Strategies to improve plasma half life time of peptide and protein drugs

Summary. Due to the obvious advantages of long-acting peptide and protein drugs, strategies to prolong plasma half life time of such compounds are highly on demand. Short plasma half life times are commonly due to fast renal clearance as well as to enzymatic degradation occurring during systemic circulation. Modifications of the peptide/protein can lead to prolonged plasma half life times. By shortening the overall amino acid amount of somatostatin and replacing l-analogue amino acids with d-amino acids, plasma half life time of the derivate octreotide was 1.5 hours in comparison to only few minutes of somatostatin. A PEG_2,40 K conjugate of INF-α-2b exhibited a 330-fold prolonged plasma half life time compared to the native protein. It was the aim of this review to provide an overview of possible strategies to prolong plasma half life time such as modification of N- and C-terminus or PEGylation as well as methods to evaluate the effectiveness of drug modifications. Furthermore, fundamental data about most important proteolytic enzymes of human blood, liver and kidney as well as their cleavage specificity and inhibitors for them are provided in order to predict enzymatic cleavage of peptide and protein drugs during systemic circulation.     

Circadian rhythm of plasma aldosterone and time dependent alterations of aldosterone regulators

In order to investigate the relative contribution of ACTH, the renin angiotensin and dopaminergic system to the circadian rhythm of plasma aldosterone, hormone levels were determined at hourly intervals over a 24 h span in four supine healthy men. Blood was withdrawn under basal conditions (control), after either dexamethasone, captopril or bromocriptine++ (CB-154) administration. Plasma aldosterone rhythmicity was abolished in dexamethasone treated groups but not in captopril or CB treated groups. Time dependent alterations of aldosterone regulators were analyzed by multiple regression methods at 3 hourly intervals. These results indicate that plasma aldosterone rhythmicity is predominantly under the control of ACTH whereas the renin angiotensin or dopaminergic system plays a little role. In supine, sodium repleted states, ACTH is a potent stimulus of aldosterone at 000-0600 h and 1700-1900 h clocktime, whereas during daytime renin-angiotensin is an additional regulator.     

Effect of time and temperature on bovine serum and plasma progesterone concentration

Whole blood, with and without anticoagulant, from 5 pregnant cows was incubated at 40°C for 0 (30 minutes after collection), 6 and 24 hours (hr) before the blood was centrifuged and the plasma or serum was frozen for later progesterone assay. Mean plasma progesterone concentration decreased from 6.6 ng/ml at 0 hr to 1.7 ng/ml at 6 hr (P < 0.01) and to 2.8 ng/ml at 24 hr (P < 0.01). Mean serum progesterone concentration decreased from 6.1 ng/ml at 0 hr to 3.9 ng/ml at 6 hr (P < 0.01) and to 4.4 ng/ml at 24 hr (P < 0.01). Whole blood samples with and without EDTA were also incubated at 4°C for 24 hr. Mean plasma progesterone concentration decreased from 6.6 ng/ml at 0 hr to 4.2 ng/ml at 24 hr (P < 0.01). Mean serum progesterone concentration decreased from 6.1 ng/ml at 0 hr to 4.7 ng/ml at 24 hr (P < 0.01). The incubation time and temperature of whole blood, from collection of blood to the separation of serum or plasma, significantly affects assayable concentration of progesterone.     

The effect of fasting time on plasma total cholesterol concentration in Mongolian gerbils

This study investigated the effect of length of fasting time on plasma total cholesterol response of male Mongolian gerbils (Meriones unguiculatus). Plasma cholesterol levels from fed and fasted gerbils were also compared with those reported for humans under similar metabolic states. Plasma total cholesterol response showed a significant quadratic relationship with time over a 15-hour period. Between 6 and 9 hours of fasting (the time during which plasma triglyceride concentration became relatively constant), the average plasma total cholesterol concentration was 178 mg/dl, compared with a zero hour (fed) cholesterol level of 265 mg/dl. The difference in plasma cholesterol levels observed in fed and fasted gerbils is unlike what has been reported for humans. Results from most human studies show no differences in plasma total cholesterol concentrations for fed and fasted subjects. Failure to consider species differences in metabolic responses may have implications when results from animal experiments are extrapolated to humans.     

Development of a plasma 17alpha-hydroxyprogesterone time resolved-fluorescence immunoassay involving a new biotinylated tracer

A biotinylated 17alpha-hydroxyprogesterone probe (3) was prepared from 17alpha-hydroxyprogesterone-3-carboxymethyloxime and conjugate obtained by acylation of biotinylaminopropylammonium trifluroacetate. This new tracer was used in the development of a 17alpha-hydroxyprogesterone time-resolved fluoroimmunoassay using streptavidin-europium. The new method was compared to a long-standing radioimmunoassay method and found to be more sensitive and economical.     

