Experience with long term use of LNAA in treatment of PKU Reuben Matalon 1, Kimberlee Michals-Matalon 2, Gita Bhatia 1, James Grady 1, Stephen Tyring 3. 1 The University of Texas Medical Branch, Galveston, Texas University of Houston, Texas 77204, 3 The University of Texas Health Science Center, Houston, Texas 77030,

Previous loading of Short term studies with Large Neutral Amino Acids (LNAA) in patients with PKU resulted in decrease of blood phenylalanine (Phe) levels. The long term safety, efficacy and acceptability of LNAA tablets (NeoPhe) have not been evaluated. In this study, four patients, three female and one male, ages 25 to 38 years, were given NeoPhe tablets, 0.5gm/kg/day in three divided doses to be taken with meals. The patients were not on medical foods, for more than 10 years previously. Their blood Phe prior to taking NeoPhe had a mean value of 1507  mol/L. Blood Phe were determined two weeks after entering the study and once a month for a period of 12 months. The mean blood Phe level declined for each of the subjects during the study period: 642  mol/L, 707  mol/L, 899  mol/L and 869  mol/L. All patients reached levels within the NIH consensus report recommendation. Patients were monitored for weight incase, LNAA was used for protein synthesis. None of the patients gained or lost any weight beyond minor fluctuation of +/- 0.2 kg. The acceptability of the pills was monitored with review on every visit and there were no complains regarding the number of pills or of abdominal discomfort, nausea or changes in bowel habits. All patients asked to continue taking NeoPhe tablets because they were happy with their blood Phe levels and indicated they felt “more focused” at work. Future studies should include larger number of patients and neuropsychological tests need to be added. Abstract

Introduction Dietary treatment of phenylketonuria (PKU) has been the corner stone for controlling blood phenylalanine (Phe) concentrations in PKU. Such control has become difficult when blood Phe concentrations need to be controlled for the life of the patient. Methods to improve dietary control include addition of tetrahydrobiopterin (BH4) to the management and reformulation of formulas to make them more palatable. In order to decrease entry of Phe to the brain of patients with PKU, large neutral amino acids (LNAA) have been added to compete with the transporter of Phe. In 1987 Lou et al. (Acta Paediatr Scand 76:560) added 160 mg/kg of tyrosine and found improved attention span in patients with PKU. Pietz et al. in 1995 (J. Pediatr. 127:936 ) found no effect on the brain of adults with high doses of tyrosine. Other formulations have been tried but none have resulted in an acceptable method of treatment. Dotremont et al, in 1995 used a formulation of LNAA 0.6 gms per kg with a low protein diet and found negative nitrogen balance, with lysine the limiting amino acid. We studied the effect of PreKUnil on PKU mice (Enu2/Enu2). A surprising finding was that such mice showed a lower concentration of blood Phe. Taking into account that the GI transporter (Table 1) for LNAA has different Km than the blood brain barrier, a new formulation was undertaken. The new formulation of LNAA (NeoPhe) was developed, by introducing changes in concentration of some amino acid and addition of Lysine. In order to correct the deficiency a new PreKUnil with lysine added has been introduced.

Table 1: Transport of LNAA to the Brain K m K m App Phenylalanine (Phe) Leucine Tyrosine Tryptophan Methionine Histidine Isoleucine Valine Threonine Pardridge, Inborn Errors of Metabolism in Humans. MTP Press, 1980.

Methods Four patients, three female and one male, ages 25 to 38 years, with PKU took part in the study at University of Texas Medical Branch, Galveston, Texas, U.S. The patients where given NeoPhe tablets, 0.5gm/kg/day in three divided doses to be taken with meals Blood Phe was measured thrice at zero time, two weeks after starting on NeoPhe and once a month for a period of 12 months. Patients were monitored for weight increase, acceptability of pills. Physical was done once a month.

Results The individual blood Phe response to 0.5 g/kg of NeoPhe is shown in Figures 1-5. The paired t-test for the decline in blood Phe is p ≤ The genotype of each individual is shown. The mean blood Phe levels declined for each of the patients during the study period: 642  mol/L, 707  mol/L, 899  mol/L, and 869  mol/L. All patients reached levels within NIH consensus report recommendation. None of the patients gained or lost any weight beyond the minor fluctuation of +/- 0.2Kg. There were no complains regarding the number of pills or abdominal discomfort, nausea or changes in bowel habits.

Figure 1 Figure 1: Effect of LNAA on 4 patients over a period of 12 months.

Figure 2 Figure 2: The patient showed a 40% decrease of blood Phe levels in the 12 month period. (Female: 34 years, Mutation: Del I94 / unknown).

Figure 3 Figure 3: The patient showed a 45% decrease of blood Phe levels in the 12 month period. (Female: 25 years, Mutation: E280K / E280K).

Figure 4 Figure 4: The patient showed a 42% decrease of blood Phe levels in the 12 month period. (Female: 38 years, Mutation: IVS12nt-1g>a / P281L).

Figure 5 Figure 5: The patient showed a 49% decrease of blood Phe levels in the 12 month period. (Male: 30 years, Mutation: G46S / unknown).

Summary of the study NeoPhe 0.5 g/kg 4 subjects Mean age 32 years 1 males, 3 females Mean decrease in blood Phe after two weeks 831 µmol/L Average decrease in blood Phe 45 %.

Discussion and Conclusions Specific composition of large neutral amino acids, as in NeoPhe can reduce blood Phe concentrations. Our studies show for the first time that LNAA can effect the blood concentrations of Phe, which is the end point of treatment of PKU. NeoPhe has been more effective than PreKUnil in lowering blood Phe concentrations. Patients were able to take NeoPhe for a period of 12 months and were happy with their blood Phe levels, they indicated that they felt “more focused” at work. Patients with high protein intake and high concentration of blood Phe will require higher amounts of NeoPhe in order to effectively compete with Phe transport. Future studies should include larger number of patients and neuropsychological tests need to be added. To avoid Lysine deficiency, any product without lysine should not be used.

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