Henry J. Esber

Chemotherapy responsiveness of human tumors as first transplant generation xenografts in the normal mouse: six-day subrenal capsule assay.

Feasibility of utilizing human tumors as first transplant generation xenografts in the normal immunocompetent mouse for determining tumor sensitivity to chemotherapeutic agents was demonstrated by applying subrenal capsule (SRC) assay methodology to fresh surgical explants in a six‐day time frame. A total of 37 human breast tumors were tested in assays in which 254 xenografts were implanted into control animals. Fifty (20%) of the controls showed some degree of partial regression in the six‐day assay period. Using a mean control growth having a positive change in tumor size as the criterion for evaluability, first transplant generation human breast tumors provided an evaluable assay rate of 86%. A tumor response profile was obtained as a result of testing seven clinically active drugs against 32 previously untreated breast cancers. The pattern of responses obtained indicated that no single agent was active against all tumors, nor were tumors which were responsive to one agent necessarily responsive to another, suggesting the feasibility of predicting individual tumor response to specific chemotherapeutic agents. Had these seven drugs been developmental agents of unknown activity which were being tested for the first time against such a panel of human tumors the results would have not only predicted their clinical activity, but the tumor response rates would have also provided an indication of the relative potential of each drug for the specific treatment of breast cancer.

Growth of Human Tumor Xenografts Implanted under the Renal Capsule of Normal Immunocompetent Mice

The subrenal capsule technique proved effective in demonstrating that the growth of human tumors in normal, immunocompetent animals for 6 days was quantifiable in ocular micrometer units. Positive growth was demonstrable not only with human tumors that had been established in serial transplantation in athymic nude mouse hosts, but also with primary surgical explants. Growth rates of transplantation-established xenograft systems were similar whether implanted in athymic nude or in normal immunocompetent animals indicating that the 6-day time-frame successfully evades growth inhibitory effects of immunologic origin. Immunosuppression with a single dose of cyclophosphamide did not appear to affect growth rate, but permitted the tumors to grow larger extending the time to reach peak size. Significantly, xenografts of primary surgical explants showed positive growth more frequently in 6 days (82%) in the immunocompetent animal than in 11 days (30%) in the immunodeficient athymic nude mouse.

Long‐term controlled delivery of levonorgestrel in rats by means of small biodegradable cylinders

Hormone release rates from biodegradable cylindrical implants consisting of a physical matrix of [14 C]levonorgestrel and copolymers of [3 H]lactic and glycolic acids have been monitored in rats. Two copolymers were evaluated: one consisted of 90 parts L‐lactide/10 parts glycolide (90L/10G) containing 33 or 50% hormone by weight, and the other of 50 parts DL‐lactide/50 parts L‐lactide (50DL/50L) containing 50% hormone. For each system, 4–6 rods (0·8 times 16 mm) providing 19 mg of steroid per rat were subcutaneously implanted into the scapular regions of 5 rats, and 14 C and 3 H in faeces and urine were determined weekly for 90–724 days. An initial burst of hormone release, peaking at approximately 90 μg day−1, occurred in the first two weeks. This was followed by an approximately zeroorder release for 14 C from all systems. Longer‐term release rates were approximately 10–30 μg day−1 for the 90L/10G system containing 33% hormone, a more uniform rate of approximately 25 μg day−1 for the 50DL/50L system and the highest rate of approximately 40 μg day−1 for the 90L/10G system containing 50% hormone. 3 H and 14 C in residual implants of 90L/10G with 33% hormone removed from animals dying of natural causes during the test were assayed. For this system 3 H activity decreased by over 50% within 250 days, compared with < 25 % loss in 14 C activity. The amounts of 3 H and 14 C released were similar over much of the subsequent test period. At the end of the test both polymer and drug were essentially depleted. All animals with the 50DL/50L system died late in the test period. Between days 609 and 724 from 13·7‐27·2% initial 14 C and from 10·0‐22·1 % initial 3 H was measured in recovered rods. Microscopic inspection of recovered rods showed a loss of core material and tissue encapsulation. There were no signs of local tissue irritation or systemic toxicity.

