# Design and synthesis of fused tetrahydroisoquinoline-iminoimidazolines

Valeria Moas Heloire, Nicolas Renault, Vânia Batalha, Angela Rincon Arias, Mathieu Marchivie, Saïd Yous, Noémie Deguine, Luc Buee, Philippe Chavatte, David Blum, et al.

► To cite this version:

Valeria Moas Heloire, Nicolas Renault, Vânia Batalha, Angela Rincon Arias, Mathieu Marchivie, et al. Design and synthesis of fused tetrahydroisoquinoline-iminoimidazolines. European Journal of Medicinal Chemistry, 2015, 106, pp.15-25. 10.1016/j.ejmech.2015.10.030. hal-02060788

HAL Id: hal-02060788

https://hal.science/hal-02060788

Submitted on 8 Mar 2019

HAL is a multi-disciplinary open access
archive for the deposit and dissemination of sci-
entific research documents, whether they are pub-
lished or not. The documents may come from
teaching and research institutions in France or
abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est
destinée au dépôt et à la diffusion de documents
scientifiques de niveau recherche, publiés ou non,
émanant des établissements d'enseignement et de
recherche français ou étrangers, des laboratoires
publics ou privés.

<figure><img src="image_1.png" /></figure>

Distributed under a Creative Commons Attribution 4.0 International License
DESIGN AND SYNTHESIS OF FUSED TETRAHYDROISOQUINOLINE-
IMINOIMIDAZOLINES

Valeria Moas-Héloire<sup>a,b</sup>, Nicolas Renault<sup>a,b</sup>, Vania Batalha<sup>c</sup>, Angela Rincon Arias<sup>e</sup>, Mathieu Marchivie<sup>d</sup>, Said Yous<sup>a,e</sup>, Noémie Deguine<sup>a</sup>, Luc Buée<sup>a,e</sup>, Philippe Chavatte<sup>a,f</sup>, David Blum<sup>a,e</sup>, Luisa Lopes<sup>c</sup>, Patricia Melnyk<sup>a,e,*</sup> and Laurence Agouridas<sup>a,e,*</sup>

**Mail address:** UFR Pharmacie, 3 rue du Pr Laguesse, BP83, 59006 Lille

**List of e-mail addresses:** valeria.heloire-2@univ-lille2.fr, nicolas.renault-3@univ-lille2.fr, vanialnbatalha@gmail.com, ararias@quim.ucm.es, mathieu.marchivie@icmcb.cnrs.fr, said.yous@univ-lille2.fr, noemie.deguine-2@univ-lille2.fr, luc.buee@inserm.fr, philippe.chavatte@univ-lille2.fr, david.blum@inserm.fr, lvlopes@medicina.ulisboa.pt, patricia.melnyk@univ-lille2.fr, laurence.agouridas@univ-lille2.fr

**Corresponding authors:**

Patricia Melnyk, "Onco and NeuroChemistry", Jean-Pierre Aubert Research Center UMR-S1172 - Faculté des Sciences Pharmaceutiques et Biologiques (Université Lille 2)
3, rue du Pr Laguesse - BP83 - 59006 Lille Cedex Tel : 33 (0)3 20 96 49 49 - fax : 33 (0)3 20 96 49 13. mail: patrica.melnyk@univ-lille2.fr

Laurence Agouridas, "Onco and NeuroChemistry", Jean-Pierre Aubert Research Center UMR-S1172 - Faculté des Sciences Pharmaceutiques et Biologiques (Université Lille 2, 3 rue du Pr Laguesse - BP83 - 59006 Lille Cedex Tel : 33 (0)3 20 96 43 68 - fax : 33 (0)3 20 96 49 13. mail: laurence.agouridas@univ-lille2.fr

<footnote><sup>a</sup> Université de Lille, F-59000 Lille, France</footnote>

<footnote><sup>b</sup> UDSL, EA 4481, UFR Pharmacie, F-59000 Lille, France</footnote>

<footnote><sup>c</sup> Instituto de Medecina Molecular, 1640-028, Lisbon, Portugal</footnote>

<footnote><sup>d</sup> Université Bordeaux, CNRS FRE3396, F-33000 Bordeaux, France</footnote>

<footnote><sup>e</sup> Inserm UMR-S1172, JPARC, F-59000 Lille, France</footnote>

<footnote><sup>f</sup> Inserm UMR 995, LIRIC, F-59000 Lille, France</footnote>
**Abstract**

In the aim of identifying new privileged structures, we describe the 5-steps synthesis of cyclic guanidine compounds "tetrahydroisoquinoline-iminoimidazolines" derived from tetrahydroisoquinoline-hydantoin core. In order to evaluate this new minimal structure and the impact of replacing a carbonyl by a guanidine moiety, their affinity towards adenosine receptor A<sub>2A</sub> was evaluated and compared to those of tetrahydroisoquinoline-hydantoin compounds.

**KEYWORDS:** guanidines, A<sub>2A</sub> receptor, privileged structure, iminoimidazoline

**Abbreviations:** Tic-H, tetrahydroisoquinoline hydantoin; A<sub>2A</sub>R, adenosine A<sub>2A</sub> receptor.
# 1. Introduction

In the field of medicinal chemistry, privileged structures are considered as a promising source of ligands able to interact with various targets.¹⁻³ In fact, these minimal structures constitute a powerful starting point to identify original compounds by playing with the nature of their substituents. They constitute an anchor point enabling to orientate substituents in various positions, giving access to potentially promising structures and to modulate their affinity and activity. Therefore, identification of novel privileged structures appears as an interesting challenge.

Our group described a series of tetrahydroisoquinoline-hydantoin (Tic-H **1**, Figure 1) derived compounds with potent affinity for the sigma-1 receptor in the nanomolar range ensuing various therapeutical *in vivo* applications.⁴⁻⁹ This heterocycle presents an interesting hydrogen bond acceptor group on the hydantoin cycle and is amenable to various decorations. In this study we proposed to identify a new potential privileged structure and decided to evaluate the consequence of replacing the hydantoin's (**1**) hydrogen bond acceptor by a hydrogen bond donor group resulting in the guanidine moiety of compound **2** (Figure 1). This new heterocycle could be the central core for the design of new active compounds. Guanidines are present in a large variety of natural products with potent biological activities in many fields¹⁰⁻¹³ and more specifically in the central nervous system area.¹⁴,¹⁵

This approach was exemplified by preparing the guanidine derivative “Tic-guanidine” and its derivatives (**2** to **13**) in order to evaluate the affinity of this new series on the adenosine receptor A₂AR.¹⁶ In fact, in addition to its affinity towards sigma-1 receptor, Tic-H **1** showed a promising affinity constant towards adenosine receptor A₂A (Kᵢ = 44 µM). This receptor is widely expressed in the central nervous system. Expressed at different levels (neurons, astrocytes, microglial cells),¹⁷⁻²¹ it acts at various levels of regulation.²²⁻²⁵ Therefore, A₂ARs are viewed as promising targets in various neurodegenerative diseases, mainly Parkinson’s and Alzheimer’s diseases.²⁶⁻²⁸

Docking studies, based on the crystal structure of A₂A bound to the high affinity antagonist (ZM241385),²⁹ showed that Tic-guanidine restored hydrogen bonds that were missing for Tic-H leading us to expect a better affinity of these compounds.³⁰

We therefore aim to develop a new series of Tic-guanidine compound. We set up an original and efficient chemical synthesis allowing pharmacomodulations. These latter concern the
nature of the substituents on the original tricyclic structure Tic-guanidine (series A, figure 1) but also modifications by opening of the central cyclic core (series B, figure 1).

Figure 1.

**2. Results and discussion**

**2.1. Modeling studies**

Jaakola *et al*. published in 2008 a crystal structure of A<sub>2</sub>A<sub>R</sub> with the high affinity antagonist ZM241385.<sup>29</sup> Tic-H and Tic-guanidine were both docked in A<sub>2</sub>A<sub>R</sub>'s binding site. Results showed both molecules place in a same manner as ZM241385 does (Figure 2). Mainly, important hydrogen bonds with Glu<sup>169</sup> and Asn<sup>253</sup> missing in the case of Tic-H are being restored with the guanidine function. Finally, upper carbonyl moiety of Tic-H was not maintained, as it doesn't seem to play an essential role in the interactions with amino acids of the binding site.

Figure 2.

Supported by our docking studies, pharmacomodulations were envisaged: various N-
substituents on the guanidine core (series A, figure 1) and opening of the central tricyclic core
(series B, figure 1). Series A enables to evaluate the effect of the modification of the benzyl
group in the hydrophobic upper pocket of the binding site. On the other hand, series B was
prepared in order to evaluate the importance of the central core's nature and especially how its
geometry impacts compounds' affinity. We wanted to establish whether a planar
conformation was essential as described in many A<sub>2</sub>A's antagonists ref or if less restricted
structures could improve affinity as compared to already published non planar A<sub>2</sub>A
antagonists ref. Finally, B series would bring additional informations on structure activity
relationships of our Tic-guanidine compounds.

**2.2. Chemical synthesis**

Synthesis of Tic-guanidine derivatives of series A and B was achieved thanks to a unique
chemical pathway as depicted in Scheme 1. This enabled us to access desired compounds
starting from commercially available amino acids and therefore to easily access various
pharmacomodulations. We optimized the synthetic pathway of compound **2** starting from commercially available Boc-protected L-tetrahydroisoquinoline carboxylic acid **I** whose chemistry is well mastered in our group.⁷,⁹,³¹ Key step of this synthesis was the final formation of the cyclic guanidine. Various methods are described in the literature to access guanidines¹² and cyclic guanidines³²⁻³⁴ but could not be applied to our strategy. Indeed, we previously described the synthesis of Tic-thiohydantoins, which could be an interesting intermediate for the synthesis of these Tic-guanidines. But their chemical and enantiomeric instability precluded their use in this study.³⁵ For final formation of guanidine cycle, direct cyclisation of the free diamine in the presence of cyanogen bromide³⁶⁻⁴² was unsuccessful. Some adjustments were then required and we finally chose to maintain Boc-protection in order to selectively functionalize the free amine of compounds **2b-13b** with cyanogene bromide.⁴³ Expected guanidines were obtained after Boc-deprotection and subsequent cyclization of intermediate.

