Biomedical subjects

R Repp

Publications and source records attributed to R Repp.

At least 37 records · Page 2Linked to original sources

PubMed2000

Triggering Fc alpha-receptor I (CD89) recruits neutrophils as effector cells for CD20-directed antibody therapy.

CD20 Abs induce clinical responses in lymphoma patients, but there are considerable differences between individual patients. In (51)Cr release assays with whole blood as effector source, RAJI cells were effectively killed by a mouse/human chimeric IgG1 construct of CD20 Ab 1F5, whereas ARH-77 proved resistant to killing by this Ab. When whole blood was fractionated into plasma, mononuclear cells, or granulocytic effector cells, RAJI cells were effectively killed in the presence of complement-containing plasma, whereas the mature B cell line ARH-77 proved complement resistant. However, with a bispecific Ab (BsAb) against the myeloid receptor for IgA (CD89; FcalphaRI) and CD20, a broad range of B cell lines were effectively killed. FcalphaRI is expressed on monocytes/macrophages, neutrophils, and eosinophils. As the numbers of these effector cells and their functional activity can be enhanced by application of G-CSF or GM-CSF, lysis via (FcalphaRI x CD20) BsAb was significantly enhanced in blood from patients during therapy with these myeloid growth factors. Interestingly, the major effector cell population for this BsAb were polymorphonuclear neutrophils, which proved ineffective in killing malignant B cells with murine, chimeric IgG1, or FcgammaRI- or FcgammaRIII-directed BsAbs against CD20. Experiments with blood from human FcalphaRI/FcgammaRI double-transgenic mice showed corresponding results, allowing the establishment of relevant syngenic animal models in these mice. In conclusion, the combination of myeloid growth factors and an (FcalphaRI x CD20) BsAb may represent a promising approach to improve effector cell recruitment for CD20-directed lymphoma therapy.

Animals↗

PubMed2000

The human GRAF gene is fused to MLL in a unique t(5;11)(q31;q23) and both alleles are disrupted in three cases of myelodysplastic syndrome/acute myeloid leukemia with a deletion 5q.

We have isolated the human GRAF gene (for GTPase regulator associated with the focal adhesion kinase pp125(FAK)). This gene was fused with MLL in a unique t(5;11)(q31;q23) that occurred in an infant with juvenile myelomonocytic leukemia. GRAF encodes a member of the Rho family of the GTPase-activating protein (GAP) family. On the protein level, it is 90% homologous to the recently described chicken GRAF gene that functions as a GAP of RhoA in vivo and is thus a critical component of the integrin signaling transduction pathway. The particular position of the human GRAF gene at 5q31 and the proposed antiproliferative and tumor suppressor properties of its avian homologue suggest that it also might be pathogenetically relevant for hematologic malignancies with deletions of 5q. To investigate this possibility, we sequenced 4-5 individual cDNA clones from 13 cases in which one allele of GRAF was deleted. We found point mutations within the GAP domain of the second GRAF allele in one patient. In two additional patients we found an insertion of 52 or 74 bp within the GRAF cDNA that generates a reading frame shift followed by a premature stop codon. GRAF maps outside the previously defined commonly deleted 5q31 region. Nevertheless, inactivation of both alleles in at least some cases suggests that deletions and mutations of the GRAF gene may be instrumental in the development and progression of hematopoeitic disorders with a del(5q).

Artificial Gene Fusion↗

PubMed2000

Flow cytometric discrimination between viable neutrophils, apoptotic neutrophils and eosinophils by double labelling of permeabilized blood granulocytes.

Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) is frequently used to detect apoptotic cells in tissues, cytospins and suspensions. Here we show that TUNEL staining of freshly isolated granulocytes results in non-specific positivity of a distinct population, which can be observed in the presence or absence of TdT. The morphological features of the false-positive cells examined in fluorescence microscopy suggest that the non-specifically stained cells are eosinophilic granulocytes. Granules of eosinophilic granulocytes were brightly stained by non-specific TUNEL reaction independent of TdT. This staining does not, therefore, indicate apoptosis and most likely reflects 'stickiness' of the permeabilized eosinophils. Immunofluorescence with phycoerythrin (PE)-labelled CD16 antibodies performed simultaneously with conventional TUNEL staining confirmed that the false-positive cells in TUNEL staining were CD16-negative eosinophils. In this report we describe a new procedure that allows: (i) the differentiation of neutrophilic and eosinophilic granulocytes in forward scatter versus log side scatter histograms after permeabilisation; (ii) the reliable discrimination between viable neutrophils, apoptotic neutrophils and eosinophilic granulocytes in cytofluorimetry.

