Logs Of Tumor Cells Research Articles

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

An Effective, Direct Immunomagnetic Procedure for Purging Acute Lymphoblastic Leukemia Cells from Human Bone Marrow.

The AB4 monoclonal antibody, which recognizes an HLA-DR epitope, was found to bind to a high percentage of malignant blast cells in samples obtained from 27 patients with ALL. These included 11 of 11 cases with c-ALL, 3 of 7 with pre-pre-B, and 8 of 9 cases with pre-B ALL. AB4 was used together with anti CD10 and anti CD19 antibodies and super-paramagnetic particles for developing a direct immunomagnetic procedure for purging human bone marrow of leukemic cells. In model experiments with KM3 cells admixed to mononuclear bone marrow cells, the individual antibodies each removed 2.8-3.1 logs and 3.6-4.1 logs of tumor cells with one and two purging cycles, respectively. In comparison, the efficacy of a mixture of the three antibodies was 4.4 logs with one treatment cycle, and > 5 logs with repeated treatments. Whereas the use of a commercially available anti-HLA-DR antibody resulted in a 90% reduction in the survival of CFU-GMs and normal blast colonies, AB4 had only a moderate effect on the progenitor cells (46% and 30% reduction). In conjunction with autologous transplantation, bone marrow from a patient was purged with the antibody mixture and 50% of the CFU-GMs and 47% of the CD34+ cells remained after treatment. The patient showed a normal engraftment, reaching a level of 0.5 x 109/l neutrophils by day 20 and 20 x 109/l platelets by day 30. It is concluded that the antibody cocktail may safely and effectively be used for rapid autograft purging in patients with c-ALL, and also in phenotypically selected cases with other subtypes of ALL.

Stem cell isolation.

High-dose chemotherapy with autologous hematopoietic progenitor cell support is increasingly used to treat a variety of malignant diseases. A drawback of this technique is the potential for infusing clonogenic tumor cells with the autograft, producing relapse of the disease in the patient. The use of positive selection techniques to isolate stem cells and thus reduce or eliminate tumor cell contamination has been extensively studied over the past few years. Preliminary clinical results have demonstrated that these procedures deplete 2 to 7 logs of tumor cells and do not impair engraftment. It is too early to assess the ultimate clinical benefit of this strategy. Additional applications of CD34-selection include ex vivo expansion of and gene transfer into hematopoietic progenitor cells and T-cell depletion of allogeneic grafts to reduce the incidence of graft-versus-host disease.

Enhancement of fast neutron beams with boron neutron capture therapy. A mechanism for achieving a selective, concomitant tumor boost.

Both fast neutron radiotherapy and boron neutron capture therapy (BNCT) have been utilized to treat malignant disease. Herein we discuss the potential of combining these treatments to enhance the effectiveness of fast neutron therapy through a concomitant BNCT boost. Using a fast neutron beam generated from a 50 MeV proton on beryllium reaction, we have determined that 0.1% of the beam per microgram of boron-10 per gram of tissue (microgram/g) can be deposited via BNCT. Our mathematical modeling predicts that BNCT enhancement of our beam will lead to an additional 1-2 logs of tumor cell kill for boron-10 concentrations of 30-50 micrograms/g. We have validated this via V-79 cell line in vitro measurements. A Poisson model estimation of how this additional cell kill will influence local tumor control, predicts that BNCT enhancement of fast neutron radiation will lead to a clinically significant improvement in outcome.

The antitumor activity of an anti-CD22 immunotoxin in SCID mice with disseminated Daudi lymphoma is enhanced by either an anti-CD19 antibody or an anti-CD19 immunotoxin

The antitumor activities of immunotoxins (ITs) constructed with deglycosylated ricin A chain (dgA) and either anti-CD19 (HD37) or anti-CD22 (RFB4) monoclonal antibodies were compared in SCID mice with disseminated human Daudi lymphoma (SCID/Daudi). As reported previously, after intravenous injection with Daudi cells, SCID mice develop disseminated lymphoma, which infiltrates the vertebral column and causes paralysis of the hind legs before death. The mean paralysis time (MPT) has been taken as an end point in this tumor model. We have previously reported that early treatment of SCID/Daudi mice with RFB4 coupled to dgA prolongs the MPT in a manner consistent with the killing of 4 logs of tumor cells. In the present study, we show that HD37-dgA kills 2 logs of tumor cells. The lower potency of the HD37-dgA is consistent with its lower IC50 on Daudi cells in vitro. We further show that the antitumor activity of a mixture of HD37-dgA and RFB4-dgA is significantly enhanced in SCID/Daudi mice and is consistent with the killing in excess of 5 logs of tumor cells. However, identical enhancement was observed when a mixture of the RFB4-dgA and the HD37 antibody was administered. In contrast, enhancement was not observed when mice were injected with a mixture of the RFB4 antibody and the HD37-dgA. The results indicate that a "cocktail" of HD37 antibody and RFB4-dgA immunotoxin can have significant antitumor activity in this mouse model of lymphoma and suggest that combinations of particular antibodies and ITs may have cooperative antitumor activity.