Synthesis of series A started from Boc-protected L-tetrahydroisoquinoline carboxylic acid **I**. Corresponding aldehyde was obtained *via* reduction of Weinreb amide **II**⁴⁴,⁴⁵ using $LiAlH_4$ in THF at 0°C.⁴⁴ Reductive amination with appropriate amine in the presence of sodium triacetoloxyborohydride in $CH_2Cl_2$ gave expected Boc-monoprotected derivatives **2b-6b**.⁴⁶ Free amine of this last compound was then functionalized using cyanogen bromide in ethanol to give corresponding nitrile derivatives.⁴³ Finally, guanidine cyclization was achieved thanks to acidic deprotection of the carbamate and *in situ* cyclization of the intermediate diamine compound with yields from 46 to 88%.

**Scheme 1.**

**Figure 3.**

Depending on the nature of the amine in the reductive amination step, compounds **2** to **6** were first obtained (scheme 1), as depicted in figure 3. On the other hand, opening of the upper part of the central cycle while maintaining *N*-benzyl substitution of the guanidine (scheme 2) core resulted from modification of starting amino acid (compounds **7-13**, Figure 4). Compound **12**, derived from tyrosine, was prepared from Boc-Tyr(*t*Bu)-OH. The tert-butyl protection was removed in the final cyclisation step.

**Scheme 2.**
Figure 4.

Structure of final compounds was confirmed by various analysis methods. LC-MS gave expected mass, discarding the formation of the dimer. FT-IR indicated the disappearance of the nitrile band (2260-2240 cm⁻¹) and appearance of the guanidine C=NH bond (1690-1640 cm⁻¹) confirming cyclisation of the compound (see supporting informations). Finally, 1D and 2D ¹H and ¹³C NMR showed appropriate signals and correlations, especially the guanidine C=NH bond.

Some of our compounds were able to crystallize. X-ray spectroscopy thus enabled us to confirm the structure and enantiopurity of compounds **2** and **3** (figure 5) but also compounds **7**, **9** and **11** whose absolute configuration was maintained.⁴⁷

On the other hand, crystallographic data showed compound **8** was present as a racemic mixture (for details, see supplementary informations). This compound differs from the other ones, as it is *N*-methylated. *N*-methylation was achieved following a protocol described in the literature to be a non-epimerizing route.⁴⁸ However, in our case, the basic conditions of methylation led to a complete racemization.

Figure 5.

Figure 6.

## 2.3. Affinity of Tic-guanidine compounds

Affinity of our compounds was evaluated on human A₂A receptor membranes stably expressed in HEK293 cells. Results (table 1) showed no improvement of binding compared to initial Tic-H compound **1** (Ki = 44 µM) except for the tricyclic guanidine substituted with phenylbutylamine **5** (Ki = 35 µM). More specifically, affinity was drastically decreased for open guanidines.

Table 1.

Except compound **5** (35 µM) that gives an affinity comparable to the one of Tic-H **1** (44 µM), binding was not improved with our Tic-guanidine compounds. For sure, rigid tricyclic structures are necessary to bind the A₂AR receptor as open structures (**7** to **13**) showed no affinity at all. X-ray analysis helped understanding this observation. In fact, as shown in
figure 5, open Tic-guanidine structures adopt a curved conformation that does not allow the binding of the molecule in the pocket of the receptor whereas tricyclic structures keep a flat conformation.⁴⁹

Concerning the tricyclic structures (2 to 6), we therefore assume the replacement of carbonyl moiety of 1 by a hydrogen bond donor does not give us expected improved affinity for the A₂AR.

## 2.4. Cytotoxicity of Tic-guanidine compounds

Cytotoxicity assays have been conducted on SY5Y cells and showed no toxicity of our compounds at 100 µM (Table 1).

## 3. Conclusion

As Tic-H core was of interest for the design of various biologically active compounds, we proposed the Tic-guanidine core as a new privileged structure. This work therefore presented a new and efficient synthesis of guanidine cycles derived from amino acids. We applied this concept to the design of A₂AR ligands. Unfortunately, binding results established that our initial hypothesis was not confirmed: replacement of the hydrogen bond acceptor moiety carbonyl of compound 1 by the hydrogen bond donor guanidine did not improve the affinity for A₂AR. Other decorations are needed to improve the affinity for A₂AR. Of particular importance is the lack of cytotoxicity of this new scaffold. Thus valorization of this new scaffold for other receptors is currently under evaluation.

# 4. Experimental section

## 4.1. General information

Chemicals and solvents were obtained from commercial sources, and used without further purification unless otherwise noted. Reactions were monitored by TLC performed on Macherey-Nagel Alugram® Sil 60/UV₂₅₄ sheets (thickness 0.2mm). Purification of products was carried out by either column chromatography or thick layer chromatography. Column chromatography was carried out on using Macherey-Nagel silica gel (230-400 mesh). Thick layer chromatography was performed on glass plates coated with Macherey-Nagel Sil/UV₂₅₄ (thickness 2 mm), from which the pure compounds were extracted with the following solvent system: DCM/MeOH(NH₃), 90:10. NMR spectra were recorded on a Bruker DRX 300 spectrometer (operating at 300 MHz for ¹Hand 75 MHz for ¹³C). Chemical shifts are
expressed in ppm relative to either tetramethylsilane (TMS) or to residual proton signal in
deuterated solvents. Chemical shifts are reported as position (δ in ppm), multiplicity (s =
singlet, d = doublet, t = triplet, q = quartet, p = pentet, dd = double doublet, br = broad and m
= multiplet), coupling constant (J in Hz), relative integral and assignment. The attributions of
protons and carbons were achieved by analysis of 2D experiments (COSY, HSQC and
HMBC). Mass spectra were recorded on a Varian triple quadrupole 1200W mass spectrometer
equipped with a non-polar C18 TSK-gel Super ODS (4.6 x 50 mm) column, using
electrospray ionization and a UV detector (diode array). HRMS-ESI spectra were recorded on
a Thermo Scientific Exactive spectrometer. The purity of final compounds was verified by
two types of high pressure liquid chromatography (HPLC) columns: C18 Interchrom
UPTISPHERE and C4 Interchrom UPTISPHERE. Analytical HPLC was performed on a
Shimadzu LC-2010AHT system equipped with a UV detector set at 254 nm and 215 nm.
Compounds were dissolved in 50 µL methanol and 950 µL buffer A, and injected into the
system. The following eluent systems were used: buffer A (H₂O/TFA, 100:0.1) and buffer B
(CH₃CN/H₂O/TFA, 80:20:0.1). HPLC retention times (HPLC tR) were obtained at a flow rate
of 0.2 mL/min for 35 min using the following conditions: a gradient run from 100% of buffer
A over 1 min, then to 100% of buffer B over the next 30 min. The melting point analyses were
performed on Barnstead Electrothermal Melting Point Series IA9200 and are uncorrected.
Infrared spectra were performed on Bruker FT-IR spectrometer model α. Preparative HPLC
were performed using a Varian PRoStar system using an OmniSphere 10 column C18 250
mm x 41.4 mm Dynamax from Varian, Inc. A gradient starting from 20% CH₃CN/80%
H₂O/0.1% formic acid and reaching 100% CH₃CN/0.1% formic acid at a flow rate of 80
mL/min was used. Optical rotations were measured at 20°C on a Perkin-Elmer 343
polarimeter.

Crystallographic data (excluding structure factors) for the structures in this paper have been
deposited with the Cambridge Crystallographic Data Centre as supplementary publication
1401476, 1401477, 1401478, 1401480, 1401481 and 1401479. Copies of the data can be
obtained, free of charge, on application to CCDC, 12 Union Road, Cambridge CB2 1EZ, UK
(fax: +44-(0)1223-336033 or email: deposit@ccdc.cam.ac.uk).

**4.2. Preparation of Weinreb amide compounds.**

4.2.1. *General method 1.* To a solution of appropriate amino protected carboxylic acid derivative (1 eq.) dissolved in a mixture of CH₂Cl₂ and CH₃CN (1:1, 3,7 mL/mmol of carboxylic acid), EDC (1.3 eq.), HOBt (1.3 eq.), NMM (6.5 eq.) and HN(Me)OMe.HCl (2.1
eq.) were added. The mixture was stirred at room temperature over 24-72 h and then evaporated. The resulting crude product was dissolved in CH₂Cl₂, washed three times with saturated NaHCO₃ solution, three times with 1M HCl and once with brine. Organic layer was dried over MgSO₄, filtered and concentrated in vacuo. Purification via flash chromatography was performed.

### 4.2.2. tert-butyl (3S)-3-[methoxy(methyl)carbamoyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate (II).
According to general method 1, **II** was obtained as a colorless oil (5.13g, 61%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm) *indicates the presence of 2 conformers:* 7.15-7.20 (m, 4H); 5.23 and 4.86 [X part of ABX system (ft, *J* = 6.0 Hz, 0.5H) and (ft, *J* = 7.5 Hz, 0.5H)]; 4.70 and 4.69 [(AB system, *J* = 16.5 Hz, 1H, Δ*υ* = 36 Hz) and (AB system, *J* = 16.5 Hz, 1H, Δ*υ* = 123 Hz)]; 3.85 and 3.78 (2s, 3H); 3.19 and 3.16 (2s, 3H); 3.19-2.94 (unresolved AB part of ABX system, m, 2H); 1.51 and 1.45 (2s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm) *indicates the presence of 2 conformers:* 175.6, 174.9, 173.0, 155.3, 154.9, 154.7, 135.5, 134.7, 134.2, 133.9, 133.0, 132.9, 132.9, 132.7, 132.1, 129.9, 129.4, 128.8, 127.9, 127.7, 127.6, 127.1, 125.8, 83.9, 81.6, 80.4, 61.7, 61.5, 61.3, 54.8, 53.5, 53.2, 52.3, 50.7, 45.4, 44.5, 43.4, 32.7, 32.5, 32.2, 30.9, 30.4, 30.1, 29.8, 28.4, 28.3, 28.1. MS (ESI⁺): m/z = 321.1 [M,H]⁺ found; C₁₇H₂₅N₂O₄ calculated m/z = 321.2[M,H]⁺.

