Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a Chemical structures of the adenosine, CF101 and CF102 are provided, highlighting modifications at the 5’-N-methylcarboxamide in the ribose group, as well as the N 6 and C2 positions of the adenosine group. The atom numbering is indicated in blue. CF101, is also named IB-MECA and N 6 -(3-iodobenzyl)adenosine-5’-N-methyluronamide. CF102, is also named Cl-IB-MECA and 2-chloro-N 6 -(3-iodobenzyl)adenosine-5’-N-methyluronamide. NanoBiT association assays monitoring ligand activity on adenosine receptors for adenosine ( b ), CF101 ( c ) and CF102 ( d ), respectively. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. Cryo-EM map ( e ) and model ( f ) of the CF101-A 3 AR-G i complex, with inset showing CF101 density. The density map in the inset is shown at 0.232 threshold. Cryo-EM map ( g ) and model ( h ) of the CF102-A 3 AR-G i complex, with inset showing CF102 density. The density map in the inset is shown at 0.17 threshold. Subunits are colored as indicated.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Activity Assay, Cryo-EM Sample Prep

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: Detailed interactions between A 3 AR and CF101 ( a ) or CF102 ( b ) from the membrane plane. Residues involved in ligand interaction are colored blue and pink in two complexes, respectively. Black dashed lines indicate hydrogen bonds. Dose-response curves of mutants of A 3 AR induced by CF101 (upper panels, c , e ) or CF102 (lower panels, d , f ) using NanoBiT assay. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Membrane

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a sequence alignment of ECL3 among adenosine receptors. The disulfide bond was shown as green linker. b Superimposed structures of adenosine receptors reveal that A 3 AR has the shortest ECL3. The residues in A 3 AR are shown in pink. The residues formed disulfide bond on ECL3 in A 1 AR were shown in green. Other TMs were omitted. c – e Assessing the effects of adenosine, CF101, and CF102 on A 1 AR, A 2A AR, and A 2B AR, along with their corresponding mutants containing the swapped ECL3 from the A 3 AR using NanoBiT assays. The results were from three independent experiments. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. f , g Assessing the effects of CF101 and CF102 on A 3 AR and its mutants with flexible ECL3 using NanoBiT assays. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Sequencing

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a Aligning the residues in the orthosteric binding pocket among the adenosine receptors. The conserved residues were colored in blue, and stars were used as markers. The unique residues in A 3 AR, distinct from other adenosine receptors subtypes were colored in orange, while residues in corresponding positions in other subtypes were colored in green. All residues were annotated based on GPCR Ballesteros-Weinstein numbering scheme. b In the superposition of adenosine receptors, the unique residues in A 3 AR, distinct from those in other adenosine receptors, are represented as yellow spheres. c , d Effects of CF101/CF102 on A 3 AR mutants containing swapped residues from other adenosine receptors by NanoBiT assay. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. e – i The binding cavities of the adenosine receptors were generated in PyMOL and depicted in gray. In A 3 AR, a subpocket is formed by His 3.37 , Ser 5.42 , and Ser 6.52 , while these positions are conserved as Gln 3.37 , Asn 5.42 , and His 6.52 in other adenosine receptor subtypes (His, H; Ser, S; Gln, Q; Asn, N). In h and i , dashed lines depict the hydrogen bonds between His 3.37 and Ser 5.42 . The names of the receptors and their associated PDB codes , , are indicated below each model.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Binding Assay, Generated

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a , b Superposition of active A 3 AR-CF101/CF102 complexes (blue/pink) with inactive A 2A AR-ZM241385 complex (gray, PDB ID 4EIY). Comparison of extracellular ( c ) and cytoplasmic ( d ) views of active A 3 AR and inactive A 2A AR. e – h Conformational changes in conserved motifs, including the toggle switch, PIF, DRY and NPxxY, upon CF101/CF102 binding to A 3 AR relative to inactive state of A 2A AR-ZM241385. Arrows indicate movement directions. In e , The sub-pocket in A 3 AR is formed by residues at position 3.37, 6.52 and 6.52. The residues at these positions from both A 3 AR and A 2A AR were labeled in green.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Comparison, Binding Assay, Labeling

