The product belongs to the class of so-called “Szeto-Schiller peptides”. Szeto-Sciller peptides or “SS peptides” are small, aromatic-cationic, water soluble, highly polar peptides, such as disclosed in U.S. Pat. No. 6,703,483 and U.S. Pat. No. 7,576,061, which can readily penetrate cell membranes. The aromatic-cationic peptides include a minimum of two amino acids, and preferably include a minimum of four amino acids, covalently joined by peptide bonds. The maximum number of amino acids is about twenty amino acids covalently joined by peptide bonds. As described by EP 2012/2436390, optimally, the number of amino acids present in the SS peptides is four.
Bendavia® is being tested for the treatment of ischemia reperfusion injury in patients with acute myocardial infarction (AMI), for the treatment of acute kidney injury (AKI) and renal microvascular dysfunction in hypertension, for the treatment of skeletal muscle dysfunction, for the treatment of mitochondrial myopathy and for the treatment of chronic heart failure. Trials are ongoing to assess the Ocuvia's potential to treat Leber's Hereditary Optic Neuropathy (LHON) a devastating inherited disease that causes sudden blindness, often in young adults.
Mitochondria are the cell's powerhouse, responsible for more than 90% of the energy our bodies need to sustain life and support growth. The energetics from mitochondria maintains healthy physiology and prevents disease. In many common and rare diseases, dysfunctional mitochondria are a key component of disease progression.
D-Arginyl-2,6-dimethyl-L-tyrosyl-L-lysyl-L-phenylalaninamide is a cell-permeable and mitochondria-targeted peptide that showed antioxidant activity and was concentrated in the inner mitochondrial membrane. Compound (<1 nM) significantly reduced intracellular reactive oxygen species, increased mitochondrial potential and prevented tBHP-induced apoptosis in both N2A and SH—SY5Y neuronal cell lines. In rats, intraperitoneal treatment (1 and 3 mg/kg) 1 day prior to unilateral ureteral obstruction and every day thereafter for 14 days significantly decreased tubular damage, macrophage infiltration and interstitial fibrosis. Compound (3 mg/kg i.p. qd for 2 weeks) also prevented apoptosis and insulin reduction in mouse pancreatic islets caused by streptozotocin.
Further studies performed in a G93A mouse model of amyotrophic lateral sclerosis (ALS) demonstrated that the compound (5 mg/kg/day i.p. starting at 30 days of age) led to a significant delay in disease onset. Potentially useful for the treatment of ALS and may be beneficial in the treatment of aging and diseases associated with oxidative stress.
In the last few years the peptide H-D-Arg-(2,6-Dimethyl)Tyr-L-Lys-L-Phe-NH2, shown in FIG. 1, and its therapeutic activity have been disclosed and claimed by in several patent applications.
EP 2436390, US 20110245182 and US 20110245183 claim topical anesthetic compositions for application to the skin for pain management or anti-skin aging agents, respectively, comprising Szeto-Schiller peptides; SS-31 is specifically claimed as active ingredient. Sequence of solid-phase synthesis is indicated as the preferred preparation process.
U.S. Pat. No. 7,718,620 claims a process of treating or preventing ischemia-reperfusion injury of the kidney in a mammal by administrating an effective amount of an aromatic-cationic peptide. SS-31 is specifically claimed as active ingredient.
WO02005/001023 discloses a generic method and carrier complexes for delivering molecules to cells comprising a molecule and an aromatic cationic peptide of type D-Arg-Dmt-Lys-Phe-NH2. The tetrapeptide SS-31 is specifically claimed as product useful for the process at claim 18.
WO2012/174117 and WO2014/210056 claim therapeutic compositions based on SS peptides and the aromatic-cationic peptide D-Arg-Dmt-Lys-Phe-NH2 as active agent.
