Recently, it has been proposed to employ three-dimensional cultivation of cell in spheroids that contributes more proper physiological conditions in matrix extracellular space architecture and cell morphology accompanied by appropriate intracellular biomechanical organization of cytoskeleton and energy generation apparatus adaptation. In this research we presented the data of proteome survey for cells growing in monolayer and in spheroids that were induced in myogenic way of differentiation and reached terminal stage in seven days. Semi-quantitative profiling of the dynamic changes in proteins abundance depicted strict advantage of spheroid manner cultivation in contrast to monolayer. Cells cultured in spheroid were significantly enriched in proteins of mitochondria biogenesis, respiratory chain proteins, extracellular proteins and cytoskeleton. Most of signal transducers involved in active proliferation and early stages of differentiation were inhibited in spheroid cultures unlike monolayer. The obtained data were broadly consistent with known molecular mechanisms of myogenesis in transcripts level and reflected strong advantage of spheroid cultivation conditions.