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a Chemical structures of the adenosine, CF101 and CF102 are provided, highlighting modifications at the 5’-N-methylcarboxamide in the ribose group, as well as the N 6 and C2 positions of the adenosine group. The atom numbering is indicated in blue. CF101, is also named IB-MECA and N 6 -(3-iodobenzyl)adenosine-5’-N-methyluronamide. CF102, is also named Cl-IB-MECA and 2-chloro-N 6 -(3-iodobenzyl)adenosine-5’-N-methyluronamide. NanoBiT association assays monitoring ligand activity on adenosine receptors for adenosine ( b ), CF101 ( c ) and CF102 ( d ), respectively. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. Cryo-EM map ( e ) and model ( f ) of the CF101-A 3 AR-G i complex, with inset showing CF101 density. The density map in the inset is shown at 0.232 threshold. Cryo-EM map ( g ) and model ( h ) of the CF102-A 3 AR-G i complex, with inset showing CF102 density. The density map in the inset is shown at 0.17 threshold. Subunits are colored as indicated.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Activity Assay, Cryo-EM Sample Prep

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: Detailed interactions between A 3 AR and CF101 ( a ) or CF102 ( b ) from the membrane plane. Residues involved in ligand interaction are colored blue and pink in two complexes, respectively. Black dashed lines indicate hydrogen bonds. Dose-response curves of mutants of A 3 AR induced by CF101 (upper panels, c , e ) or CF102 (lower panels, d , f ) using NanoBiT assay. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Membrane

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a sequence alignment of ECL3 among adenosine receptors. The disulfide bond was shown as green linker. b Superimposed structures of adenosine receptors reveal that A 3 AR has the shortest ECL3. The residues in A 3 AR are shown in pink. The residues formed disulfide bond on ECL3 in A 1 AR were shown in green. Other TMs were omitted. c – e Assessing the effects of adenosine, CF101, and CF102 on A 1 AR, A 2A AR, and A 2B AR, along with their corresponding mutants containing the swapped ECL3 from the A 3 AR using NanoBiT assays. The results were from three independent experiments. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. f , g Assessing the effects of CF101 and CF102 on A 3 AR and its mutants with flexible ECL3 using NanoBiT assays. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Sequencing

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a Aligning the residues in the orthosteric binding pocket among the adenosine receptors. The conserved residues were colored in blue, and stars were used as markers. The unique residues in A 3 AR, distinct from other adenosine receptors subtypes were colored in orange, while residues in corresponding positions in other subtypes were colored in green. All residues were annotated based on GPCR Ballesteros-Weinstein numbering scheme. b In the superposition of adenosine receptors, the unique residues in A 3 AR, distinct from those in other adenosine receptors, are represented as yellow spheres. c , d Effects of CF101/CF102 on A 3 AR mutants containing swapped residues from other adenosine receptors by NanoBiT assay. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. e – i The binding cavities of the adenosine receptors were generated in PyMOL and depicted in gray. In A 3 AR, a subpocket is formed by His 3.37 , Ser 5.42 , and Ser 6.52 , while these positions are conserved as Gln 3.37 , Asn 5.42 , and His 6.52 in other adenosine receptor subtypes (His, H; Ser, S; Gln, Q; Asn, N). In h and i , dashed lines depict the hydrogen bonds between His 3.37 and Ser 5.42 . The names of the receptors and their associated PDB codes , , are indicated below each model.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Binding Assay, Generated

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a , b Superposition of active A 3 AR-CF101/CF102 complexes (blue/pink) with inactive A 2A AR-ZM241385 complex (gray, PDB ID 4EIY). Comparison of extracellular ( c ) and cytoplasmic ( d ) views of active A 3 AR and inactive A 2A AR. e – h Conformational changes in conserved motifs, including the toggle switch, PIF, DRY and NPxxY, upon CF101/CF102 binding to A 3 AR relative to inactive state of A 2A AR-ZM241385. Arrows indicate movement directions. In e , The sub-pocket in A 3 AR is formed by residues at position 3.37, 6.52 and 6.52. The residues at these positions from both A 3 AR and A 2A AR were labeled in green.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Comparison, Binding Assay, Labeling

