**2.2.2 Romurtide (Muroctasine)**

In healthy volunteers, s.c. administration of Romurtide (Fig. 1) at a dose of 200 µg induced besides local pain and redness - an approximately 1º C increase in body temperature with great individual variability in the course of pyrogenicity curves; normalization occurred within 48 hours (Ichihara et al., 1988). Fever accompanied by chill and headache was also the most common adverse reaction in cancer patients treated with Romurtide at a dose range of 100 – 400 µg/dose s.c. for the restoraion of haemopoiesis after chemotherapy and/or radiotherapy (Tsubura et al., 1988) (Azuma & Seya, 2001; Tsubura et al., 1988).

### **2.3 nor-Muramyl glycopeptides**

We found that a combination of structural modifications both in the saccharide and peptide moiety of MDP and GMDP molecules leads to significant suppression or elimination of pyrogenicity and potentiation of immune-stimulatory activity (Fig. 2). The substitution of muramic acid with normuramic acid and L-alanine with L-2-aminobutyric acid has lead, in the case of the norAbu-MDP molecule, to a decrease of pyrogenicity and, at the same time, to the potentiation of immunoadjuvant activity. If the same structural change is carried out in GMDP molecule, a non-pyrogenic and highly immunoadjuvant analog, norAbu-GMDP is obtained. Furthermore, it has been demonstrated that by the introduction of bulky lipophilic residues into the molecules of these analogs, immunomodulatory activity can be effectively profiled, while the favourable pharmacological parameters of the parent structures are retained. These facts motivated our aims to design and prepare the new groups of lipophilic analogs of norAbu-MDP and norAbu-GMDP, which differ in the character and topology of the lipophilic residue. We primarily aimed to modify their immunopharmacologic parameters. norAbu-MDP-Lys(L18), i.e. MT05, belongs to them. In accordance with our premise, all the new compounds were nonpyrogenic (rabbit test), and the character and topology of the lipophilic residue had a significant effect on their immunologic parameters. As an example, the structural differences between norAbu-MDP-Lys(18) (MT05) and Romurtide, which

response. Since the discovery and first synthesis of MDP, about one thousand various derivatives of MDP have been designed, synthesised, and tested to develop an appropriate drug for an immunotherapeutic application that would be free of the side effect exerted by MDP. The main side effects of MDP are pyrogenicity, rigor, headache, flue-like symptoms, hypertension etc. Only several preparations reached the stage of clinical testing and only

In pyrogenicity test in rabbits, pyrogenic activity of Mifamurtide i.v. was comparable to that of MDP. In several studies with cancer patients refractory to standard therapy, infused with liposomal Mifamurtide at a dose range of 0.01 – 1.8 mg/m2/dose, dose-dependent fever (in common about 70% of patients) and rigor (about 50% of patients) were the most prominent

Mifamurtide was also assayed as an additional immunomodulator in an MF59-adjuvanted influenza virus vaccine (Keitel et al., 1993) and HIV-1 vaccine (Keefer et al., 1996); systemic symptoms including fever, chill, and nausea made these vaccines unsuitable for clinical use. Today, the main interest lies in clinical trials for liposomal Mifamurtide as a component of three-drug chemotherapy of osteosarcoma (Anderson P.M, 2006; Anderson et al., 2010).

In healthy volunteers, s.c. administration of Romurtide (Fig. 1) at a dose of 200 µg induced besides local pain and redness - an approximately 1º C increase in body temperature with great individual variability in the course of pyrogenicity curves; normalization occurred within 48 hours (Ichihara et al., 1988). Fever accompanied by chill and headache was also the most common adverse reaction in cancer patients treated with Romurtide at a dose range of 100 – 400 µg/dose s.c. for the restoraion of haemopoiesis after chemotherapy and/or radiotherapy (Tsubura et al., 1988) (Azuma & Seya, 2001; Tsubura et al., 1988).

We found that a combination of structural modifications both in the saccharide and peptide moiety of MDP and GMDP molecules leads to significant suppression or elimination of pyrogenicity and potentiation of immune-stimulatory activity (Fig. 2). The substitution of muramic acid with normuramic acid and L-alanine with L-2-aminobutyric acid has lead, in the case of the norAbu-MDP molecule, to a decrease of pyrogenicity and, at the same time, to the potentiation of immunoadjuvant activity. If the same structural change is carried out in GMDP molecule, a non-pyrogenic and highly immunoadjuvant analog, norAbu-GMDP is obtained. Furthermore, it has been demonstrated that by the introduction of bulky lipophilic residues into the molecules of these analogs, immunomodulatory activity can be effectively profiled, while the favourable pharmacological parameters of the parent structures are retained. These facts motivated our aims to design and prepare the new groups of lipophilic analogs of norAbu-MDP and norAbu-GMDP, which differ in the character and topology of the lipophilic residue. We primarily aimed to modify their immunopharmacologic parameters. norAbu-MDP-Lys(L18), i.e. MT05, belongs to them. In accordance with our premise, all the new compounds were nonpyrogenic (rabbit test), and the character and topology of the lipophilic residue had a significant effect on their immunologic parameters. As an example, the structural differences between norAbu-MDP-Lys(18) (MT05) and Romurtide, which

Mifamurtide (Fig. 1) was approved for the treatment of osteosarcoma.

from a number of acute systemic toxicities (Creaven et al., 1990).

**2.2.1 Mifamurtide (MTP-PE)** 

**2.2.2 Romurtide (Muroctasine)** 

**2.3 nor-Muramyl glycopeptides** 

influenced the profile of the effects and lead to the elimination of pyrogenicity, are depicted in Fig. 3. (Ledvina M., Turánek J., Miller A.D., Hipler K.: Compound (Adjuvants): PCT appl., WO 2009/11582 A2, 2009.)

Fig. 1. MDP derivatives developed by various pharmaceutical companies as adjuvants and immunotherapeutics.

Fig. 2. Transformation of MDP into norAbu-MDP and GMDP into norAbuGMDP and formulae of their hydrophobised derivatives suitable for development of lipid-based adjuvants.

Fig. 3. Structural differences between Romurtide and MT05.
