TARGETED DISRUPTION OF CD44 IN MDAY-D2 LYMPHOSARCOMA CELLS HAS NO EFFECT ON SUBCUTANEOUS GROWTH OR METASTATIC CAPACITY

CD44 splice variants have been shown to be involved in metastasis of c arcinomas. In addition, the standard form of CD44 has been implicated in metastasis, particularly of melanomas and lymphomas. To investigate this, we have generated a CD44-negative mutant of the highly metastat ic murine MDAY-D2 lymphosarcoma. The two CD44 alleles of this diploid cell line were sequentially disrupted by homologous recombination, usi ng isogenic CD44 genomic constructs interrupted by a neomycin or hygro mycin resistance-conferring gene. The resulting double knockout (DKO) cells had completely lost the capacity to bind to immobilized hyaluron ic acid, but did not differ from MDAY-D2 cells in integrin expression or in vitro growth. Subcutaneous (s.c.) growth potential and metastati c capacity of MDAY-D2 and DKO cells were assessed by s.c. and i.v. inj ection of the lowest cell dose (10(3) or 10(4), respectively) that gav e rise to tumor formation by MDAY-D2 cells in similar to 100% of the m ice. Quite unexpectedly, we observed no difference at all in either s. c. growth rate or local invasion into surrounding tissues between MDAY -D2 cells and the CD44-negative DKO cells. Also hematogenous metastasi s formation upon i.v. injection was similar: both parental and DKO cel ls metastasized extensively to the spleen, liver, and bone marrow. We conclude that, at least for these MDAY-D2 lymphosarcoma cells, the sta ndard form of CD44 is dispensable for tumor growth and metastasis. Our results show that targeted disruption of genes in tumor cells is a fe asible approach to study their role in tumorigenesis and metastasis.