### 4.2.3. tert-butyl N-[(1S)-1-benzyl-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (7a).
According to general method 1, **7a** was obtained as a brown oil (0.94 g, 66%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.29-7.12 (m, 5H); 5.18 and 5.03-4.86 [X part of ABX system (d, *J* = 6.0 Hz, 0.5H) and (m, 0.5 H)]; 3.65 (s, 3H); 3.16 (s, 3H); 2.95 [AB part of ABX system (dd, *J* = 12.0 and 6.0 Hz, 1H) and (dd, *J* = 15.0 and 9.0 Hz, 1H), Δ*υ* = 60 Hz]; 1.39 (s, 9H).⁵⁰

### 4.2.4. tert-butyl N-[(1S)-1-benzyl-2-[methoxy(methyl)amino]-2-oxo-ethyl]-N-methylcarbamate (8a).
According to general method 1, **8a** was obtained as a yellow oil (0.942 g, 86%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): *indicates the presence of 2 conformers.* 7.27-7.16 (m, 5H); 5.64-5.07 [unresolved X part of ABX system, m, 1H); 3.64 and 3.61 (2s, 3H); 3.16 and 3.19 (2s, 3H); 3.06-2.87 [unresolved AB part of ABX system, m, 2H]; 2.85 (s, 3H); 1.35 and 1.23 (2s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): *indicates the presence of 2 conformers.* 171.5, 155.6, 154.9, 138.0, 137.5, 129.4, 129.2, 128.4, 128.2, 126.4, 79.8, 61.3, 57.2, 54.7, 34.9, 30.2, 29.8, 28.2, 26.8. MS (ESI⁺): m/z = 323.0 [M,H]⁺ found; C₁₇H₂₆N₂O₄ calculated m/z = 323.2[M,H]⁺.

### 4.2.5. tert-butyl N-[(1S)-1-[(4-fluorophenyl)methyl]-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (9a).
According to general method 1, **9a** was obtained as a colorless oil (0.810 g, 70%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.18-7.09 (m, 2H); 7.02-6.90 (m, 2H);
5.28-5.17 (d, *J* = 12.1 Hz, 1H), 5.00-4.82 (unresolved X part of ABX system, m, 1H); 3.67 (s, 3H); 3.16 (s, 3H); 2.87 [AB part of ABX system (dd, *J* = 12.2 and 6.3 Hz, 1H) and (dd, *J* = 12.1 and 6.6 Hz, 1H), Δ*υ* = 60 Hz]; 1.38 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 172.0, 163.5, 160.2 (Jₐ-F = 234 Hz); 155.1, 132.3, 131.0, 130.9, 115.3, 115.0, 79.6, 61.6, 51.5, 38.1, 32.0, 28.3. MS (ESI⁺): m/z = 327.1 [M,H]⁺ found; C₁₆H₂₄FN₂O₄ calculated m/z = 327.2[M,H]⁺.

4.2.6. tert-butyl (2S)-2-[methoxy(methyl)carbamoyl)piperidine-1-carboxylate (10a). According to general method 1, **10a** was obtained as a yellow oil (0.986 g, 83%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 5.18-4.87 (m, 1H); 4.10-3.84 (m, 1H); 3.76 (s, 3H); 3.58-3.39 (m, 1H); 3.18 (s, 3H); 2.01 (d, *J* = 15.0 Hz, 1H); 1.75-1.57 (m, 3H); 1.44 (s, 9H); 1.34-1.20 (m, 2H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 173.5, 156.2, 79.7, 61.3, 50.8, 42.3, 32.2, 29.7, 28.5, 26.5, 19.6. MS (ESI⁺): m/z = 273.1 [M,H]⁺ found; C₁₃H₂₅N₂O₄ calculated m/z = 273.2[M,H]⁺.

4.2.7. tert-butyl N-[(1S)-2-(methoxy(methyl)amino)-1-methyl-2-oxo-ethyl]carbamate (11a). According to general method 1, **11a** was obtained as a yellow oil (0.566 g, 64%). Analyses similar to literature description.⁵¹

4.2.8. tert-butyl N-[(1S)-1-[(4-tert-butoxyphenyl)methyl]-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (12a). According to general method 1, **12a** was obtained as a yellow solid (0.722 g, 76%). Analyses similar to literature description.⁵²

4.2.9. tert-butyl N-[(1S)-2-[methoxy(methyl)amino]-1-methyl-2-oxo-ethyl]carbamate (13a). According to general method 1, **13a** was obtained as a yellow solid (0.225 g, 48%). Analyses similar to literature description.⁵³

**4.3. Preparation of amine compounds 2b-13b.**

4.3.1. General method 2. To a solution of Weinreb amide (1 eq.) in THF was added LiAlH₄ (0.9 eq. of commercial solution 1M in THF) dropwise under N₂ at 0°C and stirred for 1 h at 0°C. Then saturated KHSO₄ solution was added dropwise. THF was evaporated off. The resulting crude product was dissolved in CH₂Cl₂, washed twice with saturated NaHCO₃ solution, twice with 1M HCl, once with brine. Organic layer was dried over MgSO₄, filtered and concentrated in vacuo. No further purification was performed. To a solution of resulting aldehyde (1 eq) in CH₂Cl₂, benzylamine (1.2 eq) was added under N₂. The mixture was stirred at room temperature for 30 min, NaBH(OAc)₃ (3 eq) was added portion wise (at least for 10 min). The mixture was stirred overnight, diluted with CH₂Cl₂ (3 mL) and saturated NaHCO₃
solution (3 mL) was added. The mixture was stirred for 10 min. The organic layer was separated and the aqueous phase washed twice with CH₂Cl₂ (15 mL). Combined organic layers were dried over MgSO₄, filtered and evaporated in vacuo. Purification via flash chromatography was performed.

4.3.2. *tert-butyl (3S)-3-[(benzylamino)methyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate (2b).* According to general method 2, **2b** was obtained as a colorless oil (0.55g, 95%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.36-7.21 (m, 5H); 7.20-7.03 (m, 4H); 5.53 (s, 1H); 4.92-4.45 (m, 2H); 4.22 and 4.17 (2s, 1H); 3.85 (dd, *J* = 19.5 Hz and 13.4 Hz, 2H); 2.93 [AB part of ABX system (dd, *J* = 18.0 and 6.0 Hz, 1H) and (dd, *J* = 15.0 and 3.0 Hz, 1H), Δν = 90 Hz]; 2.72-2.58 (unresolved AB part of ABX system, m, 2H); 1.49 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 155.4, 140.5, 132.9, 129.1, 128.4, 128.0, 126.9, 126.6, 126.1, 79.9, 53.5, 50.1, 50.1, 48.3, 43.1, 31.1, 28.5. MS (ESI⁺): m/z = 353.3 [M,H]⁺ found; C₂₂H₂₉N₂O₂ calculated m/z = 353.2 [M,H]⁺.

4.3.3. *tert-butyl (3S)-3-[[2-(2-pyridyl)ethylamino]methyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate (3b).* According to general method 2, **3b** was obtained as a light brown oil (0.386g, quantitative). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 8.53 (s, 1H); 7.69-6.78 (m, 8H); 5.01-4.63 (unresolved AB system, m, 2H); 4.39-4.13 (m, 1H); 3.32-2.64 (m, 8H); 1.45 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 159.4, 136.8, 132.6, 132.0, 129.1, 126.7, 126.38, 123.5, 121.6, 80.5, 49.3, 48.6, 47.8, 31.4, 29.0, 28.6, 28.1, 27.7. MS (ESI⁺): m/z = 368.2 [M,H]⁺ found; C₂₂H₃₀N₃O₂ calculated m/z = 368.2 [M,H]⁺.

4.3.4. *tert-butyl (3S)-3-[(2-morpholinoethylamino)methyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate(4b).* According to general method 2, **4b** was obtained as a colorless oil (0.428 g, 99%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.22-7.08 (m, 4H); 4.83-4.49 (unresolved AB system, m, 2H); 4.25 [X part of ABX system (d, *J* = 18.0 Hz, 1H, CH(b)]; 3.70 (ft, *J* = 3.0 Hz, 4H); 2.92 [AB part of ABX system (dd, *J* = 16.4 and 6.7 Hz, 1H) and (d, *J* = 12.0 Hz, 1H), Δν = 96 Hz]; 2.75-2.68 (m, 2H); 2.44-2.42 (m, 8H); 1.51 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 155.3, 133.1, 129.1, 126.6, 126.2, 79.9, 67.0, 58.3, 53.7, 53.4, 50.8, 45.9, 42.7, 31.1, 28.5. MS (ESI⁺): m/z = 376.1 [M,H]⁺ found; C₂₁H₃₄N₃O₃ calculated m/z = 376.2 [M,H]⁺.

4.3.5. *tert-butyl (3S)-3-[(4-phenylbutylamino)methyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate (5b).* According to general method 2, **5b** was obtained as a colorless oil (0.416 g, 92%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 8.19 (br s, 1H); 7.30-7.10 (m, 9H); 4.65 (m, 2H); 4.30 [X part of ABX system (d, *J* = 15.0 Hz, 1H]; 3.12-2.59 (m, 8H); 1.70-1.52 (m, 4H); 1.49 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 156.0, 154.9, 141.8, 132.8, 132.4, 128.9, 128.3, 127.0, 126.6, 126.1, 125.8, 80.8, 50.4, 49.3, 48.1, 47.8, 43.9, 42.7, 35.4, 31.7,
28.4, 27.6, 27.0, 22.7. MS (ESI⁺): m/z = 395.2 [M,H]⁺ found; C₂₅H₃₅N₂O₂ calculated m/z = 395.3 [M,H]⁺.

