Oncogenes, malignant transformation, and modern medicine.

During the past decade there have been remarkable strides in the understanding of the basic mechanism of cancer. It is now clear that there is a set of genes, known as oncogenes, that can cause cells to become malignant if their expression is altered, either by mutation or overexpression. The products of these genes include growth factors, growth factor receptors, signal tranduction proteins, and DNA binding proteins. The normal cellular counterparts of these genes play very important roles in the regulation of growth and proliferation by normal cells. Another set of genes, anti-oncogenes, also play an important role in preventing abnormal cell proliferation. The remarkable explosion of understanding of the pathophysiology of malignancy has led to a common unifying concept of malignant transformation that applies to all tumors. It is likely that these new insights will lead to improved and more specific treatments for malignant disease in the next decade.

Mithramycin selectively inhibits transcription of G-C containing DNA.

Mithramycin induces a reversible inhibition of cellular RNA synthesis without affecting DNA synthesis. The authors have shown this drug induces myeloid differentiation of HL-60 promyelocytic leukemia cells and is an effective agent in certain patients with chronic granulocytic leukemia. In order to investigate the mechanism by which this drug inhibits RNA synthesis we have compared the effect of mithramycin on RNA synthesis by whole cells, isolated nuclei, and RNA synthesis by isolated E. coli RNA polymerase and eukaryotic RNA polymerase II. Exposure of HL-60 cells to mithramycin at concentrations of 4.6 X 10(-7) m or higher for 48 hours causes an almost immediate inhibition of RNA synthesis (up to 85% at 4 hours) with only modest cytotoxicity at these concentrations. Endogenous RNA synthesis by isolated nuclei can be inhibited by mithramycin only at high concentrations (greater than 10(-5) m), suggesting that mithramycin primarily may inhibit initiation, rather than elongation. Mithramycin inhibits in vitro transcription of salmon sperm DNA by E. coli RNA polymerase at DNA:drug ratios similar to those required for RNA synthesis inhibition in whole cells. Similar DNA binding studies with synthetic oligonucleotides demonstrate that mithramycin is a potent inhibitor of transcription of Poly dG.dC by E. coli RNA polymerase but has no effect on transcription of Poly dA.dT. The rapid inhibition of whole cell and isolated RNA polymerase transcription, and the relative insensitivity of isolated nuclei, suggest mithramycin may interact with specific DNA sequences in order to inhibit the initiation of RNA synthesis in intact cells.

In vivo differentiation of blast-phase chronic granulocytic leukemia. Expression of c-myc and c-abl protooncogenes.

A patient with chronic granulocytic leukemia in acute blastic transformation was treated with mithramycin, an RNA synthesis inhibitor, after in vitro exposure of her leukemic cells to mithramycin showed differentiation to normal appearing granulocytes. Mithramycin therapy in vivo resulted in a prompt and dramatic hematologic response. Before therapy, the patient's leukemic cells had high levels of transcription of the cellular myc and abl protooncogenes. After initiation of therapy, protooncogene mRNA decreased rapidly. These observations indicate that mithramycin can induce differentiation both in vitro and in vivo and suggest that such changes may be associated with altered oncogene expression.

Control of lysozyme gene expression in differentiating HL-60 cells.

We have investigated the control of lysozyme gene expression in HL-60 cells induced to differentiate into macrophage-like cells with phorbol myristate acetate (PMA). Differentiation, as evidenced by cellular adherence, and morphological changes corresponded temporally to an increase in nonspecific esterase activity. The lysozyme concentration in the medium of uninduced HL-60 cells was 10 micrograms/10(7) cells, increasing to a maximum of 46 micrograms/10(7) cells after 48 h incubation with PMA (16 nM). At 72 h the lysozyme concentration decreased to 16 micrograms/10(7) cells. Intracellular lysozyme activity remained constant throughout differentiation. If HL-60 cells were exposed to PMA for 24 h, washed, then maintained in normal medium, they differentiated normally, confirming their irreversible commitment to differentiate. The increase in lysozyme secretion by these cells, however, is markedly blunted suggesting that continued PMA treatment of differentiated cells is required for their secretion of lysozyme. There is no change in the rate of extracellular degradation of lysozyme during differentiation. The level of lysozyme mRNA does not correlate directly with the amount of lysozyme secreted into the medium. Hybridization of uninduced HL-60 cell RNA with a chicken lysozyme cDNA probe demonstrates moderate hybridization. There is a modest (five-fold) increase in lysozyme mRNA between 0 and 36 h of exposure to PMA, corresponding to the burst of lysozyme secretion by these cells. The lysozyme mRNA decreases to a level which is lower than the original baseline by 72 h, when the cells are still secreting substantial amounts of lysozyme. These data suggest that both transcriptional and post-transcriptional controls are operative in the control of lysozyme gene expression during the differentiation of HL-60 cells. They also imply that lysozyme secretion is not a necessary component in the macrophage-monocyte differentiation of these cells.

Pulmonary stenosis in infants and young children.

Twenty-four patients less than 3 years old underwent operation for pulmonary stenosis. Pulmonary dysplasia was diagnosed preoperatively in only 4 patients; in 20 patients the lesion was categorized simply as pulmonary stenosis. At operation, more severe valve deformities were often present in patients less than 2 years of age. Preoperative evaluation did not reveal the extent of the deformity in 7 additional patients. The deformities included not only valvular dysplasia (thickened redundant valve cusps) but also supravalvular and annular abnormalities. Relief of obstruction was obtained only when all components of the obstructive abnormality were relieved. Patch angioplasty of the right ventricular outflow tract was necessary in 13 patients with complex morphology. Valvotomy was effective only for pulmonary stenosis due to pure commissural fusion. A spectrum of the morphology of pulmonary stenosis is recognized, with more complex lesions than simple commissural fusion identified in younger children. The more complex lesions may require more extensive operations (outflow tract patch) to completely relieve the obstructive pathological condition in the outflow tract.

Supravalvular aortic stenosis. Repair by extended aortoplasty.

Enlargement of the aorta with a diamond-shaped patch of the noncoronary sinus of Valsalva may not be sufficient in severe cases of supravalvular aortic stenosis. This traditional reconstruction is asymmetric, and, if the fibrous supravalvular ring is thick and rigid, the aorta may not open wide with patch angioplasty, so that aortic obstruction may remain. Also, because aortic valve function may not be perfect after asymmetric reconstruction, there may be aortic valve incompetence or obstruction of coronary ostia by the valve cusps. A new reconstructive operation was designed and used in eight patients. All survived and are asymptomatic. The aortoplasty was extended so that the supravalvular ring was incised at two points in the noncoronary and in the right coronary sinuses of Valsalva. The area of stenosis was opened wide, and the cusps of the aortic valve were lengthened, which provided better approximation and function. A tubular Dacron prosthesis, tailored to reconstruct the aorta, provided a wide aortic cross-sectional area. This technique of extended aortoplasty for symmetric reconstruction of the aorta should provide more predictable relief of aortic obstruction and improved function of the aortic valve.