Prevention of liver metastases through perioperative acute CpG-C immune stimulation.

Following excision of colorectal tumors, metastatic disease is prevalent, primarily occurs in the liver, and is highly predictive of poor prognosis. The perioperative period is now recognized as critical in determining the incidence of postoperative metastases and long-term cancer outcomes. Thus, various perioperative prophylactic interventions are currently studied during this time frame. However, immune stimulation during the perioperative period has rarely been attempted due to specific contraindications to surgery and various adverse effects. Here, to prevent liver metastases, we perioperatively employed a TLR-9 agonist, CpG-C, which exhibits minimal pyrogenic and other adverse effects in patients. We found that marginating-hepatic (MH) cells in BALB/c mice contained high percentage of NK cells, but exhibited negligible NK cytotoxicity, as previously reported in humans. However, a single CpG-C administration (25-100 µg/mouse) doubled MH-NK cell numbers, increased NK cell activation and maturation markers (NKp46, CD11b), decreased the inhibitory NKG2A ligand, and dramatically increased MH-NK-cell cytotoxicity against the syngeneic CT26 colon cancer line. Moreover, in operated mice, this innocuous intervention also markedly improved resistance to CT26 and MC38 hepatic metastases in BALB/c and C57BL/6 mice, respectively. Beneficial effects of CpG-C were mediated through activation of MH-NK cells, as indicated by an in vivo NK depletion study. Last, CpG-C protected against surgery-induced suppression of MH-NK cytotoxicity and improved their activation indices. Thus, we suggest that systemic perioperative CpG-C treatment should be considered and studied as a novel therapeutic approach to improve long-term cancer outcomes in colorectal cancer patients.

Deleterious synergistic effects of distress and surgery on cancer metastasis: Abolishment through an integrated perioperative immune-stimulating stress-inflammatory-reducing intervention.

The perioperative period holds disproportionate impact on long-term cancer outcomes. Nevertheless, perioperative interventions to improve long-term cancer outcomes are not clinical routines, including perioperative stress-reducing or immune-stimulating approaches. Here, mimicking the clinical setting of pre-operative distress, followed by surgery, we examined the separate and combined effects of these events on the efficacy of pre-operative immune stimulation in rats and mice, and on post-operative resistance to tumor metastasis of the syngeneic mammary adenocarcinoma MADB106 in F344 rats and the CT26 colon carcinoma in Balb/C mice. The novel immune stimulating agents, GLA-SE or CpG-C (TLR-4 and TLR-9 agonists, respectively), were employed pre-operatively. Sixteen hours of pre-operative behavioral stressors (i) lowered CpG-C induced plasma IL-12 levels, and reduced resistance to MADB106 and CT-26 experimental metastases, and (ii) worsened the deleterious effects of laparotomy on metastasis in both tumor models. In rats, these effects of pre-operative stress were further studied and successfully abolished by the glucocorticoid receptor antagonist RU-486. Additionally, in vitro studies indicated the dampening effect of corticosterone on immune stimulation. Last, we tested a perioperative integrated intervention in the context of pre-operative stress and laparotomy, based on (i) antagonizing the impact of glucocorticoids before surgery, (ii) activating anti-metastatic immunity perioperatively, and (iii) blocking excessive operative and post-operative adrenergic and prostanoid responses. This integrated intervention successfully and completely abolished the deleterious effects of stress and of surgery on post-operative resistance to experimental metastasis. Such and similar integrated approaches can be studied clinically in cancer patients.

Maintaining unperturbed cerebral blood flow is key in the study of brain metastasis and its interactions with stress and inflammatory responses.

Blood-borne brain metastases are associated with poor prognosis, but little is known about the interplay between cerebral blood flow, surgical stress responses, and the metastatic process. The intra-carotid inoculation approach, traditionally used in animal studies, involves permanent occlusion of the common carotid artery (CCA). Herein we introduced a novel intra-carotid inoculation approach that avoids CCA ligation, namely - assisted external carotid artery inoculation (aECAi) - and compared it to the traditional approach in C57/BL6 mice, assessing cerebral blood flow; particle distribution; blood-brain barrier (BBB) integrity; stress, inflammatory and immune responses; and brain tumor retention and growth. Doppler flowmetry and two-photon imaging confirmed that only in the traditional approach regional and capillary cerebral blood flux were significantly reduced. Corticosterone and plasma IL-6 levels were higher in the traditional approach, splenic numbers of NK, CD3+, granulocytes, and dendritic cells were lower, and many of these indices were more profoundly affected by surgical stress in the traditional approach. BBB integrity was unaffected. Administration of spherical beads indicated that CCA ligation significantly limited brain distribution of injected particles, and inoculation of D122-LLC syngeneic tumor cells resulted in 10-fold lower brain tumor-cell retention in the traditional approach. Last, while most of the injected tumor cells were arrested in extra-cranial head areas, our method improved targeting of brain-tissue by 7-fold. This head versus brain distribution difference, commonly overlooked, cannot be detected using in vivo bioluminescent imaging. Overall, it is crucial to maintain unperturbed cerebral blood flow while studying brain metastasis and interactions with stress and inflammatory responses.

Perioperative treatment with the new synthetic TLR-4 agonist GLA-SE reduces cancer metastasis without adverse effects.

The use of TLR agonists as an anti-cancer treatment is gaining momentum given their capacity to activate various host cellular responses through the secretion of inflammatory cytokines and type-I interferons. It is now also recognized that the perioperative period is a window of opportunity for various interventions aiming at reducing the risk of cancer metastases-the major cause of cancer related death. However, immune-stimulatory approach has not been used perioperatively given several contraindications to surgery. To overcome these obstacles, in this study, we used the newly introduced, fully synthetic TLR-4 agonist, Glucopyranosyl Lipid-A (GLA-SE), in various models of cancer metastases, and in the context of acute stress or surgery. Without exerting evident adverse effects, a single systemic administration of GLA-SE rapidly and dose dependently elevated both innate and adaptive immunity in the circulation, lungs and the lymphatic system. Importantly, GLA-SE treatment led to reduced metastatic development of a mammary adenocarcinoma and a colon carcinoma by approximately 40-75% in F344 rats and BALB/c mice, respectively, at least partly through elevating marginating-pulmonary NK cell cytotoxicity. GLA-SE is safe and well tolerated in humans, and currently is used as an adjuvant in phase-II clinical trials. Given that the TLR-4 receptor and its signaling cascade is highly conserved throughout evolution, our current results suggest that GLA-SE may be a promising immune stimulatory agent in the context of oncological surgeries, aiming to reduce long-term cancer recurrence.

Reducing liver metastases of colon cancer in the context of extensive and minor surgeries through ß-adrenoceptors blockade and COX2 inhibition.

Liver metastases are a major cause of colorectal cancer death, and the perioperative period is believed to critically affect the metastatic process. Here we tested whether blocking excess release of catecholamines and prostaglandins during surgical procedures of different extent can reduce experimental liver metastasis of the syngeneic CT26 colon cancer in female and male BALB/c mice. Animals were either treated with the beta-blocker, propranolol, the COX-2 inhibitor, etodolac, both drugs, or vehicle. The role of NK cells in controlling CT26 hepatic metastasis and in mediating the effect of the drugs was assessed by in vivo depletion or stimulation of NK cells, using anti-asialo GM1 or CpG-C, respectively. Surgical extent was manipulated by adding laparotomy to small incision, extending surgical duration, and enabling hypothermia. The results indicated that combined administration of propranolol and etodolac, but neither drug alone, significantly improved host resistance to metastasis. These beneficial effects occurred in both minor and extensive surgeries, in both sexes, and in two tumor inoculation approaches. NK cell-mediated anti-CT26 activity is involved in mediating the beneficial effects of the drugs. Specifically, CpG-C treatment, known to profoundly activate mice marginating-hepatic NK cytotoxicity, reduced CT26 hepatic metastases; and NK-depletion increased metastases and prevented the beneficial effects of the drugs. Overall, given prevalent perioperative psychological and physiological stress responses in patients, and ample prostaglandin release by colorectal tumors and injured tissue, propranolol and etodolac could be tested clinically in laparoscopic and open colorectal surgeries, attempting to reduce patients' metastatic disease.

Improving postoperative immune status and resistance to cancer metastasis: a combined perioperative approach of immunostimulation and prevention of excessive surgical stress responses.

BACKGROUND: Surgical procedures, including primary tumor resection, have been suggested to suppress immune competence and to promote postoperative infections and cancer metastasis. Catecholamines and prostaglandins were recently implicated in these processes, and in directly promoting tumor angiogenesis and invasion. OBJECTIVE: To examine the integration of 2 complementary approaches to reduce postoperative immunosuppression and metastatic progression: (1) perioperative immunostimulation with CpG-C and (2) pharmacological blockade of the tumor-promoting and immunosuppressing effects of catecholamines and prostaglandins, using propranolol (P) and etodolac (E), respectively. METHODS: F344 rats were treated before surgery with CpG-C, P+E, both interventions, or vehicles, and were intravenously inoculated with syngeneic MADB106 mammary adenocarcinoma cells. Blood was withdrawn, marginating-pulmonary leukocytes were harvested, and NK activity and lung MADB106 tumor retention were assessed. In addition, C57BL/6 mice were implanted with syngeneic B16F10.9 melanoma cells. When tumors reached 100 mm, mice were treated with CpG-C/vehicle, and 24 hours later the tumor was excised along with P+E/vehicle treatment. Recurrence-free survival was monitored thereafter. RESULTS: Each of the regimens alone, CpG-C or P+E, showed improvement in most indices examined, including improved long-term recurrence-free survival rates. Most importantly, the combined treatment yielded additive or synergistic effects, further improving tumor clearance from the lungs and enhancing NK numbers and cytotoxicity via different, but complimentary, mechanisms. CONCLUSIONS: Treatment aimed at perioperative enhancement of CMI and simultaneous inhibition of excessive catecholamine and prostaglandin responses, employing CpG-C, propranolol, and etodolac, could be successful in limiting postoperative immunosuppression and metastatic progression, more so than each treatment alone.

Continuous stress disrupts immunostimulatory effects of IL-12.

Immune stimulation by biological response modifiers is a common approach in tumor immunotherapy. IL-12 was found effective in various animal studies, but clinical trials showed limited success. However, among other differences, animal models do not simulate psychological or physiological stress while employing IL-12, whereas cancer patients often experience distress while treated with immunostimulants. Thus, in the current study we assessed the impact of continuous stress on the efficacy of IL-12 immunostimulation. F344 rats were subjected to a pharmacological stress paradigm (continuous administration of a ß-adrenergic agonist) or to a 20 h behavioral stress paradigm (wet cage exposure) commencing 2h before IL-12 administration. Twenty-six hours after stress initiation, we studied indices known to reflect IL-12 immunostimulatory impacts, including NK cell numbers and activity in different immune compartments, and in vivo resistance to MADB106 lung tumor colonization. The results indicated that both the pharmacological and behavioral stress paradigms significantly reduced the increase in the number and activity of marginating-pulmonary NK cells evident in non-stressed IL-12 treated animals. Additionally, stressed animals exhibited a lower IL-12-induced improvement of MADB106 lung clearance, an in vivo index that markedly depends on total marginating-pulmonary NK activity. These deleterious effects of stress were more prominent in males than in females. Overall, the findings demonstrate that prolonged stress exposure can disrupt the efficacy of simultaneous immunostimulatory treatments, irrespective of stress effects on baseline immune measures. Neuroendocrine and cellular mediating mechanisms are yet unknown, but the potential clinical ramifications of these findings warrant consideration in clinical trials employing immunostimulatory agents.

Immune perturbations in patients along the perioperative period: alterations in cell surface markers and leukocyte subtypes before and after surgery.

BACKGROUND: Surgery renders patients susceptible to life-threatening complications, including infections, multiple organ failure, and presumably cancer metastases. Surgery-induced immune perturbations were suggested to contribute to such deleterious effects, but also to facilitate post-injury healing. Preoperative psychological and physiological stress responses may contribute to these immune perturbations, and could thus jeopardize patients even before surgery. The current study assessed the effects of various operations on an array of immune indices during the perioperative period. To qualify immune changes before surgery, patients' immune status was also compared to that of healthy controls. METHODS: A total of 81 subjects (operated patients and healthy controls) provided up to five daily blood samples during the perioperative period, for assessment of leukocyte subtypes (granulocytes, monocytes, Tc, Th, NK, NKT, CD4+CD25+, CD8(bright)CD4(dim), and B cells) and their surface markers (HLA-DR and LFA-1). RESULTS: Even before surgery patients displayed immune perturbations, including reduced lymphocyte HLA-DR expression and increased monocyte LFA-1 expression. Following surgery, we recorded a reduction in lymphocyte numbers that was subtype specific, increased granulocyte numbers, and reduced expression of HLA-DR by lymphocytes and monocytes. Finally, no significant associations were found between alteration in leukocyte numbers and cell surface markers (although these indices showed high correlations with other variables), implying differential mediating mechanisms. CONCLUSION: Several immune alterations are manifested prior to surgery, and contribute to the marked postoperative changes, which are commonly interpreted as immune suppression. We discuss the possible adaptive and maladaptive nature of these perturbations in the context of natural injury, stress, and surgery.

Selective Harvesting of Marginating-pulmonary Leukocytes.

Marginating-pulmonary (MP) leukocytes are leukocytes that adhere to the inner endothelium of the lung capillaries. MP-leukocytes were shown to exhibit unique composition and characteristics compared to leukocytes of other immune compartments. Evidence suggests higher cytotoxicity of natural killer cells, and a distinct pro- and anti-inflammatory profile of the MP-leukocyte population compared to circulating or splenic immunocytes. The method presented herein enables selective harvesting of MP-leukocytes by forced perfusion of the lungs in either mice or rats. In contrast to other methods used to extract lung-leukocytes, such as tissue grinding and biological degradation, this method exclusively yields leukocytes from the lung capillaries, uncontaminated with parenchymal, interstitial, and broncho-alveolar cells. In addition, the perfusion technique better preserves the integrity and the physiological milieu of MP-leukocytes, without inducing physiological responses due to tissue processing. This unique MP leukocyte population is strategically located to identify and react towards abnormal circulating cells, as all circulating malignant cells and infected cells are detained while passing through the lung capillaries, physically interacting with endothelial cells and resident leukocytes,. Thus, selective harvesting of MP-leukocytes and their study under various conditions may advance our understanding of their biological and clinical significance, specifically with respect to controlling circulating aberrant cells and lung-related diseases.

Selective Harvesting of Marginating-hepatic Leukocytes.

Marginating-hepatic (MH) leukocytes (leukocytes adhering to the sinusoids of the liver), were shown to exhibit unique composition and characteristics compared to leukocytes of other immune compartments. Specifically, evidence suggests a distinct pro- and anti-inflammatory profile of the MH-leukocyte population and higher cytotoxicity of liver-specific NK cells (namely, pit cells) compared to circulating or splenic immunocytes in both mice and rats. The method presented herein enables selective harvesting of MH leukocytes by forced perfusion of the liver in mice and rats. In contrast to other methods used to extract liver-leukocytes, including tissue grinding and biological degradation, this method exclusively yields leukocytes from the liver sinusoids, uncontaminated by cells from other liver compartments. In addition, the perfusion technique better preserves the integrity and the physiological milieu of MH leukocytes, sparing known physiological responses to tissue processing. As many circulating malignant cells and infected cells are detained while passing through the liver sinusoids, physically interacting with endothelial cells and resident leukocytes, the unique MH leukocyte population is strategically located to interact, identify, and react towards aberrant circulating cells. Thus, selective harvesting of MH-leukocytes and their study under various conditions may advance our understanding of the biological and clinical significance of MH leukocytes, specifically with respect to circulating aberrant cells and liver-related diseases and cancer metastases.

The marginating-pulmonary immune compartment in mice exhibits increased NK cytotoxicity and unique cellular characteristics.

To test whether marginating-pulmonary (MP) leukocytes in mice have a unique potential to identify and destroy aberrant circulating cells, we compared MP to circulating leukocytes with respect to natural killer (NK) cytotoxicity, proinflammatory characteristics, molecular determinants of activation, and response to IL-12 immunostimulation. Cytotoxicity was assessed employing the YAC-1, B16F10, and 3LL target lines. C57BL/6 mice were injected with either saline or murine IL-12 (0.1 or 0.5 µg/mouse), either once or three times 48-h apart. Twenty-four hours after last injection, cardiac blood was withdrawn and MP leukocytes were collected by forced lung perfusion. NK cytotoxicity, cellular composition, and surface molecular markers were studied. MP leukocytes exhibited greater NK cytotoxicity than circulating leukocytes against the syngeneic B16F10 and 3LL tumor lines, but not against the allogeneic YAC-1 line. NKG2D and IL-12 receptor expression predicted NK cytotoxicity in circulating leukocytes, but not in MP leukocytes. IFNγ-receptor, IL-12-receptor, CD69, CD11a, and CD11b showed different patterns of expression in the two leukocyte populations, suggesting pro-inflammatory characteristics of the MP compartment. IL-12 stimulation caused differential effects on these markers and also elevated cytotoxicity in both compartments, but in different effector: target ratio-dependent patterns. MP leukocytes may play a critical role in eliminating aberrant circulating cells due to their enhanced NK cytotoxicity and given their strategic location in the lungs vasculature, which forces physical interactions with all circulating aberrant cells. MP-NK cells are unique in their cytotoxic mechanisms against syngeneic targets and in their activation profile and response to immunostimulatory agents.

The marginating-pulmonary immune compartment in rats: characteristics of continuous inflammation and activated NK cells.

A significant role has been indicated for cellular immunity in controlling circulating cancer cells, but most autologous tumor cells seem resistant, in vitro, to natural killer cell (NKC) and cytotoxic T lymphocytes cytotoxicity. Addressing this apparent contradiction, we recently identified a unique leukocyte population, marginating-pulmonary (MP)-leukocytes, which exhibit potent natural killer (NK) cytotoxicity. Here, we characterize the MP-compartment in naive and immunostimulated rats, and assessed its cytotoxicity against "NK-resistant" tumors cells. Animals were treated with poly I-C (3x0.2 mg/kg) or saline, and circulating-leukocytes and MP-leukocytes were collected and analyzed in terms of cellular composition, cellular activation markers, and NK cytotoxicity of leukocytes and purified NKCs. Compared with circulating-leukocytes, MP-leukocytes showed greater proportion of granulocytes, monocytes, NKCs, and large NKCs; higher expression of activation and adhesion markers (CD25, CD11a, CD11b, and NKR-P1, IFN-gamma); and elevated NK cytotoxicity of leukocytes and purified NKCs against several syngeneic and xenogeneic NK-resistant target cells (from both F344 and BDX inbred rats). In immunostimulated animals (treated with poly I-C), but not in naive animals, purified NKCs from the MP-compartment showed markedly superior cytotoxicity, suggesting that poly I-C immunostimulation uniquely affect MP-NKCs, and that in naive animals other MP-leukocytes support NK cytotoxicity. Overall, the results suggest that the MP-compartment is characterized by a continuous activated inflammatory microenvironment uniquely affected by immunostimulation. If similarly potent MP-NKCs exist in patients, then circulating autologous tumor cells that are considered "NK-resistant" could actually be controlled by MP-NKCs. Innate immunity may assume greater role in controlling malignant spread, especially after immunostimulation.

CpG-C oligodeoxynucleotides limit the deleterious effects of beta-adrenoceptor stimulation on NK cytotoxicity and metastatic dissemination.

Suppression of natural killer (NK) cell activity is common after stress, has been reported to predict malignant recurrence in cancer patients, and was shown to underlie metastatic dissemination in animal models. We have previously reported that catecholamines play a major role in NK cell suppression, particularly in the context of physiologic stress and surgery. In the current study using Fisher 344 rats, we examined the prophylactic use of different regimens of type-C CpG oligodeoxynucleotides (CpG-C ODN) on NK activity and metastatic dissemination in the context of pharmacologic stress (using metaproterenol for beta-adrenoceptor stimulation). Our results indicated that the beneficial effects of CpG-C ODN were more profound under pharmacologic stress than under baseline conditions. A bolus of CpG-C ODN (330 microg/kg, intraperitoneally) 24 hours before metaproterenol-challenge was most effective at reducing lung tumor retention of an experimental syngeneic mammary adenocarcinoma (MADB106), although having no observable side effects. Depletion of NK cells revealed their key role in improving baseline levels of resistance to metastatic dissemination after CpG-C ODN administration. When NK cell cytotoxicity was assessed in the circulation and the marginating-pulmonary immune compartments, we found that CpG-C ODN protected individual NK cells from metaproterenol-induced suppression in both compartments. Moreover, in the critical marginating-pulmonary compartment, CpG-C ODN also elevated baseline cytotoxicity per NK cell against MADB106 tumor cells, and increased NK cell numbers in nonstressed rats. Overall, prophylactic CpG-C ODN treatment can improve immunocompetence and potentially reduce metastatic dissemination, especially in clinical settings characterized by enhanced sympathetic stress responses.

Inducing a mode of NK-resistance to suppression by stress and surgery: a potential approach based on low dose of poly I-C to reduce postoperative cancer metastasis.