Δ9‐Tetrahydrocannabinol suppression of the primary immune response in rats

The interaction of Δ9‐tetrahydrocannabinol (Δ9‐THC) with the primary immune response was investigated in female Fischer rats receiving sesame oil vehicle or Δ9‐THC in oral doses of 1, 5, or 10 mg/kg. Rats were given a single intraperitoneal injection of sheep red blood cells (SRBC) after, during, or before treatment with Δ9‐THC in order to evaluate the effect of the cannabinoid on the inductive and productive phases of the primary immune response to SRBC. At necropsy, body and spleen weights were recorded, sera were analyzed for antibody to SRBC by an hemagglutination method (HT), and splenic antibody forming cells (AFC) were determined by the localized hemolysis In gel plaque formation. Δ9‐THC inhibited the primary immune response by 33–44% at 10 mg/kg, suppressed the inductive phase by 48–78% at all doses, and impaired the productive phase by 26–59% at the higher doses. These results are the first direct demonstration of a Δ9‐THC‐induced immunosuppression of both phases of the primary immune response.

Cannabinoid‐induced hormone changes in monkeys and rats

Previous findings at various laboratories indicated that cannabinoids distribute to sexual behavior centers in the brain, and endocrine aberrations have consistently been observed in animals treated with cannabis constituents. Subacute and chronic studies were performed to monitor hormone changes in rats and monkeys exposed to marihuana smoke or pure cannabinoids. In oral studies, young Fischer rats of both sexes were given delta 9-tetrahydrocannabinol (delta 9-THC) doses of 2, 10, or 50 mg/kg for 14--180 d and pregnant rats received 1, 5 or 10 mg/kg during gestation and lactation. Other male rats were exposed to marihuana smoke at delta 9-THC doses of 2 or 4 mg/kg for 14 d. Rhesus monkeys of either sex were given oral cannabidiol doses of 30, 100, and 300 mg/kg for 90 d. Serum pituitary, steroid, and thyroid hormone levels were determined by radioimmunoassay. Marihuana smoke (and oral delta 9-THC) depressed testosterone 20--30% and triiodothyronine 17--29%. In pregnant rats, small doses of delta 9-THC suppressed luteinizing hormone, but larger doses elevated both follicle-stimulating hormone and estrogens (approximately 50--100%) without affecting progesterone levels. Prolonged oral administration of delta 9-THC to young rats tended to increase gonadotropins, to which tolerance developed in males. Cannabidiol-treated monkeys responded with slight elevations in luteinizing hormone and follicle-stimulating hormone in males, whereas steroid hormones were essentially unchanged for both sexes. Hormone imbalance may explain cannabinoid-induced embryotoxicity and impaired gonadal function.

Toxic effects of hexachlorobenzene after daily administration to beagle dogs for one year

Abstract Hexachlorobenzene (HCB) was administered daily in gelatin capsules to male and female beagles at 1000, 100, 10, and 1 mg/dog for 12 months. Mortality, anorexia, and weight loss occurred at the highest and, to a lesser degree, at the next lower dose. After 3 months, body weight stabilized or losses were regained. Clinical laboratory changes in severely affected animals which may have been related to malnutrition included anemia, hypoglycemia, hypocalcemia, and testicular degeneration. A dose-related neutrophilia appeared in the animals receiving the two high dosages. The most widespread pathological lesions were confined to the abdomen and included serositis, necrosis, fibrosis, and steatitis of the omentum. Nodular hyperplasia of gastric lymphoid tissue was found in all treated dogs including those at 1 mg/day (6.5–10.0 mg/kg). Four severely affected animals at the highest dose showed a generalized vasculitis and one had amyloidosis. One dog from each of the two highest doses had bile duct hyperplasia and pericholangitis. Bile and perirenal fat showed a time- and dose-related accumulation of HCB. No hepatic fluorescence was found at necropsy, indicating that these dogs were free of porphyria. Nevertheless, hepatic fluorescence was produced in female rats by feeding the same material used in the canine study.

Immunological deficiency associated with cigarette smoke inhalation by mice. Primary and secondary hemagglutinin response.

Inhalation of cigarette whole smoke (CWS) or its vapor phase (CVP) significantly impaired immune response capability in mice. Significant immunosuppressive effects on the humoral antibody response to a single antigenic stimulus were evident in animals exposed to smoke for seven days before or two days after administration of antigen. Impairment of the immunological response capability appeared to be temporary, with recovery about 14 days after exposure. Different lengths of exposure prior to antigenic stimulation neither produced an additive impairment of the immunological response nor rendered the experimental animals more tolerant to CWS or CVP. The immunological deficiency was specific to CWS and CVP inhalation rather than to nonspecific debilitating stress factors. The inductive phase was the period of the primary and secondary immune response most sensitive to impairment by exposure to CWS or CVP.