Apoptosis↗

PubMed2000

Nuclear gene targeting using negatively charged liposomes.

Oligonucleotides are a very useful tool to control gene activity. Oligos work by complementary base-pairing with target sequences either in the nucleus or in the cytosol (Zelphati, O., Szoka, F.C., Jr., 1996. Liposomes as a carrier for intracellular delivery of antisense oligonucleotides: a real or magic bullet? J. Contr. Rel. 41, 99-119). In a new approach using chimeric oligonucleotides (Yoon, K., Cole Strauss, A., Kmiec, E.B., 1996. Targeted gene correction of episomal DNA in mammalian cells mediated by a chimeric RNA-DNA oligonucleotide. Proc. Natl. Acad. Sci. USA 93, 2071-2076) conversion of single base mutations with help of intranuclear repair mechanisms maybe an advantageous method to cure genetic diseases which are based on single point mutations. These chimeric oligonucleotides are constructed in a way that they form an intramolecular double strand of DNA and modified RNA-bases. We used a fluorescent labelled pure 68-mer DNA-analogue of a chimeric oligonucleotides to follow the intracellular fate of these kind of genetic material. The oligos were complexed with protamine sulfate and coated with three different liposomal formulations. The AVE-3 formulation shows enhanced properties compared to a classical neutral and negatively charged formulation. Nuclear localisation of oligos could only be observed with the AVE-3 formulation. Furthermore only the negatively charged liposome formulations interact with the protamine-complexed oligonucleotides.

Biological Transport↗

PubMed2000

Nuclear transport of oligonucleotides in HepG2-cells mediated by protamine sulfate and negatively charged liposomes.

PURPOSE: The aim of this study was to characterize the intracellular fate and nuclear uptake kinetics of oligonucleotides (ON) that were complexed with protamine sulfate (PS) and negatively charged liposomes at different ratios of ON to PS. METHODS: Double-fluorescence labelling of ON and liposomal lipid was applied to simultaneously monitor the interaction as well as the individual fate of active agent and carrier upon intracellular delivery using confocal laser scanning microscopy (CLSM). A DNA-analogue of a 68-mer intramolecular double-stranded RNA:DNA-hybridoligonucleotide (chimeraplasts) with unmodified phosphate backbone was employed. This construct was condensed with PS and coated with a liposomal formulation (AVE-3 = artificial viral envelope). RESULTS: PS-ON complexes and AVE -3-coated complexes with a defined composition were very effective in nuclear transport of ON for a ON:PS charge ratio of 1:3. Nucleus:cytosol fluorescence ratios peaked at about 10 hrs and started to decrease again at 21 hrs. CONCLUSIONS: AVE associates with PS-condensed ON, and this complex is able to be taken up by cells and to deliver ON to the nucleus. PS-ON complexes are released from the liposomal formulation, mainly as an extranuclear enzymatic degradation of the liposomal phospholipids. The results of the kinetic analysis can be used to optimize transfection protocols with ON in HepG2 cells.

Anions↗

PubMed2000

Quantitative DNA fragment analysis for detecting low amounts of hepatitis B virus deletion mutants in highly viremic carriers.

Many variants of hepatitis B virus (HBV) with deletions in the viral genome have been identified. Some of these variants are indicator or even effector of a more severe course of hepatitis. These deletion mutants contribute a variable and sometimes very low proportion to the viral population. For early detection of small amounts of deletion mutants among a large number of wild-type genomes, we applied a new screening method designated quantitative fragment analysis (QFA). By QFA the whole viral genome can be scanned for the presence of deletions or insertions of >/=3 nucleotides representing more than 2% of the viral population. Using QFA we showed that an often described deletion of 8 nucleotides is packaged in viral capsids and not a polymerase chain reaction (PCR) artifact. QFA was applied to study the emergence of deletion mutants in a group of 18 pediatric patients who had been infected from a common source while being under multidrug cancer chemotherapy. All patients had developed a highly viremic asymptomatic HBV carrier state. In 3 of these patients 3 different kinds of HBV deletion mutants were found by QFA: 8 bp deletions within the core promoter, core gene deletions from 8 to 86 bp, and large deletions of up to 1,989 bp spanning the precore/core and the preS/S reading frames. PCR primers that specifically amplify deletion variants enabled the detection of additional patients harboring the investigated variant.

Adolescent↗

PubMed2000

Comparison of methylene blue, riboflavin, and N-acetylcysteine for the reduction of nitric oxide-induced methemoglobinemia.

OBJECTIVE: To investigate the treatment of nitric oxide (NO)-induced methemoglobinemia by methylene blue (MB), riboflavin, and N-acetylcysteine (NAC) in vitro. DESIGN: Prospective, controlled in vitro study. SETTING: Research laboratory in a university hospital. PARTICIPANTS: Five healthy volunteers. INTERVENTIONS: Generation of 16% to 18% of methemoglobin in red blood cells by NO and subsequent addition of MB, riboflavin, or NAC. Simultaneous NO (32 ppm) and MB or riboflavin exposure of red blood cells. Induction of 14% to 18% of methemoglobin in red blood cells by NO, subsequent addition of MB or riboflavin, and further incubation with NO (80 ppm). MEASUREMENTS AND MAIN RESULTS: After discontinuation of NO, mean half-life for methemoglobin was significantly reduced by MB from 356 mins (controls) to 5 mins (10 microM) in a dose-dependent manner (p < .001). NAC did not alter the half-life for methemoglobin, and a reduction from 356 to 168 mins was seen for 120 microM riboflavin (p < .001). Methemoglobin formation after 3 hrs of NO exposure was 4.3%+/-0.7% in controls and 0.3%+/-0.1% with 10 microM MB (p < .001); 1 microM MB attenuated methemoglobin formation to 1.9%+/-0.1% (p < .01). With riboflavin (120 microM), methemoglobin was 2.2%+/-0.5% vs. 3.2%+/-0.6% in controls (p < .001). In the presence of high methemoglobin concentrations, further methemoglobin formation was inhibited by 1 and 10 microM MB (p < .001) and attenuated by 0.1 microM MB (p < .001) but not by riboflavin. CONCLUSIONS: In vitro, NO-induced methemoglobin formation is significantly decreased by medium (1 microM) and high (10 microM) concentrations of MB and partially by high riboflavin concentrations (120 microM). NAC and low concentrations of riboflavin do not alter methemoglobin formation.

Acetylcysteine↗

PubMed1999

A DNA damage repair mechanism is involved in the origin of chromosomal translocations t(4;11) in primary leukemic cells.

Some chromosomal translocations involved in the origin of leukemias and lymphomas are due to malfunctions of the recombinatorial machinery of immunoglobulin and T-cell receptor-genes. This mechanism has also been proposed for translocations t(4;11)(q21;q23), which are regularly associated with acute pro-B cell leukemias in early childhood. Here, reciprocal chromosomal breakpoints in primary biopsy material of fourteen t(4;11)-leukemia patients were analysed. In all cases, duplications, deletions and inversions of less than a few hundred nucleotides indicative of malfunctioning DNA repair mechanisms were observed. We concluded that these translocation events were initiated by several DNA strand breaks on both participating chromosomes and subsequent DNA repair by 'error-prone-repair' mechanisms, but not by the action of recombinases of the immune system.

Adolescent↗

PubMed1999

Rapid isolation of chromosomal breakpoints from patients with t(4;11) acute lymphoblastic leukemia: implications for basic and clinical research.

Chromosomal translocations t(4;11)(q21;q23) are associated with a group of acute lymphoblastic leukemias with very poor prognosis. From the complete sequences of the breakpoint cluster regions of the human MLL and AF-4 translocation partner genes, a novel set of 66 oligonucleotides that facilitates the rapid identification of translocation breakpoints by PCR analysis of genomic DNA was designed. For each breakpoint, a pair of optimally snited primers can be assigned, which improves the monitoring of the disease during treatment. Comparison of the breakpoints with the corresponding parental sequences also contributes to our better understanding of the illegitimate recombination events leading to these translocations.

Alu Elements↗

PubMed1999

Quantitation of gene expression by real-time PCR disproves a "retroviral hypothesis" for childhood-onset diabetes mellitus.

Children with insulin-dependent diabetes mellitus (IDDM) suffer from a chronic autoimmune beta cell destruction of unknown origin, maybe due to superantigens or retroviral endogenous genes. Recently, a novel endogenous retrovirus designated as IDDMK 22 was proposed to encode for such a candidate autoimmune gene in type 1 diabetes. We therefore analyzed the expression of IDDMK 22 genes in peripheral blood leukocytes and plasma from 55 healthy children and 55 diabetic children including 11 patients with acute disease onset. In our study we applied an improved quantitative and highly specific real-time PCR assay. In contrast to previous data obtained by conventional PCR. IDDMK 22 gene expression did not differ between diabetic and nondiabetic individuals. For this reason, we propose that IDDMK 22 is an ubiquitous endogenous retroviral element in the human genome but not a candidate autoimmune gene for IDDM, especially in childhood-onset disease. Real-time PCR proved to be a highly sensitive and specific method for detection and quantitation of very low amounts of mRNA and will thereby be useful regarding the special demands in pediatric studies dealing with very low amounts of specimen.

Adolescent↗

PubMed1999

Leptin and neuropeptide Y gene expression in human placenta: ontogeny and evidence for similarities to hypothalamic regulation.

The objective of the present study was to examine the impact of preeclampsia on the relation of leptin and neuropeptide Y (NPY) gene expression in human placenta. A second goal was to monitor the change of leptin messenger RNA (mRNA) with increasing gestational age. Placental tissue was obtained from 17 premature deliveries, 18 term deliveries, and 10 mothers with preeclampsia. Gene expression of leptin, NPY, and two housekeeping genes (beta-actin and glyceraldehyde-3-phosphate dehydrogenase was quantified using real-time PCR. The leptin/beta-actin mRNA ratio was significantly higher in specimens of patients with preeclampsia than in those of gestational age-matched controls (0.63+/-0.23 vs. 0.09+/-0.04 relative U (RU); P = 0.03). NPY/beta-actin mRNA was significantly reduced in the preeclampsia group (0.003+/-0.001 vs. 0.026+/-0.008 RU in controls; P = 0.01). The NPY/leptin ratio was 0.11+/-0.09 for preeclamptic placenta samples and 1.7+/-0.6 RU for the controls (P = 0.02). The leptin/beta-actin ratio was significantly lower in placenta from premature deliveries than in term deliveries (0.02+/-0.004 vs. 0.12+/-0.05 RU; P = 0.01). Similar results were obtained for normalization to glyceraldehyde-3-phosphate dehydrogenase mRNA. Our data suggest an increase of placental leptin production with gestational age. In patients with preeclampsia, elevated leptin expression goes along with suppressed NPY expression. This resembles hypothalamic regulation.

Adult↗

PubMed1998

Evaluating antibodies for their capacity to induce cell-mediated lysis of malignant B cells.

Promising results from clinical trials have led to renewed interest in effector mechanisms operating in antibody-based therapy of leukemia and lymphoma. We tested a panel of B-cell antibodies from the Sixth Human Leukocyte Differentiation Antigen workshop for their capacity to mediate antibody-dependent cellular cytotoxicity, often considered to be one of the most potent effector mechanisms in vivo. As effector cells, mononuclear cells and polymorphonuclear (PMN) cells from healthy donors were compared with Fc gammaRI (CD64)-expressing PMN cells from patients receiving granulocyte colony-stimulating factor (G-CSF) treatment. Of the 29 IgG workshop antibodies binding most strongly to the tested malignant human B-cell lines, only 3 consistently induced target cell lysis. These three antibodies were determined to be HLA DR reactive. Experiments with a panel of HLA class II antibodies showed the involvement of individual Fc gamma receptors on effector cells to be strongly dependent on the antibody isotype. We then compared killing mediated by chimeric IgG1 antibodies with that from Fc gammaRI-directed bispecific antibodies, targeting classical HLA class II, or the Lym-1 and Lym-2 antigens. The latter two are variant forms of HLA class II, which are highly expressed on the surface of malignant B cells but which are found only at low levels in normal cells. With blood from G-CSF-treated donors, bispecific antibodies showed enhanced killing compared to their chimeric IgG1 derivatives, because they were more effective in recruiting Fc gammaRI-expressing PMN cells. G-CSF- and Fc gammaRI-directed bispecific antibodies to HLA class II, therefore, seem to be an attractive combination for lymphoma therapy.

Antibodies, Bispecific↗

PubMed1998

Fcgamma receptor IIa polymorphism in Caucasian patients with systemic lupus erythematosus: association with clinical symptoms.