Perioperative suppression of NK activity has been suggested to compromise host resistance to tumor progression. Here, we sought to develop a clinically applicable preoperative regimen to prevent immunosuppression and promotion of metastasis by stress or surgery. The synthetic ds-RNA, poly I-C, was used in vivo in F344 rats, based on its alleged in vitro ability to protect immunocytes from suppression by cAMP elevating agents. Different regimens of poly I-C were studied in controls and in rats subjected to a pharmacological stressor, swim stress, or surgical stress. Resistance to lung experimental metastasis of the syngeneic non-immunogenic MADB106 mammary adenocarcinoma was assessed. Numbers of circulating and marginating-pulmonary NK cells and their cytotoxicity against the MADB106 and YAC-1 target lines were also studied. Our findings established a regimen of repeated low-dose poly I-C administration with minimal side effects (0.2mg/kg i.p. 5, 3, and 1day before tumor inoculation). This regimen, while hardly affecting resistance levels in non-stressed animals, prevented all stressors from promoting metastases. These beneficial effects occurred in the presence of a primary tumor and in both sexes. Poly I-C increased the numbers of NK cells, and, on a per NK cell basis, while not increasing cytotoxicity, profoundly protected marginating-pulmonary NK cells from suppression by surgery. This study suggests a non-toxic clinically translatable prophylactic use of poly I-C to target the critical perioperative period. By increasing the number of marginating-pulmonary NK cells, and by transforming them into a mode of resistance to immunosuppression, this approach may reduce postoperative metastasis in cancer patients.

Amelioration of operation-induced suppression of marginating pulmonary NK activity using poly IC: a potential approach to reduce postoperative metastasis.

BACKGROUND AND OBJECTIVES: Pulmonary metastasis is a major cause of death in cases of operable cancer, and evidence suggests that postoperative immunosuppression contributes to this complication. In this study, we aimed to circumvent this risk and identify immunocytes critical in preventing pulmonary metastases. METHODS: F344 rats were treated with either vehicle or repeated low doses of poly I-C (0.2 mg/kg i.p., days 5, 3, and 1 preoperatively), a Th1-cytokine-inducing agent, then subjected or not to laparotomy. Using a non-immunogenic syngeneic mammary adenocarcinoma line (MADB106) we studied: (a) NK cytotoxicity (NKC) in marginating-pulmonary (MP) and in circulating leukocytes; (b) resistance to experimental lung metastasis; and (c) in vitro susceptibility of NKC to corticosterone and prostaglandin-E(2), substances thought to mediate postoperative immunosuppression. RESULTS: MP but not circulating leukocytes showed significant NKC against MADB106 cells. Surgery suppressed this MP-NKC per NK cell and promoted MADB106 metastasis, and poly I-C treatment completely abolished both effects. Poly I-C quadrupled the numbers of MP-NK cells without causing apparent side effects, and protected MP-NKC from in vitro suppression by corticosterone and prostaglandin-E(2). CONCLUSIONS: MP-NK cells are unique in their ability to kill this apparently immunoresistant tumor. Low doses of synthetic ds-RNA (poly I-C), and potentially Th1 cytokines, can expand this MP-NK population and protect it from immunosuppression. The novelty of such a prophylactic approach is targeting the immediate postoperative period, which is characterized by high vulnerability to residual disease, and protecting critical anti-metastatic immunity against postoperative suppression. Testing such a potentially innocuous intervention in oncology patients preparing for surgery may reduce metastatic recurrence.

Marginating pulmonary-NK activity and resistance to experimental tumor metastasis: suppression by surgery and the prophylactic use of a beta-adrenergic antagonist and a prostaglandin synthesis inhibitor.

