Immune response to L2 guinea pig leukemia-specific antigens.

Strain 2 guinea pigs were immunized with the LE-L2C cell line and challenged with either LE-L2C or BZ-L2C, a subline of L2C leukemia deficient in la gene product and C3 receptor. The LE-L2C-immune animals were completely protected from a challenge of 5 X 10(6) cells (100 times the lethal dose) of either cell line, and the delayed skin test responses for both lines of injected cells were equivalent. Thus established in vivo immune recognition of the leukemia antigen was similar for both cell lines. After repeated immunization of strain 2 animals with BZ-L2C cells, animals were challenged with 3 X 10(5) viable cells; 9 of 13 animals survived. A subsequent 5 x 10 (6) challenge lowered survival to 8 of the 13 animals. The skin test response to BZ-L2C developed slowly during the immunization period, but survivors of the viable cell challenge exhibited good responses to skin testing with BZ-L2C or LE-L2C leukemic cells. Thus the BZ-L2C cell line possesses a leukemia-specific antigen, but the immunogenicity of this mutant line is decreased when compared to that of the LE-L2C line.

Effect of immunotherapy with Corynebacterium parvum and methanol extraction residue of BCG administered intravenously on host defense function in cancer patients.

The effects of active nonspecific immunotherapy were studied in 42 patients receiving daily iv Corynebacterium parvum at 2 mg/m2 in 14-day courses and in 14 patients receiving iv methanol extraction residue of BCG (MER) at 0.5 mg/m2 weekly. The host defense evaluations included measurement of the number of adherent macrophage precursors per milliliter of blood (monocyte adherence), serum lysozyme, and antibody-dependent cell-mediated cytotoxicity (ADCC) of peripheral blood mononuclear cells to chicken red blood cells (CRBC) or human red blood cells (HRBC). During a single course of C. parvum, monocyte adherence did not rise significantly, whereas ADCC of peripheral blood mononuclear cells to CRBC and HRBC rose significantly (15.7-49.9% and 34.8-53.5% lysis of target cells, respectively). However, after a mean of 4.5 months on therapy, monocyte adherence increased an average of 7.5-fold. During weekly MER therapy, monocyte adherence, serum lysozyme, and ADCC of peripheral blood mononuclear cells to CRBC rose significantly within 4-7 days after the first dose (3.8-8.7 adherent cells/ml blood x 10(4), 7.6-10.8 microgram, and 34.4-41.4% target cell lysis, respectively). The host defense parameter, which was subnormal in the cancer patients (monocyte adherence), was boosted into the normal range in all the deficient patients by iv MER. The host defense parameters, which were normal or slightly elevated in the patients before therapy (serum lysozyme and ADCC of peripheral blood mononuclear cells to CRBC and HRBC), were hyperactivated above the upper limit of the normal range in 71.4, 71.4, and 50% of the patients, respectively, by iv MER. These methods can quantitatively reflect activation of monocytes and killer cells by C. parvum and MER and may be useful for evaluation and quantitation of both active nonspecific and immunorestorative immunotherapy in general.

Radiosensitive, thymic hormone-sensitive peripheral blood suppressor cell activity in cancer patients.

Suppressor cell activity which was radiosensitive in most subjects and thymic hormone sensitive in some was identified in patients with cancer, and compared to simultaneously studied normal controls. Suppressor cell activity was measured in cocultures of normal lymphocytes with patient lymphocytes added in microwells using the blastogenic response to phytohemagglutinin and concanavalin A as the measure of activity. Thirty-five patients (lung cancer, 21; leukemia in remission, seven; and various solid tumors, seven) and an equal number of controls were studied. Suppressor cell activity was identified in 71% of the patients. In approximately 75% of these, the suppressor cell activity was radiosensitive (4000 to 6000 rads). For the phytohemagglutinin response, suppressor cell activity was thymic hormone sensitive in approximately 40% (Thymosin Fraction 5 or thymic humoral factor), and for the concanavalin A response, it was thymic hormone sensitive in about 25% of the cases. There was a significant correlation between the presence of immunodeficiency (defined as a phytohemagglutinin response < 35,000 or a concanavalin A response < 12,000 cpm) and the presence of the suppressor cell activity. The suppressor cell activity was heterogenous relative to its radiosensitivity and thymic hormone sensitivity. Suppressor cell activity was observed in all the patient categories. These results indicate that certain available therapeutic manipulations may have significant effects on suppressor cell activity and should be an important subject for further investigation.

