cathepsin-g and Cell-Transformation--Neoplastic

Here, we evaluated whether the overexpression of transcriptionally inactive ΔNp73 cooperates with PML/RARA fusion protein in the induction of an APL-leukemic phenotype, as well as its role in vitro in proliferation, myeloid differentiation, and drug-induced apoptosis. Using lentiviral gene transfer, we showed in vitro that ΔNp73 overexpression resulted in increased proliferation in murine bone marrow (BM) cells from hCG-PML/RARA transgenic mice and their wild-type (WT) counterpart, with no accumulation of cells at G2/M or S phases; instead, ΔNp73-expressing cells had a lower rate of induced apoptosis. Next, we evaluated the effect of ΔNp73 on stem-cell self-renewal and myeloid differentiation. Primary BM cells lentivirally infected with human ΔNp73 were not immortalized in culture and did not present significant changes in the percentage of CD11b. Finally, we assessed the impact of ΔNp73 on leukemogenesis or its possible cooperation with PML/RARA fusion protein in the induction of an APL-leukemic phenotype. After 120 days of follow-up, all transplanted mice were clinically healthy and, no evidence of leukemia/myelodysplasia was apparent. Taken together, our data suggest that ΔNp73 had no leukemic transformation capacity by itself and apparently did not cooperate with the PML/RARA fusion protein to induce a leukemic phenotype in a murine BM transplantation model. In addition, the forced expression of ΔNp73 in murine BM progenitors did not alter the ATRA-induced differentiation rate in vitro or induce aberrant cell proliferation, but exerted an important role in cell survival, providing resistance to drug-induced apoptosis.

Topics: Animals; Antimetabolites, Antineoplastic; Apoptosis; Bone Marrow Transplantation; Cathepsin G; Cell Differentiation; Cell Proliferation; Cell Self Renewal; Cell Transformation, Neoplastic; Cells, Cultured; Cytarabine; Gene Expression Regulation, Leukemic; Genetic Predisposition to Disease; Leukemia; Mice, Inbred NOD; Mice, SCID; Mice, Transgenic; Neoplastic Stem Cells; Phenotype; Promyelocytic Leukemia Protein; Retinoic Acid Receptor alpha; Signal Transduction; Time Factors; Transfection; Tumor Protein p73; Up-Regulation

Recurrent chromosomal translocations involving the RAR alpha locus on chromosome 17 are the hallmark of acute promyelocytic leukemia (APL). The RAR alpha gene fuses to variable partners (PML, PLZF, NPM, NuMA and STAT5B: X genes) leading to the expression of APL-specific fusion proteins with identical RAR alpha moieties. To analyse whether the variable X moiety could affect the activity of the fusion protein in vivo, we generated and characterized, on a comparative basis, NPM/RAR alpha transgenic mice (TM) in which the fusion gene is expressed under the control of a human Cathepsin G (hCG) minigene. We compared the features of the leukemia observed in these TM with those in hCG-PML/RAR alpha and hCG-PLZF/RAR alpha TM. In all three transgenic models, leukemia developed after a variably long latency, with variable penetrance. However, the three leukemias displayed distinct cytomorphological features. hCG-NPM/RAR alpha leukemic cells resembled monoblasts. This phenotype contrasts with what was observed in the hCG-PML/RAR alpha TM model in which the leukemic phase was characterized by the proliferation of promyelocytic blasts. Similarly, hCG-PLZF/RAR alpha TM displayed a different phenotype where terminally differentiated myeloid cells predominated. Importantly, the NPM/RAR alpha oncoprotein was found to localize in the nucleolus, unlike PML/RAR alpha and PLZF/RAR alpha, thus possibly interfering with the normal function of NPM. Similarly to what was observed in human APL patients, we found that NPM/RAR alpha and PML/RAR alpha, but not PLZF/RAR alpha leukemia, was responsive to all-trans retinoic acid (ATRA) or As2O3 treatments. Taken together, our results underscore the critical relevance of the X moiety in dictating the biology of the disease and the activity of the APL fusion oncoprotein.

Topics: Animals; Antineoplastic Agents; Cathepsin G; Cathepsins; Cell Proliferation; Cell Transformation, Neoplastic; DNA-Binding Proteins; Gene Fusion; Humans; Kruppel-Like Transcription Factors; Leukemia, Promyelocytic, Acute; Mice; Mice, Transgenic; Neoplasm Proteins; Nuclear Proteins; Phenotype; Promyelocytic Leukemia Protein; Promyelocytic Leukemia Zinc Finger Protein; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Serine Endopeptidases; Transcription Factors; Translocation, Genetic; Tretinoin; Tumor Suppressor Proteins

Small, deeply invasive carcinomas invading the muscularis propria or deeper and measuring < or = 2 cm in greatest dimension (S-ADV) are rare in comparison with their larger counterparts (NS-ADV), and their clinicopathologic features are obscure.. S-ADV and NS-ADV cases as well as cases of submucosal carcinoma (SM-CA) were comparatively assessed for: 1) clinicopathologic findings; 2) Ki-67, mitotic, and apoptotic indices; 3) cathepsin G, p53, and bcl-2 immunoreactivities; and 4) c-Ki-ras mutations.. S-ADV and SM-CA, which both are significantly smaller than NS-ADV, did not differ in size, but the frequency of moderately and poorly differentiated carcinoma elements at the leading edges was observed to be higher than in the central cores only in S-ADV, as was tumor "budding" of small clusters of undifferentiated carcinoma cells. The frequency of severe lymphatic involvement in S-ADV was as high as in NS-ADV, and significantly greater than in SM-CA. The Ki-67, mitotic, and apoptotic indices for S-ADV were significantly increased compared with those for NS-ADV and/or SM-CA. Expression of cathepsin G in S-ADV tumor and stromal cells was significantly decreased compared with NS-ADV and/or SM-CA cases. No significant differences in the expression of either p53 or bcl-2 or the incidence of c-Ki-ras mutations were observed among the three groups.. S-ADV can be considered a distinct type of deeply invasive carcinoma, presenting with poor tumor differentiation at the leading edge, and with increased tumor cell proliferation despite its small size.

Topics: Biomarkers, Tumor; Cathepsin D; Cathepsin G; Cathepsins; Cell Division; Cell Transformation, Neoplastic; Colonic Neoplasms; Humans; Middle Aged; Proto-Oncogene Proteins c-bcl-2; Rectal Neoplasms; Serine Endopeptidases; Tumor Suppressor Protein p53