4.3.6. tert-butyl (3S)-3-[(hexylamino)methyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate (6b). According to general method 2, 6b was obtained as a colorless oil (0.386 g, 97%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.23-7.10 (m, 4H); 4.88-4.13 (m, 2H); 4.37-4.15 (m, 1H); 3.09 (AB part of ABX system, dd, *J* = 16.2 Hz and 6.9 Hz, 1H); 2.85-2.67 (m, 5H); 2.04 (s, 3H); 1.50 (s, 9H); 1.33-1.23 (m, 6H); 0.87 (t, *J* = 7.5 Hz, 3H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 170.0, 133.0, 132.7, 128.9, 126.9, 126.4, 126.0, 80.5, 50.9, 50.4, 48.3, 39.7, 31.5, 29.6, 28.7, 28.2, 28.0, 22.5, 13.9. MS (ESI⁺): m/z = 347.2 [M,H]⁺ found; C₂₁H₃₅N₂O₂ calculated m/z = 347.3 [M,H]⁺.

4.3.7. tert-butyl N-[(1S)-1-benzyl-2-(benzylamino)ethyl]carbamate (7b). According to general method 2, 7b was obtained as a brown solid (0.498 g, 90%). Analyses similar to literature description.⁵⁴

4.3.8. tert-butyl N-[(1S)-1-benzyl-2-(benzylamino)ethyl]-N-methyl-carbamate (8b). According to general method 2, 8b was obtained as a light yellow oil (0.120 g, 52%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): indicates the presence of 2 conformers. 7.44-7.05 (m, 10H); 4.71-4.34 (unresolved X part of ABX system, m, 1H); 3.89 and 3.71 [AB part of ABX system (d, *J* = 13.1 Hz, 1H) and (m, 1H)]; 2.93-2.55 (m, 7H); 2.00 (br s, 1H); 1.38 and 1.28 (2s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 156.3, 138.6, 129.0, 128.4, 128.2, 127.1, 126.6, 79.6, 55.6, 53.2, 50.1, 49.6, 36.9, 28.2. MS (ESI⁺): m/z = 355.1 [M,H]⁺ found; C₂₂H₃₀N₂O₂ calculated m/z = 355.2 [M,H]⁺.

4.3.9. tert-butyl N-[(1S)-1-[(benzylamino)methyl]-2-(4-fluorophenyl)ethyl]carbamate (9b). According to general method 2, 9b was obtained as a light yellow oil (0.510 g, 70%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.38-7.22 (m, 5H); 7.16-7.08 (m, 2H); 7.01-6.89 (m, 2H); 4.74 (br s, 1H), 3.99-3.85 (unresolved X part of ABX system, m, 1H); 3.77 (AB system, *J* = 12.4 Hz, Δν = 24 Hz, 2H); 2.90-2.71 (unresolved AB part of ABX system, m, 2H); 2.71-2.54 (unresolved AB part of ABX system, m, 2H); 2.10(br s, 1H); 1.41 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 163.2, 160.0 (*J*<sub>C-F</sub>= 242.2 Hz); 155.6, 139.9, 133.8, 130.8, 130.7, 128.5, 128.2, 127.1, 115.3, 115.0, 79.4, 53.7, 51.4, 51.2, 38.3, 28.4. MS (ESI⁺): m/z = 359.0 [M,H]⁺ found; C₂₁H₂₈FN₂O₂ calculated m/z = 359.2 [M,H]⁺.

4.3.10. tert-butyl (2S)-2-[(benzylamino)methyl]piperidine-1-carboxylate (10b). According to general method 2, 10b was obtained as a yellow oil (0.453 g, 62%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.37-7.20 (m, 5H); 4.45-4.32 (unresolved X part of ABX system, m, 1H); 4.02-3.87 (m, 1H); 3.83 (AB system, d, *J* = 12.0 Hz, Δν = 24.2 Hz, 1H); 2.79 [AB part of
ABX system(dd, J = 12.3 and 3.1 Hz, 1H) and (dd, J = 12.0 and 6.6 Hz, 1H), Δν = 81.2 Hz); 2.73-2.69 (m, 1H); 1.76-1.53 (m, 6H); 1.44 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 155.1, 140.4, 128.3, 128.0, 126.9, 124.8, 79.4, 53.4, 49.9, 48.0, 39.3, 28.5, 26.7, 25.4, 19.3. MS (ESI⁺): m/z = 305.0 [M,H]⁺ found; C₁₈H₂₈N₂O₂ calculated m/z = 305.2 [M,H]⁺.

4.3.11. *tert-butyl N-[2-(benzylamino)-1-methyl-ethyl]carbamate (11b).* According to general method 2, **11b** was obtained as a yellow oil (0.286 g, 19%). Analyses similar to literature description.⁵⁵

4.3.12. *tert-butyl N-[(1S)-1-[(benzylamino)methyl]-2-(4-tert-butoxyphenyl)ethyl]carbamate (12b).* According to general method 2, **12b** was obtained as a yellow oil (0.252 g, 58%).¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.33-7.20 (m, 5H); 7.14-6.98 (m, 2H); 6.91-6.82 (m, 2H); 4.98 (br s, 1H); 3.94-3.84 (unresolved X part of ABX system, m, 1H); 3.77 (AB system, J = 13.1 Hz, Δν = 26.8 Hz, 2H); 3.12-2.91 (unresolved AB part of ABX system, m, 1H); 2.90-2.69 (unresolved AB part of ABX system, m, 3H); 1.41 and 1.32 (2s, 19H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 155.8, 153.7, 139.0, 132.8, 129.7, 128.5, 127.2, 124.1, 79.2, 78.2, 53.2, 51.0, 38.5, 28.8, 28.4. MS (ESI⁺): m/z = 413.1 [M,H]⁺ found; C₂₅H₃₇N₂O₃ calculated m/z = 413.3 [M,H]⁺.

4.3.13. *tert-butyl N-[(1S)-3-(benzylamino)-1-[(4-methoxyphenyl)methyl]-2-oxopropyl]carbamate (13b).* According to general method 2, **13b** was obtained as a yellow oil (0.032 g, 22%). ¹H NMR (CDCl₃, 300 MHz), δ (ppm): 7.31-7.15 (m, 5H); 7.09 (d, J = 8.6 Hz, 2H); 6.82 (d, J = 9.5 Hz, 2H); 4.74 (br s, 1H); 3.89-3.78 (unresolved X part of ABX system, m, 1H); 3.77-3.68 (m, 5H); 2.80-2.65 (unresolved AB part of ABX system, m, 2H); 2.63-2.57 (unresolved AB part of ABX system, m, 2H); 1.75-1.66 (m, 1H, NH); 1.43 (s, 9H). ¹³C NMR (CDCl₃, 75 MHz), δ (ppm): 158.2, 155.7, 140.2, 130.3, 130.1, 128.4, 128.2, 127.0, 113.8, 79.8, 55.2, 53.8, 51.33, 42.9, 38.2, 28.4. MS (ESI⁺): m/z = 371.2 [M,H]⁺ found; C₂₂H₃₁N₂O₃ calculated m/z = 371.2 [M,H]⁺.

**4.4. Preparation of guanidine compounds 2-13.**

4.4.1. *General method 3.* To a solution of appropriate secondary amine (1 eq) in EtOH (1.6 mL/mmol of amine), NaHCO₃ (3 eq) and BrCN (1.1 eq) were added. The mixture was stirred at room temperature for 5 h. The mixture was diluted with H₂O and extracted twice with CH₂Cl₂. Combined organic layers were dried under MgSO₄. The solution was filtered and evaporated *in vacuo*. No further purification was performed unless specified. The nitrile compound (1 eq) was dissolved in 3M HCl/dioxane (0.08 M) and stirred at room temperature
overnight. Dioxane was evaporated *in vacuo*. Purification *via* flash chromatography was
performed.

4.4.2. (10aS)-2-benzyl-1,5,10,10a-tetrahydroimidazo[1,5-b]isoquinolin-3-imine monohydrochloride (2). According to general method 2, 2 was obtained (chromatography, CH₂Cl₂/methanol 94/6) as a white powder (0.13g, 46%). ¹H NMR (CD₃OD, 300 MHz), δ (ppm): 7.41-7.36 (m, 5H); 7.26-7.18 (m, 4H); 4.72-4.54 (m, 4H); 4.11-4.01 (m, 1H); 3.61 [AB part of ABX system (t, *J* = 9.0 Hz, 1H) and (t, *J* = 6.0 Hz, 1H), Δυ = 144 Hz]; 2.95 [AB part of ABX system (dd, *J* = 18.0 and 9.0 Hz, 1H) and (dd, *J* = 15.0 and 9.0 Hz, 1H), Δυ = 72 Hz]. ¹³C NMR (CD₃OD, 75 MHz), δ (ppm): 156.7, 134.4, 132.0, 129.9, 129.1, 128.8, 128.1, 127.7, 127.0, 126.8, 126.2, 53.2, 51.7, 48.5, 43.9, 33.4. HRMS (ESI⁺): m/z = 278.16548 [M,H]⁺ found; C₁₈H₁₉N₃ calculated m/z = 278.16517 [M,H]⁺. Mp: 198.5°C. αD (20°C, CH₃OH) = -0.548° (0.500 mg/mL). IR, ν (cm⁻¹): 3089, 1660 (C=N). HPLC: C₄ column: tR = 11.7 min, purity > 99% C₁₈ column: tR = 20.5 min, purity > 99%.

4.4.3. (10aS)-2-[2-(2-pyridyl)ethyl]-1,5,10,10a-tetrahydroimidazo[1,5-b]isoquinolin-3-imine dihydrochloride (3). According to general method 2, 3 was obtained (chromatography, CH₂Cl₂/methanol 90/10) as a white powder (0.10 g, 67%). ¹H NMR (CD₃OD, 300 MHz), δ (ppm): 8.84 (d, *J* = 6.0 Hz, 1H); 8.64 (t, *J* = 9.0 Hz, 1H); 8.23 (d, *J* = 6.0 Hz, 1H); 8.04 (t, *J* = 9.0 Hz, 1H); 7.23 (s, 4H); 4.67 (AB system, *J* = 15.0 Hz, Δυ = 75 Hz, 2H]; 4.15-3.91 (m, 4H); 3.63 [AB part of ABX system (t, *J* = 9.0 Hz, 1H) Δυ = 114 Hz]; 3.56 (t, *J* = 8.0 Hz, 2H); 2.99 [AB part of ABX system (d, *J* = 15.0 Hz, 1H) and (t, *J* = 9.0 Hz, 1H), Δυ = 51 Hz]. ¹³C NMR (CD₃OD, 75 MHz), δ (ppm): 156.6, 152.9, 147.1, 141.4, 131.9, 129.7, 129.1, 128.1, 127.0, 126.8, 126.1, 125.7, 53.4, 52.6, 43.8, 44.2, 33.4. HRMS (ESI⁺): m/z = 293.17595 [M,H]⁺ found; C₁₈H₁₉N₄ calculated m/z = 293.17607 [M,H]⁺. Mp: 123.8°C. αD (20°C, CH₃OH) = -0.449° (0.500 mg/mL). IR, ν (cm⁻¹): 3039, 1668 (C=N). HPLC: C₄ column: tR = 6.7 min, purity > 99% C₁₈ column: tR = 12.9 min, purity 98%.

4.4.4. (10aS)-2-(2-morpholinoethyl)-1,5,10,10a-tetrahydroimidazo[1,5-b]isoquinolin-3-imine dihydrochloride (4). According to general method 2, 4 was obtained (chromatography, CH₂Cl₂/methanol 90/10) as a brown oil (0.282 g, 69%). ¹H NMR (CD₃OD, 300 MHz), δ (ppm): 7.34-7.18 (m, 4H); 4.66 (AB system, *J* = 16.0 Hz, Δυ = 70 Hz, 2H); 4.17-3.99 (m, 1H); 3.71 (t, *J* = 4.6 Hz, 4H); 3.67-3.48 (m, 4H); 3.03 [AB part of ABX system (dd, *J* = 15.0 and 3.7 Hz, 1H) and (dd, *J* = 15.4 and 10.3 Hz, 1H), Δυ = 51 Hz]; 2.69 (t, *J* = 5.8 Hz, 2H); 2.60 (t, *J* = 4.5 Hz, 4H). ¹³C NMR (CD₃OD, 75 MHz), δ (ppm): 157.4, 132.0, 129.9, 129.1, 127.0, 126.8, 126.0, 66.4, 55.7, 53.5, 53.2, 43.7, 42.3, 30.7. HRMS (ESI⁺): m/z = 301.20265
[M,H]$^+$ found; C$_{17}$H$_{25}$N$_4$O calculated m/z = 301.20229 [M,H]$^+$. HPLC: C$_4$ column: t$_R$ = 12.3 min, purity > 94% C$_{18}$ column: t$_R$ = 4.7 min, purity 97%.

4.4.5. (10aS)-2-(4-phenylbutyl)-1,5,10,10a-tetrahydroimidazo[1,5-b]isoquinolin-3-imine hydrochloride (5). According to general method 2, 5 was obtained (chromatography, CH$_2$Cl$_2$/methanol 94/6) as a light brown oil (0.206 g, 82%). $^1$H NMR (CD$_3$OD, 300 MHz), $\delta$ (ppm): 7.29-7.13 (m, 9H); 4.62 ([AB system (dd, J = 18.0 Hz, $\Delta$ν = 81 Hz, 2H); 4.03-3.97 [unresolved X part of ABX system (m, 1H)]; 3.87 and 3.46-3.32 [AB part of ABX system (t, J = 9.0 Hz, 1H) and (m, 1H), $\Delta$ν = 144 Hz]]; 3.46-3.39 (m, 2H); 2.95 [AB part of ABX system (dd, J = 15.0 and 3.0 Hz, 1H) and (dd, J = 15.0 and 9.0 Hz, 1H), $\Delta$ν = 72 Hz]]; 2.69 (t, J = 9.0 Hz, 2H); 1.83-1.60 (m, 4H). $^{13}$C NMR (CD$_3$OD, 75 MHz), $\delta$ (ppm): 156.6, 141.8, 132.0, 129.9, 129.7, 128.8, 128.0, 127.3, 126.9, 126.8, 126.1, 125.5, 124.8, 53.1, 51.9, 45.4, 43.3, 35.0, 33.3, 28.0, 26.1. HRMS (ESI$^+$): m/z = 320.21259 [M,H]$^+$ found; C$_{21}$H$_{26}$N$_3$ calculated m/z = 320.21212 [M,H]$^+$. $\alpha_D$ (20°C, CH$_3$OH) = -0.499° (0.500 mg/mL). IR, ν (cm$^{-1}$): 2929, 1664 (C=N). HPLC: C$_4$ column: t$_R$ = 12.7 min, purity 98% C$_{18}$ column: t$_R$ = 23.7 min, purity 98%.

4.4.6. tert-butyl (3S)-3-[(hexylamino)methyl]-3,4-dihydro-1H-isoquinoline-2-carboxylate (6). According to general method 2, 6 was obtained (chromatography, CH$_2$Cl$_2$/methanol 85/15) as an orange oil (228 mg, 88%). $^1$H NMR (CD$_3$OD, 300 MHz), $\delta$ (ppm): 7.29-7.14 (m, 4H); 4.68 (AB system, J = 15.7 Hz, $\Delta$ν = 90.6 Hz, 2H); 4.03-3.94 (m, 2H); 3.64-3.32 (m, 3H); 2.97 [AB part of ABX system (dd, J = 15.5 and 2.7 Hz, 1H) and (dd, J = 15.5 and 10.0 Hz, 1H), $\Delta$ν = 87.7 Hz]; 1.66 (quin, J = 6.9 Hz, 2H); 1.49-1.31(m, 6H); 0.93 (t, J = 6.2 Hz, 3H). $^{13}$C NMR (CD$_3$OD, 75 MHz), $\delta$ (ppm): 156.6, 132.1, 130.1, 129.1, 127.0, 126.8, 126.2, 53.2, 52.0, 44.9, 43.9, 33.4, 31.3, 26.6, 25.9, 22.3, 13.1. HRMS (ESI$^+$): m/z = 272.21249 [M,H]$^+$ found; C$_{17}$H$_{26}$N$_3$ calculated m/z = 272.21212 [M,H]$^+$. HPLC: C$_4$ column: t$_R$ = 3.3 min, purity 98% C$_{18}$ column: t$_R$ = 26.9 min, purity 97%.

4.4.7. (4S)-1,4-dibenzylimidazolidin-2-imine hydrochloride (7). According to general method 2, 7 was obtained (chromatography, CH$_2$Cl$_2$/methanol 95/5) as a white powder (19 mg, 27%). $^1$H NMR (CD$_3$OD, 300 MHz), $\delta$ (ppm): 7.38-7.22 (m, 6H); 7.21-7.10 (m, 4H); 4.35 (dd, J = 28.9 and 13.5 Hz, 2H); 4.29-4.21 (unresolved X part of ABX system, m, 1H); 3.61 and 3.33-3.27 [AB part of ABX system (ft, J = 9.6 Hz, 1H) and (m, 1H), $\Delta$ν = 93 Hz]; 2.87 (d, J = 5.9 Hz, 2H). $^{13}$C NMR (CD$_3$OD, 75 MHz), $\delta$ (ppm): 157.9, 135.7, 134.2, 129.2, 128.8, 128.4, 127.9, 127.4, 126.7, 53.8, 51.1, 48.5, 39.9. HRMS (ESI$^+$): m/z = 266.16529 [M,H]$^+$ found; C$_{17}$H$_{20}$N$_3$ calculated m/z = 266.16517 [M,H]$^+$. Mp: 196.1°C. $\alpha_D$ (20°C, CH$_3$OH) = -0.006°
(0.500 mg/mL). IR, ν (cm⁻¹): 3060, 1668 (C=N). HPLC: C₄ column: tR = 13.1 min, purity > 99% C₁₈ column: tR = 24.7 min, purity > 99%.

4.4.8. **(4S)-1,4-dibenzyl-3-methyl-imidazolidin-2-imine hydrochloride (8).** According to general method 2, **8** was obtained (chromatography, CH₂Cl₂/methanol 95/5) a white powder (14 mg, 67%). ¹H NMR (CD₃OD, 300 MHz), δ (ppm): 7.39-7.23 (m, 6H); 7.17-7.04 (m, 4H); 4.39 (AB system, *J* = 15.7 Hz, Δυ = 38.9 Hz, 2H); 4.23-4.11 (unresolved X part of ABX system, m, 1H); 3.37 [AB part of ABX system (*t*, *J* = 9.7 Hz, 1H) and (dd, *J* = 9.9 and 5.7 Hz, 1H), Δυ = 91.7 Hz]; 3.10 (s, 3H); 2.94 [AB part of ABX system (dd, *J* = 13.9 and 4.4 Hz, 1H) and (dd, *J* = 7.1 and 6.8 Hz, 1H), Δυ = 61.9 Hz]. ¹³C NMR (CD₃OD, 75 MHz), δ (ppm): 157.4, 135.3, 134.2, 129.2, 128.7, 128.5, 127.9, 127.3, 126.9, 59.8, 49.4, 48.0 (CH₂(d)); 36.8 (CH₂(a)); 30.2 (CH₃(l)). HRMS (ESI⁺): m/z = 280.18080 [M,H]⁺ found; C₁₈H₂₂N₃ calculated m/z = 280.18082 [M,H]⁺. Mp: 244.5°C. αD (20°C, CH₃OH) = -0.098° (0.500 mg/mL). IR, ν (cm⁻¹): 3024, 1664 (C=N). HPLC: C₄ column: tR = 10.1 min, purity > 99% C₁₈ column: tR = 4.5 min, purity > 99%.

4.4.9. **(4S)-1-benzyl-4-[(4-fluorophenyl)methyl]imidazolidin-2-imine hydrochloride (9).** According to general method 2, **9** was obtained (chromatography, CH₂Cl₂/methanol 90/10) a white powder (217 mg, 90%). ¹H NMR (CD₃OD, 300 MHz), δ (ppm): 7.41-7.28 (m, 3H); 7.26-7.17 (m, 2H); 7.17-7.09 (m, 2H); 7.06-6.95 (m, 2H); 4.44 [AB system, *J* = 16.1Hz, Δυ = 41 Hz, 2H]; 4.32-4.19 (unresolved X part of ABX system, m, 1H); 3.62 and 3.34-3.24 [unresolved AB part of ABX system(*t*, *J* = 11.2 Hz, 1H) and (m, 1H), Δυ = 108 Hz]; 2.94-2.77 (unresolved AB part of ABX system, m, 2H). ¹³C NMR (CD₃OD, 75 MHz), δ (ppm): 163.7, 160.4 (Jc-F= 242.1 Hz); 157.9, 134.2, 131.6,131.1, 131.0, 128.7; 127.9, 127.5, 115.2, 114.9, 53.7, 50.9, 48.5, 38.8. HRMS (ESI⁺): m/z = 284.15576 [M,H]⁺ found; C₁₇H₁₉FN₃ calculated m/z = 284.15575 [M,H]⁺. Mp: 244.5°C. αD (20°C, CH₃OH) = -0.072° (0.500 mg/mL). HPLC: C₄ column: tR = 11.0 min, purity > 99% C₁₈ column: tR = 19.5 min, purity > 99%.

4.4.10. **(8aS)-2-benzyl-1,5,6,7,8,8a-hexahydroimidazo[1,5-a]pyridin-3-imine hydrochloride (10).** According to general method 2, **10** was obtained (chromatography, CH₂Cl₂/methanol 90/10) a white powder (130 mg, 29%). ¹H NMR (CD₃OD, 300 MHz), δ (ppm): 7.47-7.28 (m, 5H); 4.59 (AB system, *d*, *J* = 15.1 Hz, Δυ = 18.7 Hz, 1H]; 3.92-3.88 (m, 1H); 3.51 [unresolved AB part of ABX system(dd, *J* = 12.3 and 9.0 Hz, 1H) and (dd, *J* = 15.3 and 9.2 Hz, 1H), Δυ = 246 Hz]; 3.87-3.73 (m, 1H); 3.08 (dd, *J* = 9.0 and 3.1 Hz, 1H); 1.91-1.77 (m, 2H); 1.58-1.30 (m, 4H). ¹³C NMR (CD₃OD, 75 MHz), δ (ppm): 156.0, 134.4, 128.7, 128.0; 127.5, 59.5, 51.5, 47.8, 42.3, 29.7, 24.0, 22.2. HRMS (ESI⁺): m/z = 230.16490 [M,H]⁺ found;
$C_{14}H_{20}N_3$ calculated m/z = 230.16517 [M,H]$^+$. Mp: 188.2°C. $\alpha_D$ (20°C, $CH_3OH$) = +10.6° (0.500 mg/mL). IR, v (cm$^{-1}$): 2941, 1660 (C=N). HPLC: $C_4$ column: $t_R$ = 9.0 min, purity 98% $C_{18}$ column: $t_R$ = 16.6 min, purity > 99%.

**4.4.11. (4S)-1-benzyl-4-methyl-imidazolidin-2-imine hydrochloride (11).** According to general method 2, **11** was obtained (chromatography, $CH_2Cl_2$/methanol 90/10) a white powder (42 mg, 53%). $^1$H NMR ($CD_3OD$, 300 MHz), $\delta$ (ppm): 7.48-7.27 (m, 5H); 4.59 (br s, 1H); 4.42 (br s, 2H); 4.12-3.99 (unresolved X part of ABX system, m, 1H); 3.71 and 3.14 [AB part of ABX system (dd, $J$ = 18.9 and 9.5 Hz, 1H) and (dd, $J$ = 9.6 and 6.7 Hz, 1H) $\Delta\nu$ = 179.9 Hz); 1.25 (d, $J$ = 6.0 Hz, 3H). $^{13}$C NMR ($CD_3OD$, 75 MHz), $\delta$ (ppm): 158.0, 134.5, 128.8, 127.9, 127.5, 53.6, 48.9, 19.5. HRMS (ESI$^+$): m/z = 190.13416 [M,H]$^+$ found; $C_{11}H_{16}N_3$ calculated m/z = 190.13387 [M,H]$^+$. Mp: 194.9°C. $\alpha_D$ (20°C, $CH_3OH$) = -0.045° (0.500 mg/mL). IR, v (cm$^{-1}$): 3055, 1667 (C=N). HPLC: $C_4$ column: $t_R$ = 7.3 min, purity 98% $C_{18}$ column: $t_R$ = 8.0 min, purity > 99%.

**4.4.12. 4-[[**(4S)-1-benzyl-2-imino-imidazolidin-4-yl]methyl]phenol hydrochloride (12).** According to general method 2, **12** was obtained (chromatography, $CH_2Cl_2$/methanol 85/15) a brown solid (53 mg, 73%). $^1$H NMR ($CD_3OD$, 300 MHz), $\delta$ (ppm): 7.33 (m, 3H); 7.06 (m, 2H); 6.98 (d, $J$ = 8.5 Hz, 2H); 6.72 (d, $J$ = 8.5 Hz, 2H); 4.45 (AB system, $J$ = 15.8 Hz, 2H, $\Delta\nu$ = 42.9 Hz); 4.20 (unresolved X part of ABX system, m, 1H); 3.59 and 3.33-3.27 [AB part of ABX system (t, $J$ = 9.7 Hz, 1H) and (m, 1H), $\Delta\nu$ = 259.7 Hz]; 2.82-2.66 (unresolved AB part of ABX system, m, 2H). $^{13}$C NMR ($CD_3OD$, 75 MHz), $\delta$ (ppm): 158.0, 156.3, 134.2, 130.3, 128.7, 127.8, 127.2, 126.0, 115.1, 53.9, 50.9, 47.9, 38.9. HRMS (ESI$^+$): m/z = 282.15985 [M,H]$^+$ found; $C_{17}H_{20}N_3O$ calculated m/z = 282.16009 [M,H]$^+$. Mp: °C. $\alpha_D$ (20°C, $CH_3OH$) = -0.014° (0.500 mg/mL). IR, v (cm$^{-1}$): 3064, 1670 (C=N). HPLC: $C_4$ column: $t_R$ = 10.2 min, purity 99% $C_{18}$ column: $t_R$ = 15.8 min, purity > 99%.

**4.4.13. (4S)-1-benzyl-4-[(4-methoxyphenyl)methyl]imidazolidin-2-imine hydrochloride (13).** According to general method 2, **13** was obtained (chromatography, $CH_2Cl_2$/methanol 90/10) a brown solid (14 mg, 67%). $^1$H NMR ($CD_3OD$, 300 MHz), $\delta$ (ppm): 7.39-7.31 (m, 5H); 7.09 (d, $J$ = 8.6 Hz, 2H); 6.83 (d, $J$ = 6.6 Hz, 2H); 4.45 (AB system, 2H, $J$ = 15.8 Hz, $\Delta\nu$ = 37.3 Hz); 4.26-4.19 (unresolved X part of ABX system, m, 1H); 3.79 (s, 3H); 3.61 [AB part of ABX system (ft, $J$ = 9.7 Hz, 1H) and (m, 1H), $\Delta\nu$ = 90 Hz]]; 2.80 (unresolved AB part of ABX system, m, 2H). $^{13}$C NMR ($CD_3OD$, 75 MHz), $\delta$ (ppm): 158.9, 157.9, 134.2, 130.2, 128.7, 127.9, 127.4, 127.3, 113.8, 54.3, 53.9, 50.9, 48.4, 48.1, 47.9, 38.9. HRMS (ESI$^+$): m/z = 296.17555 [M,H]$^+$ found; $C_{17}H_{20}N_3O$ calculated m/z = 296.17574 [M,H]$^+$. Mp: °C. $\alpha_D$
(20°C, CH3OH) = -0.045° (0.500 mg/mL). IR, ν (cm⁻¹): 3064, 1670 (C=N). HPLC: C4 column: tR = 13.3 min, purity 99% C18 column: tR = 8.0 min, purity > 99%.

**4.5. In vitro testing.**

**4.5.1. Displacement binding assays.** Competition binding curves of the A₂A receptor antagonist [³H]-ZM24135 by the designed A₂A antagonists described above, were performed as before⁵⁶ in Human HEK293 A₂A R membranes (Perkin Elmer). 0.5 µL of membranes (0.5U of A₂A R) were incubated with [³H]-ZM24135 (2 nM) and increasing concentrations of the designed A₂A R antagonists (0 to 600 nM) in a final volume of 300 µL in the presence of 1U/mL of adenosine deaminase (Roche). All samples were assayed in duplicate. Non-specific binding was determined for each assay in the presence of the antagonist ZM-24135 (8.3nM). Microplates were incubated for 1 hour at room temperature and the reaction was stopped by vacuum filtration with a Skatron semi-automatic cell harvester with chilled incubation solution (pH 7.4, Tris 50mM MgCl20mM) to filtermats 1.5 µm (Molecular Devices). 3 mL of scintillation cocktail (OptiPhase ‘HiSafe’ 2, PerkinHelmer) were added and radioactivity bound to the filters was determined after 12 hours with an efficiency of 55-60% for 2 minutes. Displacement reference curves were performed with ZM-24135 (0 to 6nM in 6%, 40% or 60% of DMSO) and CGS241680 (0 to 6nM).

