In-Vitro And Preclinical Trials

Science. 1983 Aug 12;221(4611):671-3.

Naltrexone modulates tumor response in mice with neuroblastoma.

Zagon IS, McLaughlin PJ.

Naltrexone, an opiate antagonist, had both stimulatory and inhibitory effects, depending on the dosage, on the growth of S20Y neuroblastoma in A/Jax mice. Daily injections of 0.1 milligram of naltrexone per kilogram of body weight, which blocked morphine-induced analgesia for 4 to 6 hours per day, resulted in a 33 percent tumor incidence, a 98 percent delay in the time before tumor appearance, and a 36 percent increase in survival time. Neuroblastoma-inoculated mice receiving 10 milligrams of naltrexone per kilogram, which blocked morphine-induced analgesia for 24 hours per day, had a 100 percent tumor incidence, a 27 percent reduction in the time before tumor appearance, and a 19 percent decrease in survival time. Inoculation of neuroblastoma cells in control subjects resulted in 100 percent tumor incidence within 29 days. These results show that naltrexone can modulate tumor response and suggest a role for the endorphin-opiate receptor system in neuro-oncogenic events.

PMID: 6867737

Science. 1983 Sep 16;221(4616):1179-80.

Increased brain size and cellular content in infant rats treated with an opiate antagonist.

Zagon IS, McLaughlin PJ.

From birth to day 21, rat offspring received daily injections of naltrexone at a dosage that blocked morphine-induced analgesia 24 hours a day. At 21 days, body, brain, and cerebellar weights of naltrexone-injected animals were 18, 11, and 5 percent greater than corresponding control weights. In addition, morphometric analysis of the cerebrum revealed a somatosensory cortex that was 18 percent thicker than that of the controls. The cerebellum of naltrexone-treated rats was 41 percent larger in total area and contained at least 70 percent more glial cells and 30 percent more granule neurons. Neurons derived prenatally were unaffected by drug treatment. These results show that naltrexone can stimulate body and brain growth in rats and suggest a role for the endorphin and opiate receptor system in development.

PMID: 6612331

Cancer Lett. 1983 Nov;21(1):89-94.

Opioid antagonists inhibit the growth of metastatic murine neuroblastoma.

Zagon IS, McLaughlin PJ.

Naltrexone (NTX), an opiate antagonist, had an inhibitory effect on the growth of S20 Y neuroblastoma (NB) in A/Jax mice. Daily injections of 0.1 mg/kg NTX resulted in a 69% tumor take, 70% delay in time prior to tumor appearance, and a 60% increase in median survival time. Inoculation of NB in control mice resulted in 100% tumor take within 15 days. The pattern and incidence of metastases of NTX and control mice were similar. These results show that NTX has antineoplastic activity, and suggests a role for the endogenous opioid system in neuro-oncogenic events.

PMID: 6640516

Life Sci. 1983 Dec 12;33(24):2449-54.

Naltrexone modulates growth in infant rats.

Zagon IS, McLaughlin PJ.

Naltrexone, a potent opiate antagonist, had both stimulatory and inhibitory effects on somatic growth in preweaning rats depending on dose. Daily injections of 50 mg/kg naltrexone, which blocked morphine-induced analgesia for 24 hr/day, resulted in increased body and organ weights, and acceleration in the appearance of physical characteristics and maturation of spontaneous motor activity. Naltrexone in a dosage of 1 mg/kg, which blocked morphine-induced analgesia for 4 hr/day, had the opposite effects. These results show that naltrexone can modulate growth, and suggest a role for the endorphins and opiate receptors in developmental events.

PMID: 6316064

Life Sci. 1984 Jul 23;35(4):409-16.

Duration of opiate receptor blockade determines tumorigenic response in mice with neuroblastoma: a role for endogenous opioid systems in cancer.

Zagon IS, McLaughlin PJ.

The relationship between the pharmacological properties of an opioid antagonist, naltrexone (NTX), and tumor response was studied in mice with transplanted neuroblastoma (NB). Animals receiving 0.1 mg/kg NTX every 6 hr, which blocked morphine-induced analgesia for 24 hr each day, had a 100% tumor incidence, no deviation in time before tumor appearance, and a 17% decrease from control values in total survival time. In contrast, once daily injections of either 0.1 mg/kg NTX or 0.4 mg/kg NTX (the equivalent of 0.1 mg/kg given 4 times daily), which blocked morphine-induced analgesia for less than 10 hr each day, resulted in a tumor incidence of 20% and 60%, respectively, delays in time prior to tumor appearance of 90% and 65%, respectively, and an increased total survival time of 10% and 24%, respectively, for tumor-bearing mice relative to control levels. Inoculation of NB in control animals resulted in 100% tumor appearance within 16 days and a mean survival time of 36 days. These results show that tumorigenic events are dictated by the duration of opiate receptor blockade rather than the dosage of opiate antagonist, and provide compelling evidence that endogenous opioid systems play a crucial role in neuro-oncogenic expression.

PMID: 6087062

Life Sci. 1984 Nov 12;35(20):2057-64.

Naltrexone modulates body and brain development in rats: a role for endogenous opioid systems in growth.

Zagon IS, McLaughlin PJ.

Preweaning rats receiving daily injections of 20, 50, or 100 mg/kg naltrexone, a potent opiate antagonist, had body and brain weights that were increased 16-22% and 6-13%, respectively, from control levels on day 21 (weaning). All of these dosages of naltrexone blocked the opiate receptor for 24 hr/day as measured in opiate challenge experiments. Dosages of 0.1, 1, and 10 mg/kg naltrexone, which blocked the opiate receptor for less than 12 hr/day, inhibited growth. Repetitive administration of low dosages (3 mg/kg naltrexone, 3 times daily), which blocked the receptor 24 hr/day, increased body and brain development by 31% and 10%, respectively, whereas a cumulative dosage of 9 mg/kg naltrexone given once daily retarded growth. These results show that developmental events are dictated by the duration of opiate receptor blockade and provide compelling evidence that endogenous opioid systems play a crucial role in growth.

PMID: 6092812

Physiol Behav. 1985 Apr;34(4):507-11.

Opioid antagonist-induced regulation of organ development.

Zagon IS, McLaughlin PJ.

Naltrexone, a potent opioid antagonist, was given to preweaning rats in order to explore the influence of endogenous opioid systems on organogenesis. Sprague-Dawley rats were injected (SC) daily with either 1 or 50 mg/kg naltrexone to invoke a temporary or complete blockade, respectively, of opioid receptors; animals injected with sterile water served as controls. At weaning (Day 21), wet and dry weights, relative organ weight, and tissue water content were determined in 10 organ systems. Naltrexone's effects on growth depended on dosage, sex, and the organ system examined. In general, dosages of 1 and 50 mg/kg naltrexone caused significant decreases and increases, respectively, in organ weight. These changes in wet weight were not due to the state of hydration, but rather to dry weight, indicating that the content of cellular matter was altered. The changes in wet weight were similar to those for body weight, suggesting that a proportional increase or decrease in animal growth took place. Although the same organs in males and females within a dosage group were influenced by naltrexone, and usually to a similar degree, a dosage of 1 mg/kg naltrexone often affected different organ systems than the 50 mg/kg dosage. These results serve as the foundation for subsequent investigations directed towards delineating the role of endogenous opioid systems in developmental biology.

PMID: 4011730

J Natl Cancer Inst. 1987 Jan;78(1):141-7.

Modulation of murine neuroblastoma in nude mice by opioid antagonists.

Zagon IS, McLaughlin PJ.

Erratum in: J Natl Cancer Inst 1987 Mar;78(3):593.

Naltrexone, an opioid antagonist, had an inhibitory effect on the growth of murine S20Y neuroblastoma in BALB/c nude mice. Daily injections of 0.1 mg naltrexone/kg, which invoked a receptor blockade for 6-8 hours/day, resulted in 31-92% delay in latency time prior to tumor expression and a 27-49% increase in mean survival time; the magnitude of antitumor response was governed by tumor burden. Inoculation of neuroblastoma (10(6)-2.5 X 10(4) cells) resulted in measurable tumors in 10-13 days and mean survival times of 30-34 days. Immunoreactive beta-endorphin was detected in tumor tissue (39.7 pg/mg protein). Receptor binding assays revealed specific saturable binding of ligands related to delta- and kappa-binding sites, but not for the mu-binding site. These results demonstrate that opioid antagonist modulation of neuro-oncogenesis is not dependent on the integrity of T-cell-mediated immunity and suggest the feasibility of utilizing the nude mouse model in exploring the role of endogenous opioids in human cancers.

PMID: 3025501

Brain Res. 1987 May 26;412(1):68-72.

Endogenous opioid systems regulate cell proliferation in the developing rat brain.

Zagon IS, McLaughlin PJ.

