A pyrazolopyran derivative preferentially inhibits the activity of human cytosolic serine hydroxymethyltransferase and induces cell death in lung cancer cells

Abstract

Serine hydroxymethyltransferase (SHMT) is a central enzyme in the metabolic reprogramming of cancer cells, providing activated one-carbon units in the serine-glycine one-carbon metabolism. Previous studies demonstrated that the cytoplasmic isoform of SHMT (SHMT1) plays a relevant role in lung cancer. SHMT1 is overexpressed in lung cancer patients and NSCLC cell lines. Moreover, SHMT1 is required to maintain DNA integrity. Depletion in lung cancer cell lines causes cell cycle arrest and uracil accumulation and ultimately leads to apoptosis. We found that a pyrazolopyran compound, namely 2.12, preferentially inhibits SHMT1 compared to the mitochondrial counterpart SHMT2. Computational and crystallographic approaches suggest binding at the active site of SHMT1 and a competitive inhibition mechanism. A radio isotopic activity assay shows that inhibition of SHMT by 2.12 also occurs in living cells. Moreover, administration of 2.12 in A549 and H1299 lung cancer cell lines causes apoptosis at LD50 34 μM and rescue experiments underlined selectivity towards SHMT1. These data not only further highlight the relevance of the cytoplasmic isoform SHMT1 in lung cancer but, more importantly, demonstrate that, at least in vitro, it is possible to find selective inhibitors against one specific isoform of SHMT, a key target in metabolic reprogramming of many cancer types.

The catalytic activity of serine hydroxymethyltransferase is essential for de novo nuclear dTMP synthesis in lung cancer cells

“…The primers have been previously reported [4]. About 1 lg of RNA was used for the reverse transcription reaction by using a SuperScript First-Strand Synthesis System Kit (Invitrogen).…”

“…The subunit concentration of holoenzyme forms was calculated according to a molar absorptivity value of e 280 = 47 565 cm À1 ÁM À1 [14]. To determine the K m for L-serine, H 4 PteGlu was maintained at 52 lM while L-serine concentration was varied between 0.15 and 20 mM. L-serine concentration was held at 10 mM and H 4 PteGlu varied in the 3-200 mM range to determine the K m for H 4 PteGlu.…”

“…Tumour cells recycle one-carbon units from different amino acids within the folate and methionine cycles to produce the building blocks needed to sustain high proliferation rates [1][2][3]. Given its pivotal role in OCM, it is not surprising that SHMT is considered a potential drug target for cancer therapy [4][5][6][7]. Serine fuels the folate cycle through the reaction catalysed by serine hydroxymethyltransferase (SHMT) that reversibly interconverts serine and tetrahydrofolate (THF) to glycine and 5,10-methylene-THF (Fig.…”

“…1). Given its pivotal role in OCM, it is not surprising that SHMT is considered a potential drug target for cancer therapy [4][5][6][7]. SHMT is a pyridoxal-5 0 -phosphate (PLP)dependent enzyme that exists in two main isoforms, which localize in the cytosol (SHMT1) and in the mitochondria (SHMT2).…”

Potent Inhibitors of Plasmodial Serine Hydroxymethyltransferase (SHMT) Featuring a Spirocyclic Scaffold

“…Human cytosolic SHMT1 and mitochondrial SHMT2 are of emerging interest as potential anticancer targets . We therefore tested the potent spiro‐dihydroindene analogues against h SHMT1 (Table ).…”