Table 1. The LOPAC molecules showing the highest protective effect in the hypoxia screens.
Hypoxia-reoxygenation injury was induced in LLC-PK1 cells and the LOPAC library was screened for protective effect in various set-ups. From each screen the ten highest-ranking cytoprotective compounds are shown with the respective activity in the other assays and the known biological function. (I.) In the pre-treatment screen (PRE) the compounds (50 μM) were applied prior to the OGD, (II.) in the post-treatment screen (POST) the cells received the compounds after the OGD. Adenosine pretreatment (30 μM) was combined (III.) with compounds pre-treatment (ADE PRE) and (IV.) with the application of the test compounds following the hypoxia (ADE POST). The shown cytoprotective effect represents the increase in viability compared to the OGD in PRE and POST screens or compared to the adenosine-treated cells subjected to OGD (ADE) in ADE PRE and ADE POST screens. (The viability values of CTL, OGD and ADE groups were 100%, 40% and 70%, respectively.)
| Compound | Known biological activity | Cytoprotection (%) |
|||
|---|---|---|---|---|---|
| PRE | POST | ADE PRE | ADE POST | ||
|
I. Pre-treatment screen
| |||||
| Adenosine | endogenous neurotransmitter | 46.0 | 13.2 | 16.6 | −0.8 |
| Disulfiram | aldehyde dehydrogenase inhibitor | 34.5 | 2.9 | −9.2 | 0.8 |
| 2-Phenylaminoadenosine | selective adenosine A2 receptor agonist | 33.5 | 1.9 | −4.2 | 2.6 |
| Tyrphostin AG 537 | tyrosine kinase inhibitor | 32.6 | 4.1 | −14.4 | 7.1 |
| Riluzole | glutamate release inhibitor | 30.4 | −1.1 | −2.1 | 10.6 |
| Tyrphostin 1 | tyrosine kinase inhibitor | 30.3 | 2.5 | 1.4 | 2.0 |
| Zardaverine | phosphodiesterase(PDE) inhibitor | 28.3 | 4.3 | −13 | −1.5 |
| Triamcinolone | glucocorticoid steroid | 27.2 | 3.3 | 1.1 | 1.5 |
| Thapsigargin | SERCA inhibitor | 26.3 | 3.0 | −15.1 | −3.1 |
| Vinblastine sulfate | inhibitor of microtubule assembly | 23.3 | 4.3 | −25.2 | −9.3 |
|
II. Post-treatment screen | |||||
| N-Acetyldopamine monohydrate | dopamine analog possessing antitumor activity | 1.1 | 15.4 | 3.1 | −3.5 |
| Enoximone | phosphodiesterase inhibitor | 4.2 | 14.0 | −6.0 | 4.5 |
| R(-)-N-Allylnorapomorphine HBr | dopamine receptor anatagonist | 4.1 | 13.5 | 7.8 | 10.4 |
| L-Aspartic acid | amino acid, participant in gluconeogenesis | 1.0 | 13.5 | 8.3 | 24.8 |
| Amperozide HCl | serotonin receptor antagonist | −6.4 | 13.3 | −73.0 | 6.5 |
| 5α-Androstane-3α,17β-diol | testosterone metabolite | 2.3 | 13.2 | −2.6 | −1.4 |
| Adenosine | endogenous neurotransmitter | 46.0 | 13.2 | 16.6 | −0.8 |
| 4-DAMP methiodide | M3 muscarinic acetylcholine receptor antagonist | −4.6 | 13.1 | −45.0 | −1.6 |
| Aminoguanidine HCl | inhibitor of nitric oxide synthase (NOS) | 3.7 | 13.0 | 6.8 | 8.0 |
| AEBSF | serine protease inhibitor | 1.0 | 12.9 | 7.3 | −1.3 |
|
III. Adenosine combined pre-treatment screen | |||||
| γ-D-Glutamylaminomethylsulfonic acid | kainate/quisqualate glutamate receptor antagonist | 1.4 | 2.0 | 27.0 | 3.2 |
| Hydrocortisone 21-hemisuccinate | glucocorticoid steroid | 1.6 | 3.0 | 25.1 | 11.7 |
| Tyrphostin AG 126 | TNF-α and NO synthesis inhibitor | 16.5 | −0.9 | 18.1 | 3.6 |
| 4-Amino-1,8-naphthalimide | poly(ADP-ribose) polymerase (PARP) inhibitor | 8.9 | 4.6 | 16.9 | 21.9 |
| Adenosine | endogenous neurotransmitter | 46.0 | 13.2 | 16.6 | −0.8 |
| Caffeic acid phenethyl ester | NF- κ B inhibitor | 1.7 | 3.6 | 12.4 | 22.5 |
| 2-Methylthioadenosine diphosphate | P2Y purine receptor agonist | 18.7 | 2.9 | 11.1 | 8.4 |
| Bestatin hydrochloride | aminopeptidase inhibitor | 5.1 | 2.6 | 10.9 | −3.8 |
| Thiocitrulline | inhibitor of neuronal and endothelial NOS | 8.0 | 3.8 | 9.3 | 15.2 |
| R(+)-6-Bromo-APB hydrobromide | D1 dopamine receptor agonist | 0.5 | 3.8 | 8.8 | 14.3 |
|
IV. Adenosine combined post-treatment screen | |||||
| Arecaidine propargyl ester HBr | M2 muscarinic acetylcholine receptor agonist | −1.4 | 3.5 | −16.1 | 35.6 |
| L-Aspartic acid | amino acid, participant in gluconeogenesis | 1.0 | 13.5 | 8.3 | 24.8 |
| Centrophenoxine HCl | cholinergic nootropic agent | 1.0 | 6.1 | 8.5 | 24.0 |
| Caffeic acid phenethyl ester | NF-kB inhibitor | 1.7 | 3.6 | 12.4 | 22.5 |
| 4-Amino-1,8-naphthalimide | poly(ADP-ribose) polymerase (PARP) inhibitor | 8.9 | 4.6 | 16.9 | 21.9 |
| KN-62 | Ca2+/calmodulin-dependent protein kinase inhibitor | 2.2 | 2.1 | −40.2 | 18.0 |
| Dipyridamole | PDE and thromboxane synthase inhibitor | 2.2 | 8.1 | 3.4 | 16.8 |
| Thiocitrulline | inhibitor of neuronal and endothelial NOS | 8.0 | 3.8 | 9.3 | 15.2 |
| R(+)-6-Bromo-APB hydrobromide | D1 dopamine receptor agonist | 0.5 | 3.8 | 8.8 | 14.3 |
| Hydrocortisone 21-hemisuccinate | glucocorticoid steroid | 1.6 | 3.0 | 25.1 | 11.7 |
Abbreviations: SERCA: sarco/endoplasmic reticulum Ca2+-ATPase, DAMP: 4-Diphenylacetoxy-N-methylpiperidine methiodide, AEBSF: 4-(2-Aminoethyl) benzenesulfonyl fluoride hydrochloride, R(+)-6-Bromo-APB hydrobromide: (+)-6-Chloro-7,8-dihydroxy-3-allyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrobromide