Effect of cisplatin upon expression of in vivo immune tumor resistance.

The major intent of cancer treatment with cytotoxic drugs is direct tumor cell damage, but some of these drugs have been shown to be immunomodulatory. Cisplatin is a widely used cytotoxic drug that has been combined with biological response modifiers in recent clinical trials. To evaluate further whether cisplatin may independently alter the level of host resistance against tumor growth, the drug was tested in the Mc7 sarcoma rat tumor model. The expression of in vivo tumor resistance against Mc7 sarcoma in syngeneic Wistar rats is mediated by circulating non-cytotoxic T lymphocytes. These cells interact specifically with tumor cells to generate cytotoxic effectors locally at the site of a tumor challenge. Activities of these components of expression of tumor resistance were measured in vivo after administration of cisplatin and dose-dependent effects were found. Low-dose cisplatin (0.3 mg/kg) increased the activity of the circulating lymphocytes that mediate tumor resistance, and high-dose cisplatin (9 mg/kg) suppressed both mediator lymphocyte activity and the generation of antitumor effector mechanisms. These studies suggest that low-dose cisplatin may be immunomodulatory and combining it with biological response modifiers might be a useful strategy. However, high-dose cisplatin given with biological response modifiers may negate potential immunomodulatory activities of such agents.

In vivo activity of solid phase interleukin 2.

Interleukin 2 (IL-2) coupled to polystyrene beads to form a solid phase of the cytokine was able to increase the cytotoxic activity of rat spleen cells in vitro. A single injection of the IL-2 coupled beads into the peritoneal cavities of normal rats also resulted in the in vivo activation of cytotoxicity of peritoneal exudate cells, whereas a single i.p. injection of the same amount of soluble IL-2 was not effective. When IL-2 coupled beads were mixed with methylcholanthrene-induced Mc7 or Mc107 sarcoma cells and injected into normal syngeneic Wistar rats, the growth of the tumor was suppressed. This effect was localized to the site of the injection. The in vivo activities were achieved with modest amounts of IL-2, less than has been previously reported to be well tolerated in rodents. IL-2 coupled to a solid matrix may be useful for delivering increased concentrations of the lymphokine to tumor dominant regions while maintaining low systemic levels and thereby increasing the therapeutic index.

Intraperitoneal lymphokine-activated killer-cell and interleukin-2 therapy for malignancies limited to the peritoneal cavity.

Autologous lymphokine-activated killer (LAK) cells and recombinant human interleukin-2 (rIL-2) were administered intraperitoneally (IP) to 24 patients with malignancies limited to the peritoneal space. Ten patients had ovarian cancer, 12 had colorectal cancer, and one patient each had endometrial carcinoma and primary small-bowel adenocarcinoma. All ovarian cancer patients, three of twelve colorectal cancer patients, and one patient with endometrial carcinoma had received prior therapy. Patients received IL-2 100,000 U/kg every 8 hours intravenously (IV) for 3 days, and 2 days later underwent daily leukapheresis for 5 days. LAK cells were generated in vitro by incubating the peripheral blood mononuclear cells in IL-2 for 7 days and were then administered IP daily for 5 days through a Tenckhoff catheter (Davol, Inc, Cranston, RI) together with IL-2 25,000 U/kg IP every 8 hours. All but one patient completed at least one cycle of therapy. Toxic side effects included minor to moderate hypotension, fever, chills, rash, nausea, vomiting, abdominal pain and distension, diarrhea, oliguria, fluid retention, thrombocytopenia, and minor elevations of liver function tests; all of these rapidly improved after discontinuation of IL-2. One patient had a grand mal seizure, and one suffered a colonic perforation; these were felt to be treatment-related. IP fibrosis developed in 14 patients and limited repeated cyclic administration of this therapy in five patients. Two of 10 (20%) ovarian cancer patients and five of 12 (42%) colorectal cancer patients had laparoscopy- or laparotomy-documented partial responses. We conclude that LAK cells and rIL-2 can be administered IP to cancer patients, resulting in moderate to severe short-term toxicity and modest therapeutic efficacy. Further investigation of this form of adoptive immunotherapy modified to address the problem of IP fibrosis and with lower IP IL-2 doses is justified by these initial results.

Treatment of myelodysplastic syndromes with low-dose oral 6-thioguanine.

Several cytotoxic agents when used in vitro in very low concentrations have been shown to induce differentiation of leukemic cells. We treated 18 patients with myelodysplastic syndromes with low-dose oral 6-thioguanine (6-TG; 20 to 60 mg daily). Four patients demonstrated significant improvement in peripheral blood counts. An additional three patients had significant reductions in the percentage of leukemic myeloblasts in the marrow without a corresponding improvement in peripheral counts. With the exception of a fall in the peripheral neutrophil count in four patients requiring dose reductions, no toxicity was observed. Low-dose oral 6-TG gives a response rate in myelodysplastic syndromes similar to that of parenteral agents such as cytosine arabinoside. Given the ease of administration and lack of toxicity, oral 6-TG may be a useful treatment modality for these syndromes either alone or in combination with other differentiation-enhancing agents.

Specific induction of local antitumor effector cells mediated in vivo by the circulating lymphocyte pool.

Wistar rats are specifically resistant to the growth of the chemical carcinogen induced syngeneic tumors, Mc7 and Mc107 sarcoma, after being immunized with s.c. implants of irradiated tumor tissue. The central lymph of such immunized rats contains cells able to systemically transfer resistance against tumor growth to normal or irradiated recipient rats. The thoracic duct lymphocytes (TDL) from tumor immune donors are not directly cytotoxic against tumor targets. However recipients of i.v. infused immune TDL develop cytotoxic activity in the peritoneal cavity when challenged with the immunizing tumor at that site. Although the induction of maximum cytotoxicity is tumor specific, peritoneal lavage cells are cytotoxic against both Mc7 and Mc107 tumor targets in the 51Cr release assay. Inhibition of cytotoxicity in the assay by addition of unlabeled Mc7 or Mc107 sarcoma cells to labeled tumor targets suggests that there is both specific and nonspecific activity in these peritoneal lavage cells. Resistance to in vivo tumor growth in adoptively immunized recipients of TDL challenged with both Mc7 and Mc107 is specific for the immunizing tumor. However growth of a mixture of Mc7 and Mc107 sarcoma cells is inhibited in recipients of immune TDL. The results support the notion that mediator lymphocytes circulate in tumor immunized rats in a noncytotoxic state, specifically recognize tumor cells at a challenge site, and mediate induction of effector cells locally. These effectors are at least in part nonspecific in their cytotoxic activity.

Well-differentiated lymphocytic lymphoma with peripheral blood involvement, osteolytic bone lesions, and hypercalcemia. A case report and review of the literature.

It is rare for small lymphocytic B-cell malignancies to be associated with osteolytic bone lesions and/or hypercalcemia. The authors present an unusual case of well-differentiated lymphocytic lymphoma (DWDL) in a 70-year-old man who had osteolytic bone lesions and subsequently developed severe refractory hypercalcemia. The possible etiologic mechanisms responsible for these findings are discussed, and a brief review of the literature is presented.

Subsets of circulating T-lymphocytes mediating resistance to in vivo growth of a carcinogen-induced syngeneic rat tumor.

Wistar rats immunized by s.c. implantation of irradiated Mc7 sarcoma tissue were resistant to the growth of a challenge of this chemical carcinogen-induced syngeneic tumor. Lymphocyte populations enriched in each of the two major T-cell lineages recognized by mouse monoclonal antibodies were prepared from the thoracic duct lymphocyte pool of tumor-resistant rats by affinity chromatography and infused i.v. into normal syngeneic rats. The recipients of either one donor equivalent of "helper" T-cells identified by the monoclonal antibody W3/25 or one donor equivalent of "nonhelper" T-cells identified by the monoclonal antibody OX-8 were resistant to a challenge of Mc7 sarcoma cells. Contaminating cells do not appear to account for the activities of each enriched population, indicating that lymphocytes contributing to expression of resistance to in vivo growth of Mc7 sarcoma must be present in both the helper and the nonhelper T-cell lineages. No direct cytotoxic activity by the thoracic duct lymphocyte populations against the Mc7 sarcoma cells could be demonstrated in vitro. These lymphocytes were generated in vivo and delivered to the systemic circulation of tumor-resistant hosts, implying they play a role in the expression of antitumor resistance in the intact immunized donor.

The subset of T lymphocytes mediating delayed-type hypersensitivity in the rat.

Populations enriched in each of the major rat T-cell subsets recognized by mouse monoclonal antibodies were prepared from thoracic-duct lymphocytes by affinity chromatography. Such enriched populations obtained from donor rats immunized with BCG were tested for their capacity to mediate delayed-type hypersensitivity (DTH) against purified protein derivative by adoptive transfer to recipient rats. The mediator activity was found in the populations enriched in T cells recognized by the monoclonal antibody W3/25. Thus, the results formally demonstrate that mediating DTH is another function of W3/25-positive T lymphocytes. In addition, the studies show that affinity chromatography is an effective method for preparing bulk quantities of functional lymphocyte populations enriched in the major rat T-cell subsets.