OBJECTIVE: The class II human leukocyte Fcy receptor for IgG (FcgammaRIIa) occurs in 2 codominantly expressed allelic forms (R131 and H131). Cells expressing IIa-H131 interact much more effectively with complexed IgG2 and IgG3 than do cells with IIa-R131. This might be linked to variability in immune complex handling, and therefore related to disease pathogenesis. The present study examines these possibilities in a cohort of Caucasian patients with systemic lupus erythematosus (SLE). METHODS: One hundred eight Caucasian patients were diagnosed with SLE according to the American College of Rheumatology criteria. The SLE patients and 187 Caucasian controls were genotyped for the FcgammaRIIa polymorphism, and associations between FcgammaRIIa genotypes, selected HLA haplotypes, and clinical as well as laboratory features were analyzed. RESULTS: No significant skewing of the FcgammaRIIa polymorphism was observed in the SLE cohort. Various clinical and serologic parameters were found more frequently or at a younger age in patients homozygous for the genotype IIa-R/R131 compared with those with the genotype IIa-H/H131. In patients with the genotype IIa-R/R131, significantly higher frequencies of proteinuria, hemolytic anemia, anti-nuclear RNP antibodies, and hypocomplementemia were found. The only clinical symptom observed more frequently in patients homozygous for IIa-H/H131 was livedo. Patients with the IIa-R/R131 genotype were significantly younger at disease onset and had an earlier incidence of arthritis, sicca syndrome, nephritis, lymphadenitis, hematologic abnormalities, immunologic abnormalities, lupus anticoagulant, cryoglobulinemia, and hypocomplementemia. HLA-DR3 was found in 41.7% of SLE patients, but was not associated with clinical symptoms, serologic abnormalities, or the homozygous genotypes of the FcgammaRIIa, although an association with a significantly later onset of SLE was found. CONCLUSION: The FcgammaRIIa polymorphism constitutes an additional factor that might influence the clinical manifestations and course of SLE, but does not represent a genetic risk factor for the occurrence of SLE.

Adolescent↗

PubMed1998

Autologous PBPCT in a patient with lymphoma and Sjögren's syndrome: complete remission of lymphoma without control of the autoimmune disease.

A patient with primary Sjögren's syndrome developed massive gland enlargement and was diagnosed a MALT-type lymphoma stage IIA. After initial radiotherapy the patient relapsed with high grade immunoblastic lymphoma. Chemotherapy led to remission of the lymphoma and improvement of the autoimmune disease. A peripheral blood stem cell harvest was performed. However, subsequent to standard chemotherapy the patient experienced an exacerbation of arthralgia and vasculitis while his lymphoma remained in partial remission. With additional chemotherapy and steroid treatment complete remission of the lymphoma and autoimmune disease was achieved and high-dose chemotherapy followed by PBPCT with an unmanipulated graft was performed. Engraftment was prompt with no signs of active autoimmune disease after the transplantation. Two months later, signs of autoimmune disease slowly recurred. Steroid treatment improved this, but the patient remained steroid-dependent and later died from therapy-resistant Pneumocystis carinii pneumonia while the lymphoma remained in complete remission. In this patient with systemic Sjögren's disease, PBPCT completely controlled the aggressive lymphoma but was not permanently effective for the autoimmune disease.

Adult↗

PubMed1998

A new fingerprint method for sequence analysis of chromosomal translocations at the genomic DNA level.

Chromosomal rearrangements constitute a significant feature of leukemogenesis and malignant transformation in general. Nucleotide patterns in the immediate vicinity of the break point may provide important information about the underlying causalities, eg illegitimate recombination events mediated by topoisomerase II, Alu repeats, or VDJ recombinase. In order to facilitate the determination of those DNA patterns, we developed a new fingerprint approach. In a first step, two DNA fragments were independently amplified by long distance PCR: the genomic region carrying the break point and the normal nonrearranged counterpart. Subsequently, both PCR products were digested with restriction enzymes, end-labelled with a fluorescent dye, and subjected to high resolution polyacrylamide gel electrophoresis. By comparing the restriction patterns of the rearranged and the nonrearranged PCR fragments, the break points could be easily localized within a size range coverable by a single sequencing reaction. Finally, the exact DNA sequence across the break point was directly determined. The 'fingerprint' technique is fast, reliable and enables the assay of multiple samples in parallel.

Base Sequence↗

PubMed1998

Coiling phagocytosis discriminates between different spirochetes and is enhanced by phorbol myristate acetate and granulocyte-macrophage colony-stimulating factor.

