**5. Citrullinemia**

**4. Preeclampsia**

464 Drug Discovery

of inflammatory processes [68].

Preeclampsia, a pregnancy-specific syndrome characterized by hypertension, proteinuria and edema, causes fetal and maternal morbidity and mortality with high incidence in devel‐ oping countries [55]. Symptoms of preeclampsia are currently combated by sodium restric‐ tion, rest and medication for blood pressure control to avoid complications for the mother and prolong the pregnancy for fetal maturation [56-58]. However, this attempt is rather un‐ specific with possible side effects for the developing fetus [59, 60]. Currently, the only thera‐ py of preeclamsia involves placenta removal resulting in pre-term birth [61]. Therefore,

novel drug development for pregnancy-specific conditions remains a challenge [59].

The pathology of preeclampsia involves systemic inflammation, oxidative stress, alterations in the levels of angiogenic factors, and vascular reactivity leading to hypertension of the mother and metabolic alterations in the fetus [61, 62]. A number of evidence suggests that clinical manifestations are caused by endothelial malfunction including insufficient produc‐ tion of NO [63, 64]. Levels of eNOS, the enzyme responsible for NO synthesis in the endo‐ thelium from L-arginine, are decreased in human umbilical vein endothelial cells from pregnant women suffering from preeclampsia [62] together with impaired AS expression [65]. The low availability of L-arginine uncouples eNOS activity, decreases NO production and increases eNOS-dependent superoxide generation [62, 66], consequently resulting in re‐ duced vasodilatation or in inflammatory processes observed in preeclampsia [66, 67]. There‐ fore, it is expected that the sustained concentration of L-arginine in endothelial cells is likely to play a critical role not only in the control of systemic blood pressure, but also in inhibition

Recently, we have reported that a *Bj*-PRO containing ten amino acid residues (*Bj*-PRO-10c), activating AS, is able to correct dysfunction of human umbilical vein endothelial cells from

*Bj*-PRO-10c, besides augmenting the activity of AS both *in vitro* and *in vivo* [14] increases sig‐ nificantly eNOS expression of human umbilical vein endothelial cells obtained from preg‐ nant women suffering from preeclampsia [65]. It was observed that the increase in NO levels induced by *Bj*-PRO-10c diminished the oxidative stress of the endothelial cells of pree‐

Most importantly, *Bj*-PRO-10c promoted NO production only in endothelial cells from pa‐ tients suffering from the disorder and not in normotensive pregnant women. In agreement, *Bj*-PRO-10c is a molecule endowed with antihypertensive activity that reduced blood pres‐ sure in hypertensive but not in normotensive rats [69]. These observations led to suggest that *Bj*-PRO-10c promotes its anti-hypertensive effect in mothers with preeclampsia without any effect on the blood pressure of the fetus, a problem with drugs currently used for mini‐ mizing health problems arising from preeclampsia. Taken together, *Bj*-PRO-10c becomes a

clamptic women, shown as a 50% reduction in superoxide levels [65] (Figure 1).

potential tool for the development of an efficient drug for preeclampsia treatment.

pregnant women suffering from preeclampsia [65] (Figure 1).

Citrullinemia, a disorder causing serious episodes of neurological symptoms associated with hyperammonemia involving disorientation, abnormal behaviors (aggression, irritabili‐ ty, and hyperactivity), seizures, coma, and potentially death from brain edema [70], occurs in two variants: CTLN1 (MIM#215700) or classical neonatal onset, and CTLN2 (MIM#603471) or adult-onset [71-74]. Classical citrullinemia in children is associated with a mutation in the AS gene [75]. However, in CTLN2 the enzyme reveals normal kinetic prop‐ erties and is quantitatively deficient only in the liver of adult-onset citrullinemia patients [71-74]. The most successful therapy of CTLN2 has been liver transplantation [76, 77] be‐ cause this treatment prevents hyperammonemic crises and corrects consequent metabolic disturbances [70].