Radioimmunoassay of androstenedione and testosterone in cow plasma at the time of luteolysis and oestrus

Androsteredione and testosterone were measured by radioimmunoassay after chromatography on micro-columns of Lipidex-5000 in jugular plasma samples taken every 2-3 h at the time of luteolysis and oestrus in 3 dairy cows. The concentrations of androstenedione and testosterone varied between 5 and 60 pg/ml and 5 and 80 pg/ml respectively and no consistent pattern in the fluctuations of either steroid was observed.     

Left ventricular systolic time intervals and plasma noradrenaline concentrations during acute hypovolaemia in newborn infants

Left ventricular systolic time intervals and plasma noradrenaline concentrations were determined in twelve newborn infants during acute moderate hypovolaemia. Rapid withdrawal of 8.5 ml of blood per kg body weight shortened the left ventricular ejection time from 204 to 176 msec (P less than 0.001) and increased the plasma noradrenaline concentration from 3.4 to 4.0 nmol/l (P less than 0.05). Systemic blood pressure was unchanged. It is concluded that even a moderate reduction in blood volume is associated with marked changes in left ventricular dynamics. The shortening of left ventricular ejection time during hypovolaemia probably reflects left ventricular adaptation to decreased volumes and response to increased noradrenaline release.     

Vanadyl ions increase the order parameter of plasma membranes without changing the rotational relaxation time

Differential polarized phase fluorometry of 1,6-diphenyl-1,3,5-hexatriene showed that vanadyl ions (VO2+) increased its limiting anisotropy (order parameter) in crude plasma membranes from brown adipose tissue of the golden hamster (Mesocricetus auratus). This was about 10(3) times larger than the effect of Ca2+ and was several times greater than the action of Co2+. Vanadate anions were without any effect. During the membrane treatment with VO2+, the rotational relaxational time of diphenylhexatriene did not change. This results suggest a possible positive influence of tetravalent vanadium on the stability of cell membranes.     

The time of increase in plasma progesterone during pregnancy in mink (Mustela vison )

In 1980 and 1981, respectively, 12 and 19 one-year-old mink females of standard and jetblack breeds were used to determine the progesterone level approximately at the time of expected implantation. Blood samples were collected every 2 to 3 days or daily in order to accurately estimate the time of increase in plasma progesterone levels. The results indicate that the progesterone level increased to above 10 ng/ml, 31.6 (sigma = 1.3) days prior to parturition on the average. This was supported by physiological explanations. The date of mating in mink had an effect on the date of increase in plasma progesterone. Matings at a late date in the estrous period, reduced the period of delay before implantation. Nevertheless, both the dates of implantation and parturition were delayed compared with the results of earlier matings in mink.     

Physiological changes in androgen plasma levels with elapsing of time from castration in adult male rats

In the present study we investigated in adult male rats the effects of castration on Dehydroepiandrosterone (DHEA), Androstenedione (delta 4), Testosterone (T) and Dihydrotestosterone (DHT) plasma levels: five days (group II), seven weeks (group III) and eleven weeks (group IV) after orchiectomy. The same hormone assays were performed in rats approximately 60 days of age which underwent a sham-operation for orchiectomy (group I). Our data show that five days following orchiectomy (group II) delta 4, T and DHT were decreased with respect to sham-operated rats. (Group I: delta 4: 83.3 +/- 14.9 (SEM) ng/dl (n = 12); T: 435.32 +/- 51.45 (n = 12); DHT: 51.47 +/- 6.54 (n = 12); Group II: delta 4: 44.81 +/- 6.09 (n = 12) P = 0.05; T: 25.54 +/- 2.88 (n = 12) P less than 0.01; DHT: 12.9 +/- 2.51 (n = 12) P less than 0.01). Seven weeks afterwards T and DHT remained significantly lower (group III: T: 54.37 +/- 12.21, n = 16) (P less than 0.01; DHT: 33.22 +/- 4.49 (n = 16) P less than 0.01) while eleven weeks after all steroids were significantly decreased with respect to the values observed in sham-operated rats. (Group IV) delta 4: 32.01 +/- 5.7 (n = 10) P less than 0.01: T: 27.29 +/- 7.05 (n = 10) P less than 0.01; DHT: 29.03: 5.34 (n = 10) P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)