Therapeutic Targeting of P53: A Comparative Analysis of APR-246 and COTI-2 in Human Tumor Primary Culture 3-D Explants

Abstract

Background:TP53 is the most commonly mutated gene in human cancer with loss of function mutations largely concentrated in "hotspots" affecting DNA binding. APR-246 and COTI-2 are small molecules under investigation in P53 mutated cancers. APR binds to P53 cysteine residues, altering conformation, while COTI-2 showed activity in P53 mutant tumors by a computational platform. We compared APR-246 and COTI-2 activity in human tumor explants from 247 surgical specimens. Methods: Ex vivo analyses of programmed cell death measured drug-induced cell death by delayed-loss-of-membrane integrity and ATP content. The LC50s were compared by Z-Score. Synergy was conducted by the method of Chou and Talalay, and correlations were performed by Pearson moment. Results: APR-246 and COTI-2 activity favored hematologic neoplasms, but solid tumor activity varied by diagnosis. COTI-2 and APR-246 activity did not correlate (R = 0.1028) (NS). COTI-2 activity correlated with nitrogen mustard, cisplatin and gemcitabine, doxorubicin and selumetinib, with a trend for APR-246 with doxorubicin. For ovarian cancer, COTI-2 showed synergy with cisplatin at 25%. Conclusions: COTI-2 and APR-246 activity differ by diagnosis. A lack of correlation supports distinct modes of action. Cisplatin synergy is consistent with P53's role in DNA damage. Different mechanisms of action may underlie disease specificity and offer better disease targeting.

Keywords: APR-246; COTI-2; TP53; anti-cancer drug.

Figure 1

Lethal concentration (LC50) values were compared using Z-scores calculated by the formula Z = X − U/S, where X = Mean value, U = Sample value and S = Standard deviation. Results falling to the left of mean are more sensitive. Results falling to the right of mean are more resistant: Heme = Hematologic tumors; Adr = Adrenocortical; Colr = Colorectal; Panc = Pancreatic; Sar = Sarcoma; Ova = Ovarian; NSCLC = Non-small cell lung cancer; Gas = Gastric; Bre = Breast.

Figure 2

Lethal concentration (LC50) values were compared using Z-scores calculated by the formula Z = X − U/S, where X = Mean value, U = Sample value and S = Standard deviation. Results falling to the left of mean are more sensitive. Results falling to the right of mean are more resistant: Heme = Hematologic tumors; Endo = Endometrial; Adr = Adrenocortical; Colr = Colorectal; Panc = Pancreatic; SAR = Sarcoma; Ova = Ovarian; NSCLC = Non-small cell lung cancer; Gas = Gastric; Bre = Breast; Ren = Renal; Glio = Glioblastoma.

Figure 3

Z-Score analysis by P53 mutational status compares APR and COTI activity against wild-type versus mutated and amorphic (low or no functional protein) versus hypomorphic (altered protein structure) P53 subtypes. Results falling to the left of mean are more sensitive. Results falling to the right of mean are more resistant.

Figure 4

(a,b)—provide representative dose–response curves for APR-246 and COTI-2. Following 72 h drug exposure, tumor cell percent viability (0–100) measured against saline controls are provided in 5-point serial dilutions. Drug concentrations are provided as mMolar. Lethal concentration 50% (LC50) are interpolated using best line fit analyses.