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Biomedical subjects

H G Klingemann

Publications and source records attributed to H G Klingemann.

At least 19 recordsLinked to original sources
PubMed2001

Factors regulating the cytotoxic activity of the human natural killer cell line, NK-92.

NK-92, a highly cytotoxic, interleukin-2 (IL-2)-dependent human natural killer (NK) cell line, has been of interest for basic and translational research. We report on a comprehensive analysis of NK-92 for factors implicated in NK cytotoxicity to elucidate factors underlying NK-92's high cytolytic activity and target range. Thus, we hope to develop a method to identify patients best suited to NK-92 immunotherapy. In addition, as a model system, we hope to increase understanding of the basis for the elevated activity exhibited by activated NK (ANK) cells. NK-92 exhibits an unusual receptor expression profile, expressing a relatively large number of activating (NKp30, NKp46, 2B4, NKGD, E, CD28) receptors. Conversely, it expresses few inhibitory receptors (NKGA/B, low levels of KIR2DL4, ILT-2), lacking most of the killer inhibitory receptors (KIRs) clonally expressed on normal NK cells. In addition, NK-92 expresses high levels of molecules involved in the perforin-granzyme cytolytic pathway as well as additional cytotoxic effector molecules including tumor necrosis factor (TNF)-superfamily members FasL, TRAIL, TWEAK, TNF-alpha, indicating the ability to kill via alternative mechanisms. NK-92 also expresses other molecules implicated immune effector cell regulation (CD80, CD86, CD40L, TRANCE) whose relevance in NK killing is unclear. This study provides initial data to develop a method to identify NK-92 susceptible cells (cells expressing ligands for NK-92 activating receptors ie CD48 for 2B4 and CD80/86 for CD28). Furthermore, this work suggests mechanisms that may contribute to ANK cell activity, including modulation of receptor expression to favor activation, up-regulation of cytotoxic effector molecules, and acquisition of new cytolytic pathways.

Cell Adhesion Molecules↗
PubMed2000

Allogeneic bone marrow transplantation for low-grade lymphoma and chronic lymphocytic leukemia.

Twenty-six patients with low-grade lymphoma (LGL) (n = 18) or chronic lymphocytic leukemia (CLL) (n = 8) received allogeneic BMTs between 1985 and 1998. Median age was 42 years, median interval from diagnosis to transplant 22 months and median number of prior treatments three. Twenty (77%) had stage IV disease; 22 (85%) had never achieved CR. Donor source was HLA matched sibling (n = 19, 73%), matched unrelated (n = 6, 23%) or syngeneic (n = 1). Conditioning therapy included total body irradiation in 23 patients and busulphan in three. Twenty-five received GVHD prophylaxis with cyclosporine A; + methotrexate (n = 19), + methylprednisolone (n = 2) or + T cell depletion of allograft +/- methotrexate (n = 4). Sixteen patients are alive, a median of 2.4 years post BMT. Death occurred due to transplant complications (n = 7) or underlying disease (n = 3). Eighteen (12 LGL, six CLL) of 22 evaluable patients (82%) achieved CR post BMT. Cumulative incidence of refractory/recurrent disease was 18% (95% confidence interval (CI) 7-42%). Overall and event-free survivals were 58% (95% CI 35-75%) and 54% (95% CI 32-72%), respectively. Allogeneic BMT for young patients with advanced LGL or CLL is feasible and can result in long-term disease-free survival.

Adult↗
PubMed2000

Prolonged survival after intensive therapy and purged ABMT in patients with multiple myeloma.

Despite numerous strategies, the cure of multiple myeloma remains a difficult challenge. Recent approaches have involved dose-intensive therapy followed by stem cell transplantation, most often with autologous stem cells (ASCT). Although ASCT is of benefit, it is not considered curative. Between 1988 and 1995, we utilized an aggressive three-drug conditioning regimen followed by ABMT using marrow purged with either 4-hydroperoxycyclophosphamide (4-HC) or mafosphamide (MAF). Twenty-nine of 42 patients who had first received VAD (14 patients) or VAD followed by cyclophosphamide (7 g/m2 i.v.) + dexamethasone (40 mg/day p.o. x4) + GM-CSF (15 patients) met the eligibility criteria needed to undergo bone marrow harvest and ABMT, ie < or =10% marrow plasma cells and > or =50% decrease in paraprotein level. Alpha-interferon maintenance therapy was given post ABMT. Median follow-up is 7.5 years (range 5.0-11.25). Six early and two late non-relapse deaths occurred; 15 patients have relapsed. Seven patients remain in continuous CR (five) or PR (two), including three with stage IIIB disease at diagnosis. One patient developed a soft tissue sarcoma 8 years post ASCT. Although this protocol produced excessive toxicity compared with current approaches, the results demonstrate that dose-intensive therapy and ASCT can produce durable remission in this disease. Further development of dose-intensive strategies is warranted.

Adult↗
PubMed2000

Treatment of steroid-resistant acute graft-versus-host disease with rabbit antithymocyte globulin.

Acute graft-versus-host disease (A-GVHD) is a life-threatening complication of allogeneic stem cell transplantation (SCT), and primary therapy consists of high-dose corticosteroids. Patients who fail to respond adequately to corticosteroids require salvage treatment, with anti-T cell antibodies being the most commonly utilized group of agents. We report our institution's experience treating steroid-resistant GVHD in 36 adult patients (median age 39 years, range 24-55) with a rabbit antithymocyte globulin product (thymoglobulin). Eleven patients had undergone sibling SCT (10 histocompatible, 1 one-antigen mismatched) and 25 patients had received unrelated donor bone marrow (17 matched, 8 one-antigen mismatched); 32 patients (89%) had grade III or IV A-GVHD. Thymoglobulin was administered in two different regimens; group 1 patients (n = 13) received 2.5 mg/kg/day x 4-6 consecutive days with maintenance of all other immunosuppressives. Group 2 patients (n = 21) were given the same dose of thymoglobulin on days 1, 3, 5, and 7 with discontinuation of cyclosporine for 14 days, during which the corticosteroid dose was held at 2-3 mg/kg/day. Two patients had severe adverse reactions to thymoglobulin (hypoxemia and hypotension) and could not complete treatment, however, in the other patients, aside from transient leukopenia (25%) and and hepatic dysfunction (25%), the antibody preparation was well tolerated. Of the 34 evaluable patients, 13 patients had a complete response (38%) and 7 patients (21%) had a partial response, for an overall response rate of 59%. Response rate was higher in group 1 patients (77%) compared to group 2 patients (48%), (p = 0.15); skin GVHD was more responsive (96% of patients) than gut GVHD (46% of patients) or hepatic GHVD (36% of patients). Opportunistic infections were a significant complication, with 11 patients developing systemic fungal infections and 9 patients serious viral infections; there were seven episodes of bacteremia following thymoglobulin treatment and one fatal protozoal infection. There were 9 patients (25%) who developed post-SCT lymphoproliferative disorder (PTLD) and 4 patients who had a relapse of underlying primary malignancy; none of these patients survived. Of the 36 patients entered on the study, only 2 patients (6%) survive, at 15+ and 34+ months post-unrelated donor SCT. Although thymoglobulin is associated with an impressive response rate when administered for advanced steroid-resistant GVHD, long-term survival is uncommon, even in responders, primarily due to the high risk of developing either an opportunistic infection or a PTLD.

Acute Disease↗
PubMed2000

Allografting for indolent lymphoid neoplasms.

BACKGROUND: Allogeneic bone marrow transplantation (BMT) has been used in patients with low-grade lymphoma (LGL) and chronic lymphocytic leukemia (CLL) with the goal of achieving long-term disease-free survival. PATIENTS AND METHODS: Twenty-nine patients with these diagnoses (LGL = 19, CLL = 10) received allogeneic BMT between September 1995 and January 1999. Median age was 42 (range 20-52) years. Twenty-three of twenty-nine patients (79%) were Ann Arbor or Rai stage IV at the time of transplant; twenty-four (83%) had never achieved complete remission (CR). Donor source was HLA-matched sibling (20), unrelated (8) and syngeneic (1). RESULTS: Seventeen patients are currently alive, a median of 29 months (range 1-85) post-BMT with a median KPS of 90%. Twenty-three of twenty-seven evaluable patients (85%) achieved CR post-BMT. Six patients had refractory/recurrent disease. Death occurred related to transplant complications in eight patients and underlying disease in four. Overall and event-free survival for the whole group is 51% and 44%, respectively. CONCLUSIONS: Allogeneic BMT for young patients with advanced stage LGL or CLL is a feasible strategy that can result in achievement of long-term disease-free survival.

Adult↗
PubMed1999

Characterization of genetically altered, interleukin 2-independent natural killer cell lines suitable for adoptive cellular immunotherapy.

NK-92 is a highly cytotoxic natural killer (NK) tumor cell line that possesses properties that make it an excellent candidate for adoptive cellular immunotherapy. However, the cytotoxicity of NK cells is dependent on cytokines such as interleukin 2 (IL-2). Although NK-92 cells maintain cytotoxicity for a time after withdrawal of IL-2, clinical use will probably require prolonged treatment with fully activated cells to eliminate disease effectively. The ability to support cytotoxic cells with exogenously administered IL-2 is limited by associated toxicity. Therefore, we describe the transfection of the IL-2-dependent NK-92 cell line with human IL-2 (hIL-2) cDNA by particle-mediated gene transfer to create two IL-2-independent variants, NK-92MI and NK-92 CI, and describe their characterization and comparison with parental cells. Both variants were shown to contain, express, and synthesize the hIL-2 cDNA. IL-2 synthesis was higher in NK-92MI cells compared with NK-92CI cells, with no expression in parental cells. Functionally, the cytotoxicity of all three cell lines was similar and coincubation with IL-2-independent variants did not affect hematopoietic progenitor cells. NK-92MI and NK-92CI cells were more radiosensitive than NK-92 cells, with proliferation inhibited at lower radiation doses and increased morality and decreased cytotoxicity compared with parental cells. Data presented here show that we have created by particle-mediated gene transfer two IL-2-independent variants of NK-92 that are identical to parental cells in virtually all respects, including high cytotoxic activity. The nonviral transfection of these cells makes them suitable for clinical applications. These IL-2-independent cells should allow prolonged treatment with fully active natural killer cells without the need for exogenous IL-2 support.

Cell Division↗
PubMed1999

Regimen-related toxicity and non-relapse mortality with high-dose cyclophosphamide, carmustine (BCNU) and etoposide (VP16-213) (CBV) and CBV plus cisplatin (CBVP) followed by autologous stem cell transplantation in patients with Hodgkin's disease.

This analysis compares the regimen-related toxicity (RRT) and overall non-relapse mortality (NRM) in Hodgkin's disease patients conditioned with either CBV (cyclophosphamide, BCNU (carmustine), and VP16-213 (etoposide)) (26 patients) or CBVP (CBV + cisplatin) (68 patients) followed by autologous stem cell transplantation (ASCT). CBVP included a continuous infusion rather than intermittent doses of etoposide, a lower BCNU dose and the addition of cisplatin. RRT and NRM were determined for each regimen and compared; risk factors for each were examined by multivariate analysis. Grade IV (fatal) RRT occurred in five patients (pulmonary in two, cardiac in two, and central nervous system in one). Eighteen patients experienced grade II-III pulmonary RRT, consistent with BCNU damage in 15. Prior nitrosourea exposure was the main risk factor for pulmonary RRT. Grade II mucosal and hepatic RRT occurred less often after CBVP vs CBV (P = 0.031 and 0.0003, respectively). In addition, three other early and eight late non-relapse deaths were seen. Median follow-up of the entire group is 5.1 (range 2.8-10.2) years. The probability of overall NRM was 26% (95% confidence interval (CI) 13-50%) with CBV vs 23% (95% CI 12-41%) with CBVP (P = 0.40). The progression-free survival and relapse rates were similar. Although the rates of fatal RRT, pulmonary RRT and overall NRM were similar with CBV or CBVP, CBVP produced less mucosal and liver RRT with a comparable antitumor effect. As many autografted patients are cured, future efforts should include measures to decrease NRM.

Adult↗
PubMed1999

Antileukemic effect of interleukin-2-transduced murine bone marrow after autologous transplantation.

Myeloablative chemotherapy or radiation therapy supported by autologous stem cell transplantation (SCT) for the treatment of hematologic malignancies such as acute leukemia, lymphoma, and myeloma is associated with high rates of relapse. The reasons for this are 1) autologous transplantation lacks the in vivo graft-vs.-tumor (GVT) effect associated with allogeneic SCT, which is effective in controlling or eliminating residual malignant cells remaining in the body after high-dose therapy, and 2) contaminating malignant cells in the autologous graft are reinfused into the body. Some researchers have attempted to administer immunomodulatory cytokines to simulate a GVT effect, and although this has shown some efficacy, it has several disadvantages. These include high toxicity associated with systemic administration, a short in vivo half-life, and insufficient levels reaching the site of residual disease. As an alternative, we investigated whether delivery of the cytokine interleukin (IL)-2 to the bone marrow can exert an antileukemic effect while avoiding the problems associated with systemic administration. We describe the delivery of IL-2 to the bone marrow by transplantation of syngeneic bone marrow, retrovirally transduced with the gene for IL-2, into lethally irradiated mice. We were able to efficiently transduce murine bone marrow with the IL-2 gene without adversely affecting clonogenic output from hematopoietic progenitors, and we were able to achieve expression of the transgene in transplanted animals. However, IL-2 transduction inhibited hematopoietic reconstitution in lethally irradiated mice. Marrow transduced with high-titer, high-expressing IL-2 retrovirus failed to engraft, and a low-titer, low-expressing IL-2 retrovirus also demonstrated reduced engraftment, although engraftment was sufficient to support survival of transplanted mice. Long-term, low-level expression of the IL-2 transgene was detectable in these mice and was effective in exerting an antileukemic effect. Mice transplanted with control marrow and challenged with leukemic cells suffered 100% mortality within 70 days, whereas mice transplanted with IL-2-transduced marrow exhibited 50% survival over the 175-day duration of this study. The work shows that delivery of immunomodulatory cytokines to the bone marrow can be achieved by transplantation of genetically modified hematopoietic cells. Furthermore, low-level IL-2 expression can exert an antileukemic effect. These data suggest that this may be an effective immunotherapeutic strategy to reduce relapse after autologous transplantation, but the selection and expression of the cytokine must be carefully considered to minimize adverse effects on hematopoiesis.

3T3 Cells↗
PubMed1999

Antileukemic effect of interleukin-2-transduced murine bone marrow after autologous transplantation.

