E. coli, S. aureus and K. pneumoniae, antifungal activity against A. flavus, F. oxysporum, A. niger and T. viridae and insecticidal activity against Periplaneta americana. The most potent tested compounds were 4-Cl-, 2-NO 2 -, 4-NO 2 -substituted cinnoline based chalcones as well as 3-Cl-, 2-NO 2 -and 4-OH-substituted cinnoline based pyrazolines. In addition, all chloro-substituted derivatives of series 7 and hydroxy-substituted derivatives of series 8 exhibited better insecticidal activity in comparison to the standard drug [23]. Cinnoline derivatives bearing sulphonamide moiety 6 ( Figure 5) were synthesized as potential antimicrobial and antifungal agents. Evaluation of their activity against a panel of bacteria strains including P. aeruginosa, E. coli, B. subtilis, S. aureus and fungi C. albicans and A. niger revealed that the combination of two active moieties in one molecule resulted in significant activity improvement. Halogen substituted derivatives showed potent activity at lesser concentrations with approximately the same zone of inhibition as the reference drug [22]. Some new cinnoline based chalcones 7 and cinnoline based pyrazoline derivatives 8 ( Figure 6) were evaluated for their antibacterial activity against B. subtilis, E. coli, S. aureus and K. pneumoniae, antifungal activity against A. flavus, F. oxysporum, A. niger and T. viridae and insecticidal activity against Periplaneta americana. The most potent tested compounds were 4-Cl-, 2-NO2-, 4-NO2substituted cinnoline based chalcones as well as 3-Cl-, 2-NO2-and 4-OH-substituted cinnoline based pyrazolines. In addition, all chloro-substituted derivatives of series 7 and hydroxy-substituted derivatives of series 8 exhibited better insecticidal activity in comparison to the standard drug [23]. In the search for potent antibacterial and antimalarial drugs, Unnissa and co-workers synthesized pyrazole based cinnoline derivatives 9 ( Figure 7). All compounds demonstrated significant antitubercular and antifungal activity. Compound 10 4-methyl-3-[5-(4-hydroxy-3methoxyphenyl)-4,5-dihydro-1H-pyrazol-3-yl]cinnoline-6-sulphonamide ( Figure 7) was found to be the most potent with promising activity against resistant strains of M. tuberculosis and various pathogenic fungi [24], as well as against protozoan parasite P. falciparum [25]. In the search for potent antibacterial and antimalarial drugs, Unnissa and co-workers synthesized pyrazole based cinnoline derivatives 9 ( Figure 7). All compounds demonstrated significant antitubercular and antifungal activity. Compound 10 4-methyl-3-[5-(4-hydroxy-3methoxyphenyl)-4,5-dihydro-1H-pyrazol-3-yl]cinnoline-6-sulphonamide ( Figure 7) was found to be the most potent with promising activity against resistant strains of M. tuberculosis and various pathogenic fungi [24], as well as against protozoan parasite P. falciparum [25].  Saxena and co-workers obtained a series of substituted 4-(p-aminopiperazine)cinnoline-3carboxamide derivatives 13 ( Figure 9). The in vitro antimicrobial screening against G+ B. subtilis and S. aureus and G− E. coli and P. aeruginosa revealed the MIC of the synthesized compounds in the range of 12.5-50 μg/mL, whereas the zone of inhibition was between 6-29 mm. A. niger and C. albicans were used for evaluation of the antifungal activity. The MIC of the tested compounds was found to be in the range of 12.5-50 μg/mL, whereas the zone of inhibition was between 8-25 mm. The most potent antimicrobial agents in comparison to standard drugs were 6-chloro, 7-chloro and 7-bromo substituted derivatives [27]. Parasuraman et al. described 7-substituted 4-aminocinnoline-3-carboxamide derivatives that were evaluated against a panel of Gram+ and Gram− bacteria. All the synthesized compounds exhibited moderate to good antibacterial activity. The MIC (Minimal inhibitory concentration) of tested compounds against V. cholera, E. coli, B. substills, B. linctus, M. luteus, S. aureus, K. pneumoniae, Corynebacterium and S. albus was found to be in the range of 6.25-25 µg/mL. The most active compounds, 11 and 12 (Figure 8), demonstrated larger or approximately the same zone of inhibition as the reference drug ciprofloxacin. In addition, the synthesized compounds exhibited moderate to good antifungal activity against A. fumigatus, S. griseus, A. niger, A. parasitus, C. albicans and M. ruber, with the zone of inhibition between 8-27 mm. MIC values were found to be in the range of 6.25-25 µg/mL [26].  [26]. Saxena and co-workers obtained a series of substituted 4-(p-aminopiperazine)cinnoline-3carboxamide derivatives 13 ( Figure 9). The in vitro antimicrobial screening against G+ B. subtilis and S. aureus and G− E. coli and P. aeruginosa revealed the MIC of the synthesized compounds in the range of 12.5-50 μg/mL, whereas the zone of inhibition was between 6-29 mm. A. niger and C. albicans were used for evaluation of the antifungal activity. The MIC of the tested compounds was found to be in the range of 12.5-50 μg/mL, whereas the zone of inhibition was between 8-25 mm. The most potent antimicrobial agents in comparison to standard drugs were 6-chloro, 7-chloro and 7-bromo substituted derivatives [27]. Saxena and co-workers obtained a series of substituted 4-(p-aminopiperazine)cinnoline-3carboxamide derivatives 13 ( Figure 9). The in vitro antimicrobial screening against G+ B. subtilis and S. aureus and G− E. coli and P. aeruginosa revealed the MIC of the synthesized compounds in the range of 12.5-50 µg/mL, whereas the zone of inhibition was between 6-29 mm. A. niger and C. albicans were used for evaluation of the antifungal activity. The MIC of the tested compounds was found to be in the range of 12.5-50 µg/mL, whereas the zone of inhibition was between 8-25 mm. The most potent antimicrobial agents in comparison to standard drugs were 6-chloro, 7-chloro and 7-bromo substituted derivatives [27].  As a continuation of previous studies, they obtained a new series of cinnoline-3-carboxamide derivatives with five-membered (thiophene 14, furan 15, pyrazole 16, imidazole 17) or six-membered heterocycle (piperazine 18) substitutions at the 4-amino group of cinnoline core ( Figure 10). Compounds were evaluated for antibacterial, antifungal and anti-inflammatory activity. They exhibited antibacterial activity against B. subtilis, S. aureus, E. coli and P. aeruginosa. However, the potency of tested compounds differed depending on the substituent at the cinnoline nucleus. The most potent compounds in comparison to the standard drug norfloxacin were 6-chloro substituted compounds. Antifungal activity against C. albicans and A. niger was observed for all series, but the most potent antifungal agents were the 7-chloro substituted cinnoline thiophene derivative and the 6-chloro substituted cinnoline furan derivative. In all five series, halogen substituted compounds were found to be the most active, followed by methyl substituted and nitro substituted derivatives [28].  As a continuation of previous studies, they obtained a new series of cinnoline-3-carboxamide derivatives with five-membered (thiophene 14, furan 15, pyrazole 16, imidazole 17) or six-membered heterocycle (piperazine 18) substitutions at the 4-amino group of cinnoline core ( Figure 10). Compounds were evaluated for antibacterial, antifungal and anti-inflammatory activity. They exhibited antibacterial activity against B. subtilis, S. aureus, E. coli and P. aeruginosa. However, the potency of tested compounds differed depending on the substituent at the cinnoline nucleus. The most potent compounds in comparison to the standard drug norfloxacin were 6-chloro substituted compounds. Antifungal activity against C. albicans and A. niger was observed for all series, but the most potent antifungal agents were the 7-chloro substituted cinnoline thiophene derivative and the 6-chloro substituted cinnoline furan derivative. In all five series, halogen substituted compounds were found to be the most active, followed by methyl substituted and nitro substituted derivatives [28]. As a continuation of previous studies, they obtained a new series of cinnoline-3-carboxamide derivatives with five-membered (thiophene 14, furan 15, pyrazole 16, imidazole 17) or six-membered heterocycle (piperazine 18) substitutions at the 4-amino group of cinnoline core ( Figure 10). Compounds were evaluated for antibacterial, antifungal and anti-inflammatory activity. They exhibited antibacterial activity against B. subtilis, S. aureus, E. coli and P. aeruginosa. However, the potency of tested compounds differed depending on the substituent at the cinnoline nucleus. The most potent compounds in comparison to the standard drug norfloxacin were 6-chloro substituted compounds. Antifungal activity against C. albicans and A. niger was observed for all series, but the most potent antifungal agents were the 7-chloro substituted cinnoline thiophene derivative and the 6-chloro substituted cinnoline furan derivative. In all five series, halogen substituted compounds were found to be the most active, followed by methyl substituted and nitro substituted derivatives [28].   Figure 11. Cinnoline nucleoside analog acting as a siderophore biosynthesis inhibitor. The cinnoline ring system was also used in designing compounds active against tropical protozoan infections. Devine and co-workers synthesized a panel of compounds with different heterocyclic scaffolds (quinoline, isoquinoline, cinnoline, phthalazine, 3-cyanoquinoline). Cinnoline derivative 20 ( Figure 12) displayed potent proliferation inhibition for L. major and P. falciparum (Half maximal effective concentration EC50 value = 0.24 μM and 0.003 μM, respectively). In addition, the cinnoline derivative exhibited increased potency against amastigotes (0.24 μM) but with a significant decrease in potency against the promastigote form [12]. Some cinnoline derivatives 21 ( Figure 13) were patented as compounds active against resistance developing bacteria. Glinka and co-workers described the invention related to efflux pump inhibitor (EPI) compounds having polybasic functionalities. The compounds inhibited bacterial efflux pumps and could be used in combination with an antibacterial agent to treat or prevent bacterial infections. [30]. Figure 13. Example of a cinnoline derivative with polybasic functionalities patented as an efflux pump inhibitor. Figure 11. Cinnoline nucleoside analog acting as a siderophore biosynthesis inhibitor. The cinnoline ring system was also used in designing compounds active against tropical protozoan infections. Devine and co-workers synthesized a panel of compounds with different heterocyclic scaffolds (quinoline, isoquinoline, cinnoline, phthalazine, 3-cyanoquinoline). Cinnoline derivative 20 ( Figure 12) displayed potent proliferation inhibition for L. major and P. falciparum (Half maximal effective concentration EC 50 value = 0.24 µM and 0.003 µM, respectively). In addition, the cinnoline derivative exhibited increased potency against amastigotes (0.24 µM) but with a significant decrease in potency against the promastigote form [12].  Figure 11. Cinnoline nucleoside analog acting as a siderophore biosynthesis inhibitor. The cinnoline ring system was also used in designing compounds active against tropical protozoan infections. Devine and co-workers synthesized a panel of compounds with different heterocyclic scaffolds (quinoline, isoquinoline, cinnoline, phthalazine, 3-cyanoquinoline). Cinnoline derivative 20 ( Figure 12) displayed potent proliferation inhibition for L. major and P. falciparum (Half maximal effective concentration EC50 value = 0.24 μM and 0.003 μM, respectively). In addition, the cinnoline derivative exhibited increased potency against amastigotes (0.24 μM) but with a significant decrease in potency against the promastigote form [12]. Some cinnoline derivatives 21 ( Figure 13) were patented as compounds active against resistance developing bacteria. Glinka and co-workers described the invention related to efflux pump inhibitor (EPI) compounds having polybasic functionalities. The compounds inhibited bacterial efflux pumps and could be used in combination with an antibacterial agent to treat or prevent bacterial infections. [30].  Some cinnoline derivatives 21 ( Figure 13) were patented as compounds active against resistance developing bacteria. Glinka and co-workers described the invention related to efflux pump inhibitor (EPI) compounds having polybasic functionalities. The compounds inhibited bacterial efflux pumps and could be used in combination with an antibacterial agent to treat or prevent bacterial infections. [30].  Figure 11. Cinnoline nucleoside analog acting as a siderophore biosynthesis inhibitor. The cinnoline ring system was also used in designing compounds active against tropical protozoan infections. Devine and co-workers synthesized a panel of compounds with different heterocyclic scaffolds (quinoline, isoquinoline, cinnoline, phthalazine, 3-cyanoquinoline). Cinnoline derivative 20 ( Figure 12) displayed potent proliferation inhibition for L. major and P. falciparum (Half maximal effective concentration EC50 value = 0.24 μM and 0.003 μM, respectively). In addition, the cinnoline derivative exhibited increased potency against amastigotes (0.24 μM) but with a significant decrease in potency against the promastigote form [12]. Some cinnoline derivatives 21 ( Figure 13) were patented as compounds active against resistance developing bacteria. Glinka and co-workers described the invention related to efflux pump inhibitor (EPI) compounds having polybasic functionalities. The compounds inhibited bacterial efflux pumps and could be used in combination with an antibacterial agent to treat or prevent bacterial infections. [30].