The role of endogenous opioid systems in modulating the proliferation of developing cerebellar cells was examined autoradiographically in 6-day-old rats. The blockade of endogenous opioid-opioid receptor interaction by naltrexone, a potent opioid antagonist, was accompanied within 1-2 h by an increased proportion of cells incorporating [3H]thymidine. When high doses of naltrexone (50 mg/kg) were administered this index was still elevated 12 h later; however, when low doses of naltrexone (1 mg/kg) were administered the index of labeled cells was decreased markedly. Injection of methionine-enkephalin, an endogenous opioid peptide, also resulted in a decrease in the proportion of cells incorporating [3H]thymidine. Concomitant injection of 1 mg/kg naloxone, however, blocked the inhibitory effects of methionine-enkephalin on cell division but did not itself affect cell generation. These studies demonstrate that endogenous opioid systems can regulate the proliferation of cell populations in the developing nervous system and do so through an inhibitory mechanism.

PMID: 3607463

Life Sci. 1987 Sep 21;41(12):1465-72.

Modulation of human neuroblastoma transplanted into nude mice by endogenous opioid systems.

McLaughlin PJ, Zagon IS.

The role of endogenous opioid systems (endogenous opioids and opioid receptors) in human cancer was explored using an opioid antagonist paradigm and neuroblastoma cells (SK-N-MC) transplanted into nude mice. Mice inoculated with 2.5 X 10(6) neuroblastoma cells received daily injections of either 0.1 or 10 mg/kg naltrexone (=0.1 and 10 NTX groups) which blocked the opioid receptor for 6-8 hr/day or the entire 24 hr/day, respectively, or sterile water. The latency for appearance of a measurable tumor (5 mm diameter) in the 0.1 NTX group was 27% longer than controls (11 days), and the first death in this group occurred 33% later than controls (day 27). Mice inoculated with tumor cells in the 10 NTX group had an acceleration (18%) in the latency of tumor appearance and, 2 weeks after cell inoculation, 70% of the mice in this group had tumors, in contrast to 10% of the controls. At the termination of the experiment (day 45), only 33% of the 10 NTX group were alive, in contrast to 90% of the controls. Receptor binding assays using DAGO, DADLE, or EKC revealed specific saturable binding only for DADLE and EKC. NTX administration resulted in a 148-186% increase in density for both binding sites, but no changes in binding affinity. Measures of opioid levels showed that tumor tissue levels of both beta-endorphin and methionine-enkephalin were elevated 2.5 to 6.5 fold from control values in both NTX groups, whereas plasma beta-endorphin was subnormal by 4 to 6 fold. These results indicate that endogenous opioid systems regulate human neuro-oncogenesis, with opioids being active inhibitors of growth. Opioid antagonists up-regulate receptors and increase tissue levels of endogenous opioids and, under conditions in which the opioid antagonist is short-acting (e.g., 0.1 NTX), can have an exaggerated antitumor effect during the interval when the antagonist is no longer present.

PMID: 3041143

Life Sci. 1988;43(16):1313-8.

Endogenous opioids and the growth regulation of a neural tumor.

Zagon IS, McLaughlin P.

Department of Anatomy, Milton S. Hershey Medical Center, Pennsylvania State University, Hershey 17033.

Endogenous opioid systems (endogenous opioids and their receptors) are known to participate in the regulation of tumor growth. The present study was conducted to examine whether [Met5]-enkephalin influences the growth of transplanted neuroblastoma, and to explore the role of other opioid peptides in carcinogenesis. A/Jax mice were inoculated with 10(6) S20Y cells and received daily injections of [Met5]-enkephalin. Dosages of 0.5 to 30 mg/kg delayed tumor appearance and prolonged survival of these mice; antitumor effects were blocked by concomitant injections of naloxone. Daily administration (10 mg/kg) of [Leu5]-enkephalin had no effect on neurotumor growth. [D-Ala2, D-Leu5]-enkephalin and ethylketocyclazocine, ligands selective for delta and kappa receptors, respectively, also did not influence neuro-oncogenesis. These results demonstrated the potent growth inhibiting effects of the naturally occurring opioid pentapeptide, [Met5]-enkephalin, and substantiate reports identifying and characterizing an opioid receptor (i.e., zeta) for which [Met5]-enkephalin is the most potent ligand.

PMID: 2845218

Brain Res. 1989 Feb 20;480(1-2):16-28.

Opioid antagonist modulation of murine neuroblastoma: a profile of cell proliferation and opioid peptides and receptors.

Zagon IS, McLaughlin PJ.

Department of Anatomy, M.S. Hershey Medical Center, Pennsylvania State University, Hershey 17033.

