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We have examined the role of T cells activated with interleukin-2 (IL-2) in vitro and subsequently irradiated (2500 rads), in stimulating murine hematopoiesis in a syngeneic system. Our data suggest that activated, irradiated T (AIT) cells significantly increased the progenitor cell activity of T cell-depleted bone marrow (BM) both in vitro and in vivo as compared with controls (P < 0.001). The efficacy of AIT cells was comparable to that of activated, nonirradiated T (AT) cells (P > 0.05). Optimal stimulation of BM progenitor cell activity was seen when T cells were activated for 4 days and used in a BM to T cell ratio of 1:2 or 1:5. The effect of these activated cells was related to the release of factors with ability to enhance hematopoiesis. These observations may have implications in enhancing the engraftment of T cell-depleted BM in allogeneic transplantation.

Ceftazidime has been reported to cause myelosuppression both in vitro and in vivo. Granulocyte colony-stimulating factor (G-CSF) is known to enhance the proliferation and differentiation of myeloid cells. The present study was carried out to define the role of G-CSF in preventing the ceftazidime-induced suppression of BM progenitor cells in vitro, and to define the mechanisms involved in ceftazidime-induced myelosuppression. Our results show that G-CSF was able to maintain the proliferative activity of BM cells in the presence of ceftazidime if it was added to the culture medium during the early phase of exposure of BM to ceftazidime. Monoclonal antibody to TNF completely inhibited the ceftazidime-induced myelosuppression. The suppressive effect on BM was mediated via CD3+ T cells whereas macrophages conferred protection against this suppression. TNF-induced suppression of BM was inhibited by G-CSF. These data suggest that G-CSF prevents the ceftazidime-induced myelosuppression by antagonizing the suppressive effect of TNF and by enhancing the proliferative activity of BM.

We have previously shown that administration of cyclosporine A (CsA) plus interferon-gamma (IFN) after chemotherapy to mice bearing B16 melanoma results in the generation of cells with major histocompatibility complex (MHC)-unrestricted cytotoxicity in vitro and in vivo; the antitumor effect of these cells could be attenuated by normal spleen cells. This study shows that antitumor effect after treatment with CsA plus IFN after chemotherapy was mediated by T and natural killer (NK) cells, both in vitro and in vivo. Infusion of purified T or NK cells into secondary recipients after chemotherapy resulted in a significant control in the dissemination of tumor as compared to chemotherapy alone. The antitumor potential of NK cells was at least 10 times greater than that of T cells. The effector cells could inhibit the proliferation of tumor cells in vitro without a contact between the effector and tumor cells, suggesting that antitumor effect in this system was partly related to the secretion of cytotoxic factors by the effector cells. Infusion of normal spleen cells inhibited the antitumor effect of adoptively transferred effector cells. This study defines the nature of effector cells involved in mediating the antitumor effect in this model; the optimal efficacy of these cells in the recipient is possibly related to the abolition of a suppressor system by chemotherapy.

Amphotericin B causes suppression of bone marrow (BM) progenitor cells in vitro. Granulocyte colony stimulating factor (G-CSF) enhances the proliferation of myeloid cells. The present study defines the role of G-CSF in preventing amphotericin B-induced myelosuppression. G-CSF increased the proliferative potential of BM and protected against amphotericin B-induced myelosuppression if it was added to the medium during the early phase of exposure of BM to amphotericin B. Monoclonal antibodies to tumour necrosis factor-alpha (TNF) or interferon-gamma (IFN) inhibited the myelosuppression partially; simultaneous presence of both these antibodies completely abrogated this suppression, suggesting that both TNF alpha and IFN gamma were involved in amphotericin-induced myelosuppression. TNF- or IFN-induced suppression of BM was also inhibited by G-CSF. These data suggest that G-CSF prevents the amphotericin B-induced myelosuppression by antagonizing the suppressive effects of TNF and IFN and by enhancing the proliferative activity of BM.