Myeloablative chemotherapy or radiation therapy supported by autologous stem cell transplantation (SCT) for the treatment of hematologic malignancies such as acute leukemia, lymphoma, and myeloma is associated with high rates of relapse. The reasons for this are 1) autologous transplantation lacks the in vivo graft-vs.-tumor (GVT) effect associated with allogeneic SCT, which is effective in controlling or eliminating residual malignant cells remaining in the body after high-dose therapy, and 2) contaminating malignant cells in the autologous graft are reinfused into the body. Some researchers have attempted to administer immunomodulatory cytokines to simulate a GVT effect, and although this has shown some efficacy, it has several disadvantages. These include high toxicity associated with systemic administration, a short in vivo half-life, and insufficient levels reaching the site of residual disease. As an alternative, we investigated whether delivery of the cytokine interleukin (IL)-2 to the bone marrow can exert an antileukemic effect while avoiding the problems associated with systemic administration. We describe the delivery of IL-2 to the bone marrow by transplantation of syngeneic bone marrow, retrovirally transduced with the gene for IL-2, into lethally irradiated mice. We were able to efficiently transduce murine bone marrow with the IL-2 gene without adversely affecting clonogenic output from hematopoietic progenitors, and we were able to achieve expression of the transgene in transplanted animals. However, IL-2 transduction inhibited hematopoietic reconstitution in lethally irradiated mice. Marrow transduced with high-titer, high-expressing IL-2 retrovirus failed to engraft, and a low-titer, low-expressing IL-2 retrovirus also demonstrated reduced engraftment, although engraftment was sufficient to support survival of transplanted mice. Long-term, low-level expression of the IL-2 transgene was detectable in these mice and was effective in exerting an antileukemic effect. Mice transplanted with control marrow and challenged with leukemic cells suffered 100% mortality within 70 days, whereas mice transplanted with IL-2-transduced marrow exhibited 50% survival over the 175-day duration of this study. The work shows that delivery of immunomodulatory cytokines to the bone marrow can be achieved by transplantation of genetically modified hematopoietic cells. Furthermore, low-level IL-2 expression can exert an antileukemic effect. These data suggest that this may be an effective immunotherapeutic strategy to reduce relapse after autologous transplantation, but the selection and expression of the cytokine must be carefully considered to minimize adverse effects on hematopoiesis.

Animals↗
PubMed1999

Expression of SCM-1alpha/lymphotactin and SCM-1beta in natural killer cells is upregulated by IL-2 and IL-12.

Recruitment of lymphocytes is an important feature of the host immune response against pathogens. However, the mechanisms by which lymphocytes are attracted are not yet fully understood. Recently, the cDNA of a lymphocyte-specific chemokine, lymphotactin (Lptn), was isolated from murine and human T cells and was also found to be expressed in murine NK cells and human NK cell clones. This study investigated the influence of interleukin (IL)-2 and IL-12 on the expression of Lptn, also known as SCM (single cysteine motif)-1alpha, and SCM-1beta, a 97% homolog of Lptn, in freshly isolated human NK cells and the human NK cell line NK-92. Northern blot analysis and RT-PCR confirmed that nonactivated human NK cells expressed both genes at low level. After activation with IL-2 or IL-12, the expression of both Lptn and SCM-1beta was upregulated within hours. NK-92 cells maintained in medium supplemented with IL-2 constitutively expressed SCM-1 mRNA. However, after 24 h of IL-2 starvation and subsequent culturing at various IL-2 concentrations, the expression of Lptn/SCM-1alpha was upregulated in a dose-dependent manner, whereas the expression of SCM-1beta remained consistently high. These observations indicate that NK cells, in addition to T lymphocytes, express Lptn/SCM-1alpha and SCM-1beta after cytokine activation. The upregulation of these chemokines in NK cells on activation likely acts to increase the number of effector cells reaching the site of an immune response such as inflammation.

Blotting, Southern↗
PubMed1999

Immunotherapy of malignant melanoma in a SCID mouse model using the highly cytotoxic natural killer cell line NK-92.

In this work, we evaluated the potential of the natural killer (NK) cell line NK-92 and its IL-2-independent variants NK-92MI and CI, as immunotherapy for melanoma. In vitro, we found that NK-92 was much more cytotoxic to a number of human melanoma cell lines than lymphokine-activated killer (LAK) cells, particularly at low effector/target (E:T) ratios. In vivo treatment of mice challenged with MEWO melanoma cells with i.v. administered NK-92 and NK-92-MI resulted in a 1.5-2.5-fold increase in average length of survival. NK-92, MI, and CI were also effective against the WM1341 cell line, causing a 2-5-fold increase in survival when administered before the malignant cells. With s.c. injection, MEWO and WM1341 caused a primary tumor mass, secondary tumors, and metastatic cells. NK-92 cells reduced WM1341 primary tumor size by 40-90% and MEWO tumors by 30-75%. Similar results were seen with NK-92MI and CI. These data show that NK-92 cells are highly cytotoxic to human melanoma cells in vitro and in vivo and suggest that treatment with NK-92 cells may be a potentially effective immunotherapeutic modality in melanoma.

Animals↗