Periorbital ecchymoses as the initial sign in multiple myeloma.

A 61-year-old man with bilateral recurrent periorbital ecchymoses was followed up for 36 months, at which time the diagnosis of multiple myeloma was established. However, in vitro tests of blood coagulation and platelet function gave normal findings. A biopsy specimen of the ecchymotic area showed no evidence of amyloid. Clinical and laboratory evaluation did not suggest another cause. The mechanism of the localized bleeding tendency could not be elucidated.

Lymphocytes that mediate systemic resistance to in vivo growth of a carcinogen-induced syngeneic tumor.

Wistar rats immunized with irradiated syngeneic methylcholanthrene-induced sarcoma, Mc7, are able to suppress a subsequent challenge of this tumor. Cells obtained from thoracic-duct lymph of these tumor-immunized animals and infused intravenously into normal syngeneic rats transfer specific tumor resistance to the recipients. These mediators of tumor resistance have the characteristics of T lymphocytes that are in a nonproliferating state and have an effective life span at least 2 weeks in normal recipient rats.

Cytarabine-induced anaphylaxis. Demonstration of antibody and successful desensitization.

An anaphylactic episode occurred in a patient receiving cytarabine (Ara-C) for acute myeloid leukemia. Specific allergy of the patient to cytarabine was demonstrated in vivo by intradermal testing and in vitro by four-hour passive cutaneous anaphylaxis in guinea pigs. Desensitization to cytarabine was successfully performed. This case represents the first known report of both cytarabine anaphylaxis and densensitization, and provides immunologic evidence implicating an allergic reaction to cytarabine.

Granulocytopenia with marked lymphocytosis manifesting Sjogren syndrome.

Sjogren syndrome is a multi-system disease leading to diverse organ involvement during its course [1, 2]. Hematologic abnormalities described in Sjogren syndrome include anemia, mild leukopenia [3, 4], eosinophilia, elevated erythrocyte sedimentation rate, hypergammaglobulinemia, mixed cryoglobulinemia, and a variety of autoantibodies [5]. Marked lymphocytosis with granulocytopenia is distinctly unusual and has not been previously reported. We report a case of Sjogren syndrome who presented with constellation of the latter problems without prominent sicca manifestations.

Transfer of immunity against Listeria monocytogenes by T cells purified by a positive selection technique.

Affinity columns prepared with rabbit antibody to the F(ab')(2) fragment of rat immunoglobulin were used to separate rat thoracic duct lymphocytes into sub-populations that differ with respect to the density of their surface membrane immunoglobulin. Using this technique, it was shown that lymphocytes in the DNA synthetic (S) phase of the mitotic cycle are added in increased number to the lymph of rats infected with Listeria monocytogenes. The great majority of these S-phase cells lacked a high density of surface immunoglobulin as indicated by their failure to bind to the immunoabsorbent. Cells which can protect recipient rats against a challenge infection with L. monocytogenes also segregated with nonadherent thoracic duct lymphocytes obtained from Listeria-immune donors. These protective cells realized their full immunological potential only in recipients that shared histocompatibility-gene-coded structures with the immune lymphocyte donors. The above findings accord with the view that immunity to L. monocytogenes is mediated in rats by activated T cells which are formed as part of the animal's cell-mediated response to infection. Although Listeria-protective lymphocytes concentrate in the nonadherent, T-cell-enriched fraction, it was consistently observed that the adherent, B-cell-enriched fractions of immune donor thoracic duct lymphocytes also could transfer a low level of antimicrobial resistance. This immunity was restricted in allogeneic recipients, a finding which implies that the protection afforded by the adherent population is related to its content of T cells. Nonadherent S-phase lymphoblasts moved in substantial numbers from the blood into peritoneal inflammatory exudates induced by L. monocytogenes. The above finding encourages the belief that recently activated T cells realize their protective function locally in centers of infection where they have secondary effects on macrophages.

Immunity to Trichinella spiralis. I. Transfer of resistance by two classes of lymphocytes.