IL-2 has been used after autologous BMT (ABMT) with the aim of inducing graft versus leukemia (GVL) effect. Our studies in mice have shown that IL-2 therapy induces GVL effect when employed after BMT with bone marrow (BM) that has been activated with IL-2 in vitro (ABM). The present study was carried out to define the time of optimal GVL effect after BMT so that the immunomodulatory approaches could be concentrated at the time of maximum GVL effect. Our data show that GVL effect was induced if IL-2 was instituted immediately after BMT with ABM in mice with acute myeloid leukemia; institution of IL-2 1 week after BMT with ABM did not induce GVL effect. IL-2 therapy instituted immediately or 1 week after BMT with fresh bone marrow (FBM) did not induce any GVL effect. A significant increase in the NK activity was noticed whether IL-2 was instituted immediately or 1 week after BMT, either with FBM or with ABM. To evaluate the ability of IL-2 in the eradication of residual disease from the autograft and the host, BM with variable infiltration with leukemia was activated with IL-2 and used for BMT in leukemic mice. The GVL effect of BM with minimal leukemic infiltration (absence of morphologically demonstrable disease) was comparable to the GVL effect of normal BM. These findings suggest that: (a) maximum GVL effect after BMT with ABM is concentrated in the early post-transplant period possibly because of minimal residual disease during this time; (b) an increase in the NK activity induced by IL-2 therapy may not predict for an improved GVL effect; and (c) for optimum GVL effect, BM with minimal leukemic infiltration should be activated with IL-2 before BMT.

Interleukin-2 (IL-2) activation of fresh or frozen bone marrow (BM) in vitro generates killer cells with potent anti-tumor effect both in vitro and in vivo. The IL-2-activated BM (ABM) retains the capacity to reconstitute the hematopoietic system in an autologous bone marrow transplantation (ABMT) setting. The killer cells lose their cytotoxicity if the ABM undergoes the procedures of freezing and thawing. Therefore, for clinical application, the ABM has to be generated after thawing a frozen stock of BM before ABMT. The thawed BM cells are fragile and may undergo lysis, resulting in clump formation and cell loss. The frozen autograft also contains components of cryoprotectant mixture whose effects on the generation of ABM have not been defined. The present studies have been carried out to optimize a technique of handling the frozen BM for immunomodulation with IL-2 for 24 h at 37 degrees C prior to ABMT, with minimal loss of cells. IL-2-activation of BM was carried out in bags containing serum free medium which were designed to permit gaseous exchange. Addition of deoxyribonuclease (DNAse) (100 micrograms/ml of BM concentrate) immediately after thawing and the presence of heparin (20 units/ml) in the medium completely abrogated immediate or delayed clumping of cells. The presence of DNAse and/or heparin during in vitro culture did not affect the cell viability, cytotoxicity against tumor cells or the progenitor cell activity of the ABM; all these functions were well maintained even when BM was placed in culture immediately after thawing (without washing). There was no microbial contamination.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin-2 results in the generation of lymphokine activated killer cells which exhibit a potent effect against a wide variety of tumours. Consequently, interleukin-2 therapy has been used to induce a graft versus tumour effect following autologous bone marrow transplantation. Preclinical studies have shown that this results in successful engraftment, and an enhanced reconstitution of the immune system.

Interferon (IFN) increases the expression of major histocompatibility (MHC) antigens on the surface of tumor cells. Cyclosporine A (CsA) administration following myeloablative therapy and syngeneic bone marrow transplantation results in the generation of cells with autoreactive and antitumor effects that are related to the expression of MHC antigens. We used these two agents following conventional chemotherapy in a non-bone marrow transplantation setting for a melanoma in a murine model. Treatment with IFN alone or CsA alone was ineffective in controlling the dissemination of melanoma. A combination therapy with both these agents resulted in a significant control in the dissemination of the tumor, prolonged the survival of the tumor-bearing mice over that with chemotherapy alone, and generated cells with potent MHC-unrestricted cytotoxic potential in vitro. Adoptive transfer of these cells to secondary tumor bearers treated with chemotherapy showed potent antitumor effect; in the absence of chemotherapy, these cells had no antitumor effect in the secondary recipients. The antitumor effect of cells generated by IFN plus CsA therapy following chemotherapy could be blocked by normal spleen cells. These data suggest that treatment with IFN plus CsA following nonmyeloablative chemotherapy generates cells with MHC-unrestricted cytotoxicity; this effect may be related to abolition of suppressor influences by the chemotherapy.