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a Chemical structures of the adenosine, CF101 and CF102 are provided, highlighting modifications at the 5’-N-methylcarboxamide in the ribose group, as well as the N 6 and C2 positions of the adenosine group. The atom numbering is indicated in blue. CF101, is also named IB-MECA and N 6 -(3-iodobenzyl)adenosine-5’-N-methyluronamide. CF102, is also named Cl-IB-MECA and 2-chloro-N 6 -(3-iodobenzyl)adenosine-5’-N-methyluronamide. NanoBiT association assays monitoring ligand activity on adenosine receptors for adenosine ( b ), CF101 ( c ) and CF102 ( d ), respectively. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. Cryo-EM map ( e ) and model ( f ) of the CF101-A 3 AR-G i complex, with inset showing CF101 density. The density map in the inset is shown at 0.232 threshold. Cryo-EM map ( g ) and model ( h ) of the CF102-A 3 AR-G i complex, with inset showing CF102 density. The density map in the inset is shown at 0.17 threshold. Subunits are colored as indicated.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Activity Assay, Cryo-EM Sample Prep

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: Detailed interactions between A 3 AR and CF101 ( a ) or CF102 ( b ) from the membrane plane. Residues involved in ligand interaction are colored blue and pink in two complexes, respectively. Black dashed lines indicate hydrogen bonds. Dose-response curves of mutants of A 3 AR induced by CF101 (upper panels, c , e ) or CF102 (lower panels, d , f ) using NanoBiT assay. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Membrane

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a sequence alignment of ECL3 among adenosine receptors. The disulfide bond was shown as green linker. b Superimposed structures of adenosine receptors reveal that A 3 AR has the shortest ECL3. The residues in A 3 AR are shown in pink. The residues formed disulfide bond on ECL3 in A 1 AR were shown in green. Other TMs were omitted. c – e Assessing the effects of adenosine, CF101, and CF102 on A 1 AR, A 2A AR, and A 2B AR, along with their corresponding mutants containing the swapped ECL3 from the A 3 AR using NanoBiT assays. The results were from three independent experiments. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. f , g Assessing the effects of CF101 and CF102 on A 3 AR and its mutants with flexible ECL3 using NanoBiT assays. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Sequencing

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a Aligning the residues in the orthosteric binding pocket among the adenosine receptors. The conserved residues were colored in blue, and stars were used as markers. The unique residues in A 3 AR, distinct from other adenosine receptors subtypes were colored in orange, while residues in corresponding positions in other subtypes were colored in green. All residues were annotated based on GPCR Ballesteros-Weinstein numbering scheme. b In the superposition of adenosine receptors, the unique residues in A 3 AR, distinct from those in other adenosine receptors, are represented as yellow spheres. c , d Effects of CF101/CF102 on A 3 AR mutants containing swapped residues from other adenosine receptors by NanoBiT assay. Data shown are mean ± S.E.M. of three independent experiments ( n = 3). Source data are provided as a Source Data file. e – i The binding cavities of the adenosine receptors were generated in PyMOL and depicted in gray. In A 3 AR, a subpocket is formed by His 3.37 , Ser 5.42 , and Ser 6.52 , while these positions are conserved as Gln 3.37 , Asn 5.42 , and His 6.52 in other adenosine receptor subtypes (His, H; Ser, S; Gln, Q; Asn, N). In h and i , dashed lines depict the hydrogen bonds between His 3.37 and Ser 5.42 . The names of the receptors and their associated PDB codes , , are indicated below each model.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Binding Assay, Generated

Journal: Nature Communications

Article Title: Cryo-EM structures of adenosine receptor A 3 AR bound to selective agonists

doi: 10.1038/s41467-024-47207-6

Figure Lengend Snippet: a , b Superposition of active A 3 AR-CF101/CF102 complexes (blue/pink) with inactive A 2A AR-ZM241385 complex (gray, PDB ID 4EIY). Comparison of extracellular ( c ) and cytoplasmic ( d ) views of active A 3 AR and inactive A 2A AR. e – h Conformational changes in conserved motifs, including the toggle switch, PIF, DRY and NPxxY, upon CF101/CF102 binding to A 3 AR relative to inactive state of A 2A AR-ZM241385. Arrows indicate movement directions. In e , The sub-pocket in A 3 AR is formed by residues at position 3.37, 6.52 and 6.52. The residues at these positions from both A 3 AR and A 2A AR were labeled in green.