**4.5.2. Cell culture and cytotoxicity assay.** The human neuroblastoma cell line (SY5Y) was cultured in DMEM (Dulbecco's Modified Eagle Medium) (Gibco) supplemented with 2 mM L-glutamine, 100 µg/ml streptomycin, 100 IU/mL penicillin, 1 mM non-essential amino acids and 10% (v/v) heat-inactivated foetal bovine serum (Sigma Aldrich), and grown at 37°C in a humidified incubator with 5% CO₂.

Cells were seeded at 2,000 cells per well onto 96-well plates in DMEM medium. Cells were starved for 24 hours to obtain synchronous cultures, and were then incubated in culture medium that contained various concentrations of test compounds, each dissolved in less than 0.1% DMSO. After 72 hours of incubation, cell growth was estimated by the colorimetric MTT (thiazolyl blue tetrazolium bromide) assay.

**Acknowledgments**

We express our thanks to Mariam Lamarti, Phillip James Brennan and Alice Cao for their
contribution in organic synthesis and Amélie Barczyk for some *in vitro* cytotoxicity
experiments. The 300 MHz NMR facilities were funded by the Région Nord-Pas de Calais
(France), the Ministère de la Jeunesse, de l'Éducation Nationale et de la Recherche (MJENR)
and the Fonds Européens de Développement Régional (FEDER).

This work was supported by Lille 2 University, ANR « Adoratau », PRES Univ Lille Nord de France. Valeria Moas Heloire is the recipient of a fellowship from Lille 2 University.

References and notes

1. Welsch, M. E.; Snyder, S. A.; Stockwell, B. R. *Curr. Opin. Chem. Biol.* **2010**, *14*, 347.

2. In *Scaffold Hopping in Medicinal Chemistry*; Brown, N., Ed.; Wiley-VCH Verlag GmbH & Co: 2014, p 1.

3. Evans, B. E.; Rittle, K. E.; Bock, M. G.; DiPardo, R. M.; Freidinger, R. M.; Whitter, W. L.; Lundell, G. F.; Veber, D. F.; Anderson, P. S.; L., C. R. S.; Lotti, V. J.; Cerino, D. J.; Kling, P. J.; Kunkel, K. A.; Springer, J. P.; Hirshfield, J. *J. Med. Chem.* **1988**, *31*, 2235.

4. Oxombre, B.; Lee-Chang, C.; Duhamel, A.; Toussaint, M.; Giroux, M.; Donnier-Marechal, M.; Carato, P.; Lefranc, D.; Zephir, H.; Prin, L.; Melnyk, P.; Vermersch, P. *Br. J. Pharmacol.* **2015**, *172*, 1769.

5. Venna, V. R.; Deplancke, D.; Melnyk, P.; Bordet, R. *Fundam. Clin. Pharmacol.* **2008**, *22*, 1.

6. Charton, J.; Cazenave Gassiot, A.; Girault-Mizzi, S.; Debreu-Fontaine, M. A.; Melnyk, P.; Sergheraert, C. *Bioorg. Med. Chem. Lett.* **2005**, *15*, 4833.

7. Toussaint, M.; Mousset, D.; Foulon, C.; Jacquemard, U.; Vaccher, C.; Melnyk, P. *Eur. J. Med. Chem.* **2010**, *45*, 256.

8. Toussaint, M.; Delair, B.; Foulon, C.; Lempereur, N.; Vaccher, C.; Maurice, T.; Melnyk, P. *Eur. Neuropsychopharmacol.* **2009**, *19*, 504.

9. Cazenave Gassiot, A.; Charton, J.; Girault-Mizzi, S.; Gilleron, P.; Debreu-Fontaine, M. A.; Sergheraert, C.; Melnyk, P. *Bioorg. Med. Chem. Lett.* **2005**, *15*, 4828.

10. Ma, Y.; De, S.; Chen, C. *Tetrahedron* **2015**, *71*, 1145.

11. Berlinck, R. G.; Trindade-Silva, A. E.; Santos, M. F. *Nat. Prod. Rep.* **2012**, *29*, 1382.

12. Berlinck, R. G.; Burtoloso, A. C.; Trindade-Silva, A. E.; Romminger, S.; Morais, R. P.; Bandeira, K.; Mizuno, C. M. *Nat. Prod. Rep.* **2010**, *27*, 1871.

13. Berlinck, R. G.; Burtoloso, A. C.; Kossuga, M. H. *Nat. Prod. Rep.* **2008**, *25*, 919.

14. Verbeken, M.; Wynendaele, E.; Mauchauffee, E.; Bracke, N.; Stalmans, S.; Bojnik, E.; Benyhe, S.; Peremans, K.; Polis, I.; Burvenich, C.; Gjedde, A.; Hernandez, J. F.; De Spiegeleer, B. *Peptides* **2015**, *63C*, 10.

15. Anzini, M.; Chelini, A.; Mancini, A.; Cappelli, A.; Frosini, M.; Ricci, L.; Valoti, M.; Magistretti, J.; Castelli, L.; Giordani, A.; Makovec, F.; Vomero, S. *J. Med. Chem.* **2010**, *53*, 734.

16. de Lera Ruiz, M.; Lim, Y. H.; Zheng, J. *J. Med. Chem.* **2014**, *57*, 3623.

17. Rebola, N.; Rodrigues, R. J.; Lopes, L. V.; Richardson, P. J.; Oliveira, C. R.; Cunha, R. A. *Neuroscience* **2005**, *133*, 79.

18. Blum, D.; Hourez, R.; Galas, M. C.; Popoli, P.; Schiffmann, S. N. *Lancet Neurol.* **2003**, *2*, 366.

19. Chen, J. F.; Sonsalla, P. K.; Pedata, F.; Melani, A.; Domenici, M. R.; Popoli, P.; Geiger, J.; Lopes, L. V.; de Mendonca, A. *Prog. Neurobiol.* **2007**, *83*, 310.

20. Milne, G. R.; Palmer, T. M. *The Scientific World Journal* **2011**, *11*, 320.

21. Fredholm, B. B.; Chern, Y.; Franco, R.; Sitkovsky, M. V. *Prog. Neurobiol.* **2007**, *83*, 263.
22. Popoli, P.; Blum, D.; Martire, A.; Ledent, C.; Ceruti, S.; Abbracchio, M. P. *Prog Neurobiol* **2007**, *81*, 331.

23. Matos, M.; Augusto, E.; Santos-Rodrigues, A. D.; Schwarzschild, M. A.; Chen, J. F.; Cunha, R. A.; Agostinho, P. *Glia* **2012**, *60*, 702.

24. Ribeiro, J. A.; Sebastião, A. M. *Acta Physiol. 2010*, **199**, 161.

25. Orr, A. G.; Orr, A. L.; Li, X. J.; Gross, R. E.; Traynelis, S. F. *Nat. Neurosci.* **2009**, *12*, 872.

26. Gomes, C. V.; Kaster, M. P.; Tome, A. R.; Agostinho, P. M.; Cunha, R. A. *Biochim. Biophys. Acta* **2011**, *1808*, 1380.

27. Popoli, P.; Blum, D.; Domenici, M. R.; Burnouf, S.; Chern, Y. *Curr. Pharm. Des.* **2008**, *14*, 1500.

28. Preti, D.; Baraldi, P. G.; Moorman, A. R.; Borea, P. A.; Varani, K. *Med. Res. Rev.* **2015**, *0*, 1.

29. Jaakola, V. P.; Griffith, M. T.; Hanson, M. A.; Cherezov, V.; Chien, E. Y.; Lane, J. R.; Ijzerman, A. P.; Stevens, R. C. *Science* **2008**, *322*, 1211.

30. Katritch, V.; Jaakola, V. P.; Lane, J. R.; Lin, J.; Ijzerman, A. P.; Yeager, M.; Kufareva, I.; Stevens, R. C.; Abagyan, R. *J. Med. Chem.* **2010**, *53*, 1799.

31. Charton, J.; Gassiot, A. C.; Melnyk, P.; Girault-Mizzi, S.; Sergheraert, C. *Tetrahedron Lett.* **2004**, *45*, 7081.

32. Ishikawa, T. *Chem. Pharm. Bull.* **2010**, *58*, 1555.

33. Katritzky, A. R.; Rogovoy, B. V. *Arkivoc* **2005**, *iv*, 49.

34. Suhs, T.; Konig, B. *Mini-Rev. Org. Chem.* **2006**, *4*, 315.

35. Cabordery, A.-C.; Toussaint, M.; Azaroual, N.; Bonte, J.-P.; Melnyk, P.; Vaccher, C.; Foulon, C. *Tetrahedron: Asymmetry* **2011**, *22*, 125.

36. Ma, D.; Cheng, K. *Tetrahedron: Asymmetry* **1999**, *10*, 713.

37. Lovick, H. M.; Michael, F. E. *Tetrahedron Lett.* **2009**, *50*, 1016.

38. Han, X.; Michne, J. A.; Pin, S. S.; Burris, K. D.; Balanda, L. A.; Fung, L. K.; Fiedler, T.; Browman, K. E.; Taber, M. T.; Zhang, J.; Dubowchik, G. M. *Bioorg. Med. Chem. Lett.* **2005**, *15*, 3870.