Rats can be solidly immunized against Trichinella spiralis by a series of methyridine-terminated oral infections with T. spiralis larvae. Injections of thoracic duct lymphocytes (TDL) obtained from such animals can protect normal rats against a Trichinella challenge. The protective cells belong to two populations which differ with respect to their adherence to affinity columns prepared with rabbit antibody to rat F(ab')2. Immune lymphocytes in the column-adherent B cell fraction are inhibited by vinblastine, whereas those in the non-adherent, T cell fraction are resistant to this drug. The above observations suggest that acquired resistance to T. spiralis is mediated by two classes of lymphocytes: B cells which are delivered to the thoracic duct and hence to the blood while still in active cycle, and T cells which have a potentially long life-span and presumably belong to a pool of recirculating small lymphocytes.

Immunity to Trichinella spiralis. II. Expression of immunity against adult worms.

Thoracic duct lymphocytes or purified T or B cells obtained from specifically immunized (Lewis x DA)F1 hybrid rats can protect normal recipients against an oral challenge infection with T. spiralis. The immune cells increase the rate of expulsion of adult worms from the small intestine. Immune TDL do not affect adult worm fecundity, as they do in other strains of rats, or the penetration and development of newborn larvae in muscle cells. Irradiated F1 rats reconstituted with either immune TDL or class-enriched populations of immune T or immune B cells also expel adult Trichinella more rapidly than do unprotected controls. However, unfractionated TDL and inocula enriched in B cells are more efficient than T cells in promoting worm expulsion. The above finding, taken in conjunction with the tissue disposition of labelled lymphocytes in the tissues of recipient rats, implies that immune T cells have a 'helper' function in promoting the formation of protective B cells.

Induction of tumor resistance with BCG-associated tumor antigen.

Soluble material enriched in tumor-associated antigen was prepared by affinity chromatography from a KCl extract of the chemically-induced D-23 rat hepatoma. Microgram quantities of the above material bound spontaneously to living BCG when the two were incubated briefly in vitro. When injected into normal syngeneic rats, the BCG-associated tumor antigen induced a measure of resistance against challenge with D-23 tumor cells. Peritoneal exudate cells (PEC) obtained from such actively immunized subjects were able to suppress the growth of D-23 tumor cells at a test site in muscle. In contrast, immunization with either BCG alone, tumor protein alone, or tumor protein admixed with BCG in circumstances designed to impede association of the protein, failed to provoke the formation of tumor suppressor PEC. The results encourage of the belief that binding of tumor antigen to BCG favors the induction of a cell-mediated tumor suppressive response.

Induction of delayed-type hypersensitivity with BCG-associated proteins.

Microgram quantities of bovine serum albumin (BSA) and hen egg albumin (OA) associate spontaneously with living BCG in aqueous suspension. The association is stable insofar as bound protein cannot be readily dissociated from the organism by repeated washing. Association of BSA or OA is dependent upon the concentration of specific protein or competing protein in the incubation mixture, pH and the amount of protein already bound to the organisms. Washed BCG "complexes" containing either BSA or OA are potent inducers of delayed-type hypersensitivity (DTH). The same soluble proteins are far less effective inducers of DTH, even when injected with BCG. The capacity of OA-BCG complexes to provoke a cell-mediated response seems to be related, at least in part, to "stabilization" of the antigen on an appropriate carrier and its concentration at the site of BCG injection.

Fibrinogen Cleveland II. An abnormal fibrinogen with defective release of fibrinopeptide A.

An abnormal fibrinogen (fibrinogen Cleveland II) was detected in the plasma of a 23-yr-old white man with a mild bleeding diathesis. The one-stage prothrombin time, thrombin time, and Reptilase time were all prolonged. 16 of 24 tested relatives had the defect, which appeared to be transmitted as an autosomal dominant characteristic. The thrombin time of normal plasma was slightly inhibited by the proband's plasma. The abnormally long thrombin time of fibrinogen Cleveland II was partially corrected by addition of calcium ions. Fibrinogen Cleveland II was indistinguishable from normal fibrinogen by immunoelectrophoresis, DEAE-cellulose column chromatography, or polyacrylamide gel electrophoresis of reduced fibrinogen in sodium dodecyl sulfate. The major defect detected appeared to be impaired release of fibrinopeptide A when fibrinogen Cleveland II was incubated with thrombin. This defect was localized to the NH(2)-terminal disulfide knot portion of the molecule. An abnormality of polymerization of fibrin monomers was also present, but the abnormal fibrin demonstrated relatively normal crosslinking. Despite these defects, fibrinogen Cleveland II achieved a degree of coagulability similar to normal fibrinogen and appeared to incorporate some molecules of fibrin with intact fibrinopeptide A into the clot. The fibrin clot that was formed appeared to be abnormal by electron microscopy. These functional defects and other descriptive characteristics appear to distinguish fibrinogen Cleveland II from other inherited abnormal fibrinogens.