The mechanisms involved in coiling phagocytosis are not yet known, and it is not even clear whether this phenomenon is either an incidental event or a specific response. Therefore, the phagocytic uptake of Borrelia burgdorferi and other spirochetes by human monocytes in vitro was used to investigate the involvement of both sides--microbes and phagocytes--in coiling phagocytosis. As seen with electron microscopy, morphologically similar Borrelia, Leptospira and Treponema strains induced markedly different frequencies of coiling phagocytosis. The monocytes used coiling phagocytosis for both live (motile) and killed (nonmotile) B. burgdorferi, but pseudopod coils were observed neither with fragmented B. burgdorferi nor with cell-free supernatant from B. burgdorferi cultures. Investigation of the relationship of coiling phagocytosis with other pseudopod-based cellular mechanisms revealed that the use of bioreagents that inhibit conventional phagocytosis also inhibited coiling phagocytis but did not affect membrane ruffling. Bioreagents that increase membrane ruffling did not affect phagocytosis of B. burgdorferi, except for granulocyte-macrophage colony-stimulating factor and phorbol myristate acetate, which increased coiling phagocytosis selectively. These results demonstrate that coiling phagocytosis is not induced by microbial motility, viability, or a certain morphology and that it is not a random event. Rather, it is a selective uptake mechanism actively driven by the phagocytes. However, whether coiling phagocytosis represents an independent alternative to conventional phagocytosis or, alternatively, a fault in conventional phagocytosis remains to be determined.

Borrelia burgdorferi Group↗

PubMed1997

Backtracking leukemia to birth: identification of clonotypic gene fusion sequences in neonatal blood spots.

Epidemiological evidence has suggested that some pediatric leukemias may be initiated in utero and, for some pairs of identical twins with concordant leukemia, this possibility has been strongly endorsed by molecular studies of clonality. Direct evidence for a prenatal origin can only be derived by prospective or retrospective detection of leukemia-specific molecular abnormalities in fetal or newborn samples. We report a PCR-based method that has been developed to scrutinize neonatal blood spots (Guthrie cards) for the presence of numerically infrequent leukemic cells at birth in individuals who subsequently developed leukemia. We demonstrate that unique or clonotypic MLL-AF4 genomic fusion sequences are present and detectable in neonatal blood spots from individuals who were diagnosed with acute lymphoblastic leukemia at ages 5 months to 2 years and, therefore, have arisen during fetal hematopoiesis in utero. This result provides unequivocal evidence for a prenatal initiation of acute leukemia in young patients. The method should be applicable to other fusion genes in children with common subtypes of leukemia and will be of value in attempts to unravel the natural history and etiology of this major subtype of pediatric cancer.

Artificial Gene Fusion↗

PubMed1997

FcalphaRI (CD89) as a novel trigger molecule for bispecific antibody therapy.

Promising results from clinical trials with unconjugated antibodies stimulated renewed interest in immune effector mechanisms of monoclonal antibodies (MoAbs). We investigated the potential of IgA as antibody isotype for cell- or complement-mediated tumor cell lysis and assessed the potential of its myeloid Fc receptor, FcalphaRI (CD89), as trigger molecule for bispecific antibody (BsAb)-mediated immunotherapy. Comparing hapten-directed antibodies of human IgA2 with IgG1 or IgG3 isotypes, we found all three to mediate effective killing of sensitized tumor target cells in whole blood assays. Analysis of effector mechanisms showed IgG-mediated lysis to be predominantly complement-dependent, whereas IgA-dependent killing was primarily effector cell-mediated. A comparison of effector cell populations in antibody-dependent cell-mediated cytotoxicity (ADCC) showed neutrophils to be most important for IgA-dependent tumor cell killing, involving FcalphaRI as shown with Fc receptor blocking antibodies. Reverse ADCC experiments against target cells sensitized with Fc receptor antibodies, or assays with FcalphaRI-directed bispecific antibodies confirmed FcalphaRI as effective trigger molecule in polymorphonuclear neutrophil (PMN)-mediated lysis. During granulocyte colony-stimulating factor (G-CSF ) therapy, (FcalphaRI x HER-2/neu) bispecific antibodies induced enhanced killing of HER-2/neu positive SK-BR-3 breast cancer cells in whole blood assays. This enhanced cytotoxicity was paralleled by increased PMN counts, which lead to higher effector to target cell ratios in G-CSF-primed blood. Furthermore, bispecific antibodies, directed to FcalphaRI and Candida albicans, enhanced neutrophils' phagocytosis of fungi. In summary, these results identify IgA as an effective antibody isotype for immunotherapy, working primarily via FcalphaRI on neutrophils. They suggest FcalphaRI-directed bispecific antibodies and G-CSF to be an attractive combination for malignant or infectious diseases.

Antibodies, Bispecific↗