Fibroblast cell lines were established from mouse embryos homozygous for a targeted disruption of the lgflr gene, encoding the type 1 receptor for insulin-like growth factor I (IGF-I) and from their wild-type littermates. The cells from the wild-type embryos (W cells) grow in serum-free medium supplemented with platelet-derived growth factor, epidermal growth factor, and IGF-I, whereas the cells from Igflr(-/-) embryos (R-cells) do not, although they grow at a reduced rate in 10% fetal calf serum. The simian virus 40 (SV40) large T antigen, expressed from a transfected plasmid, can trnsform W cells, which form foci in monolayer cultures and colonies in soft agar (anchorage-independent growth). In contrast, the SV40 large tumor antigen, although normally expressed from the transfected template, is unable to transform R-cells, which remain contact-inhibited and fail to grow in soft agar. The transformed phenotype is restored if the R-cells carrying the SV40 large tumor antigen are stably transfected with a plasmid expressing the human IGF-I receptor. These results demonstrate that signaling via the IGF-I receptor is an indispensable component ofthe SV40 transformation pathway. This conclusion is further supported from the results of antisense RNA experiments with tumor cell lines showing that interference with the function of the IGF-I receptor has a profound effect on anchorage-independent growth, even under conditions that only modestly affect growth in monolayers.Interaction of the insulin-like growth factor type I (IGF-I) receptor (IGF-IR) with its ligands (IGF-I, IGF-II, and insulin at supraphysiological concentrations) plays a pivotal role in embryonal development (1) and in the proliferation of several types of cells in culture (2-5). The simian virus 40 (SV40) large tumor antigen (TAg) is one of the most effective transforming agents of mouse cells in culture (6, 7), lowering the growth factor requirements and inducing the ability to grow in soft agar (6,(8)(9)(10)(11) (1), using DNA prepared from their tails. Wild-type and homozygous Igflr(-/-) mutant littermates were used to establish primary cultures of embryonic fibroblasts as described (14). Briefly, the embryos were minced, and after treatment with trypsin for 15 min, the cells of the resulting suspension were plated onto 100-mm culture dishes and cultured in Dulbecco's modified Eagle's medium (DMEM; GIBCO/BRL)/10% fetal bovine serum. The cultures were maintained at subconfluent levels by treating with trypsin every 3 days and reseeding at a density of 1.5 x 103 cells per cm2, following the same protocol used to generate 3T3 cell lines (15). Primary cultures underwent crisis after 2-4 weeks in culture. R-cultures entered crisis later than the wild-type cells due to the relatively slow doubling rate.T98G cells (16) are a human glioblastoma cell line that produces large amounts of IGF-I. Two other cell lines were generated from T98G cells, expressing, respectively, a sense and an antisense RNA to the human IGF-IR RNA. Preparation of the express...

The type 1 insulin-like growth factor receptor (IGF-1R), activated by its ligands, protects several cell types from a variety of apoptotic injuries. The main signaling pathway for IGF-1R-mediated protection from apoptosis has been previously elucidated and rests on the activation of phosphatidylinositol 3-kinase, Akt/protein kinase B, and the phosphorylation and inactivation of BAD, a member of the Bcl-2 family of proteins. In 32D cells (a murine hemopoietic cell line devoid of insulin receptor substrate 1 [IRS-1]), the IGF-1R activates alternative pathways for protection from apoptosis induced by withdrawal of interleukin-3. One of these pathways leads to the activation of mitogen-activated protein kinase, while a third pathway results in the mitochondrial translocation of Raf and depends on the integrity of a group of serines in the C terminus of the receptor that are known to interact with 14.3.3 proteins. All three pathways, however, result in BAD phosphorylation. The presence of multiple antiapoptotic pathways may explain the remarkable efficacy of the IGF-1R in protecting cells from apoptosis.

The type 1 insulin-like growth factor receptor (IGF-1R) plays an important role in the establishment and maintenance of the transformed phenotype. It also has a strong antiapoptotic activity and has a significant influence on the control of cell and body size. Downregulation of the IGF-1R leads to massive apoptosis of cancer cells. These characteristics make it an attractive target for anticancer therapy.