Lymphokine activated killer cells have potent antitumor effect both in vitro and in vivo. They have been reported to suppress bone marrow (BM) progenitor cell activity (PCA) in vitro, thus raising concern about the feasibility of their use after autologous bone marrow transplantation. The present study was carried out to evaluate the effect of LAK cells on BM engraftment in a syngeneic BMT setting in mice. LAK cells supplemented with or without exogenous interleukin-2 therapy did not impair the hematopoietic reconstitution or survival of mice undergoing BMT. LAK cells also did not reduce the PCA of the engrafted BM. LAK cell therapy did not cause graft-versus-host disease. Finally, LAK cells supplemented with IL-2 therapy improved the graft-versus-leukemia effect. These findings suggest that LAK cells plus IL-2 therapy after BMT does not impede hematopoiesis and should be evaluated as an adjuvant therapy with the aim of eradication of minimal residual disease after autologous BMT.

Interleukin-2 (IL-2) promotes the generation and proliferation of killer cells in the peripheral blood and bone marrow (BM) both in vitro and in vivo. When employed in a syngeneic bone marrow transplantation (BMT) setting and followed by IL-2 therapy, murine BM cells activated with IL-2 in vitro (ABM) demonstrate potent graft-versus-leukemia (GVL) and anticytomegalovirus effects. ABM cells retain the capacity to reconstitute the hemopoietic system both in normal and leukemic mice. This therapy does not cause graft-versus-host disease (GVHD). Human ABM cells carry out purging of leukemia without loss of progenitor cell activity in vitro. The purging ability of ABM can be augmented by interleukin-1, interferon, and tumor necrosis factor. IL-2 therapy stimulates the veto suppressor cell activity of T cell-depleted BM, and has reduced GVHD and permitted engraftment of mismatched allogeneic BM in murine models. Future studies should determine the optimum treatment schedules with IL-2 for improving the GVL effect in autologous BMT, and for abolishing GVHD in allogeneic BMT settings.

Incubation of bone marrow (BM) with interleukin-2 (IL-2) in vitro results in generation of killer cells providing a tool for enhancing the graft-versus-tumor effect in transplantation. We have evaluated the influence of IL-2 on the progenitor cell activity (PCA), homing pattern of BM and hemopoiesis in a syngeneic bone marrow transplantation (BMT) model in mice. The PCA index and homing pattern of BM activated with IL-2 in vitro for 24 h (ABM) were similar to those of fresh bone marrow (FBM). In vitro culture of BM for more than 1 day resulted in progressive decline in its PCA index; this was not related to the presence or absence of IL-2 in the culture medium. Toxicity of IL-2 was related to the dose and not the time of institution of IL-2 therapy after BMT. Maximum tolerated dose of IL-2 instituted immediately after BMT was 10 times higher than the dose in a non-BMT setting. The pattern of marrow reconstitution following BMT with ABM was comparable to that with FBM. This study shows that BMT with BM activated with IL-2 for 24 h results in normal hemopoiesis, and IL-2 therapy instituted immediately after BMT with ABM does not cause additional toxicity.

Twenty-five patients with acute nonlymphoblastic leukemia undergoing 41 cycles of chemotherapy with daunorubicin/cytosine arabinoside (ara-C) or with etoposide/ara-C received metoclopramide (MCP; 0.5 mg/kg 6 hourly i.v.) or MCP (same dose) plus oral lorazepam (1 mg/d) during and 24 hours following the chemotherapy as antiemetic medication. Control of vomiting was achieved is 55% (complete 5%, partial 50%) of the patients receiving MCP alone and in 100 percent (complete 76.1%; partial 23.8%) of those receiving MCP plus lorazepam (p less than 0.001). Eighteen of the 21 patients (85.7%) receiving MCP plus lorazepam opted for the same antiemetic regimen as compared to six of the 20 (30%) receiving MCP alone (p less than 0.01). One patient in each group developed mild sedation during the treatment. It is concluded that oral lorazepam is an effective and safe adjuvant to MCP for the control of vomiting during cancer chemotherapy.

Low uptake of monoclonal antibodies (MAbs) in cancer lesions is a significant problem in cancer therapy. Recent studies have shown that antibody uptake in tumor is controlled in large part by the tumor blood flow and the vascular permeability of the tumor endothelium. We have hypothesized that these physiological properties of tumor vessels may be altered by pretreatment with vasoactive drugs or peptides linked to tumor-specific MAbs. To test this hypothesis, two MAbs, Lym-1 directed against human malignant lymphomas and B72.3 reactive with the TAG-72 antigen expressed in solid tumors, were chemically conjugated with human recombinant interleukin 2 (IL-2). IL-2 has been used in humans to activate lymphokine-activated killer cells for the treatment of cancer but is also known to produce a generalized vascular permeability by an unknown mechanism when used systemically. Chemical conjugation of IL-2 to MAbs appears to destroy its cytokine function as shown by T-cell proliferation studies in vitro. Despite this finding, MAb/IL-2 immunoconjugates retain their ability to produce an enhanced vascular permeability when injected i.v. into nude mice bearing relevant tumor models only. Biodistribution studies using 125I-labeled tracer Lym-1 have demonstrated that the Lym-1/IL-2 immunoconjugate can increase antibody uptake in tumor by a factor of 4 in a time (2.5-h pretreatment)- and dose (30 micrograms/mouse)-dependent manner. In contrast, treatment of mice with free IL-2 and antibody showed this effect in all organs of the mouse including the tumor. Bidirectional crossover imaging studies in individual tumor-bearing nude mice showed improved uptake and decreased blood pool when the MAb/IL-2 immunoconjugates were used compared to controls. Finally, tumor blood flow and vascular permeability studies demonstrate that the physiological effect of the MAb/IL-2 is due to a reversible and specific vascular leakage at the tumor site. These studies indicate that pretreatment with this novel immunoconjugate may enhance the diagnostic and therapeutic potential of MAbs, drugs, and other macromolecules for the treatment of cancer.

The present work is a continuation of our studies to improve the graft versus leukemia (GVL) effect in autologous bone marrow transplantation. We have recently shown that the GVL effect of bone marrow transplantation (BMT) with interleukin 2 (IL-2)-activated bone marrow (ABM) followed by IL-2 therapy immediately after BMT is superior to the GVL effect of BMT with fresh, syngeneic bone marrow, with or without IL-2 therapy, in mice with acute myeloid leukemia. The present studies show that institution of IL-2 treatment 1, 2, or 3 weeks after BMT with ABM resulted in shortening of survival and fall in cure rate as compared to IL-2 therapy instituted immediately after BMT with ABM. Increasing the dose of IL-2 did not improve results. However, reducing the frequency of IL-2 administration to once a day instead of twice a day affected the results adversely. Commencing IL-2 therapy 1, 2, or 3 weeks after BMT with fresh, syngeneic bone marrow did not improve the GVL effect as compared to IL-2 therapy started immediately after BMT with fresh, syngeneic bone marrow. Cryopreserved bone marrow was effectively activated with IL-2 and used successfully for BMT after thawing. The animals cured of leukemia by BMT with ABM and and IL-2 therapy were not resistant to leukemia and died when reinfused with leukemic cells. Our findings suggest that for optimum GVL effect, activation of bone marrow is necessary and IL-2 therapy should be started immediately after BMT with ABM.

We have shown that incubation of bone marrow (BM) with interleukin 2 (IL-2) generates activated bone marrow cells (ABM) with potent tumoricidal activity in vitro and in vivo. The present study was carried out to define the interaction of other cytokines with IL-2 in generation of ABM. Our data show that interleukin 1 (IL-1), interferon (IFN)- both gamma and alpha, and tumor necrosis factor (TNF-alpha) significantly increased the cytolytic potential of ABM. Interleukin 3, interleukin 4, transforming growth factor-beta and adherent cells were reduced, while granulocyte-macrophage colony-stimulating factor had no influence on the generation of cytolytic activity. IL-1 was enhanced while TNF-alpha depressed the BM progenitor cell activity in vitro. The IL-2-induced purging ability of BM contaminated with leukemic cells was increased by IL-1, TNF-alpha and IFN-gamma. This study shows that biomodulation of BM with combination of cytokines in vitro can be useful in purging a large leukemic burden.

The effects of certain antibiotics on the colony forming activity of human bone marrow cells in semisolid methylcellulose medium in vitro and on murine BM cells in spleen colony forming units (cfu-s) in vivo were evaluated. Amikacin, gentamicin, piperacillin, co-trimoxazole and pentamidine had little or no effect on human bone marrow progenitor cell function; amphotericin B, aztreonam, ceftazidime and imipenem caused significant suppression of human colony forming unit-erythroid (cfu-e), burst forming unit-erythroid (bfu-e) and colony forming unit-granulocyte macrophage (cfu-gm) at both peak and trough serum concentrations. At molar equivalent concentrations ceftazidime, cefotaxime and cefoperazone caused significant decreases in human cfu-e, bfu-e and cfu-gm in vitro (P less than 0.01) and murine cfu-s in vivo (P less than 0.05); cefoxitin, cefuroxime, ceftizoxime and ceftriaxone did not suppress human bone marrow progenitor cell activity. Gentamicin, piperacillin and ceftriaxone had no effect on murine cfu-s formation. Further studies to evaluate the effect of these antibiotics on human bone marrow in vivo are suggested.

Four cases of primary non-Hodgkin's lymphoma of the lung are described. Two cases had low and two intermediate grade lymphoma at the time of diagnosis. The patient who had disease for long duration and received pulmonary radiotherapy developed intractable chest infection and died six months after diagnosis; the three patients having short history of disease and treated with surgery and/or chemotherapy have been doing well for 4 to 77 months after the diagnosis. It is concluded that diagnosis of primary pulmonary lymphoma should be suspected in patients with nodular or interstitial lung disease and bronchoalveolar lavage with aspiration cytology should be done to make an early diagnosis.

Aggressive chemotherapy regimens and supportive measures in haemato-oncology patients demand reliable venous access. Experience with this method in India has been limited. During a period of six months, we have used 42 subclavian indwelling catheters and 31 cubital Cavafix long lines. The mean age of patients in the two groups was 32 years and 7 years respectively. Subclavian catheters had a median duration of catheter placement of 46 days (range 4-145) and total 1494 catheter days, while cubital longlines yielded a median duration of insertion of 14 days (range 4-27) and total 508 catheter days. Catheter related complications were infection in 25% of patients, thrombophlebitis in 22%, blockade in 12% and misplacement in 17% in both groups taken together. The patients and families were extremely satisfied with the devices. Our experience supports further use of durable venous access in cancer patients. Implanted central venous catheters should be preferred whenever feasible.

The cytotoxic potential of interleukin 2 (IL-2) activated bone marrow (ABM) was compared with that of IL-2 activated peripheral blood lymphocytes (LAK cells) against three hematologic tumor cell lines (K-562, CEM, Daudi) and fresh lymphoid blasts in short-term chromium release assays. ABM was found to be superior to LAK cells against all tumor cells tested. The recovery of bone marrow (BM) cells dropped with passage of time in culture but their clonogenic potential was not impaired (with or without IL-2). BM contaminated with CEM cells and treated with IL-2 showed significant ability to purge itself of the leukemic cells in semisolid agar culture; the purging ability of 3- and 1-day ABM was comparable. IL-2 alone or BM alone had no influence on the growth of CEM cells. This study suggests that BM can be activated with IL-2 in vitro to generate the ability to eliminate contaminating leukemic cells without affecting its progenitor cell function in vitro.