It has been reported that administration of L-arginine to CTLN2 patients is effective in de‐ creasing blood ammonia concentration [78, 79]. For ammonia detoxification, arginine needs to enter the liver via the portal vein where is metabolized by mitochondrial arginase to pro‐ vide ornithine for citrulline and aspartate synthesis and for the priming of the urea cycle [80]. However, care must be taken when administering L-arginine, as fatal cases caused by L-arginine hydrochloride overdose have been reported [81]. In general, the dose of L-argi‐ nine supplementation used in the treatment of hyperammoaemia is in the high range be‐ tween 100 and 700 mg/kg body weight per day [82-85]. In animal models the effects of hyperargininaemia can be observed, reflecting toxic effects of high L-arginine concentration and making it possible to predict side effects of L-arginine supplementation including cogni‐ tive deficits, epilepsy and a progressive spastic diplegia [86]. Therefore, drugs augmenting AS activity, the step-limiting enzyme of urea cycle, may be a promising strategy for CTLN2 therapy, since AS is the step-limiting enzyme of the urea cycle. A consequence of increased AS activity, a final common pathway is triggered resulting in the excretion of waste nitrogen as urea [87].

carboxypeptidases and endopeptidases [95]. Nevertheless, cyclodextrins or nanocompounds could provide carriers for *Bj*-PROs, since drug release could be controlled in accordance

Applications of Snake Venom Proline-Rich Oligopeptides (Bj-PROs) in Disease Conditions Resulting from...

http://dx.doi.org/10.5772/52509

467

H.U. acknowledges grant support from Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) and (Conselho Nacional de Desenvolvimento Científico e Tecnológico

Department of Biochemistry, Institute of Chemistry, University of São Paulo, São Paulo, Brazil

[1] Tanen DA, Ruha AM, Graeme KA, Curry SC, Fischione MA. Rattlesnake envenoma‐ tions: unusual case presentations. Arch Intern Med. 2001 Feb 12;161(3):474-9.

[2] Walter FG, Bilden EF, Gibly RL. Envenomations. Crit Care Clin. 1999 Apr;15(2):

[3] Kini RM. Molecular moulds with multiple missions: functional sites in three-finger toxins. Clinical and experimental pharmacology & physiology. 2002 Sep;29(9):815-22.

[4] Hayashi MA, Murbach AF, Ianzer D, Portaro FC, Prezoto BC, Fernandes BL, et al. The C-type natriuretic peptide precursor of snake brain contains highly specific in‐ hibitors of the angiotensin-converting enzyme. J Neurochem. 2003 May;85(4):969-77.

[5] Murayama N, Hayashi MA, Ohi H, Ferreira LA, Hermann VV, Saito H, et al. Cloning and sequence analysis of a Bothrops jararaca cDNA encoding a precursor of seven bradykinin-potentiating peptides and a C-type natriuretic peptide. Proceedings of the National Academy of Sciences of the United States of America. 1997 Feb 18;94(4):

[6] Ferreira SH. A Bradykinin-Potentiating Factor (Bpf) Present in the Venom of Bo‐

[7] Wei L, Alhenc-Gelas F, Corvol P, Clauser E. The two homologous domains of human angiotensin I-converting enzyme are both catalytically active. The Journal of biologi‐

throps jararca. Br J Pharmacol Chemother. 1965 Feb;24:163-9.

cal chemistry. 1991 May 15;266(14):9002-8.

(CNPq), Brazil. C.L. is grateful for a postdoctoral fellowship awarded by FAPESP.

with therapeutic propose [96, 97].

Claudiana Lameu and Henning Ulrich

**Acknowlegments**

**Author details**

**References**

353-86, ix.

1189-93.

As previously mentioned, CTLN2 is not associated with genetic mutation of the AS gene; however, Saheki et al. identified the SLC25A13 gene as being defective in CTLN2 patients. This gene encodes for a Ca2+-dependent mitochondrial solute carrier, designated citrin [88]. According to Saheki et al., it is difficult to predict disease-causing effects of citrin deficiency in CTLN2, since children carrying citrin gene mutations may suffer from CTLN2 after more than 10 years or several decades of being asymptomatic [70]. In view of that, an option to prevent CLTN2 in infants with mutation of the citrin gen is being sought, having in mind that the nutritional management with appropriate intake of proteins only avoids accumula‐ tion of nitrogen [70].

Based on information discussed here, the strategy to increase AS activity in the liver could be an effective treatment for CTLN2 as well to prevent that children diagnosed as carrying SLC25A13 mutations from developing CTLN2 in the future [89]. We believed that direct pharmacological and clinical studies with *Bj*-PROs for these proposals, could turn them into a powerful therapeutic tool. The efficiency of *Bj*-PRO action can yet be improved by the ra‐ tional design of a compound which in the liver accelerates the urea cycle for eliminating am‐ monia or even preventing its accumulation.