Specific and Nonspecific Immune Resistance Enhancing Activity of Staphage Lysate

The immunopotentiating activity of staphage lysate (SPL) was evaluated in terms of its immune protection against lethal bacterial infection and its antitumor activity. Mice were pretreated weekly with 10(8) viable, Staphylococcus aureus, strain 18Z for 3 weeks (Induction), followed by intraperitoneal SPL injections (Elicitation) at various times in relation to infectious challenge or tumor implantation. Induction without elicitation, or elicitation alone failed to provide protection against Klebsiella pneumoniae infection and resulted in only 30-40% survival against homologous infection with pathogenic S. aureus type III, whereas combined induction and elicitation produced enhanced resistance induction and elicitation regimens resulted in 50% and 80-100% survival in mice inoculated with K. pneumoniae and S. aureus, respectively. SPL had no antitumor effect in mice implanted with median survival time resulting from induction and elicitation in animals implanted which Ehrlichs ascites. This enhancement of immune resistance may possibly be related to activation of thymus-modulated lymphocytes and macrophages by SPL.

Assessment of Tolerance to Immunosuppressive Activity of Δ9-Tetrahydrocannabinoll in Rats

Immunosuppression evoked by delta 9-tetrahydrocannabinol (delta 9-THC) has been a consistent finding in rats but the development of tolerance to this phenomenon has not been explored. Therefore, Fischer rats of both sexes were orally given delta 9-THC at 6 or 12 mg/kg or sesame oil as vehicle control for 5-26 days before and after I.P. antigenic stimulation with sheep red blood cells (SRBC). delta 9-THC doses were relevant to those of man and produced mild CNS-inhibition followed by CNS-stimulation, tolerance developing to both behavioral phases. The primary immune response was evaluated by determining splenic antibody-forming cells (AFC), hemagglutinin (HT) and/or hemolysin (HS) titers. Simultaneous administration of delta 9-THC and SE induced dose-related splenic atrophy and reduced AFC proliferation as well as HT and HS responses. These changes were not elicited by sesame oil. Tolerance did not develop to immunosuppression during 26 days of cannabinoid treatment. delta 9-THC given 3 days post SRBC inoculation induced immunosuppression at 12 but not 6 mg/kg. Immunosuppression was directly related to delta 9-THC rather than to non-specific debilitating factors since body weights are stable. The inductive phase of the primary immune response was most sensitive to impairment although the reproductive phase was also affected at the high dose level.

Effects of hysterectomy on milk secretion and serum levels of prolactin, growth hormone, estrogen, and progesterone in rhesus monkeys with hormone-induced uterine hypertrophy☆

Abstract Lactation and uterine hypertrophy were induced and maintained in 5 nonpregnant female rhesus monkeys by exogenous estradiol-17β (E 2 ) and progesterone (P). Hypertrophic uteri were then removed surgically. Quantitation of weekly milk secretion and radioimmunoassays of serum estrone (E 1 ), E 2 , P, prolactin, and growth hormone (GH) were carried out from the fifth week prior to hysterectomy and through the eighth week after hysterectomy. The effects of hysterectomy were studied by comparing the serum hormonal levels and milk secretion of the posthysterectomy period with those of the prehysterectomy period. Following removal of the hypertrophic uteri, milk secretion increased from 266 ± 38 to 400 ± 37 mg.; serum levels of E 1 increased from 0.20 ± 0.01 to 0.41 ± 0.02 ng. per milliliter, and E 2 increased from 1.40 ± 0.09 to 1.78 ± 0.12 ng. per milliliter. Slight decreases were observed in serum levels of P (from 36.6 ± 7.6 to 32.3 ± 2.7 ng. per milliliter), prolactin (from 12.3 ± 1.5 to 9.4 ± 1.0 ng. per milliliter), and GH (from 28.8 ± 3.8 to 27.2 ± 6.2 ng. per milliliter.) These decreases were statistically not significant. The increased mammary secretory activity was correlated with the increased levels of circulating estrogen following hysterectomy. Such increase indicates significant competition for estrogen between the uterus and mammary glands.