{'fraction_non_alphanumeric': 0.01796023091725465, 'fraction_numerical': 0.006414368184733804, 'mean_word_length': 5.190476190476191, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 1, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Development of an LC‐MS/MS based biochemical assay to evaluate Isoprenylcysteine carboxyl methyltransferase (ICMT) as a cancer target

We developed a highly sensitive LC‐MS/MS based biochemical assay to quantitate the enzymatic activity of ICMT. The activity of endogenous ICMT was first determined in two kras dependent cancer cell lines and then following treatment with various siRNA constructs. The degree of knockdown was quantiated by comparison of siRNA treated cells with control treated cells. Enzyme activity loss correlated well with loss of messenger RNA. Time course experiments were also performed to evaluate the turnover time for endogenous ICMT in these cell lines. These data together with anchorage‐independent proliferation experiments allowed us to conclude that 80–90% loss of ICMT activity was insufficient to slow the growth of 10 different disease relevant cancer cell lines. Furthermore, ICMT knockout cell lines were engineered to evaluate whether 10–15% remaining ICMT activity was sufficient to enable cell growth. The LC‐MS/MS assay was unable to detect ICMT enzymatic activity in these cells. The lack of cell growth inhibition in these cells suggests that ICMT is not critical for kras localization and/or activity in these kras mutant cell lines. The use of LC‐MS/MS as a biochemical assay product detection method enabled us to monitor enzymatic activity to quantitate loss of protein in knockdown studies and loss of function in knockout studies. Our assay enabled the evaluation of ICMT as a Ras pathway therapeutic target.