A preliminary experience in the use of central venous catheters, including Hickman (15), subclavian jugular (Desseret) (50) and Secalon Hydrocath (3), in cancer chemotherapy is presented. There were no technical complications and no mortality attributable to catheterisation. Infections formed the major complication in Hickman (30%) and subclavian jugular (Desseret) (36%) catheters, while the use of Secalon Hydrocath was event-free. The commonest organism isolated was P aeruginosa (60%), followed by staphylococci (30%). Prolonged venous access was achieved with minimum morbidity. It is concluded that central venous catheters are safe and convenient both for the patient and the treating team, and should be used in all patients undergoing protracted chemotherapy and supportive care.

The effect of hydroxyurea on foetal haemoglobin (HbF) levels was evaluated in 36 patients of myeloproliferative and myelodysplastic disorders. In 17 (47.2%) patients, HbF levels increased from 1.40 +/- 1.17 to 3.03 +/- 1.97 per cent after 4 wk therapy with hydroxyurea. In the responders this increase was highly significant (P less than 0.001). The rise in the HbF levels after hydroxyurea therapy was significant in patients with chronic myeloid leukaemia but not in the other groups.

Thrombocytopenia is a rare side effect of amphotericin B. Two patients with acute leukaemia in remission who developed severe thrombocytopenia during amphotericin B therapy are reported. Thrombocytopenia recovered after the withdrawal of the drug in one patient and after reducing the dose in the other patient.

Twenty-nine children (age range 1-14, median 8 years) with acute non-lymphoblastic leukemia (ANLL) were induced in remission with daunorubicin and cytosine arabinoside. Twenty-three (79.3%) patients achieved complete remission (CR) and were administered two cycles of the same drugs as consolidation therapy; no maintenance treatment was given. Three (10.3%) patients died during induction; 3 (10.3%) patients were resistant to therapy. Multivariate analysis showed that female sex, TLC less than 50 X 10(9)/L, absence of in ection, albumin greater than 3.5 g/dl and high myeloperoxidase activity had a favourable influence on achievement of CR. TLC less than 50 X 10(9)/L and albumin greater than 3.5 g/dl also had a favourable prognostic value. Eight patients are alive between 13 and 32 months with overall survival at 2 years being 27.5%; four patients are free of disease with projected DFS at 2 years being 13.7%. The present data indicates the need for newer approaches to improve the long term survival in childhood ANLL.

We report a patient with Poland's syndrome who developed acute lymphoblastic leukemia (ALL) at 28 years of age. This is the first time that such an association has been reported from outside the United States or in an adult. The relevant literature is reviewed.

Thirty thrombocytopaenic patients of acute leukaemias and myelodysplastic syndrome were transfused platelets collected from ABO-matched donors using Haemonetics V30 and V50 blood processors. Twenty-seven patients had septicaemia and/or splenomegaly; 2 patients had disseminated intravascular coagulation (DIC). Pre-transfusion platelet count was 11.0 +/- 4.0 X 10(9)/L. The mean corrected count increments (CCI) 1 hour and 18 hours post-transfusion were 13.02 X 10(9)/L and 3.88 X 10(9)/L respectively, in the absence of DIC. Active bleeding stopped when platelet count was above 15.0 X 10(9)/L. There was no difference between the platelet yield from two blood processors.

A case of patient developing epithelial ovarian cancer 15 years after carcinoma of cervix treated successfully with radiotherapy, is reported. The patient has shown good initial response to chemotherapy and surgery.

Case records of 78 patients of acute lymphoblastic leukemia have been reviewed. Complete remission occurred in seven cases following an episode of septicemia and supportive care.