Analgesic and Antiinflamatory Activities
In an effort to find dual acting compounds, Chaudhary et al. designed a series of cinnoline derivatives with pyrazoline 23 or without a pyrazoline nucleus 22 ( Figure 14) as anti-inflammatory and antibacterial agents. It has been shown that cinnolines bearing pyrazoline ring 23 ( Figure 14) and having electron donating functional groups at the phenyl moiety (methoxyl and hydroxyl) exhibited the highest anti-inflammatory activity. In case of antibacterial activity, an electron withdrawing substituent at the phenyl group of cinnoline derivatives without pyrazoline ring 22 (Figure 14), as well as hydroxyl substitution of the phenyl ring of cinnoline derivatives with a pyrazoline moiety, were associated with increased activity against G+ (S. aureus, B. subtilis) and G− bacteria (E. coli) [31].

Analgesic and Antiinflamatory Activities
In an effort to find dual acting compounds, Chaudhary et al. designed a series of cinnoline derivatives with pyrazoline 23 or without a pyrazoline nucleus 22 ( Figure 14) as anti-inflammatory and antibacterial agents. It has been shown that cinnolines bearing pyrazoline ring 23 ( Figure 14) and having electron donating functional groups at the phenyl moiety (methoxyl and hydroxyl) exhibited the highest anti-inflammatory activity. In case of antibacterial activity, an electron withdrawing substituent at the phenyl group of cinnoline derivatives without pyrazoline ring 22 (Figure 14), as well as hydroxyl substitution of the phenyl ring of cinnoline derivatives with a pyrazoline moiety, were associated with increased activity against G+ (S. aureus, B. subtilis) and G− bacteria (E. coli) [31]. A series of dual acting pyrazolo [4,3-c]cinnoline derivatives were also obtained by Tonk and coworkers. It was elucidated that compounds with an electron donating group in the benzoyl ring exhibited higher anti-inflammatory activity than compounds with a benzoyl ring substituted by electron withdrawing groups. Moreover, a methylene spacer between the phenyl group and the carbonyl carbon increased anti-inflammatory activity, whereas the O-CH2 group caused a considerable decrease in activity. Compounds that exhibited excellent protection against inflammation 24 and 25, depicted in Figure 15, also showed a strong cyclooxygenase-2 (COX-2) binding profile. They were considered safer in terms of gastric ulcerogenicity and lipid peroxidation activity than the standard drug naproxen. In case of antibacterial activity, compounds with a 4-nitro-(26) or 2,4-dichloro (27) substituent at the benzoyl group exhibited significant activity against G− (E. coli and P. aeruginosa) and G+ (S. aureus) bacterial strains. However, compounds with an unsubstituted phenyl ring and methylene spacer 28 ( Figure 15) were found to be the best dual antiinflammatory and antibacterial agent (with significant activity against all three strains) [32]. Cinnoline derivatives have been also reported as phosphodiesterase 4 (PDE4) inhibitors [33]. PDE4 is the predominant isoenzyme in almost all immune and inflammatory cells and is an important regulator of cyclic adenosine monophosphate (cAMP) content in airway smooth muscle. Inhibition of PDE4 leads to bronchodilation and the reduction in the production of inflammatory mediators such as tumor necrosis factor (TNF-α) by cAMP down regulation. A PDE4 inhibitor could be used as a potential anti-inflammatory agent in chronic obstructive pulmonary disease (COPD), asthma, rhinitis and rheumatoid arthritis [34]. Structurally related to quinoline PDE4 inhibitors, 3amido-4-anilinocinnoline 29 has been designed by Lunniss et al. in order to overcome the poor A series of dual acting pyrazolo [4,3-c]cinnoline derivatives were also obtained by Tonk and co-workers. It was elucidated that compounds with an electron donating group in the benzoyl ring exhibited higher anti-inflammatory activity than compounds with a benzoyl ring substituted by electron withdrawing groups. Moreover, a methylene spacer between the phenyl group and the carbonyl carbon increased anti-inflammatory activity, whereas the O-CH 2 group caused a considerable decrease in activity. Compounds that exhibited excellent protection against inflammation 24 and 25, depicted in Figure 15, also showed a strong cyclooxygenase-2 (COX-2) binding profile. They were considered safer in terms of gastric ulcerogenicity and lipid peroxidation activity than the standard drug naproxen. In case of antibacterial activity, compounds with a 4-nitro-(26) or 2,4-dichloro (27) substituent at the benzoyl group exhibited significant activity against G− (E. coli and P. aeruginosa) and G+ (S. aureus) bacterial strains. However, compounds with an unsubstituted phenyl ring and methylene spacer 28 ( Figure 15) were found to be the best dual anti-inflammatory and antibacterial agent (with significant activity against all three strains) [32].