Surgery is imperative for cancer treatment, but was suggested to suppress immunity and facilitate metastasis. Here we study the involvement of catecholamines and prostaglandins (PG) in such outcomes, and the role played by marginating-pulmonary (MP)-NK cells in controlling MADB106 metastasis. Non-operated and laparotomized F344 rats were injected postoperatively with a PG synthesis inhibitor (indomethacin, 4 mg/kg i.p.), a beta-blocker (nadolol, 0.6 mg/kg s.c.), both drugs, or vehicle. Rats were then inoculated intravenously with non-immunogenic syngeneic MADB106 cells, and 24 h later lung tumor retention was assessed, or 3 weeks later lung metastases were counted. Additionally, 12 h after surgery we harvested MP-NK cells and circulating-NK cells and compared their numbers and cytotoxicity against MADB106 cells and standard YAC-1 target cells. Surgery significantly increased MADB106 metastasis. Nadolol and indomethacin reduced this effect by approximately 50% when used alone, and significantly more (75%) when used together. Only MP-leukocytes exhibited NK cytotoxicity against MADB106 cells. Surgery markedly suppressed it, and nadolol and indomethacin additively restored it. Similar effects were observed assessing MP-NK and circulating-NK cytotoxicity against YAC-1 target cells. Alterations in the numbers of NK cells were partly associated with alterations in total MP-NK activity, but not with circulating-NK activity. Last, administrating nai ve rats with physiologically relevant doses of a beta-adrenergic agonist (metaproterenol), and/or with PGE2, additively and independently of each other promoted MADB106 metastasis, simulating the effects of surgery. These findings point at potential prophylactic measures in cancer patients undergoing surgery, and suggest a role for MP-NK cells in resisting metastasis of apparently insensitive tumors.

Suppression of natural killer cell activity and promotion of tumor metastasis by ketamine, thiopental, and halothane, but not by propofol: mediating mechanisms and prophylactic measures.

UNLABELLED: Postoperative immunosuppression is partly ascribed to anesthesia and has been suggested to compromise patients' resistance to infection and tumor metastasis. We compared the effects of various anesthetics on natural killer (NK) cell activity and on resistance to experimental metastasis, and studied mediating mechanisms and prophylactic measures. Fischer 344 rats served as controls or were anesthetized for 1 h with ketamine, thiopental, halothane, or propofol. Anesthetized rats were either maintained in normothermia or left to spontaneously reach 33 degrees C-35 degrees C. Rats were then injected IV with MADB106 tumor cells, and 24 h later lung tumor retention was assessed, or 3 wk later, lung metastases were counted. Additionally, the number and activity of circulating NK cells were assessed after anesthesia. All anesthetics, except propofol, significantly reduced NK activity and increased MADB106 lung tumor retention or lung metastases. Hypothermia had no significant effects. Ketamine increased metastasis most potently, and this effect was markedly reduced in rats pretreated with a beta-adrenergic antagonist (nadolol) or with chronic small doses of an immunostimulator (polyriboinosinic:polyribocytidylic acid). Overall, the marked variation in the NK-suppressive effects of anesthetics seems to underlie their differential promotion of MADB106 metastasis. Prophylactic measures may include perioperative immunostimulation and the use of beta-blockers. IMPLICATIONS: This study in a rat model of pulmonary metastasis demonstrates that some anesthetics, but not others, increase susceptibility to tumor metastasis, apparently by suppressing natural killer cell activity. Ketamine was most deleterious, and its effects were prevented by peripheral blockade of beta-adrenoceptors combined with low levels of immunostimulation.

Prostaglandin e(2) suppresses NK activity in vivo and promotes postoperative tumor metastasis in rats.

BACKGROUND: Prostaglandins (PGs) were shown in vitro to suppress several functions of cellular immunity. It is unclear, however, whether physiological levels of PGs can suppress cellular immunity in vivo and whether such suppression would compromise postoperative host resistance to metastasis. METHODS: Fischer 344 rats were administered PGE(2) in doses (18 to 300 micro g/kg subcutaneously) that increased the serum levels approximately 2- to 4-fold. We then assessed the number and activity of circulating natural killer (NK) cells, as well as rats' resistance to experimental metastasis of a syngeneic NK-sensitive tumor (MADB106). To study whether endogenously released PGs after surgery compromise these indices, we tested whether laparotomy adversely affects them and whether a cyclooxygenase-synthesis inhibitor, indomethacin (4 mg/kg), attenuates these effects. RESULTS: PGE(2) dose-dependently suppressed NK activity per NK cell and dose-dependently increased 4- and 24-hour MADB106 lung tumor retention (LTR); 240 micro g/kg of PGE(2) quadrupled the number of lung metastases counted 3 weeks later. Selective depletion of NK cells abrogated the promotion of LTR by PGE(2). Surgery significantly suppressed NK activity and increased MADB106 LTR, and indomethacin halved these effects without affecting nonoperated rats. CONCLUSIONS: PGE(2) is a potent in vivo suppressor of NK activity, and its postoperative release may promote tumor recurrence.