WO 2013/086020, WO 2004/070054 and WO 2005/072295 provide processes for preventing mitochondrial permeability transition and reducing oxidative damage in a mammal, a removed organ, or a cell in need thereof and specifically claims the process wherein the peptide does not have mu-opioid receptor agonist activity, i.e., D-Arg-Dmt-Lys-Phe-NH2.
WO 2009/108695 discloses a process for protecting a kidney from renal injury which may be associated with decreased or blocked blood flow in the subject's kidney or exposure to a nephrotoxic agent, such as a radiocontrast dye. The processes include administering to the subject an effective amount of an aromatic-cationic peptide to a subject in need thereof and one of the selected peptide is D-Arg-Dmt-Lys-Phe-NH2.
U.S. Pat. No. 6,703,483 discloses a detailed procedure for the preparation of novel analogs of DALDA [H-Tyr-D-Arg-Phe-Lys-NH2], namely H-Dmt-D-Arg-Phe-Lys-NH2 using the solid-phase techniques and p-methylbenzhydrylamine resin and protocols that have been extensively used by inventor's laboratory.
Most prior art processes for preparing the compound typically comprise conventionally performed peptide solid-phase synthesis with further purification by chromatography in order to obtain the requested purity for therapeutic use.
It is well known that solid-phase synthesis followed by chromatographic purification is time consuming, very expensive and very difficult to be scaled up on industrial scale, so the need of developing a process for large scale production is obvious. The compound is isolated as organic acid salt, as acetate or trifluoro acetate.
Reddy et al., Adv. Exp. Med. Biol., 2009, 611, 473 generally describe the liquid-phase synthesis of antioxidant peptides of FIG. 1 and similar others (SS-02, SS-20), involving routinely used side chain protecting groups for amino acid building blocks. The guanidine group was protected with NO2 and the ε-NH2 of Lys was protected by Cbz or 2-Cl-Cbz. These peptides were synthesized using Boc/Cbz chemistry and BOP reagent coupling. Starting with the C-terminal Lys residue protected as H-Lys(2-Cl-Cbz)-NH2, (prepared from the commercially available Boc-Lys(2-Cl-Cbz)-OH in two steps by amidation with NH4HCO3 in the presence of DCC/HOBt following a literature procedure [Ueyama et all, Biopolymers, 1992, 32, 1535, PubMed: 1457730], followed by exposure to TFA). Selective removal of the 2-Cl-Cbz in the presence of the NO2 group was accomplished using catalytic transfer hydrogenolysis (CTH) [Gowda et al., Lett. Pept. Sci., 2002, 9, 153].
A stepwise procedure by standard solution peptide synthesis for preparation of potent μ agonist [Dmt]DALDA and its conversion into a potent δ antagonist H-Dmt-Tic-Phe-Lys(Z)—OH by substitution of D-Arg with Tic to enhance the δ opioid agonist activity is described by Balboni et al., J. Med. Chem., 2005, 48, 5608]. A general synthetic procedure for a similar tetrapeptide ([Dmt-D-Arg-Phe-Lys-NH2 is described by Ballet et al., J. Med. Chem. 2011, 54, 2467].
Similar DALDA analog tetrapeptides were prepared by the manual solid-phase technique using Boc protection for the α-amino group and DIC/HOBt or HBTU/DIEA as coupling agent [Berezowska et al., J. Med. Chem., 2009, 52, 6941; Olma et al., Acta Biochim. Polonica, 2001, 48, 4, 1121; Schiller at al., Eur. J. Med. Chem., 2000, 35, 895.
Despite the high overall yield in the solid-phase approach, it has several drawbacks for the scale-up process such as:                a. use of the highly toxic and corrosive hydrogen fluoride for cleavage of the peptide from the resin,        b. low loading (0.3-0.35 mmol/g of resin) proved necessary for successful end-step, and        c. use of excess amounts of reagents (3-fold of DIC, 2.4-fold of HOBt, etc.) on each step [Ryakhovsky et al., Beilstein J. Org. Chem., 2008, 4(39), 1, doi: 10.376/bjoc.4.39]        