The endogenous opioids and their receptors are known to play a major role in neoplasia. In the present study, naltrexone (NTX), a potent opioid antagonist, was utilized to explore the interactions of opioids and opioid receptors in mice with transplanted neuroblastoma (S20Y). Tumors from mice subjected to either intermittent (4-6h/day; 0.1 mg/kg NTX) or complete (24 h/day; 10 mg/kg NTX) opioid receptor blockade exhibited an up-regulation of DADLE and Met-enkephalin binding sites, as well as tissue levels of beta-endorphin and Met-enkephalin. Binding affinity to [D-Ala2,D-Leu5]enkephalin (DADLE) or ethylketocyclazocine (EKC), the levels of plasma beta-endorphin, and the anatomical location and quantity of Met- and Leu-enkephalin and cytoskeletal components (i.e. tubulin, actin, brain spectrin (240/235) were similar in NTX and control tumor-bearing animals. Tissue viability of the 0.1 NTX group was increased compared to controls. Both mitotic and labeling indexes were increased during the period of opioid receptor blockade, but decreased in the period subsequent to receptor blockade. NTX treatment produced a 2-fold increased in sensitivity to opioids. Met-enkephalin (10 mg/kg) produced a depression in both mitotic and labeling indexes in tumor-bearing mice that could be reversed by naloxone (10 mg/kg) administration. Thus, the endogenous opioids are trophic agents that inhibit growth by suppressing cell proliferation. The duration of receptor blockade by opioid antagonists modulates these actions, affecting both tumor incidence and survival time. Complete opioid receptor block prevents the interaction of increased levels of putative growth-related peptides with a greater number of opioid receptors, thereby increasing cell proliferation and accelerating tumor growth. With intermittent blockade, an enhanced opioid-receptor interaction occurs during the interval when the opioid antagonist is no longer present, producing an exaggerated inhibitory action on cell proliferation and the repression of tumorigenic events.

PMID: 2540873

Neuroscience. 1990;37(1):223-6.

Opioid antagonist (naltrexone) stimulation of cell proliferation in human and animal neuroblastoma and human fibrosarcoma cells in culture.

Zagon IS, McLaughlin PJ.

Department of Anatomy, M. S. Hershey Medical Center, Pennsylvania State University, Hershey 17033.

Endogenous opioids play a role in carcinogenic events by serving as inhibitory growth factors that alter cell proliferative events by interaction with opioid receptors. The present study addresses the question of whether endogenous opioid systems function tonically in tissue culture. Using S20Y neuroblastoma, a cell line that produces a growth-related opioid peptide (i.e.[Met5]enkephalin) and contains the zeta receptor known to be associated with growth, the effects of opioid receptor blockade by naltrexone, a potent opioid antagonist, was examined. Drug concentrations of 10(-4) to 10(-8) M naltrexone stimulated cell proliferation, with 32-86% more cells found in the naltrexone groups than control from 12 to 48 h after initiating drug exposure; drug concentrations of 10(-9) to 10(-13) M had no effect on growth. Evaluation of labeling and mitotic indices revealed that both DNA synthesis and mitosis were increased by naltrexone, as were the number of cells with process lengths greater than 40 microns. Naltrexone (10(-6) M) also stimulated the growth of N115 murine neuroblastoma, SK-N-MC human neuroblastoma, and HT-1080 human fibrosarcoma. These results indicate that endogenous opioids function in vitro to regulate growth by inhibitory mechanisms, and do so actively. This autocrine mechanism in tissue culture also occurs in other animal neuroblastoma cell lines, as well as for human neuroblastoma and fibrosarcoma cell lines.

PMID: 2243594

Cancer Lett. 1996 Mar 29;101(2):159-64.

Inhibition of human colon cancer by intermittent opioid receptor blockade with naltrexone.

Hytrek SD, McLaughlin PJ, Lang CM, Zagon IS.

Department of Comparative Medicine, Pennsylvania State University, College of Medicine, Hershey 17033, USA.

Nude mice inoculated with human colon cancer (HT-29) and receiving 0.1 mg/kg naltrexone (NTX) beginning immediately after tumor cell injection exhibited a marked retardation in tumorigenicity. This dosage of NTX, which blocked opioid receptors for 6-8 h/day, resulted in a delay of 2.4-fold in tumor appearance compared to control subjects. At the time (10 days) when all control mice had tumors, 80% of the mice in the 0.1 mg/kg NTX group had no signs of neoplasia. Binding capacity, but not affinity, of [3H][Met5]-enkephalin was reduced 85% of control levels in tumor tissue from mice of the 0.1 NTX group. Plasma, but not tumor tissue levels of [Met5]-enkephalin were elevated (2.5-fold) in contrast to control values. These results suggest that daily intermittent opioid receptor blockade with NTX provokes the interaction of opioids and receptors in the interval following drug availability, with opioids serving to inhibit tumorigenicity of human colon cancer.

PMID: 8620464

Clinical Trials

In-Vitro And Preclinical Trials