Article Snippet: The purification procedures of CF102-A 3 AR-G i complex were almost the same as in CF102-A 3 AR-G i complex preparation, while the CF101 compounds was replaced by CF102 (TargetMol, T6884).

Techniques: Comparison, Binding Assay, Labeling

Journal: Purinergic Signalling

Article Title: Targeting the A3 adenosine receptor to treat hepatocellular carcinoma: anti-cancer and hepatoprotective effects

doi: 10.1007/s11302-023-09925-2

Figure Lengend Snippet: Namodenoson chemical structure

Article Snippet: Namodenoson (CF102, CL-IB-MECA, Can-Fite BioPharma, Petah Tikva, Israel) is a 2-chloro analog of the protoypical agonist CF101.

Techniques:

Journal: Purinergic Signalling

Article Title: Targeting the A3 adenosine receptor to treat hepatocellular carcinoma: anti-cancer and hepatoprotective effects

doi: 10.1007/s11302-023-09925-2

Figure Lengend Snippet: Binding of Namodenoson to Human Receptors, as determined with radioligand binding assays

Article Snippet: Namodenoson (CF102, CL-IB-MECA, Can-Fite BioPharma, Petah Tikva, Israel) is a 2-chloro analog of the protoypical agonist CF101.

Techniques: Binding Assay, Activity Assay

Journal: Purinergic Signalling

Article Title: Targeting the A3 adenosine receptor to treat hepatocellular carcinoma: anti-cancer and hepatoprotective effects

doi: 10.1007/s11302-023-09925-2

Figure Lengend Snippet: Effect of namodenoson on the growth of HCC tumors in a rat orthotopic model

Article Snippet: Namodenoson (CF102, CL-IB-MECA, Can-Fite BioPharma, Petah Tikva, Israel) is a 2-chloro analog of the protoypical agonist CF101.

Techniques:

Journal: Purinergic Signalling

Article Title: Targeting the A3 adenosine receptor to treat hepatocellular carcinoma: anti-cancer and hepatoprotective effects

doi: 10.1007/s11302-023-09925-2

Figure Lengend Snippet: Mechanism of action of namodenoson

Article Snippet: Namodenoson (CF102, CL-IB-MECA, Can-Fite BioPharma, Petah Tikva, Israel) is a 2-chloro analog of the protoypical agonist CF101.

Techniques:

Journal: Purinergic Signalling

Article Title: Targeting the A3 adenosine receptor to treat hepatocellular carcinoma: anti-cancer and hepatoprotective effects

doi: 10.1007/s11302-023-09925-2

Figure Lengend Snippet: Comparison of 12-month overall survival (OS) rate in patients with Child–Pugh score 7 (namodenoson 25 mg BID vs placebo) in the phase II study

Article Snippet: Namodenoson (CF102, CL-IB-MECA, Can-Fite BioPharma, Petah Tikva, Israel) is a 2-chloro analog of the protoypical agonist CF101.

Techniques: Comparison

Journal: Purinergic Signalling

Article Title: Targeting the A3 adenosine receptor to treat hepatocellular carcinoma: anti-cancer and hepatoprotective effects

doi: 10.1007/s11302-023-09925-2

Figure Lengend Snippet: Best Observed responses (RECIST 1.1) by treatment arm in the phase II trial [ 52 ]

Article Snippet: Namodenoson (CF102, CL-IB-MECA, Can-Fite BioPharma, Petah Tikva, Israel) is a 2-chloro analog of the protoypical agonist CF101.

Techniques: