SIGMA RECEPTORS LIGANDS WITH ANTI-APOPTOTIC AND/OR PRO-APOPTOTIC PROPERTIES, OVER CELLULAR MECHANISMS, EXHIBITING PROTOTYPICAL CYTOPROTECTIVE AND ALSO ANTICANCER ACTIVITY

Abstract

The present invention involves new and original sigma receptors Ligands: (Mono-or dialkylaminoalkyl)-γ-butyro-lactones, their analogues aminotetrahydroturanes, the (1-adamantyl) benzene alkylamines, the N,N Dialkyl α-[(adamantyl-1)ben-zyloxy-2] alkylamines and the 3-cyclopentyl adamantyl-amines or alkylamines or-alkyl phenylamines, their enantiomers or di-astereoisomers, their pharmaceutically acceptable salts and Quinacrine Me-thylene blue, Astemizole and their relative analogues with pro-apoptotic and/or anti-apoptotic properties over cellular biochemical mechanisms, with prototypical anti-cancer, an-timetastatic and antiviral activities associated with antagonism of the neuropatic pain and, at very low doses, with cytoprotectve and cytoregenerative activity against the cytodegenerative diseases.

Description

The present invention concerns the prototypical a receptor ligands (Mono- or Dialky-laminoalkyl)-γ-butyrolactones (AL), their analogues aminotetrahydrofuranes (AE), with various substitutes: phenyl-, diphenyl-, phenoxymethyl, fluorenyl or adamantyl, the (1-adamantyl) phenyl(s) alkylamines-(AdBAA), the N,N Dialkyl α-[(adamantyl-1) benzyloxy-2] alkylamines-(AdBOAA) and the 3-cyclopentyl-adamantyl-amines or alkylamines or alkyiphenyl amines (AdCP), halogenated, or methoxylated on their substitutes and their pharmaceutically acceptable salts of the racemics, enantiomers and diastereoisomers and also the characterized by the inventor as a ligands generics Quinacrine (QNC), Methylene blue (MB), Astemizole (AST) and their related analogues,with prototypical anticancer and antimetastatic properties associated with analgesic activity against the neuropathic pain, generally, and also those induced by the clinically used anticancer drugs. At lower doses the above compounds and drugs exhibited prototypical cytoprotective properties associated with cytoregenerative activity developed by antagonism of the endoplasmic reticulum stress (ER stress) via activation of the sigma receptors in the connecting membrane between ER and mitochondrion.

The present invention concretize the concept of bio-modulatory activity with the original σ-receptor ligands AL, AE, AdBAA, AdBOAA, AdCP and the, recently characterized by the inventor as σ ligands, generics: QNC,MB,AST and also their related analogues with a new and sophisticated mechanism including the disclosed orthosteric and allosteric modulations but, principally, the activation of their σ-1 receptors (chaperones) in the dysregulated cells of the degenerative diseases characterized by hyper activation of the apoptotic process or, in cancer cells, the deactivation of these chaperones, at high doses of the above ligands, where they abolish the defensive inhibition of the apoptotic processus in the cancer cells.

In vitro and in vivo experiments clearly demonstrated the principal interaction of AL, AE, AdBAA, AdBOAA, AdCP and also QNC,MB and AST over the σ-1, σ-2 receptors from sub-nanomolar up to micromolar levels of affinity along with their regulatory roles over the G-protein coupled receptors (GPCRs) and sodium (Na), calcium (Ca), potassium (K) and chloride (Cl) ion channels and, principally, on the pathways of the ER and mitochondrion which determinate the protection of the cells in the degenerative diseases and the apoptosis of the cancer cells.

Thus, in contrast to the precedent patents of the inventor, the above invention displaced the area of the putative application of these molecules from the symptomatic to the therapeutic use because, specially for the cytoprotecting and cytoregenerating properties of the following claims the activity of the molecules concern the intracellular targets and pathways of pathogenesis and not the activity of superficial cell membrane receptors, as it is usually the case with the symptomatic drugs: neuroleptics, anti depressive drugs and cholinergics for, respectively, psychosis, depression and Alzheimer.