39. Li, Y.; Liang, J.; Siu, T.; Hu, E.; Rossi, M. A.; Barnett, S. F.; Defeo-Jones, D.; Jones, R. E.; Robinson, R. G.; Leander, K.; Huber, H. E.; Mittal, S.; Cosford, N.; Prasit, P. *Bioorg. Med. Chem. Lett.* **2009**, *19*, 834.

40. Latli, B.; D'Amour, K.; Casida, E. *J. Med. Chem.* **1999**, *42*, 2227.

41. Ma, Z.; Day, C. S.; Bierbach, U. *J. Org. Chem.* **2007**, *72*, 5387.

42. Wu, Y. Q.; Hamilton, S. K.; Wilkinson, D. E.; Hamilton, G. S. *J. Org. Chem.* **2002**, *67*, 7553.

43. Kubota, H.; Nakamura, Y.; Higashijima, T.; Yamamoto, Y.; Oka, K.; Igarashi, S. 2005; Vol. WO2005095395 (A2).

44. Ho, J. Z.; Braun, M. P.; Subramanian, R.; Gao, Y. D.; Dean, D. C.; Melillo, D. G. *Helv. Chim. Acta* **2004**, *87*, 674.

45. Hutchinson, J. H.; Halczenko, W.; Brashear, K. M.; Breslin, M. J.; Coleman, P. J.; Duong, L. T.; Fernandez-Metzler, C.; Gentile, M. A.; Fisher, J. E.; Hartman, G. D.; Huff, J. R.; Kimmel, D. B.; Leu, C. T.; Meissner, R. S.; Merkle, K.; Nagy, R.; Pennypacker, B.; Perkins, J. J.; Prueksaritanont, T.; Rodan, G.; Varga, S. L.; Wesolowski, G. A.; Zartman, A. E.; Rodan, S. B.; Duggan, M. E. *J. Med. Chem.* **2003**, *46*, 4790.

46. Ryckebusch, A.; Deprez-Poulain, R.; Maes, L.; Debreu-Fontaine, M. A.; Mouray, E.; Grellier, P.; Sergheraert, C. *J. Med. Chem.* **2003**, *46*, 542.

47. Dolomanov, O. V.; Bourhis, L. J.; Gildea, R. J.; Howard, J. A. K.; Puschmann, H. *J. Appl. Crystallogr.* **2009**, *42*, 339.

48. Malkov, A. V.; Vrankova, K.; Cerny, M.; Kocovsky, P. *J. Org. Chem.* **2009**, *74*, 8425.
49. Xu, Z.; Cheng, F.; Da, C.; Liu, G.; Tang, Y. *J Mol Model* **2010**, *16*, 1867.

50. McHardy, T.; Caldwell, J. J.; Cheung, K. M.; Hunter, L. J.; Taylor, K.; Rowlands, M.; Ruddle, R.; Henley, A.; de Haven Brandon, A.; Valenti, M.; Davies, T. G.; Fazal, L.; Seavers, L.; Raynaud, F. I.; Eccles, S. A.; Aherne, G. W.; Garrett, M. D.; Collins, I. *J. Med. Chem.* **2010**, *53*, 2239.

51. Cheung, M.; Kuntz, K. W.; Pobanz, M.; Salovich, J. M.; Wilson, B. J.; Andrews, C. W., 3rd; Shewchuk, L. M.; Epperly, A. H.; Hassler, D. F.; Leesnitzer, M. A.; Smith, J. L.; Smith, G. K.; Lansing, T. J.; Mook, R. A., Jr. *Bioorg. Med. Chem. Lett.* **2008**, *18*, 6214.

52. Arnaud, O.; Koubeissi, A.; Ettouati, L.; Terreux, R.; Alame, G.; Grenot, C.; Dumontet, C.; Di Pietro, A.; Paris, J.; Falson, P. *J. Med. Chem.* **2010**, *53*, 6720.

53. Velmourougane, G.; Harbut, M. B.; Dalal, S.; McGowan, S.; Oellig, C. A.; Meinhardt, N.; Whisstock, J. C.; Klemba, M.; Greenbaum, D. C. *J. Med. Chem.* **2011**, *54*, 1655.

54. Sorto, N. A.; Painter, P. P.; Fettinger, J. C.; Tantillo, D. J.; Shaw, J. T. *Org. Lett.* **2013**, *15*, 2700.

55. Reginato, G.; Di Credico, B.; Andreotti, D.; Mingardi, A.; Paio, A.; Donati, D. *Tetrahedron: Asymmetry* **2007**, *18*, 2680.

56. Lopes, L. V.; Cunha, R. A.; Ribeiro, J. A. *J. Neurophysiol.* **1999**, *82*, 3196.
**Table 1.** <sup>a</sup>Yield of the cyclization step. <sup>b</sup>A<sub>2a</sub>R's agonist CGS-21680 used as a reference (Ki = 0.503 µM). Displacement of specific [<sup>3</sup>H]-ZM 241385 binding in membranes obtained from hA<sub>2a</sub> receptor stably expressed in HEK293 cells. <sup>c</sup>Cytotoxicity assays on SY5Y cells. <sup>c</sup> Ki > 450 µM.

**Figure 1.**

**Figure 2.** A<sub>2a</sub>R's crystal structure with (i) high-affinity antagonist ZM241385, (ii) Tic-H 1 and (iii) Tic-guanidine 2. Hydrogen bonds with Glu<sup>169</sup> and Asn<sup>253</sup> missing with Tic-H (ii) are being restored with the guanidine function (iii).

**Figure 3.** Pharmacomodulations of compounds **2-6** of the Tic-guanidine core using various amines in the reductive amination step.

**Figure 4.** Opening of the central Tic-guanidine core.

**Figure 5.** Thermal ellipsoid drawing<sup>47</sup> (30% probability) of the asymmetric unit of compound **2** (left) and **3** (right) showing the molecular structure and the labeling scheme.

**Figure 6.** Thermal ellipsoid drawing<sup>47</sup> (30% probability) of the asymmetric unit of compound **7-9** and **11** showing the molecular structure and the labeling scheme.

**Scheme 1.** i) HNMe(OMe).HCl, EDC, HOBt, NMM, CH<sub>3</sub>CN, CH<sub>2</sub>Cl<sub>2</sub>, 61%; ii) LiAlH<sub>4</sub>, THF 0°C then aq. KHSO<sub>4</sub>, 66%; iii) R-NH<sub>2</sub>, NaHB(OAc)<sub>3</sub>, CH<sub>2</sub>Cl<sub>2</sub> then aq. NaHCO<sub>3</sub>, 92%-quant; iv) BrCN, NaHCO<sub>3</sub>, EtOH, 54-97%; v) HCl, dioxane, 46-88%.

**Scheme 2.** i) HNMe(OMe).HCl, EDC, HOBt, NMM, CH<sub>3</sub>CN, CH<sub>2</sub>Cl<sub>2</sub>, 36-86%; ii) LiAlH<sub>4</sub>, THF 0°C then aq. KHSO<sub>4</sub>, 20-97%; iii) Bn-NH<sub>2</sub>, NaHB(OAc)<sub>3</sub>, CH<sub>2</sub>Cl<sub>2</sub> then aq. NaHCO<sub>3</sub>, 19-99%; iv) BrCN, NaHCO<sub>3</sub>, EtOH, 44-94%; v) HCl, dioxane, 23-90%.
<table><caption>Table 1.</caption><thead><tr><th>Entry</th><th>Cpd</th><th>Yield<sup>a</sup></th><th>Ki<br>(μM)<sup>b</sup></th><th>CC<sub>50</sub><br>(μM)<sup>c</sup></th></tr></thead><tbody><tr><td>1</td><td>1</td><td>/</td><td>44</td><td>&gt;100</td></tr><tr><td>2</td><td>2</td><td>46%</td><td>64</td><td>&gt;100</td></tr><tr><td>3</td><td>3</td><td>67%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>4</td><td>4</td><td>69%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>5</td><td>5</td><td>82%</td><td>35</td><td>&gt;100</td></tr><tr><td>6</td><td>6</td><td>88%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>7</td><td>7</td><td>27%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>8</td><td>8</td><td>67%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>9</td><td>9</td><td>90%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>10</td><td>10</td><td>29%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>11</td><td>11</td><td>53%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>12</td><td>12</td><td>73%</td><td>&gt;200</td><td>&gt;100</td></tr><tr><td>13</td><td>13</td><td>67%</td><td>&gt;200</td><td>&gt;100</td></tr></tbody></table>

<figure><img src="image_2.png" /><figcaption>Figure 1.</figcaption></figure>
<figure><img src="image_3.png" /><figcaption>(i) ZM241385</figcaption></figure>

<figure><img src="image_4.png" /><figcaption>(ii) Tic-H 1</figcaption></figure>

<figure><img src="image_5.png" /><figcaption>(iii) Tic-guanidine 2</figcaption></figure>

<figure><img src="image_6.png" /><figcaption>Figure 2.</figcaption></figure>

2

<figure><img src="image_7.png" /></figure>

3

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

4

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

5

<figure><img src="image_10.png" /></figure>

6

<figure><img src="image_11.png" /><figcaption>Figure 3.</figcaption></figure>

7

<figure><img src="image_12.png" /></figure>

8

<figure><img src="image_13.png" /></figure>

9

<figure><img src="image_14.png" /></figure>

10

<figure><img src="image_15.png" /></figure>

11

<figure><img src="image_16.png" /></figure>

12 R = H

13 R = Me
<figure><img src="image_17.png" /><figcaption>Figure 4.</figcaption></figure>

<figure><img src="image_18.png" /><figcaption>Figure 5.</figcaption></figure>

<figure><img src="image_19.png" /></figure>

<figure><img src="image_20.png" /></figure>

<figure><img src="image_21.png" /></figure>

<figure><img src="image_22.png" /><figcaption>Figure 6.</figcaption></figure>
<figure><img src="image_23.png" /><figcaption>Scheme 1.</figcaption></figure>

<figure><img src="image_24.png" /><figcaption>Scheme 2.</figcaption></figure>