Analgesic and Antiinflamatory Activities
In an effort to find dual acting compounds, Chaudhary et al. designed a series of cinnoline derivatives with pyrazoline 23 or without a pyrazoline nucleus 22 ( Figure 14) as anti-inflammatory and antibacterial agents. It has been shown that cinnolines bearing pyrazoline ring 23 ( Figure 14) and having electron donating functional groups at the phenyl moiety (methoxyl and hydroxyl) exhibited the highest anti-inflammatory activity. In case of antibacterial activity, an electron withdrawing substituent at the phenyl group of cinnoline derivatives without pyrazoline ring 22 (Figure 14), as well as hydroxyl substitution of the phenyl ring of cinnoline derivatives with a pyrazoline moiety, were associated with increased activity against G+ (S. aureus, B. subtilis) and G− bacteria (E. coli) [31]. A series of dual acting pyrazolo [4,3-c]cinnoline derivatives were also obtained by Tonk and coworkers. It was elucidated that compounds with an electron donating group in the benzoyl ring exhibited higher anti-inflammatory activity than compounds with a benzoyl ring substituted by electron withdrawing groups. Moreover, a methylene spacer between the phenyl group and the carbonyl carbon increased anti-inflammatory activity, whereas the O-CH2 group caused a considerable decrease in activity. Compounds that exhibited excellent protection against inflammation 24 and 25, depicted in Figure 15, also showed a strong cyclooxygenase-2 (COX-2) binding profile. They were considered safer in terms of gastric ulcerogenicity and lipid peroxidation activity than the standard drug naproxen. In case of antibacterial activity, compounds with a 4-nitro-(26) or 2,4-dichloro (27) substituent at the benzoyl group exhibited significant activity against G− (E. coli and P. aeruginosa) and G+ (S. aureus) bacterial strains. However, compounds with an unsubstituted phenyl ring and methylene spacer 28 ( Figure 15) were found to be the best dual antiinflammatory and antibacterial agent (with significant activity against all three strains) [32]. Cinnoline derivatives have been also reported as phosphodiesterase 4 (PDE4) inhibitors [33]. PDE4 is the predominant isoenzyme in almost all immune and inflammatory cells and is an important regulator of cyclic adenosine monophosphate (cAMP) content in airway smooth muscle. Inhibition of PDE4 leads to bronchodilation and the reduction in the production of inflammatory mediators such as tumor necrosis factor (TNF-α) by cAMP down regulation. A PDE4 inhibitor could be used as a potential anti-inflammatory agent in chronic obstructive pulmonary disease (COPD), asthma, rhinitis and rheumatoid arthritis [34]. Structurally related to quinoline PDE4 inhibitors, 3amido-4-anilinocinnoline 29 has been designed by Lunniss et al. in order to overcome the poor Cinnoline derivatives have been also reported as phosphodiesterase 4 (PDE4) inhibitors [33]. PDE4 is the predominant isoenzyme in almost all immune and inflammatory cells and is an important regulator of cyclic adenosine monophosphate (cAMP) content in airway smooth muscle. Inhibition of PDE4 leads to bronchodilation and the reduction in the production of inflammatory mediators such as tumor necrosis factor (TNF-α) by cAMP down regulation. A PDE4 inhibitor could be used as a potential anti-inflammatory agent in chronic obstructive pulmonary disease (COPD), asthma, rhinitis and rheumatoid arthritis [34]. Structurally related to quinoline PDE4 inhibitors, 3-amido-4-anilinocinnoline 29 has been designed by Lunniss et al. in order to overcome the poor pharmacokinetic profile in the cynomolgus monkey [33]. Compound 29 ( Figure 16) retained excellent in vitro potency and >100-fold selectivity versus other PDE isoenzymes with improved pharmacokinetics in the monkey in comparison to the quinoline analog [33]. pharmacokinetic profile in the cynomolgus monkey [33]. Compound 29 ( Figure 16) retained excellent in vitro potency and >100-fold selectivity versus other PDE isoenzymes with improved pharmacokinetics in the monkey in comparison to the quinoline analog [33]. Vanilloid receptor subtype VR1 (TRPV1) present in various brain regions, the spinal cord, peripheral sensory neurons and non-neuronal tissues is considered as a new target for pain management but all natural vanilloid receptor agonists such as capsaicin cause an initial burning effect. TRPV1 competitive antagonists, which lack excitatory effects, were designed and evaluated in vivo in animal pain models. Urea derivative bearing cinnoline group 30 ( Figure 17) was synthesized among other compounds with various bicyclic heteroaromatic pharmacophores as novel potential analgesics acting through the TRPV1 receptor antagonism ( [35] and references therein). Since Bruton's tyrosine kinase (BTK) is a kinase implicated in autoimmune disorders, BTK inhibition is considered as an attractive approach for the treatment of autoimmune diseases such as rheumatoid arthritis [36]. In 2013, scientists from Takeda Pharmaceutical Company Ltd. patented cinnoline derivatives of general formula 31 depicted in Figure 18 as BTK inhibitors [37]. In addition, the discovery of a series of 4-aminocinnoline-3-carboxamides that exhibited BTK inhibition were reported by Smith et al. A fragment-based screening approach incorporating X-ray cocrystallography was used to identify a cinnoline fragment and characterize its binding mode. Optimization of the fragment hit resulted in the identification of compound 32 (Figure 18), an orally absorbed, noncovalent BTK inhibitor reducing paw swelling in a dose-and exposure-dependent fashion in a rat model of collagen-induced arthritis [38]. Vanilloid receptor subtype VR1 (TRPV1) present in various brain regions, the spinal cord, peripheral sensory neurons and non-neuronal tissues is considered as a new target for pain management but all natural vanilloid receptor agonists such as capsaicin cause an initial burning effect. TRPV1 competitive antagonists, which lack excitatory effects, were designed and evaluated in vivo in animal pain models. Urea derivative bearing cinnoline group 30 ( Figure 17) was synthesized among other compounds with various bicyclic heteroaromatic pharmacophores as novel potential analgesics acting through the TRPV1 receptor antagonism ( [35] and references therein). pharmacokinetic profile in the cynomolgus monkey [33]. Compound 29 ( Figure 16) retained excellent in vitro potency and >100-fold selectivity versus other PDE isoenzymes with improved pharmacokinetics in the monkey in comparison to the quinoline analog [33]. Vanilloid receptor subtype VR1 (TRPV1) present in various brain regions, the spinal cord, peripheral sensory neurons and non-neuronal tissues is considered as a new target for pain management but all natural vanilloid receptor agonists such as capsaicin cause an initial burning effect. TRPV1 competitive antagonists, which lack excitatory effects, were designed and evaluated in vivo in animal pain models. Urea derivative bearing cinnoline group 30 ( Figure 17) was synthesized among other compounds with various bicyclic heteroaromatic pharmacophores as novel potential analgesics acting through the TRPV1 receptor antagonism ( [35] and references therein). Since Bruton's tyrosine kinase (BTK) is a kinase implicated in autoimmune disorders, BTK inhibition is considered as an attractive approach for the treatment of autoimmune diseases such as rheumatoid arthritis [36]. In 2013, scientists from Takeda Pharmaceutical Company Ltd. patented cinnoline derivatives of general formula 31 depicted in Figure 18 as BTK inhibitors [37]. In addition, the discovery of a series of 4-aminocinnoline-3-carboxamides that exhibited BTK inhibition were reported by Smith et al. A fragment-based screening approach incorporating X-ray cocrystallography was used to identify a cinnoline fragment and characterize its binding mode. Optimization of the fragment hit resulted in the identification of compound 32 (Figure 18), an orally absorbed, noncovalent BTK inhibitor reducing paw swelling in a dose-and exposure-dependent fashion in a rat model of collagen-induced arthritis [38]. Since Bruton's tyrosine kinase (BTK) is a kinase implicated in autoimmune disorders, BTK inhibition is considered as an attractive approach for the treatment of autoimmune diseases such as rheumatoid arthritis [36]. In 2013, scientists from Takeda Pharmaceutical Company Ltd. patented cinnoline derivatives of general formula 31 depicted in Figure 18 as BTK inhibitors [37]. In addition, the discovery of a series of 4-aminocinnoline-3-carboxamides that exhibited BTK inhibition were reported by Smith et al. A fragment-based screening approach incorporating X-ray co-crystallography was used to identify a cinnoline fragment and characterize its binding mode. Optimization of the fragment hit resulted in the identification of compound 32 (Figure 18), an orally absorbed, noncovalent BTK inhibitor reducing paw swelling in a dose-and exposure-dependent fashion in a rat model of collagen-induced arthritis [38]. Cinnoline derivatives were also evaluated as human neutrophil elastase (HNE) inhibitors. Excessive HNE activity is connected with many inflammatory disorders and compounds which are able to inhibit the proteolytic activity of HNE represent promising therapeutic agents for the treatment of diseases involving its excessive activity. Potential HNE inhibitors bearing cinnoline scaffolds were designed by transformation of indazole into the cinnoline by enlargement of the pyrazole ring of the N-benzoylindazoles reported earlier [39,40]. Studies revealed that although cinnoline derivatives (33 and 34 were the most potent) ( Figure 19) were reversible competitive inhibitors of HNE with increased stability in aqueous solution, they exhibited lower potency in comparison to N-benzoylindazoles ( [41] and references therein). A cinnoline fused Mannich base with a large hydrophobic diphenyl substituent at amino group 35 ( Figure 20) exhibited higher analgesic activity when compared to diclofenac at 120 min and 180 min. In addition, its dose level (50 mg/kg) resulted in similar anti-inflammatory activity in comparison to celecoxib (20 mg/kg). What is more, compound 35 as well as 36 (with a dicyclohexane moiety) (Figure 20) also exhibited antibacterial activity with a larger zone of inhibition when compared to streptomycin in S. aureus and E. coli, respectively [42]. Cinnoline derivatives were also evaluated as human neutrophil elastase (HNE) inhibitors. Excessive HNE activity is connected with many inflammatory disorders and compounds which are able to inhibit the proteolytic activity of HNE represent promising therapeutic agents for the treatment of diseases involving its excessive activity. Potential HNE inhibitors bearing cinnoline scaffolds were designed by transformation of indazole into the cinnoline by enlargement of the pyrazole ring of the N-benzoylindazoles reported earlier [39,40]. Studies revealed that although cinnoline derivatives (33 and 34 were the most potent) ( Figure 19) were reversible competitive inhibitors of HNE with increased stability in aqueous solution, they exhibited lower potency in comparison to N-benzoylindazoles ( [41] and references therein). Cinnoline derivatives were also evaluated as human neutrophil elastase (HNE) inhibitors. Excessive HNE activity is connected with many inflammatory disorders and compounds which are able to inhibit the proteolytic activity of HNE represent promising therapeutic agents for the treatment of diseases involving its excessive activity. Potential HNE inhibitors bearing cinnoline scaffolds were designed by transformation of indazole into the cinnoline by enlargement of the pyrazole ring of the N-benzoylindazoles reported earlier [39,40]. Studies revealed that although cinnoline derivatives (33 and 34 were the most potent) (Figure 19) were reversible competitive inhibitors of HNE with increased stability in aqueous solution, they exhibited lower potency in comparison to N-benzoylindazoles ( [41] and references therein). A cinnoline fused Mannich base with a large hydrophobic diphenyl substituent at amino group 35 ( Figure 20) exhibited higher analgesic activity when compared to diclofenac at 120 min and 180 min. In addition, its dose level (50 mg/kg) resulted in similar anti-inflammatory activity in comparison to celecoxib (20 mg/kg). What is more, compound 35 as well as 36 (with a dicyclohexane moiety) (Figure 20) also exhibited antibacterial activity with a larger zone of inhibition when compared to streptomycin in S. aureus and E. coli, respectively [42]. Cinnoline derivatives were also evaluated as human neutrophil elastase (HNE) inhibitors. Excessive HNE activity is connected with many inflammatory disorders and compounds which are able to inhibit the proteolytic activity of HNE represent promising therapeutic agents for the treatment of diseases involving its excessive activity. Potential HNE inhibitors bearing cinnoline scaffolds were designed by transformation of indazole into the cinnoline by enlargement of the pyrazole ring of the N-benzoylindazoles reported earlier [39,40]. Studies revealed that although cinnoline derivatives (33 and 34 were the most potent) (Figure 19) were reversible competitive inhibitors of HNE with increased stability in aqueous solution, they exhibited lower potency in comparison to N-benzoylindazoles ( [41] and references therein). A cinnoline fused Mannich base with a large hydrophobic diphenyl substituent at amino group 35 ( Figure 20) exhibited higher analgesic activity when compared to diclofenac at 120 min and 180 min. In addition, its dose level (50 mg/kg) resulted in similar anti-inflammatory activity in comparison to celecoxib (20 mg/kg). What is more, compound 35 as well as 36 (with a dicyclohexane moiety) (Figure 20) also exhibited antibacterial activity with a larger zone of inhibition when compared to streptomycin in S. aureus and E. coli, respectively [42].

Potential for Neurological Disorders
Compounds bearing a cinnoline nucleus fused with various heterocyclic scaffolds were also designed as potential therapeutic agents aiming at treating many neurological and psychiatric disorders e.g., Huntington's [43] or Alzheimer's disease [44].
Amer et al. synthesized dibenzopyrazolocinnolines and evaluated their antiparkinsonian activity. The pharmacological screening revealed that the most active compounds 37 and 38, depicted in Figure 21, exhibited antiparkinsonian activity comparable to benzatropine [45].

Potential for Neurological Disorders
Compounds bearing a cinnoline nucleus fused with various heterocyclic scaffolds were also designed as potential therapeutic agents aiming at treating many neurological and psychiatric disorders e.g., Huntington's [43] or Alzheimer's disease [44].
Amer et al. synthesized dibenzopyrazolocinnolines and evaluated their antiparkinsonian activity. The pharmacological screening revealed that the most active compounds 37 and 38, depicted in Figure 21, exhibited antiparkinsonian activity comparable to benzatropine [45]. Mutations in the leucine-rich repeat kinase 2 (LRRK2) protein have been associated with Parkinson's disease. Inhibition of LRRK2 kinase activity by a selective small-molecule inhibitor has been proposed as a potential treatment for this disease [46]. Scientists from Elan Pharmaceuticals worked on a series of cinnoline LRRK2 small-molecule inhibitors identified from a kinase-focused high throughput screening (HTS) of an in-house library [47]. In addition, Garofalo et al. reported 4aminocinnoline-3-carboxamide derivatives 39, 40 ( Figure 22) potent against both wild-type and mutant LRRK2 kinase activity in biochemical and cellular assays. In addition, these compounds exhibited excellent central nervous system penetration. Unfortunately, due to disappointing kinase specificity, they were no longer studied [48]. The phosphodiesterase 10A (PDE10A) enzyme is involved in cellular signaling pathways in schizophrenia. As a consequence, inhibitors of PDE10A offer a promising therapeutic approach for the treatment or prevention of psychiatric disorders, especially schizophrenia and related diseases [49].  Mutations in the leucine-rich repeat kinase 2 (LRRK2) protein have been associated with Parkinson's disease. Inhibition of LRRK2 kinase activity by a selective small-molecule inhibitor has been proposed as a potential treatment for this disease [46]. Scientists from Elan Pharmaceuticals worked on a series of cinnoline LRRK2 small-molecule inhibitors identified from a kinase-focused high throughput screening (HTS) of an in-house library [47]. In addition, Garofalo et al. reported 4-aminocinnoline-3-carboxamide derivatives 39, 40 ( Figure 22) potent against both wild-type and mutant LRRK2 kinase activity in biochemical and cellular assays. In addition, these compounds exhibited excellent central nervous system penetration. Unfortunately, due to disappointing kinase specificity, they were no longer studied [48].

Potential for Neurological Disorders
Compounds bearing a cinnoline nucleus fused with various heterocyclic scaffolds were also designed as potential therapeutic agents aiming at treating many neurological and psychiatric disorders e.g., Huntington's [43] or Alzheimer's disease [44].
Amer et al. synthesized dibenzopyrazolocinnolines and evaluated their antiparkinsonian activity. The pharmacological screening revealed that the most active compounds 37 and 38, depicted in Figure 21, exhibited antiparkinsonian activity comparable to benzatropine [45]. Mutations in the leucine-rich repeat kinase 2 (LRRK2) protein have been associated with Parkinson's disease. Inhibition of LRRK2 kinase activity by a selective small-molecule inhibitor has been proposed as a potential treatment for this disease [46]. Scientists from Elan Pharmaceuticals worked on a series of cinnoline LRRK2 small-molecule inhibitors identified from a kinase-focused high throughput screening (HTS) of an in-house library [47]. In addition, Garofalo et al. reported 4aminocinnoline-3-carboxamide derivatives 39, 40 ( Figure 22) potent against both wild-type and mutant LRRK2 kinase activity in biochemical and cellular assays. In addition, these compounds exhibited excellent central nervous system penetration. Unfortunately, due to disappointing kinase specificity, they were no longer studied [48]. The phosphodiesterase 10A (PDE10A) enzyme is involved in cellular signaling pathways in schizophrenia. As a consequence, inhibitors of PDE10A offer a promising therapeutic approach for the treatment or prevention of psychiatric disorders, especially schizophrenia and related diseases [49].  The phosphodiesterase 10A (PDE10A) enzyme is involved in cellular signaling pathways in schizophrenia. As a consequence, inhibitors of PDE10A offer a promising therapeutic approach for the treatment or prevention of psychiatric disorders, especially schizophrenia and related diseases [49]. Since some 6,7-dimethoxy-4-(pyridine-3-yl)cinnolines also exhibited PDE3 activity (a risk of offtarget effects), optimization of structure 43 ( Figure 24) led to the discovery of compounds 44 and 45 ( Figure 24) with significantly improved selectivity against PDE3 but maintaining their PDE10A inhibitory activity and in vivo metabolic stability comparable to 43 ( Figure 24) [52]. Recently, Geneste et al. described the optimization of HTS hit structure 46 ( Figure 25) supported by X-ray crystal structure analysis and molecular modeling which gave 3H-pyrazolo[3,4-c]cinnolines 47 and 48 (Figure 25), which are potent, selective and brain-penetrant PDE10A inhibitors with an improved pharmacokinetic profile in rats [53]. Preparation of 3H-pyrazolo[3,4-c]cinnoline derivatives as PDE10A inhibitors was also the subject of patent WO2014/027078 [54]. Since some 6,7-dimethoxy-4-(pyridine-3-yl)cinnolines also exhibited PDE3 activity (a risk of off-target effects), optimization of structure 43 ( Figure 24) led to the discovery of compounds 44 and 45 ( Figure 24) with significantly improved selectivity against PDE3 but maintaining their PDE10A inhibitory activity and in vivo metabolic stability comparable to 43 ( Figure 24) [52]. Since some 6,7-dimethoxy-4-(pyridine-3-yl)cinnolines also exhibited PDE3 activity (a risk of offtarget effects), optimization of structure 43 ( Figure 24) led to the discovery of compounds 44 and 45 ( Figure 24) with significantly improved selectivity against PDE3 but maintaining their PDE10A inhibitory activity and in vivo metabolic stability comparable to 43 ( Figure 24) [52]. Recently, Geneste et al. described the optimization of HTS hit structure 46 ( Figure 25) supported by X-ray crystal structure analysis and molecular modeling which gave 3H-pyrazolo[3,4-c]cinnolines 47 and 48 (Figure 25), which are potent, selective and brain-penetrant PDE10A inhibitors with an improved pharmacokinetic profile in rats [53]. Preparation of 3H-pyrazolo[3,4-c]cinnoline derivatives as PDE10A inhibitors was also the subject of patent WO2014/027078 [54].  (Figure 25), which are potent, selective and brain-penetrant PDE10A inhibitors with an improved pharmacokinetic profile in rats [53]. Preparation of 3H-pyrazolo [3,4-c]cinnoline derivatives as PDE10A inhibitors was also the subject of patent WO2014/027078 [54]. The cinnoline scaffold turned out to be a useful building block in designing compounds targeting histamine receptor H3. Involvement of the H3 receptor subtype in the presynaptic regulation of the release of various neurotransmitters in the central nervous system makes it an attractive target for treating diseases such as attention-deficit hyperactivity disorder, Alzheimer's disease, mild cognitive impairment and schizophrenia. Josef and co-workers obtained compounds with the tricyclic benzocinnolinone pyridazinone core as analogues of irdabisant. The compounds 2Hbenzo[h]cinnolin-3-ones and 3H-benzo[f]cinnolin-2-ones exhibited high H3 receptor binding affinity with excellent selectivity against the H1R, H2R and H4R subtypes of histamine receptor. Modification to the linker/amine region of the pharmacophore resulted in ±49 as a mixture of diastereoisomers (Figure 26), which showed improved metabolic stability and rat pharmacokinetics following oral administration ( [55] and references therein). Cinnoline derivatives are also enumerated among non-benzodiazepine modulators of γaminobutyric acid receptor A (GABA A) [56]. Astra Zeneca works on an orally bioavailable positive modulator of the GABA A α2 and α3 subunits and developed novel compound 50 depicted in Figure  27 as a possible treatment or prophylaxis of anxiety disorders, cognitive disorders, and/or mood disorders [57]. AZD7325 (51) and AZD6280 (52) depicted in Figure 27 were identified as positive modulators at α2/α3 and negative modulators at α5 GABA A receptors and exhibited a potent anxiolytic-like effect without sedation or cognitive impairment [13,58]. These compounds have undergone clinical trial phase I [59,60]. Moreover, AZD7325 was studied in two phase II proof-ofconcept trials in patients with general anxiety disorders (NCT 00807937 and NCT00808249) as well as in a phase II proof-of-mechanism in patients with autism spectrum disorders (NCT01966679). In addition, the diverse metabolite profile of AZD7325 was investigated [61]. In vivo studies in rat and in vitro studies in human, rat, mouse, rabbit and dog liver microsomes were performed with radiolabeled AZD7325, revealing approximately 40 metabolites [61,62]. The cinnoline scaffold turned out to be a useful building block in designing compounds targeting histamine receptor H 3 . Involvement of the H 3 receptor subtype in the presynaptic regulation of the release of various neurotransmitters in the central nervous system makes it an attractive target for treating diseases such as attention-deficit hyperactivity disorder, Alzheimer's disease, mild cognitive impairment and schizophrenia. Josef and co-workers obtained compounds with the tricyclic benzocinnolinone pyridazinone core as analogues of irdabisant. The compounds 2H-benzo[h]cinnolin-3-ones and 3H-benzo[f ]cinnolin-2-ones exhibited high H 3 receptor binding affinity with excellent selectivity against the H 1 R, H 2 R and H 4 R subtypes of histamine receptor. Modification to the linker/amine region of the pharmacophore resulted in ±49 as a mixture of diastereoisomers (Figure 26), which showed improved metabolic stability and rat pharmacokinetics following oral administration ( [55] and references therein). The cinnoline scaffold turned out to be a useful building block in designing compounds targeting histamine receptor H3. Involvement of the H3 receptor subtype in the presynaptic regulation of the release of various neurotransmitters in the central nervous system makes it an attractive target for treating diseases such as attention-deficit hyperactivity disorder, Alzheimer's disease, mild cognitive impairment and schizophrenia. Josef and co-workers obtained compounds with the tricyclic benzocinnolinone pyridazinone core as analogues of irdabisant. The compounds 2Hbenzo[h]cinnolin-3-ones and 3H-benzo[f]cinnolin-2-ones exhibited high H3 receptor binding affinity with excellent selectivity against the H1R, H2R and H4R subtypes of histamine receptor. Modification to the linker/amine region of the pharmacophore resulted in ±49 as a mixture of diastereoisomers (Figure 26), which showed improved metabolic stability and rat pharmacokinetics following oral administration ( [55] and references therein). Cinnoline derivatives are also enumerated among non-benzodiazepine modulators of γaminobutyric acid receptor A (GABA A) [56]. Astra Zeneca works on an orally bioavailable positive modulator of the GABA A α2 and α3 subunits and developed novel compound 50 depicted in Figure  27 as a possible treatment or prophylaxis of anxiety disorders, cognitive disorders, and/or mood disorders [57]. AZD7325 (51) and AZD6280 (52) depicted in Figure 27 were identified as positive modulators at α2/α3 and negative modulators at α5 GABA A receptors and exhibited a potent anxiolytic-like effect without sedation or cognitive impairment [13,58]. These compounds have undergone clinical trial phase I [59,60]. Moreover, AZD7325 was studied in two phase II proof-ofconcept trials in patients with general anxiety disorders (NCT 00807937 and NCT00808249) as well as in a phase II proof-of-mechanism in patients with autism spectrum disorders (NCT01966679). In addition, the diverse metabolite profile of AZD7325 was investigated [61]. In vivo studies in rat and in vitro studies in human, rat, mouse, rabbit and dog liver microsomes were performed with radiolabeled AZD7325, revealing approximately 40 metabolites [61,62]. Cinnoline derivatives are also enumerated among non-benzodiazepine modulators of γ-aminobutyric acid receptor A (GABA A) [56]. Astra Zeneca works on an orally bioavailable positive modulator of the GABA A α2 and α3 subunits and developed novel compound 50 depicted in Figure 27 as a possible treatment or prophylaxis of anxiety disorders, cognitive disorders, and/or mood disorders [57]. AZD7325 (51) and AZD6280 (52) depicted in Figure 27 were identified as positive modulators at α2/α3 and negative modulators at α5 GABA A receptors and exhibited a potent anxiolytic-like effect without sedation or cognitive impairment [13,58]. These compounds have undergone clinical trial phase I [59,60]. Moreover, AZD7325 was studied in two phase II proof-of-concept trials in patients with general anxiety disorders (NCT 00807937 and NCT00808249) as well as in a phase II proof-of-mechanism in patients with autism spectrum disorders (NCT01966679). In addition, the diverse metabolite profile of AZD7325 was investigated [61]. In vivo studies in rat and in vitro studies in human, rat, mouse, rabbit and dog liver microsomes were performed with radiolabeled AZD7325, revealing approximately 40 metabolites [61,62]. Cinnolinones (53, 54) ( Figure 28) as diaza analogues of known aminobutyrophenones were designed as potential atypical psychotics. Determination of the binding affinities towards the serotonin receptors 5-HT2A and 5-HT2C, and the dopamine D2 receptors revealed that these compounds lacked appreciable affinity for the dopamine D2 receptors, and as a consequence, they were not suited as potential psychotics. However, they displayed the highest affinity for the 5-HT2C receptor [63].

Anticancer Properties
Cinnoline derivatives were also designed as potential anticancer drugs. Extensive studies have been performed to assess the topoisomerase 1-targeting (TOP1-targeting) activity and cytotoxicity of substituted dibenzo[c,h]cinnolines 55, 56 ( Figure 29) as non-CPT (camptothecin) TOP1 inhibitors. Structure-activity relationship (SAR) studies of dibenzo[c,h]cinnolines revealed that removal of the methylenedioxy group on the D ring or its replacement by other substituents (methoxy-, benzyloxyor hydroxy-groups) resulted in a substantial loss of TOP1-targeting activity. The presence of 2,3dimethoxy substituents in ring A was also determined as a crucial structural element for retaining TOP1 activity and cytotoxicity. Although the substituted dibenzo[c,h]cinnolines with significant TOP1-targeting activity exhibited cross-resistance in camptothecin-resistant cell lines, their cytotoxicity was not diminished in cells overexpressing multidrug resistance protein 1 MDR1 [64]. Cinnolinones (53, 54) ( Figure 28) as diaza analogues of known aminobutyrophenones were designed as potential atypical psychotics. Determination of the binding affinities towards the serotonin receptors 5-HT 2A and 5-HT 2C , and the dopamine D 2 receptors revealed that these compounds lacked appreciable affinity for the dopamine D 2 receptors, and as a consequence, they were not suited as potential psychotics. However, they displayed the highest affinity for the 5-HT 2C receptor [63]. Cinnolinones (53, 54) ( Figure 28) as diaza analogues of known aminobutyrophenones were designed as potential atypical psychotics. Determination of the binding affinities towards the serotonin receptors 5-HT2A and 5-HT2C, and the dopamine D2 receptors revealed that these compounds lacked appreciable affinity for the dopamine D2 receptors, and as a consequence, they were not suited as potential psychotics. However, they displayed the highest affinity for the 5-HT2C receptor [63].

Anticancer Properties
Cinnoline derivatives were also designed as potential anticancer drugs. Extensive studies have been performed to assess the topoisomerase 1-targeting (TOP1-targeting) activity and cytotoxicity of substituted dibenzo[c,h]cinnolines 55, 56 ( Figure 29) as non-CPT (camptothecin) TOP1 inhibitors. Structure-activity relationship (SAR) studies of dibenzo[c,h]cinnolines revealed that removal of the methylenedioxy group on the D ring or its replacement by other substituents (methoxy-, benzyloxyor hydroxy-groups) resulted in a substantial loss of TOP1-targeting activity. The presence of 2,3dimethoxy substituents in ring A was also determined as a crucial structural element for retaining TOP1 activity and cytotoxicity. Although the substituted dibenzo[c,h]cinnolines with significant TOP1-targeting activity exhibited cross-resistance in camptothecin-resistant cell lines, their cytotoxicity was not diminished in cells overexpressing multidrug resistance protein 1 MDR1 [64].