The Antitumor Activity of an Anti-CD22 Immunotoxin in SCID Mice With Disseminated Daudi Lymphoma Is Enhanced by Either an Anti-CD19 Antibody or an Anti-CD19 Immunotoxin

Abstract The antitumor activities of immunotoxins (ITs) constructed with deglycosylated ricin A chain (dgA) and either anti-CD19 (HD37) or anti- CD22 (RFB4) monoclonal antibodies were compared in SCID mice with disseminated human Daudi lymphoma (SCID/Daudi). As reported previously, after intravenous injection with Daudi cells, SCID mice develop disseminated lymphoma, which infiltrates the vertebral column and causes paralysis of the hind legs before death. The mean paralysis time (MPT) has been taken as an end point in this tumor model. We have previously reported that early treatment of SCID/Daudi mice with RFB4 coupled to dgA prolongs the MPT in a manner consistent with the killing of 4 logs of tumor cells. In the present study, we show that HD37-dgA kills 2 logs of tumor cells. The lower potency of the HD37-dgA is consistent with its lower IC50 on Daudi cells in vitro. We further show that the antitumor activity of a mixture of HD37-dgA and RFB4-dgA is significantly enhanced in SCID/Daudi mice and is consistent with the killing in excess of 5 logs of tumor cells. However, identical enhancement was observed when a mixture of the RFB4-dgA and the HD37 antibody was administered. In contrast, enhancement was not observed when mice were injected with a mixture of the RFB4 antibody and the HD37-dgA. The results indicate that a “cocktail” of HD37 antibody and RFB4-dgA immunotoxin can have significant antitumor activity in this mouse model of lymphoma and suggest that combinations of particular antibodies and ITs may have cooperative antitumor activity.

Elimination of myeloma cells from bone marrow by using monoclonal antibodies and magnetic immunobeads

The efficacy of immunomagnetic beads to purge human myeloma cells from bone marrow ex vivo was evaluated. The optimal conditions for purging were studied first by using three myeloma cell lines: RPMI-8226, SKO- 007, and SKMM-2. Myeloma cells labeled with the vital fluorescent dye Hoechst 33342 were admixed with normal bone marrow cells, and two monoclonal antibodies reactive with the myeloma cells (PCA-1 and BL-3) were added alone or in combination with the cells. Magnetic beads coated with goat antimouse immunoglobulin G were then added, and the tumor cells to which beads were attached were separated from the mixture with a magnet. The efficacy of tumor cell removal was dependent on the bead-to-tumor ratio; a ratio of more than 500 was optimal in the presence of excess normal marrow cells. The combination of monoclonal antibodies PCA-1 and BL-3 increased the tumor cell removal as compared with either antibody alone. Two cycles of treatment were more effective than one cycle was. Under optimal conditions, 2.3 to 4 logs of tumor cells could be removed from the mixture containing 10% myeloma cells without a significant loss of normal hematopoietic progenitors as measured by CFU-GM, CFU-GEM, and BFU-E. When the efficacy of this procedure was tested on fresh bone marrow from patients with multiple myeloma (MM) by using the combination of PCA-1, BL-3, and J-5, 1.6 to 2.5 logs of tumor cells could be removed by one cycle of treatment, even from marrows containing less than 10% myeloma cells. These observations support the use of monoclonal antibody combinations and immunobeads as a reliable and nontoxic method to eliminate contaminating myeloma cells ex vivo in preparation for autologous bone marrow transplantation in patients with MM.

Human leukemia cell binding and killing by anti-CD5 radioimmunotoxins

We have synthesized a reagent for antibody directed cell targeting composed of the monoclonal antibody (MoAb) T101 linked to the potent toxin ricin. The immunotoxin (IT) was subsequently radiolabeled by a cyclic anhydride procedure with 90Yttrium ( 90Y) to construct a radioimmunotoxin (RIT) that may have potential for cancer therapy. We evaluated the reagent for selectivity in binding and protein synthesis inhibition (PSI) assays. The RIT selectively bound antigen positive leukemia T-cell lines, with minimal binding to antigen negative control lines. The IT inhibited 87% or greater protein synthesis activity at 1 μ g ml and exhibited an IC 50 (the dose inhibiting 50% activity) of 0.18 ± 0.08 μ g ml in the presence of lactose. RIT and nonlabeled IT showed comparable degrees of PSI at 1 μ g ml and 10 μ g ml , suggesting that labeling had little overall effect on the activity of the immunoconjugate. However, indirect evidence showed that the galactose binding site of ricin was inhibited 10-fold by its exposure to 90Y. Control RIT were minimally inhibitory. IT labeled with 131Iodine ( 131I) by an iodine monochloride technique also retained its capability to selectively inhibit protein synthesis. When RIT were tested for potency in a clonogenic assay against human leukemia T-cell lines, they inhibited 3.61 logs of tumor cell growth at μ g ml ml. This did not represent an improvement over the log elimination with radiolabeled antibody alone, which showed 4.19 log elimination of tumor cells. Our observation that the 90Y-labeled RIT and labeled antibody can selectively eliminate about four logs of tumor cells in an in vitro clonogenic assay is unique. The ability of RIT to kill several logs of tumor cells in vitro renders RIT interesting anti-tumor reagents.