Elimination of neuroblastoma and small-cell lung cancer cells with an anti-neural cell adhesion molecule immunotoxin.
BACKGROUND
The development of immunotoxins has been hampered by difficulties, particularly in solid tumors, of finding appropriate target antigens and of linking sufficiently potent toxins.


OBJECTIVE
We evaluated the tissue specificity of an immunotoxin, N901-blocked ricin (N901-bR), and assessed its potential for eliminating neural cell adhesion molecule (NCAM)-positive tumor cells in conditions appropriate for in vitro purging, prior to autologous stem cell transplantation, and its potential for myelosuppression. N901-bR consists of a monoclonal antibody (MAb), N901, directed against CD56, an antigen of the family of NCAMs, covalently linked to blocked ricin as the cytotoxic effector moiety.


METHODS
The tissue specificity of the N901 MAb and the N901-bR immunotoxin was tested against a wide array of human tumor tissues and normal human tissues by immunohistochemical staining. The cytotoxic activity of N901-bR was tested against both small-cell lung cancer (SCLC) cells and neuroblastoma cells, either alone or among normal bone marrow mononuclear cells, and the efficacy of this treatment to specifically eliminate these cells was evaluated in a limiting dilution assay. In addition, normal bone marrow mononuclear cells were incubated with N901-bR, and the toxic effects of the immunotoxin on normal hematopoietic progenitors was evaluated.


RESULTS
N901 and N901-bR exhibited specificity for several neoplasms of neuroectodermal origin, including SCLC and neuroblastoma. Staining of normal tissues was essentially limited to various neuroendocrine cells, cardiac muscle cells, and cells in peripheral nerve tissue. We observed a time- and dose-dependent elimination of tumor cells in vitro, with three logs (i.e., > 99.9%) of malignant cells being killed following only 5 hours of exposure to 10 nM N901-bR. Unconjugated N901 MAb specifically blocked the elimination of NCAM-positive cells by N901-bR, whereas neither an isotype-matched control MAb nor galactose (the ligand of native ricin) had any effect on the activity of the immunotoxin, confirming the specificity of its cytotoxic activity. Importantly, N901-bR used under optimal conditions for in vitro tumor cell depletion was not toxic to hematopoietic precursors.


CONCLUSIONS
N901-bR has the properties required to target CD56, an antigen present not only on cells from a large number of cancers of neuroendocrine origin, but also on some important normal tissues. In addition, treatment with this immunotoxin results in the highly effective and specific elimination of neuroblastoma and SCLC cells and does not affect normal hematopoietic progenitors.


CONCLUSIONS
N901-bR may have clinical utility for purging of neuroblastoma cells and SCLC cells before autologous stem cell transplantation. Further toxicology studies are warranted to assess the potential of N901-bR for in vivo administration.
Roy
D C
DC
Maisonneuve-Rosemont Hospital Research Center, Department of Medicine, University of Montreal, Canada.
Ouellet
S
S
Le Houillier
C
C
Ariniello
P D
PD
Perreault
C
C
Lambert
J M
JM
D000911:Antibodies, Monoclonal / Q000627:therapeutic use*; D016465:Bone Marrow Purging / Q000379:methods; D018288:Carcinoma, Small Cell / Q000276:immunology*; D004305:Dose-Response Relationship, Drug; D006412:Hematopoietic Stem Cells; D006801:Humans; D018796:Immunoconjugates; D007150:Immunohistochemistry; D016130:Immunophenotyping; D000922:Immunotoxins / Q000627:therapeutic use*; D008175:Lung Neoplasms / Q000276:immunology*; D019006:Neural Cell Adhesion Molecules / Q000276:immunology*; D009447:Neuroblastoma / Q000188:drug therapy*; D009928:Organ Specificity; D012276:Ricin / Q000031:analogs & derivatives* / Q000627:therapeutic use; D013997:Time Factors
D000911:Antibodies, Monoclonal; D018796:Immunoconjugates; D000922:Immunotoxins; C080540:N901 MoAb-blocked ricin conjugate; D019006:Neural Cell Adhesion Molecules; D012276:Ricin