Anticancer Properties
Cinnoline derivatives were also designed as potential anticancer drugs. Extensive studies have been performed to assess the topoisomerase 1-targeting (TOP1-targeting) activity and cytotoxicity of substituted dibenzo[c,h]cinnolines 55, 56 ( Figure 29) as non-CPT (camptothecin) TOP1 inhibitors. Structure-activity relationship (SAR) studies of dibenzo[c,h]cinnolines revealed that removal of the methylenedioxy group on the D ring or its replacement by other substituents (methoxy-, benzyloxy-or hydroxy-groups) resulted in a substantial loss of TOP1-targeting activity. The presence of 2,3-dimethoxy substituents in ring A was also determined as a crucial structural element for retaining TOP1 activity and cytotoxicity. Although the substituted dibenzo[c,h]cinnolines with significant TOP1-targeting activity exhibited cross-resistance in camptothecin-resistant cell lines, their cytotoxicity was not diminished in cells overexpressing multidrug resistance protein 1 MDR1 [64].  As a continuation of studies on the most potent derivative 57, 5,6,11-triazachrysen-12-ones with various substituents at 11-position were synthesized [65]. Compound 58 (ARC-31, Figure 30) exhibited an enhanced ability to induce DNA cleavage in the presence of TOP1 and exceptional cytotoxic activity with IC50 values below 2 nM against the human lymphoblastoma cell line (RPMI8402) but dose limiting toxicity limited in vivo efficacy in the human tumor xenograft athymic nude mouse model (MDA-MB-435 breast tumor cell line) [66]. In an effort to obtain a less toxic analog with improved efficacy, a number of compounds related to 58 ( Figure 30) were synthesized where the 11-ethyl group was substituted at its 2-position with various polar moieties (N-methylamino-, Nisopropylamino-, hydroxy-and hydroxylamino-groups). These analogs were prepared via the trimethylammonium derivatives of ARC-31 according to methods described in [67]. All analogs exhibited high cytotoxic activity. Although, derivatives with N-methylamine 59 and Nisopropylamine 60 ( Figure 31) exhibited greater cytotoxic activity in vitro in comparison to ARC-31, evaluation in vivo in athymic nude mice showed minimal differences in efficacy in comparison to ARC-31 without therapeutic index improvement [11].  As a continuation of studies on the most potent derivative 57, 5,6,11-triazachrysen-12-ones with various substituents at 11-position were synthesized [65]. Compound 58 (ARC-31, Figure 30) exhibited an enhanced ability to induce DNA cleavage in the presence of TOP1 and exceptional cytotoxic activity with IC 50 values below 2 nM against the human lymphoblastoma cell line (RPMI8402) but dose limiting toxicity limited in vivo efficacy in the human tumor xenograft athymic nude mouse model (MDA-MB-435 breast tumor cell line) [66]. In an effort to obtain a less toxic analog with improved efficacy, a number of compounds related to 58 ( Figure 30) were synthesized where the 11-ethyl group was substituted at its 2-position with various polar moieties (N-methylamino-, N-isopropylamino-, hydroxy-and hydroxylamino-groups). These analogs were prepared via the trimethylammonium derivatives of ARC-31 according to methods described in [67]. All analogs exhibited high cytotoxic activity. Although, derivatives with N-methylamine 59 and N-isopropylamine 60 ( Figure 31) exhibited greater cytotoxic activity in vitro in comparison to ARC-31, evaluation in vivo in athymic nude mice showed minimal differences in efficacy in comparison to ARC-31 without therapeutic index improvement [11]. As a continuation of studies on the most potent derivative 57, 5,6,11-triazachrysen-12-ones with various substituents at 11-position were synthesized [65]. Compound 58 (ARC-31, Figure 30) exhibited an enhanced ability to induce DNA cleavage in the presence of TOP1 and exceptional cytotoxic activity with IC50 values below 2 nM against the human lymphoblastoma cell line (RPMI8402) but dose limiting toxicity limited in vivo efficacy in the human tumor xenograft athymic nude mouse model (MDA-MB-435 breast tumor cell line) [66]. In an effort to obtain a less toxic analog with improved efficacy, a number of compounds related to 58 ( Figure 30) were synthesized where the 11-ethyl group was substituted at its 2-position with various polar moieties (N-methylamino-, Nisopropylamino-, hydroxy-and hydroxylamino-groups). These analogs were prepared via the trimethylammonium derivatives of ARC-31 according to methods described in [67]. All analogs exhibited high cytotoxic activity. Although, derivatives with N-methylamine 59 and Nisopropylamine 60 ( Figure 31) exhibited greater cytotoxic activity in vitro in comparison to ARC-31, evaluation in vivo in athymic nude mice showed minimal differences in efficacy in comparison to ARC-31 without therapeutic index improvement [11].  As a continuation of studies on the most potent derivative 57, 5,6,11-triazachrysen-12-ones with various substituents at 11-position were synthesized [65]. Compound 58 (ARC-31, Figure 30) exhibited an enhanced ability to induce DNA cleavage in the presence of TOP1 and exceptional cytotoxic activity with IC50 values below 2 nM against the human lymphoblastoma cell line (RPMI8402) but dose limiting toxicity limited in vivo efficacy in the human tumor xenograft athymic nude mouse model (MDA-MB-435 breast tumor cell line) [66]. In an effort to obtain a less toxic analog with improved efficacy, a number of compounds related to 58 ( Figure 30) were synthesized where the 11-ethyl group was substituted at its 2-position with various polar moieties (N-methylamino-, Nisopropylamino-, hydroxy-and hydroxylamino-groups). These analogs were prepared via the trimethylammonium derivatives of ARC-31 according to methods described in [67]. All analogs exhibited high cytotoxic activity. Although, derivatives with N-methylamine 59 and Nisopropylamine 60 ( Figure 31) exhibited greater cytotoxic activity in vitro in comparison to ARC-31, evaluation in vivo in athymic nude mice showed minimal differences in efficacy in comparison to ARC-31 without therapeutic index improvement [11]. Zoidis and co-workers obtained tetra-and pentacyclic cinnoline based compounds indeno [1,2c]cinnoline and benzo[h]indeno [1,2-c]cinnoline, respectively, bearing protonable amino groups. All tested compounds inhibited proliferation of human cervical carcinoma (HeLa) and human breast adenocarcinoma (MCF-7) cell lines as well as displayed intercalating properties on different nucleic  Figure 32) caused the highest thermal stabilization in melting studies and exerted acceptable inhibitory activity on human topoisomerase IIα [68]. Borowski and co-workers obtained a series of anthrapyridazone derivatives 63 ( Figure 33) bearing one or two basic side chains at various positions of the tetracyclic core [69,70]. The compounds 2,7-dihydro-3H-dibenzo[de,h]cinnoline-3,7-diones 64 and 65 ( Figure 33) exhibited in vitro cytotoxic activity against murine (L1210) and human (K562) leukemia cell lines. In addition, they were active against human leukemia multi-drug-resistant (K562/DX) cell lines. The most active compounds 64 and 65 ( Figure 33) were also tested in vivo against murine P388 leukemia and showed activity comparable to mitoxantrone [71]. Borowski and co-workers obtained a series of anthrapyridazone derivatives 63 ( Figure 33) bearing one or two basic side chains at various positions of the tetracyclic core [69,70]. The compounds 2,7-dihydro-3H-dibenzo[de,h]cinnoline-3,7-diones 64 and 65 ( Figure 33) exhibited in vitro cytotoxic activity against murine (L1210) and human (K562) leukemia cell lines. In addition, they were active against human leukemia multi-drug-resistant (K562/DX) cell lines. The most active compounds 64 and 65 ( Figure 33) were also tested in vivo against murine P388 leukemia and showed activity comparable to mitoxantrone [71]. . Particular efficacy of tested compounds was observed against the leukemia subpanel. In addition, they were also found to be active in cells overexpressing MDR1. The compounds caused apoptosis, mitochondrial depolarization, generation of reactive oxygen species, and the activation of caspase-3, caspase-8, and caspase-9. Moreover, they acted as topoisomerase I inhibitors [72]. Barlaam and co-workers, while working on the optimization of selective quinoline based inhibitors of ataxia teleangiectasia mutated (ATM) kinase involved in the repair of DNA double strand breaks, synthesized a series of cinnoline-3-carboxamides as suitable replacements of quinoline carboxamides. Compound 67 (Figure 35) was identified as a potent ATM inhibitor with excellent kinase selectivity and good physicochemical and pharmacokinetic properties. Monotherapy with ATM inhibitor 67 did not cause tumor regression in the SW620 colorectal tumor xenograft model, whereas combination with irinotecan resulted in significantly greater tumor growth inhibition in . Particular efficacy of tested compounds was observed against the leukemia subpanel. In addition, they were also found to be active in cells overexpressing MDR1. The compounds caused apoptosis, mitochondrial depolarization, generation of reactive oxygen species, and the activation of caspase-3, caspase-8, and caspase-9. Moreover, they acted as topoisomerase I inhibitors [72]. . Particular efficacy of tested compounds was observed against the leukemia subpanel. In addition, they were also found to be active in cells overexpressing MDR1. The compounds caused apoptosis, mitochondrial depolarization, generation of reactive oxygen species, and the activation of caspase-3, caspase-8, and caspase-9. Moreover, they acted as topoisomerase I inhibitors [72]. Barlaam and co-workers, while working on the optimization of selective quinoline based inhibitors of ataxia teleangiectasia mutated (ATM) kinase involved in the repair of DNA double strand breaks, synthesized a series of cinnoline-3-carboxamides as suitable replacements of quinoline carboxamides. Compound 67 (Figure 35) was identified as a potent ATM inhibitor with excellent kinase selectivity and good physicochemical and pharmacokinetic properties. Monotherapy with ATM inhibitor 67 did not cause tumor regression in the SW620 colorectal tumor xenograft model, whereas combination with irinotecan resulted in significantly greater tumor growth inhibition in Barlaam and co-workers, while working on the optimization of selective quinoline based inhibitors of ataxia teleangiectasia mutated (ATM) kinase involved in the repair of DNA double strand breaks, synthesized a series of cinnoline-3-carboxamides as suitable replacements of quinoline carboxamides. Compound 67 (Figure 35) was identified as a potent ATM inhibitor with excellent kinase selectivity and good physicochemical and pharmacokinetic properties. Monotherapy with ATM inhibitor 67 did not cause tumor regression in the SW620 colorectal tumor xenograft model, whereas combination with irinotecan resulted in significantly greater tumor growth inhibition in comparison to irinotecan alone [14,73]. The 1,3-dihydroimidazo [4,5-c]cinnoline-2-one derivatives of general formula 68 ( Figure 35) were patented as ATM modulators used to treat or prevent ATM mediated diseases, including cancer [74].
comparison to irinotecan alone [14,73]. The 1,3-dihydroimidazo [4,5-c]cinnoline-2-one derivatives of general formula 68 ( Figure 35) were patented as ATM modulators used to treat or prevent ATM mediated diseases, including cancer [74]. Colony-stimulating factor-1 (CSF-1) through binding to its receptor (CSF-1R )regulates the migration, proliferation, function, and survival of macrophages [75]. Since CSF-1R is overexpressed in many tumors and at sites of inflammation, CSF-1R inhibitors seem to be an attractive therapeutic strategy for cancer as well as autoimmune and inflammatory diseases. The 3-amido-4anilinocinnolines of general formula 69 ( Figure 36) were reported as potent, highly selective CSF-1R inhibitors [76,77]. They were designed in order to overcome the cardiovascular liability of potent and selective 3-amido-4-anilinoquinoline CSF-1R inhibitor (AZ683), which was able to reduce the level of tumor-associated macrophages in a breast cancer xenograft model. The 3-amido-4-anilinocinnoline compound with 1-hydroxyethylpiperazine substituent at 7 position of cinnoline scaffold 70 (AZD7507, Figure 36) was a potent CSF-1R inhibitor demonstrating good oral pharmacokinetic profile as well as reduced risk of cardiotoxicity in comparison to AZ683 [78]. c-Met receptor tyrosine kinase is another cellular target for compounds designed as potential anticancer agents because it has been found to be overexpressed or mutated in various human cancer cells [79]. Some 4-(2-fluorophenoxy)quinoline derivatives bearing a 4-oxo-1,4-dihydrocinnoline-3carboxamide moiety 71 ( Figure 37) were designed as c-Met inhibitors and evaluated against five c- Colony-stimulating factor-1 (CSF-1) through binding to its receptor (CSF-1R) regulates the migration, proliferation, function, and survival of macrophages [75]. Since CSF-1R is overexpressed in many tumors and at sites of inflammation, CSF-1R inhibitors seem to be an attractive therapeutic strategy for cancer as well as autoimmune and inflammatory diseases. The 3-amido-4-anilinocinnolines of general formula 69 ( Figure 36) were reported as potent, highly selective CSF-1R inhibitors [76,77]. They were designed in order to overcome the cardiovascular liability of potent and selective 3-amido-4-anilinoquinoline CSF-1R inhibitor (AZ683), which was able to reduce the level of tumor-associated macrophages in a breast cancer xenograft model. The 3-amido-4-anilinocinnoline compound with 1-hydroxyethylpiperazine substituent at 7 position of cinnoline scaffold 70 (AZD7507, Figure 36) was a potent CSF-1R inhibitor demonstrating good oral pharmacokinetic profile as well as reduced risk of cardiotoxicity in comparison to AZ683 [78].