On the other hand, recent experiments in vivo demonstated the prototypical profile of the above σ ligands which associated the anticancer activity with the antagonism of pain or neu-ropathic pain and also those originated from the anticancer treatments with the clinically used anticancer drugs and specially with taxanes, platinum agents and vicristine. Thus, the inven-tion of this prototypical activity, supported the synergistical use of the above σ ligands with the clinically active anticancer drugs—as it was recently demonstrated with xenograft expe-riments on SCID mice and also, with neuropathic pain protocols, on CD-1 and Balb-C mice—with simultaneous antagonism, by the above Cr ligands, of the painful effects and disability in-duced by the clinically used anticancer drugs.

Finally, their ability to regulate cellular apoptotic processes, through ER-mitochondrial σ-1 and σ-2 receptors, associated with the stereochemical re-articulation of the membrane sphingolipides, induced by the above ligands, supported the antiviral activity of the above compounds (more specially those containing adamantyl(s)),which was demonstrated experimentally in vitro.

EXAMPLES

A) Sigma (σ)-1 Agonists with Bio-Modulatory Properties and with Antagonistic Action on the Apoptotic Processes of the Cells.

• • AE14: TETRAHYDRO-N,N,-DIMETHYL-5,5-DIPHENYL-3-FURANOMETHANAMINE

AE14 is a selective ligand for σ-1 receptors with a nanomolar affinity (pKi=7,6) while the affinity for σ-2 receptors is more than 100 times lower [over 10 micro-Moles (10 μM): pKi<5]. The above molecular profile is the basis for AE14 modulatory role over the G-protein coupled receptors (GPCRs) and its pharmacological properties: anti-amnesic action (anta-gonistic action against scopolamine and dizocilpine (MK-801) induced amnesia or amyloid peptide β25-35 (Aβ25-35), at low doses (from 0.03 mg/kg administered per os (po), in mice). This unique new profile of AE14: anti-apoptotic via agonistic sigma-1 regulation of the volume regulated chloride channels (VRCC), anti-oxidative stress via agonistic action on the sigma-1 receptors of the ER-mitochondrion.

Recent experiments amplified the above disclosed results and demonstrated the putative cytoprotective and cytoregenerative activity of AE14 and the relative analogues, by triggering the activity of the σ chaperones in the ER-mitochondrion,in protocols of acute or chronic cellular stress (in vitro and in vivo on mice) using oligopeptides or streptozocin which initiate ER stress and cytotoxic activity by apoptosis. In the above protocols, AE14 antagonized the ER stress extending its application area to the intimate causes: dysregulation of the σ chaperones regulated alarm pathways to mitochondrion,and therefore to all the cytodegenerative diseases.

• • TETRAHYDRO-N,N-DTIVIETHYL-2,2-DINIENYL-3-FURANOMETHANAMINE

Similar pharmacological profile as to AE14: smaller affinity for σ-1 receptors (pKi=6,3) and with no affinity for σ-2 receptors.

Therefore, AE37, as a selective σ-1 agonist with anti-apoptotic action, was recently studied in vitro and in vivo on mice in protocols of chronic ER and cellular stress and exhibited high anti-apoptotic properties and putative cytoprotective and cytoregenerative activity.

• • AE37Met:TETRAHYDRO-N-METHYL-2,2-DIPHENYL-3-FURANOMETHANAMINE

AE37 Met is the only metabolite of AE37 and is slightly more active than AE37 for the properties mentioned above but it mainly concerns the cytoprotective action, especially against acute intracellular (ER-mitochondrion) stress and then against the ischemic syndromes (brain, heart), in relevance with its exceptional protection against ER-mitochondrion anoxia and oxidative stress caused by pentylenetetrazole (PTZ), in vitro or in vivo, on mice, following its protection against the PTZ-induced tonic crises.

B) Weak σ-1 Agonists or σ-1 Antagonists with Agonistic σ-2 Component and Agonistic Action to the Apoptotic Processes of the Cells.

• • The following molecules are representatives of the chemical families that concern the present invention:
  • 1) MAL: DIHYDRO-4-(D1METHYLAMINOMETHYL) SPIRO {FURANE-2(5H),2′-TRICYCLO[3.3.1.1_3,7] DECAN}-5-ONE 6—
  • 2) AdAE: (DIHYDRO-4-(DIMETHYLAMINOMETHYL)SPIRO {FURANE-2(3H),2′-TRICYCLO[3.3.1.1_3,7] DECANE}
  • 3) AdPhAL: 5-(TRICYCLO[3.3.1.1_3,7] DEC-1-YL)-DIHYDRO-3-[DIMETHYLAMINOMETHYL)-5-PHENYLFLTRAN-2(3H)-ONE—
  • 4) AdPhAE: 5-(TRICYCLO[3.3.1.1_3,7] DEC-1-YL)-TETRAHYDRO-N,N-DIMETHYL-5-PHENYL-3-FURANEMETHANAMINE—
  • 5) AL142Me:(α-(DINIETHYLAMENOETHYL)-γ,γ-DIPHENYL-γ-BUTYROLACTONE)
  • 6) AdBPA:γ-(1-ADAMANTYL)PRENYL-N,N-DIMETHYLPROPANAMINE
  • 7) AdBPP: γ-(1-ADAMANTYL)PHENYL-PROPYLPIPERIDINE
  • 8) AdBOPP: [α-(ADAMANTYL-1)BENZYLOXY-2] PROPYLPIPERIDINE
  • 9) AdBOEA (Me): N,N DIMETHYL-[α-(ADAMANTYL-1)BENZYDRYLOXY-2-ETHYLAMINE
  • 10) -Ad(Me)CP: 3-CYCLOPENTYL-N-METYL-1-TRICYCLO[3.3.1.1_3,7]DECANEMETHANAMINE
  • 11) Quinacrine (QNC)
  • 12) Methylene blue (MB)
  • 13) Astemizole (AST)
  • (see also, in appendix, undescribed derivatives of the above chemical families)

All molecules present strong chemical affinities (sub-nanomolar to micromolar) for σ-1 receptors with antagonistic action (except AdAE and AdPhAE which are weak agonists) and with nanomolar to micromolar affinities for σ-2 receptors and agonistic action. Therefore, all molecules above and their derivatives are pro-apoptotic and only at very low concentrations or doses are anti-apoptotic. All molecules above are toxic to the cancer cells : colorectal, prostate, ovarian renal, pancreas, lung, gliomas, glioblastomas leukemia, lymphomas, melanomas, sarcomas and hepatoma, with antiproliferative and cytotoxic concentrations usually lower than 10 microMolar (μM),for more than 50 of the above derivatives. Outstanding cytotoxicity, in vitro and the corresponding in vivo anticancer activity in mice xenografts, is for the first time disclosed for QNC and AST : 3.0-5.0 μM for all the above cancer cells and an outstanding in vivo activity of QNC on primary (drug resistant) melanoma, in which Dacarbazine is inactive, or MB : lower than 1.0 μM in vitro for primary (drug resistant) melanoma or leukaemia and lung and close to 2.0 μM in breast, colon and glioblastomas confirmed in vivo, specially in primary melanoma. All these molecules make excellent and prototypical anticancer drug candidates due to their aptitude to, simultaneously, antagonize the neurogenic and, more exceptional, the neuropathic and inflammatory pain as recently demonstrated by the appropriate experimental protocols: hot plate, formalin, von Frey fibers and, principally, the protocols based on previous administration of paclitaxel, oxaliplatine and streptozocin which concern more specifically the neuropathic pain originating from chronic administration of anticancer drugs, from diabetes or other neurotoxic stimulus. The pharmacological profile of the above molecules is totally prototypical and original giving the possibility of a therapeutic synergy with the clinically used anticancer drugs with simultaneous antagonism and protection against the most grave secondary effects i.e., neuropathic pain and disability induced by the clinically used anticancer drugs.

• • 1) Pouli N., Fytas G., Foscolos G., Kolocouris N., Marakos P., and Vamvakides A. Synthese et etude pharmacologique des adaman-tylbenzenepropanamines et propenamines. Annales Pharmaceutiques Francaises, 1995, 53, No 4, pp 163-169
  • 2) Fytas G., Marakos P., Kolocouris N., Foscolos G., Pouli N., Vamvakides A., Ikeda S., De Clercq E. 3-cyclopentyl-1-aclamantamines and adamantanemethanamines. Antiviral activity evaluation and convulsion studies. Farmaco 1994, 49, No 10, pp 641-647
  • 3) Vamvakides Alexandre: Tetrahydro-N,N-dimethyl-2,2-diphenyl-3-furanemethanamine, its enantiomers and their pharmaceutically acceptable acid addition salts. WO97/30983 and patent 1002616 (GR)

Claims

1-8. (canceled)

9. A method of using sigma(σ) receptor ligands, said method comprising the steps of:

providing a compound comprising a sigma(σ)-receptor ligands and a sigma(σ) ligands generics, said sigma(σ) ligands generics being selected from the group consisting of Quinacrine, analogues of Quinacrine, Methylene blue, analogues of Methylene blue, Astemizole, and analogues of Astemizole; and

using said compound for preparation of pharmaceuticals with anticancer, antimetastatic and antiviral activity associated with analgesic properties.

10. The method according to claim 9, wherein said sigma(σ)-receptor ligands being selected from the group consisting of (mono- or di-alkylaminoalkyl)-γ-butyrolactones), aminotetrahydrofuranes, enantiomers or diastereoisomers of aminotetrahydrofuranes, (1-adamantyl) benzene alkylamines, enantiomers or diastereoisomers of (1-adamantyl) benzene alkylamines, N,N di-alkyl α-[(adamantyl-1) benzyloxy-2] alkylamines, enantiomers or diastereoisomers of N,N di-alkyl α-[(adamantyl-1) benzyloxy-2] alkylamines, 3-cyclopentyl adamantyl-amines or -alkylamines or -alkyl phenylamines, enantiomers or diastereoisomers of 3-cyclopentyl adamantyl-amines or -alkylamines or -alkyl phenylamines, and their pharmaceutically acceptable salts.

11. The method according to claim 10, wherein said compound further comprising at least one pharmaceutically acceptable excipient.

12. A method of using sigma(σ) receptor ligands, said method comprising the steps of:

providing a compound comprising a sigma(σ)-receptor ligands and a sigma(σ) ligands generics, said sigma(σ) ligands generics being selected from the group consisting of Quinacrine, analogues of Quinacrine, Methylene blue, analogues of Methylene blue, Astemizole, and analogues of Astemizole; and

using said compound for preparation of pharmaceuticals with analgesic activity against neuropathic pain.

13. The method according to claim 12, wherein said sigma(σ)-receptor ligands being selected from the group consisting of (mono- or di-alkylaminoalkyl)-γ-butyrolactones), aminotetrahydrofuranes, enantiomers or diastereoisomers of aminotetrahydrofuranes, (1-adamantyl) benzene alkylamines, enantiomers or diastereoisomers of (1-adamantyl) benzene alkylamines, N,N di-alkyl α-[(adamantyl-1) benzyloxy-2] alkylamines, enantiomers or diastereoisomers of N,N di-alkyl α-[(adamantyl-1) benzyloxy-2] alkylamines, 3-cyclopentyl adamantyl-amines or -alkylamines or -alkyl phenylamines, enantiomers or diastereoisomers of 3-cyclopentyl adamantyl-amines or -alkylamines or -alkyl phenylamines, and their pharmaceutically acceptable salts.

14. The method according to claim 13, wherein said compound further comprising at least one pharmaceutically acceptable excipient.

15. A method of using sigma(σ) receptor ligands, said method comprising the steps of:

providing a compound comprising a sigma(σ)-receptor ligands and a sigma(σ) ligands generics, said sigma(σ) ligands generics being selected from the group consisting of Quinacrine, analogues of Quinacrine, Methylene blue, analogues of Methylene blue, Astemizole, and analogues of Astemizole; and

using said compound for preparation of pharmaceuticals selected from the group consisting of pharmaceuticals acting synergistically with clinically used anticancer drugs and antagonizing neuropathic pain induced by the anticancer drugs, pharmaceuticals with cytoprotective activity against pathogenesis of cytodegenerative diseases, pharmaceuticals with cytoprotective and cytoregenerative activity, and pharmaceuticals with protective activity against the pathogenesis of inflammatory and neuropathic pain.

16. The method according to claim 15, wherein said sigma(σ)-receptor ligands being selected from the group consisting of (mono- or di-alkylaminoalkyl)-γ-butyrolactones), aminotetrahydrofuranes, enantiomers or diastereoisomers of aminotetrahydrofuranes, (1-adamantyl) benzene alkylamines, enantiomers or diastereoisomers of (1-adamantyl) benzene alkylamines, N,N di-alkyl α-[(adamantyl-1) benzyloxy-2] alkylamines, enantiomers or diastereoisomers of N,N di-alkyl α-[(adamantyl-1) benzyloxy-2] alkylamines, 3-cyclopentyl adamantyl-amines or -alkylamines or -alkyl phenylamines, enantiomers or diastereoisomers of 3-cyclopentyl adamantyl-amines or -alkylamines or -alkyl phenylamines, and their pharmaceutically acceptable salts.

17. The method according to claim 16, wherein said compound further comprising at least one pharmaceutically acceptable excipient.