Growth of a human carcinoma (HEp3) in nude mice: Rapid and efficient metastasis

Abstract

Our aim was to identify conditions which would permit the development of spontaneous metastasis of a human tumor in nude mice in a rapid and predictable manner and to explore ways to quantitate metastasis. Using a human squamous carcinoma--HEp3--we determined that invasiveness and metastasis were influenced by the host. HEp3 cells grew very rapidly and without a significant lag period in Balb/c and NIH(S)-II nude mice kept in aseptic conditions; a much longer lag period was observed in NIH-Swiss mice kept in conventional conditions. The HEp3 tumor displayed a highly invasive behavior in N-NIH(S)-II mice, in which it invaded the body wall, gaining access to the peritoneal cavity. Microinvasion was noted in all strains of mice inoculated with HEp3 cells. To prolong survival of the mice until metastases became evident, primary tumors were excised when they weighed 1-2 gm. N-NIH(S)-II and Balb/c nude mice, maintained in germ-free conditions, were most receptive to the development of metastases-lung metastases developed in 80% of these mice. Over 60% of all metastases were present within 4 weeks following the removal of the primary. Only 26% of tumor bearing NIH-Swiss developed lung metastases. Lung metastases were observed in some mice in the absence of local microinvasion, local tumor recurrence, and regardless of the presence of lymph node involvement. They were also noted in mice from which primary tumors were not excised. We compared three methods of lung metastasis detection: histology, detection of tumor cells in the cultures of lung minces, and the measurement of the levels of human urokinase-type plasminogen activator directly in the lysates of lungs. The detection of tumor cells in cultures of lung minces appeared to be the most sensitive of these methods and the determination of enzyme in lung lysates seemed to hold most promise for a quantitative approach. These data indicate that, the type of tumor, as well as the genetic background and the maintenance conditions of the host, have to be carefully selected to ensure the successful outcome of the particular tumor-host interaction being studied. Adherence to these guidelines allowed us, in the case of the HEp3 tumor, to develop a rapid, predictable, and efficient model in which to study factors affecting metastasis of this human tumor.

Green Fluorescent Protein Tagging of Extracellular Signal-Regulated Kinase and p38 Pathways Reveals Novel Dynamics of Pathway Activation during Primary and Metastatic Growth

“…These results provide a functional link to our previous findings showing an increased S-phase fraction of T-HEp3 cells and a rapid G 0 -G 1 arrest of D-HEp3 cells, respectively, 48 hours after in vivo inoculation (10). These conclusions, reached through the study of a primary tumor, were also true for metastasis where we found ERK/Elk activation in growth-permissive metastatic sites at times when micrometastatic growth is known to begin (15). In both the primary and the metastatic sites, a proportion of the T-Elk cells had undetectable GFP levels, which suggested that the threshold of ERK signaling that is required to induce GFP is higher than that required to promote cell cycle progression, or indicated a more complete silencing of ERK signaling and induction of dormancy.…”

“…6A). In the nude mouse, a permissive organ for HEp3 metastasis is the lung, in which we have shown that at the time of primary tumor removal 10% of mice have lung metastases and, 4 weeks later, 61% of the mice have lung metastases (15,19). In contrast, the liver and spleen are sites known not to harbor overt HEp3 metastases in nude mice.…”

“…For metastasis studies in the chick embryo, lungs were excised, minced, and inoculated onto a new CAM for expansion of cells lodged in the lung (14). Cells were grown in nude mice as described previously (15). Briefly, 1 ϫ 10 6 cells in 100 L of PBS were inoculated subcutaneously into the interscapular region of BALB/c nu/nu mice (Charles River, Wilmington, MA, or Taconic Farms, Germantown, NY).…”

Secreted Protein Acidic and Rich in Cysteine (SPARC) Mediates Metastatic Dormancy of Prostate Cancer in Bone

“…Although several in vitro models have been described previously, in vivo working models for dormant and recurrent growth have yet to be developed. Recent attempts have characterized syngeneic head and neck squamous cell carcinoma cells (T-Hep and D-Hep) or a pair of breast cancer cell lines (D2.OR and D2A1 cells) that recapitulates dormant growth in vivo (13,(15)(16)(17). However, there is still no appropriate in vivo model that mimics dormancy and recurrence for prostate cancer, especially one that replicates the phenomenon of bone recurrence in patients.…”

Inhibition of migration and invasion of carcinoma cells by urokinase‐derived antagonists of αvβ5 integrin activation

“…Human epidermoid carcinoma HEp3 (T-HEp3), 25 tumorigenic and metastatic in the chick embryo and in nude mice, 26,27 was serially transplanted on chorioallantoic membranes (CAMs) and used as a source of tumorigenic cells. T-HEp3, HEK-293 (human embryonic kidney) and the derived stably transfected cell lines were cultured in DMEM with 10% FBS.…”