Miscellaneous
A compound with cinnoline moiety 78 ( Figure 40) is patented by Stein and co-authors for very interesting uses. Among other hydrazone derivatives, it was tested as inhibitor of a transient receptor potential cation channel, subfamily M, member 5 (TRPM5) protein which has been shown to be essential for taste transduction. Such compounds could be used as taste inhibitors when administered as a component of pharmaceutical or food products to improve acceptance. Moreover, these agents are intended for use in treating diabetes mellitus, obesity, insulin resistance syndrome and many more [89]. Cinnoline derivatives were also patented as thyroid hormone receptor agonists [90], orexin receptor antagonists [91][92][93][94][95], histone deacetylase (HDAC) inhibitors [96], liver X receptors β selective modulators for the treatment of atherosclerosis [97], somatostatin regulators [98] or cannabinoid-1 receptor inverse agonists [99] and many more.

Conclusions
In this paper, we have presented a review of studies focused on the biological activity of cinnoline derivatives conducted by many research groups worldwide between 2005 and 2019. The provided information clearly indicates the enormous significance of the cinnoline framework as a building block of many valuable compounds. Compounds bearing the cinnoline scaffold are able to interact with a variety of molecular targets including receptors such as GABA A, CSF-1R, H3R and enzymes such as cyclooxygenase-2, topoisomerases, phosphodiesterase, human neutrophil elastase, Bruton's tyrosine kinase involved in pathogenesis of many diseases. As a consequence, they are intended to be used as antibacterial, antifungal, antimalarial, anti-inflammatory, analgesic, anxiolytic and antitumor agents. Some cinnoline derivatives are under evaluation in clinical trials. There is no doubt that development of cinnoline based molecules constitutes a significant contribution to the

Miscellaneous
A compound with cinnoline moiety 78 ( Figure 40) is patented by Stein and co-authors for very interesting uses. Among other hydrazone derivatives, it was tested as inhibitor of a transient receptor potential cation channel, subfamily M, member 5 (TRPM5) protein which has been shown to be essential for taste transduction. Such compounds could be used as taste inhibitors when administered as a component of pharmaceutical or food products to improve acceptance. Moreover, these agents are intended for use in treating diabetes mellitus, obesity, insulin resistance syndrome and many more [89].

Miscellaneous
A compound with cinnoline moiety 78 ( Figure 40) is patented by Stein and co-authors for very interesting uses. Among other hydrazone derivatives, it was tested as inhibitor of a transient receptor potential cation channel, subfamily M, member 5 (TRPM5) protein which has been shown to be essential for taste transduction. Such compounds could be used as taste inhibitors when administered as a component of pharmaceutical or food products to improve acceptance. Moreover, these agents are intended for use in treating diabetes mellitus, obesity, insulin resistance syndrome and many more [89]. Cinnoline derivatives were also patented as thyroid hormone receptor agonists [90], orexin receptor antagonists [91][92][93][94][95], histone deacetylase (HDAC) inhibitors [96], liver X receptors β selective modulators for the treatment of atherosclerosis [97], somatostatin regulators [98] or cannabinoid-1 receptor inverse agonists [99] and many more.

Conclusions
In this paper, we have presented a review of studies focused on the biological activity of cinnoline derivatives conducted by many research groups worldwide between 2005 and 2019. The provided information clearly indicates the enormous significance of the cinnoline framework as a building block of many valuable compounds. Compounds bearing the cinnoline scaffold are able to interact with a variety of molecular targets including receptors such as GABA A, CSF-1R, H3R and enzymes such as cyclooxygenase-2, topoisomerases, phosphodiesterase, human neutrophil elastase, Bruton's tyrosine kinase involved in pathogenesis of many diseases. As a consequence, they are intended to be used as antibacterial, antifungal, antimalarial, anti-inflammatory, analgesic, anxiolytic and antitumor agents. Some cinnoline derivatives are under evaluation in clinical trials. There is no doubt that development of cinnoline based molecules constitutes a significant contribution to the Cinnoline derivatives were also patented as thyroid hormone receptor agonists [90], orexin receptor antagonists [91][92][93][94][95], histone deacetylase (HDAC) inhibitors [96], liver X receptors β selective modulators for the treatment of atherosclerosis [97], somatostatin regulators [98] or cannabinoid-1 receptor inverse agonists [99] and many more.

Conclusions
In this paper, we have presented a review of studies focused on the biological activity of cinnoline derivatives conducted by many research groups worldwide between 2005 and 2019. The provided information clearly indicates the enormous significance of the cinnoline framework as a building block of many valuable compounds. Compounds bearing the cinnoline scaffold are able to interact with a variety of molecular targets including receptors such as GABA A, CSF-1R, H 3 R and enzymes such as cyclooxygenase-2, topoisomerases, phosphodiesterase, human neutrophil elastase, Bruton's tyrosine kinase involved in pathogenesis of many diseases. As a consequence, they are intended to be used as antibacterial, antifungal, antimalarial, anti-inflammatory, analgesic, anxiolytic and antitumor agents. Some cinnoline derivatives are under evaluation in clinical trials. There is no doubt that development of cinnoline based molecules constitutes a significant contribution to the identification of lead compounds with optimized pharmacodynamic and pharmacokinetic properties.