Increasing the affinity for tumor antigen enhances bispecific antibody cytotoxicity.

We tested the hypothesis that bispecific Abs (Bsab) with increased binding affinity for tumor Ags augment retargeted antitumor cytotoxicity. We report that an increase in the affinity of Bsab for the HER2/neu Ag correlates with an increase in the ability of the Bsab to promote retargeted cytotoxicity against HER2/neu-positive cell lines. A series of anti-HER2/neu extracellular domain-directed single-chain Fv fragments (scFv), ranging in affinity for HER2/neu from 10(-7) to 10(-11) M, were fused to the phage display-derived NM3E2 human scFV: NM3E2 associates with the extracellular domain of human FcgammaRIII (CD16). The resulting series of Bsab promoted cytotoxicity of SKOV3 human ovarian carcinoma cells overexpressing HER2/neu by human PBMC preparations containing CD16-positive NK cells. The affinity for HER2/neu clearly influenced the ability of the Bsab to promote cytotoxicity of (51)Cr-labeled SKOV3 cells. Lysis was 6.5% with an anti-HER2/neu K(D) = 1.7 x 10(-7) M, 14.5% with K(D) = 5.7 x 10(-9) M, and 21.3% with K(D) = 1.7 x 10(-10) M at 50:1 E:T ratios. These scFv-based Bsab did not cross-link receptors and induce leukocyte calcium mobilization in the absence of tumor cell engagement. Thus, these novel Bsab structures should not induce the dose-limiting cytokine release syndromes that have been observed in clinical trials with intact IgG BSAB: Additional manipulations in Bsab structure that improve selective tumor retention or facilitate the ability of Bsab to selectively cross-link tumor and effector cells at tumor sites should further improve the utility of this therapeutic strategy.

Effect of calcium/phosphorus ratio on mineral retention in parenterally fed premature infants.

We hypothesized that retention of parenterally delivered calcium (Ca) and phosphorus (P) is affected by the ratio of the delivered minerals and that a 1.7:1 ratio would be optimal since this is the ratio of retention of these minerals by the fetus. Forty-one very low birth weight (VLBW) infants were randomly assigned to one of three total parenteral nutrition (TPN) solutions that were different only in their Ca:P ratios: 2:1 (76 mg/kg/day Ca and 38 mg/kg/day of P), and 1.3:1 (58 mg/kg/day Ca and 45 mg/kg/day P), and 1.3:1 (58 mg/kg/day of Ca and 45 mg/kg/day of P). Serum levels of calcium, phosphorus, and alkaline phosphatase, retentions of calcium and phosphorus and urinary cyclic AMP levels were measured after 48 h on the assigned Ca to P ratio. Calcium retentions were higher with the 2:1 and 1.7:1 ratios and phosphorus retentions were higher with the 1.3:1 and 1.7:1 ratios. The 1.7:1 ratio allowed for the highest absolute retention of both minerals and was the closest to published in utero accretion of calcium and phosphorus. The serum and urine studies demonstrated no abnormalities on any of the three ratios. Cyclic AMPs were not different among groups and were not elevated compared to previous reports suggesting that none resulted in parathyroid hormone (PTH) stimulation. We conclude that the 1.7:1 ratio is better than higher or lower ratios for delivery of calcium and phosphorus in TPN solutions at the quantities studied.

Effect of high calcium and phosphorus intake on mineral retention in very low birth weight infants chronically treated with furosemide.

The treatment of premature infants with the diuretic furosemide appears to be a contributory factor in the development of metabolic bone disease presumably because of furosemide-induced hypercalciuria. In this study, we measured calcium and phosphorus balance in furosemide-treated very low birth weight infants (VLBW) infants with bronchopulmonary dysplasia (BPD) who were fed a specialized premature formula containing increased amounts of calcium and phosphorus. Furosemide-treated infants received 166 +/- 37 mg/kg/day and retained 80 +/- 34 mg/kg/day of calcium, and 87 +/- 19 mg/kg/day and retained 52 +/- 14 mg/kg/day of phosphorus. The amounts retained were approximately 65% of the calcium and 72% of the phosphorus requirements for in utero mineral accretion. Compared to a group of similarly fed VLBW infants without BPD and not treated with the diuretic, the furosemide-treated infants excreted a larger percent of the calcium intake in the urine but had similar total urinary calcium and phosphorus losses (mg/kg/day) and serum calcium, phosphorus, alkaline phosphatase, and parathyroid hormone (PTH) levels. From the latter two findings, we suggest that the extra mineral content of the formula may have promoted bone mineralization and prevented the occurrence of secondary hyperparathyroidism.

Simultaneous infusion of calcium and phosphorus in parenteral nutrition for premature infants: use of physiologic calcium/phosphorus ratio.

