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DNA flow cytometric analysis was performed for prediction of malignancy potential, and hence of outcome, in 17 patients aged 29-76 (mean 52) years with cartilaginous tumour of the chest wall. Histologically there were nine chondromas and eight chondrosarcomas. The chondrosarcomas (3 grade I, 4 grade II, 1 grade III) were significantly larger than the chondromas (13.6 +/- 7.8 vs 4.2 +/- 2.0 cm, p < 0.001). DNA flow cytometry was performed on paraffin-embedded blocks of resected tumour. In eight of the 17 tumours (6 chondromas, 1 grade I and 1 grade II chondrosarcoma) measurement of DNA content failed because of low cellularity and severe calcification. Aneuploid DNA content was detected only in three chondrosarcomas (2 grade II, 1 grade III). The synthetic phase fraction was less than 15% in all the studied chondromas, but exceeded 25% in four of six chondrosarcomas. This case series though small, may indicate a trend towards association between higher histologic grading and greater incidence of aneuploidy with higher synthetic phase fraction. Tumour size, histologic grade, DNA ploidy and adequacy of surgical resection are the main influences on prognosis.

Neoplastic transformation of rat liver cells in vitro by DNA-mediated gene transfer with an oncogene, hhcM, derived from human (Mahlavu) hepatocellular carcinoma, is described and compared with that of NIH3T3 cells. hhcM was cloned in a neomycin-resistant simian virus 40 promoter vector (pNeor/S) and was designated pNrpM-1. BRL-1 or NIH3T3 cells, transfected with pNrpM-1 DNA, showed significant morphological changes, loss of contact inhibition, and anchorage-independent growth. They became highly tumorigenic in nude rats and nu/nu mice. Control BRL-1 and NIH3T3 cells, whether transfected with pNeor/S DNA or not, remained contact inhibited and nontumorigenic. Both the transformants and the tumor cells contained integrated hhcM DNA as shown by Southern blot hybridization. The complete nucleotide sequence of the hhcM 3.0-kilobase DNA was also determined, and it consisted of a possible open reading frame for a protein of 52 kilodaltons (467 amino acids). The high-level production of a slightly modified form of this 52-kilodalton protein in a bacterial expression system has been successfully achieved. The bacteria-produced protein was similar in electrophoretic behavior to the 52- to 53-kilodalton protein synthesized in a cell-free translation system using rabbit reticulocyte lysate programmed with hybrid-selected hhcM-specific mRNA from Mahlavu hepatocellular carcinoma cells.

A B-cell lymphoma was induced in athymic NIH Swiss nu/nu mice by challenging the animals with NIH3T3 cells, previously transfected with a recombinant DNA carrying a human oncogene hhcM, ligated to an SV40 promoter with a neomycin-resistance marker. The gross pathology of the tumor-bearing animal revealed generalized lymphadenopathy and the histopathology indicated widespread infiltration of lymphocytes into organs, such as brain, liver, kidney and lung, in addition to the lymphoid tissues and spleen; the appearance was consistent with diffuse histiocytic lymphoma. Direct immunofluorescence assays with specific typing anti-sera on live cells prepared from spleen and various lymph nodes in short-term culture, suggested the cells were B-cells. This B-cell lymphoma provides an experimental model, not only for studying possible oncogene activation but also for studying the various interactions involved in signal transduction essential for the activation of B-cell proliferation and differentiation.

Several related human transforming DNA sequences, hhc, and a putative normal liver homologue, c-hhc, have been molecularly cloned from the genomic DNAs of individual African and Asian hepatomas and from normal liver respectively. hhcM (Mahlavu) and hhcK3 (Korean), but not c-hhc, transformed NIH3T3 cells in DNA-mediated gene transfer assays. Transformed cells were found tumorigenic in athymic NIH Swiss nu/nu mice. In view of recent epidemiological studies implicating hepatitis B virus (HBV) infection early in life as causative for the eventual development of primary hepatocellular carcinoma in humans in Southeast Asia, the Far-East, and certain areas of Africa, we hereby analyzed the relationship between these hhcs and HBV in a survey of 20 hepatomas for DNA sequences homologous to hhcM and HBV by sequential hybridizations against [32p]hhcM and [32p]HBV probes. hhcM related DNA sequence were found highly amplified in 80% of the 20 hepatomas but HBV DNA sequence was rare or low. hhcM lends itself as a marker for human hepatomas. However, overall results indicated that patients with integrated HBV DNA sequences showed high copy number of hhcM sequence. Furthermore, EcoR1-restricted hepatoma DNAs showed that HBV and hhcM DNA sequences resided at different fragments in hepatomas. Our results suggest that HBV contributes to hepatocarcinogenesis probably via an activation mechanism involving possibly an integration or transient interaction of HBV DNA with hepatocyte DNA sequences, leading to recombination and eventual amplifications of the hhcM sequence in Mahlavu.

The nascent DNA transcript of intracisternal A particle RNA-directed DNA polymerase appeared to be covalently linked to an RNA primer. Fidelity of transcription is evident since the DNA transcript hybridized specifically back to 35-70 S RNA of intracisternal A particles but not with heterologous RNAs. This DNA transcript has an approximate molecular weight of 145 000, estimating 350 polynucleotides and base ratios with an excess of dGMP.