Table 1.

Analysis of the basis of PALA resistance in human cell lines

Cell line	Karyotype	Rearranged chromosome 2	Selective concentration of PALA, μM*	Rate of PALA resistance, ×10−5	Colonies examined by FISH, n	Colonies with the specified mechanism of PALA resistance, %†
						Chromosomal amplification	Isochromosomes 2p or 2p arms	More copies of chromosome 2	No increase in CAD copies
HT-1080	pseudodiploid;	no	20–60 (rate)	5	23	4	39	0	57
	few rearrangements		40 (frequency)		15	7	27	0	67
MDAHO41	hypertetraploid	no	170–190 (rate)	16	13	0	15	0	85
			170 (frequency)		14	7	14	0	79
LIM-1215	pseudodiploid; few rearrangements	no	50–60 (rate)	7	14	0	7‡	100‡	0
Saos-2	hypotriploid; >60% of chromosomes rearranged	yes	130–170 (rate)	140§	11	100	0	0	0
U-2 OS	hypertriploid; >70% of chromosomes rearranged	yes	400 (rate)	1.4¶	15	33	0	ND	ND

Before selection with PALA, 500-cell aliquots were grown to ≈10⁵ cells each and dispersed into 10-cm dishes. Selections were performed using PALA concentrations near 3 × ID₅₀ determined separately for each cell line. Independent PALA-resistant colonies, isolated from separate plates, were grown to ≈5 × 10⁵ cells in PALA before analysis. ND, not determined. 

^*The colonies examined were obtained in either rate or frequency selections, as noted. 

^†The predominant mechanism is shown. Clones often contained a small number of cells that displayed a different mechanism. 

^‡One clone displayed both an isochromosome and an additional copy of chromosome 2. 

^§We had to select Saos-2 cells with a high concentration of PALA (8–10 × ID₅₀). There were too many colonies at 3 × ID₅₀ because the parental cell line included many cells that have already amplified the CAD gene. FISH analysis revealed unselected cells with one or two extra copies of CAD on a rearranged marker chromosome, similar to the pattern in PALA-resistant clones. 

^¶The selective concentration of PALA was ≈6 × ID₅₀ in this case.