Cost-effective imaging approach to the nonbilious vomiting infant.

OBJECTIVE: To develop a cost- and time-effective algorithm for differentiating hypertrophic pyloric stenosis (HPS) from other medical causes of emesis in infants referred from community-based pediatricians and family practitioners to the imaging department of a tertiary children's care facility. METHODS: Eighty-nine vomiting infants (22 females, 67 males) between the ages of 11 and 120 days (mean, 43.5 days) had received nothing by mouth for at least 1 hour before the study. Each child was assessed for duration of vomiting, status of body weight, time and volume of last ingestion, and time of last emesis. A #8 French (Sherwood Medical, St Louis, MO) nasogastric feeding tube was placed in the child's stomach. The contents were aspirated and measured to determine likelihood of HPS. An aspirated volume >/=5 mL implicated gastric outlet obstruction, and ultrasonography (US) was performed. If this study was positive for HPS, the patient was referred for surgery. If US was negative, an upper gastrointestinal series (UGI) was performed. An aspirated stomach contents volume <5 mL suggested a medical cause for the emesis, and UGI was performed. Pediatric surgeons with no knowledge of the volume results palpated the abdomens of 73 of 89 infants (82%). RESULTS: Twenty-three of 89 patients (25%) had HPS. The aspirate criteria for HPS had a sensitivity of 91%, a specificity of 88%, and an accuracy of 89%. Of the false-positive studies (total = 8), six were related to recent significant ingestion (within 2 hours of the study), and two were attributable to antral dysmotility. The surgeons palpated the mass in 10 of 19 patients (53%). Sensitivity and specificity were 53% and 93%, respectively. Only 6 of 89 infants (7%) required both US and UGI to determine the etiology of the nonbilious vomiting. By performing the UGI in 66 patients, it was also found that 14% had slow gastric emptying and 79% had gastroesophageal reflux. Eighty-one percent of the gastroesophageal reflux was significant. CONCLUSION: The volumetric method of determining the proper imaging study is cost- and time-effective in the evaluation of the nonbilious vomiting infant for pyloric stenosis. If US was performed initially in all patients referred for imaging, two studies would have been performed in 68 of 89 patients (76%) to define the etiology of the emesis. Because we used the volumetric method, 62 fewer imaging studies were performed, representing a savings of $4464 and 30 hours of physician time. If children are given nothing by mouth for 3 to 4 hours before gastric aspiration, the specificity of the volumetric method improves to 94%, and the accuracy improves to 96%.

Kinetics of protein A activation of mononuclear cells from patients with chronic lymphocytic leukemia--I. CLL B-cells are not intrinsically unresponsive to staphylococcal protein A.

The response of lymphocyte subpopulations to staphylococcal Protein A (SPA) was studied in patients with B-cell chronic lymphocytic leukemia (CLL) and normal volunteers. The kinetics of the proliferative response, optimal dose and sera requirements were determined. Of 92 experiments conducted in 60 patients, SPA induced peripheral blood mononuclear cells (PBMC) proliferation in 81.5% of cases studied. The mean proliferative response of CLL PBMC was significantly lower than normal PBMC, 5823 vs. 30,364 cpm. However, enriched CLL B-cells failed to respond to SPA compared to normal enriched B-cells, with mean cpm 444 vs. 6438 respectively. As PBMC from CLL consist of increased proportions of B-cells and decreased proportions of T-cells, enriched CLL B-cells were cultured at 1: 1 ratio with autologous or allogeneic normal T-cell enriched fractions. CLL B-lymphocytes were stimulated by SPA when cultured in the presence of T-lymphocytes, indicating a T-helper defect in the B-CLL proliferative response to SPA, rather than an intrinsic inability of CLL B-cells to proliferate. These observations are of import for further studies of CLL B-lymphocyte function, cytogenetics, and establishment of CLL B-cell lines in culture.

A comparative study of the effectiveness of different mitogens for karyotypic analysis in chronic B cell leukemia.

In order best to study the karyotypic abnormalities of nine patients with chronic lymphocytic leukemia of B cell origin, we used a combination of T and/or B cell mitogens so as to achieve an optimal proliferative response. As assessed by DNA synthesis, and subsequent cytogenetic study, pokeweed mitogen (PWM) and protein A Sepharose (PROT A) appeared to hold promise as potent stimulators of neoplastic B lymphocytes. Lipopolysaccharide from E. coli (LPS) proved to be an ineffective mitogen. Phytohemagglutinin (PHA), while producing the highest stimulation indices by DNA assay, appeared to induce predominantly diploid metaphases. Pseudodiploid clonal abnormalities were noted in three out of nine patients, while another four patients showed random structural abnormalities, including translocations, which were indicative of damage. Presence of the A33 antigen, as determined by HLA typing, was noted in five out of nine patients. The increased frequency of this antigen in patients with chronic B cell leukemia is discussed.

Isolated congenital esophageal stenosis.

The incidence of congenital esophageal stenosis (CES) is approximately 1 in 25,000 to 50,000 live births. There is associated esophageal atresia in one third of cases; the remainder are classified as isolated CES. Histologically, the anomaly may include tracheobronchial remnants, a membranous diaphragm, or diffuse fibrosis of the muscularis and submucosa. The authors report their experience with three patients who had isolated CES. The patients were free of symptoms for the first 6 months of life. All had difficulty with feeding from 6 to 12 months of age, which corresponded with the introduction of solids. Most solid feedings were regurgitated. Evaluation consisted of cine-esophagogram, pH monitoring, manometry, and endoscopy, with biopsies to exclude the diagnosis of gastroesophageal reflux. All patients underwent hydrostatic dilatation, but the benefit was only transient. The patients were referred for surgical correction when symptoms recurred. Limited resection of the esophageal stenosis with primary anastomosis was performed on all three patients via a left thoracotomy. The stenoses were located in the distal third of esophagus, near the junction with the middle third. Pathological examination showed tracheobronchial remnants in one patient and fibrotic muscle in the other two. The average age at the time of surgery was 19 months. The hospital stay averaged 8 days. The contrast study 1 week postthoracotomy showed esophageal patency in all patients, with no leakage. All were discharged from the hospital, tolerating solid food. CES should be sought for in patients who present with dysphagia to solid food that begins after the first 6 months of life.(ABSTRACT TRUNCATED AT 250 WORDS)

Tetanus toxin and antigenic derivatives. II. Effect of protein and formaldehyde concentration on toxoid formation.

Tetanus toxoid formation was examined under varying conditions. Products of the reaction depended upon the concentration of both formaldehyde and toxin. High concentrations of protein and formaldehyde favored the formation of large polymers, whereas low concentrations yielded smaller polymers and monomer. The monomer had an observed S value of 7.1, whereas the polymers ranged from 10.1S to 110S. The cross-linking between toxin molecules to form toxoid polymers appeared to be random.

Tetanus toxin and antigenic derivatives. I. Purification of the biologically active monomer.

A procedure for the isolation of pure tetanus toxin in a lethal monomeric form was developed based on the extraction of whole cells and chromatographic techniques. A crude extract of toxin was obtained by hypertonic extraction of cells from a 72-hr culture of Clostridium tetani Massachusetts strain. The extract was precipitated with ammonium sulfate and further purified by sequential use of ion-exchange chromatography and gel filtration. The degree of purification obtained by the fractionation procedures was monitored by polyacrylamide gel electrophoresis. The pure toxin has an average specific activity of 150 x 10(6) mouse MLD per mg of N and 3,000 Lf per mg of N. Immunological purity was demonstrated by a single line on both immunoelectrophoresis and agar double diffusion. One band was obtained on polyacrylamide electrophoresis, as was a single symmetrical peak in the ultracentrifuge and on Sephadex G-100 chromatography. The pure protein has an absorbancy ratio (280/260 mmu) of 2.1 in phosphate buffer (pH 7.5).

Histochemical patterns of human T lymphocyte subpopulations with nonspecific esterase staining.

Alpha-naphthyl-acetate-esterase (ANAE) activity was histochemically demonstrated in human T lymphocytes and their subpopulations. Examination of T lymphocytes bearing Fc receptors for IgM showed the presence of circumscribed deposits of esterase activity, while T cells with FC receptors for IgG showed of predominantly diffuse granular pattern. The difference in the topography of the enzymatic activity was shown to be related to cellular activation resulting from exposure to IgG-sensitized bovine erythrocytes or to strong polyclonal activation by lectin mitogens. The ANAE stain also characterized monocytes, which exhibited an intense nongranular cytoplasmic stain. Non-T cells showed no detectable ANAE activity when tested by this method.

Effect of microbial fractions and vehicle on survival of mice bearing Lewis lung carcinoma.

Mice bearing pulmonary metastases of the Lewis lung carcinoma were treated iv with a nonviable microbial vaccine following amputation of the primary inoculation site. The vaccine consisted of BCG cell wall skeleton, trehalose, dimycolate, and endotoxin attached to mineral oil microdroplets. Single and repeated doses ranging from 15 to 675 micrograms were tested. Survival was modestly but consistently prolonged by vaccination. A portion of the activity appeared to be due to the Tween saline-oil vehicle. A single low dose (15 micrograms) was as efficacious as higher or repeated doses.

Active specific immunotherapy of stage IV renal carcinoma with aggregated tumor antigen adjuvant.

Active-specific immunotherapy of human malignancy with a vaccine consisting of admixtures of modified tumor antigens and an adjuvant such as tuberculin has not been fully explored, despite preliminary reports of clinical success and conceptual support from animal studies. Three years ago, we designed a prospective study using aggregated soluble tumor antigens admixed with tuberculin or phytohemagglutinin as an adjuvant (TAA) in order to treat patients with Stage IV renal carcinoma. Autologous tumor vaccines were used initially in 24 patients with operatively accessible tumor, although most patients eventually were switched to allogeneic preparations. Scarifications with Bacillus-Calmette Guerin were used in order to ensure maximum reactivity to tuberculin and patients received no other therapy while in the study. Two patients achieved complete remission and 2, partial remission. The overall survival rate for the 30 patients entered is equivalent to reported survival rates for patients with extensive disease treated with aggressive surgery with or without chemotherapy. We believe these results offer strong preliminary evidence of efficacy of this particular type of therapy in an advanced human malignancy.

Aminoglycoside pharmacokinetics: dosage requirements and nephrotoxicity in trauma patients.

We evaluated prospectively gentamicin and tobramycin pharmacokinetics in 37 patients with multiple system trauma and seven patients with isolated closed head trauma. The mean apparent volume of distribution (Vd) was 0.38 +/- 0.10 and 0.27 +/- 0.04 L/kg actual body weight (ABW) in patients with multiple trauma and closed head trauma, respectively. The difference in Vd between the two groups of patients was significant (p less than .002). Vd was not predictable on the basis of age, sex, weight, trauma score, or hospital day that therapy was initiated. Mean aminoglycoside clearance (Cl) was 123 +/- 46 ml/min. Neither serum creatinine nor estimated creatinine Cl predicted aminoglycoside Cl with sufficient accuracy to be clinically useful (r = .33 and .67, respectively). The mean daily dose was 6.1 +/- 1.6 mg/kg. The mean peak serum level was 5.8 +/- 1.3 micrograms/ml. Only one patient developed clinically significant renal dysfunction. Our data indicate that a loading dose of gentamicin or tobramycin of 3 mg/kg ABW in patients with multiple trauma and 2.5 mg/kg ABW in patients with isolated head trauma will obtain a mean initial peak serum level of 6.6 micrograms/ml. Although adequate maintenance dosing requires individualization based on pharmacokinetic analyses, large aminoglycoside doses can be used safely in patients with blunt trauma if appropriate monitoring is employed.

Endotoxin-free biologically active component of Escherichia coli.

The proteinaceous component of gram-negative bacteria, which has been termed "protodyne," enhances nonspecific host resistance while eliciting a slight pyrogenic response equivalent to 0.2% that of a typical endotoxin. Since this material still contains small amounts of carbohydrate and lipid, it was imperative to establish that its biological activities are not the result of endotoxin contamination. Evidence that the protective activity of protodyne does not result from endotoxin contamination has now been obtained by an evaluation of the Pronase digestion products of this substance. These digestion products were found to be nonpyrogenic and to contain no measurable amount of 2-keto-3-deoxyoctonate, an essential component of bacterial lipopolysaccharides.

Antibody-dependent cell-mediated cytotoxicity in human cancer: characterization of patient leukocyte activity and treatment effects.

Antibody-dependent cell-mediated cytotoxicity (ADCC), medicated by peripheral blood Hypaque-Ficoll separated mononuclear cells, was studied in humans using chicken erythrocytes (CRBC) incubated in a 1:1200 dilution of rabbit anti-CRBC and human B erythrocytes (HRBC) incubation in a 1:20 dilution of isoantibody. At the optimal target effector ratio of 3:1, ADCC to both CRBC and HRBC was significantly higher than normal in 27 lung cancer, 18 malignant melanoma, and seven colon cancer patients, but not in 20 breast cancer patients. Chemotherapy (single-agent or combination) in 12 patients did not effect ADCC in vitro but significantly suppressed ADCC to both targets after only four or five days of therapy in vivo (ADCC to CRBC, 47.4 to 24.1% lysis: ADCC to HRBC, 48.1 to 16.3% lysis). Immunotherapy with intravenous (IV) corynebacterium parvum or IV methanol extraction residue of BCG (MER) boosted ADCC to both targets within four to seven days of the first dose. It was found that ADCC to HRBC but not to CRBC was completely absent in three cases of active hairy cell leukemia but was present in two cases in remission. The ADCC to HRBC showed an age-dependent increase in both the 51 normal subject and the cancer patients. This was not observed for ADCC to CRBC. The ADCC to CRBC was mediated mainly by an Fc-receptor-positive, nonadherent, small lymphocyte, and ADCC to HRBC was mediated entirely by an adherent monocyte. The ADCC did not correlate significantly with the H3 thymidine incorporation of peripheral blood mononuclear cells, cultured without stimulation for either one or seven days. It also did not correlate with the number of residual granulocytes in the mononuclear cell suspensions. Measurement of ADCC is a useful method of characterizing host defense in malignant disease and its modification by therapy.

Natural history of central nervous system acute leukemia in adults.

Central nervous system (CNS) involvement occurred in 45 of 222 (20.3%) leukemic adults achieving bone marrow (BM) complete remission (CR), including 12 of 23 (52%) acute undifferentiated leukemia (AUL), 12 of 32 (39%) lymphoma leukemia, 5 of 26 (19%) acute lymphoblastic leukemia, and 16 of 142 (11%) acute myelogenous leukemia. Risk factors for CNS disease were lactic dehydrogenase (LDH) greater than or equal to 25,000/mm3. AUL morphology, age less than 20 years, and extramedullary involvement were most significant. Pattern of CNS involvement varied with morphology. Survival after CNS relapse depended most on BM status and symptoms. Duration of CNS CR was longest for asymptomatic patients with low CSF cell counts. Also important were duration of first BM CR, ease of achievement of initial BM CR, and leukocyte count (original and at most closely antecedent BM involvement), reflecting the common origin of BM and CNS leukemic cells. Central nervous system relapse generally did not shorten BM CR or survival, although early primary CNS relapse was associated with early BM relapse.