We hypothesized that parenteral delivery of calcium and phosphorus in a ratio of 1.7:1 would promote retention of these minerals and decrease urinary phosphorus excretion, and that delivery of increased amounts of this ratio would result in higher retentions. Serum levels and retention of calcium and phosphorus were measured as calcium intake was increased from 36 to 76 mg/kg/day in 10 mg increments and as phosphorus intake was adjusted to maintain the 1.7:1 ratio. Five different infants were studied at each of the five levels. The amounts of calcium and phosphorus retained increased steadily and at level 5 were 71.8 +/- 1.2 mg/kg/day and 40.9 +/- 1.7 mg/kg/day, respectively. Over the five levels the average percent calcium retention was 91.4 +/- 4.2 and the average percent phosphorus retention was 89.1 +/- 7.7. The provision of parenteral calcium and phosphorus in a 1.7:1 ratio resulted in a balanced retention of both minerals over the range studied. The use of this calcium/phosphorus ratio appears to be appropriate for the preterm infant receiving total parenteral nutrition.

Growth and phosphorus metabolism in premature infants fed human milk, fortified human milk, or special premature formula. Use of serum procollagen as a marker of growth.

Human milk promotes less than optimal growth and is associated with phosphorus deficiency and decreased bone mineralization in very-low-birth-weight (VLBW) infants. In this study, the effects of feeding premature infants either human milk (HM), fortified human milk (FHM), or special premature formula (Similac Special Care [SSC]) on growth, phosphorus metabolism, and serum type I procollagen (pColl-I-C) were evaluated. Infants fed FHM exhibited a rate of weight gain and an increase in head circumference comparable with infants fed SSC and significantly greater than infants fed HM, despite the fact that both the FHM group and the HM group demonstrated biochemical evidence of phosphorus deficiency. The pColl-I-C concentrations in VLBW infants were tenfold to 20-fold greater than concentrations in normal children older than 2 years of age. The pColl-I-C levels correlated positively with weight gain and were significantly greater in the FHM and SSC groups than in the HM group. By contrast, serum alkaline phosphatase levels did not correlate with weight gain and were significantly lower in the rapidly growing SSC group than in either of the two groups with phosphorus deficiency and presumed poor bone mineralization. We conclude that the serum pColl-I-C concentration is a biochemical marker of growth in VLBW infants and may prove useful as a predictor of growth responses to various nutritional and therapeutic interventions.

Achievement of in utero retention of calcium and phosphorus accompanied by high calcium excretion in very low birth weight infants fed a fortified formula.

Calcium and phosphorus retention was evaluated in 13 very low birth weight infants who were fed an experimental formula designed to deliver quantities of calcium and phosphorus sufficient to meet the intrauterine accretion rates for these minerals. Retention of calcium and phosphorus in slight excess of these rates was achieved without any apparent difficulties for the infants. Biochemical measurements demonstrated normal serum calcium (9.8 +/- 8 mg/dL) and alkaline phosphatase (242 +/- 51.6 IU) values. However, there was evidence of high tubular reabsorption of phosphate (98.1% +/- 3.3%), hypercalciuria (7.2 +/- 3.8 mg/kg/d), and a relatively low serum phosphorus concentration (5.7 +/- 0.6 mg/dL). This biochemical picture is similar to that seen in phosphorus deficiency except for the low alkaline phosphatase activity. The latter finding, in concert with the high retention of calcium and phosphorus in these balance studies, makes such a diagnosis unlikely. We speculate that this biochemical picture is the result of an inappropriately high calcium/phosphorus ratio.

Hypophosphatemia and hypercalciuria in small premature infants fed human milk: evidence for inadequate dietary phosphorus.

Phosphorus and calcium balance was measured prospectively in stable premature infants (less than or equal to 1600 gm) fed human milk or a standard commercial formula. Throughout the study, the P and Ca intakes of the infants fed human milk were two to three times less than those of infants fed formula. Infants fed human milk showed low serum P and normal serum Ca concentrations, complete renal reabsorption of P, and elevated renal Ca excretion. The net effect in infants fed human milk was a 50% reduction in the P and Ca retention, compared with the formula-fed group. Despite the unfavorable P and Ca balance in the group fed human milk, the only evidence of rickets was elevated alkaline phosphatase activity. Nevertheless, based on the biochemical changes in these infants, low serum P values, and excess urinary calcium losses, we conclude that the stable small premature infant fed human milk exclusively is deficient in phosphorus and only slightly more sufficient in calcium.

Comparisons of antidotal efficacy of chelating drugs upon acute toxicity of Ni(II) in rats.

Six chelating drugs were administered to rats by im injection at equimolar dosages in order to compare their relative effectiveness in prevention of death after a single parenteral injection of NiCl2. Triethylenetetramine and d-penicillamine were most effective antidotes for acute Ni (II)-toxicity. In order of decreasing antidotal effectiveness, diglycyl-L-histidine-N-methylamide, sodium diethyldithiocarbamate and calcium disodium versenate significantly reduced the acute mortality of rats following ip injection of Ni (II). Alpha-Lipoic acid was not effective as an antidote for acute Ni (II)-toxicity.