By combining in-cell stable isotope incorporation assay with quantitative metabolomics we show: cellular uptake of external purines and their internal generation are equivalent; an upregulation in lysosome biogenesis that functions in response to purine deficiency caused by methotrexate (MTX) and lometrexol (LTX) treatment. This leads to increased RNA digestion as visualized by a newly developed intracellular RNA-FRET oligo assay. Interestingly, downregulation of lysosomal RNase activity through knockdown of RNAseT2 increased RNA accumulation and a compensating increase in DNPB.

These findings establish purine metabolic enzyme inhibition as a viable therapeutic strategy to induce differentiation and attenuate tumor progression in high-risk MYCN-amplified NBs.

Furthermore, a GART inhibitor, pemetrexed (PEM), effectively suppressed cell proliferation and tumor growth in a human cell line-derived xenograft model (CDX). In conclusion, our findings demonstrate that GART promotes MM cell proliferation and tumor stemness by activating the HSP90α/CDK6/β-catenin axis. Targeting GART may be a promising strategy for developing and improving MM treatments.

Our findings suggest that HB patients harboring activated β-catenin mutations and consequent DNPS upregulation, may be treated efficaciously with DNPS enzyme inhibitors like lometrexol. These novel findings bear major therapeutic implications for targeted precision medicine of HB.

In addition, UCK2-Ser254 phosphorylation level displays a positive relationship with GART-Ser440 phosphorylation level and its enhancement is correlated with poor prognosis of human hepatocellular carcinoma (HCC) patients. These findings reveal a non-canonical role of GART in regulating pyrimidine salvage synthesis under nutrient stress, and raise the potential for alternative treatments in targeting pyrimidine salvage-dependent tumor growth.

Similar scaffold commercial drugs leucal (LEU), epirubicin (EPI), and lometrexol also displayed strong binding to the active site of MTHFD2...The interaction of breast cancer serum with high expression of MTHFD2 also showed an increase in binding of epirubicin in the presence of leucovorin...Further experimental studies are required to establish the potential mode of inhibition of the novel small ligands. Future in vivo trials may validate the effectiveness of the combinatorial therapy.

Pemetrexed (PEM), a compound targeting GART, combined with other chemotherapy drugs greatly suppresses tumor growth both in a PDX model and in CRC patients. The present study demonstrates a novel methyltransferase function of GART and the role of the GART/RUVBL1/β-catenin signaling axis in promoting CRC stemness. PEM may be a promising therapeutic agent for the treatment of CRC.

Finally, we demonstrated that tumors with MLL3 and/or MLL4 mutations were highly sensitive to lometrexol in vivo, both in culture and in animal models of cancer. Our results depicted a targetable metabolic dependency arising from epigenetic factor deficiency, providing molecular insight to inform therapy for cancers with epigenetic alterations secondary to MLL3/4 COMPASS dysfunction.

Moreover, the GART inhibitor lometrexol (LMX) and drugs approved for clinical treatment of primary and metastatic BC (4OH-tamoxifen and the CDK4/CDK6 inhibitors) exert synergic antiproliferative effects in BC cells. In conclusion, GART inhibition by LMX or other inhibitors of the de novo purine biosynthetic pathway could be a novel effective strategy for the treatment of primary and metastatic BCs.

The role of some of these genes was previously reported in the resistance to chemotherapy in other diseases. This can be used as clues to detect treatment-resistant (drug-resistant) cases in the early stages of diseases.

Using the cMap, candidate drugs screened with potential efficacy for ICC included three tyrosine kinase inhibitors (dasatinib, NVP-BHG712, tivantinib), two cannabinoid receptor agonists (palmitoylethanolamide, arachidonamide), two antibiotics (moxifloxacin, amoxicillin), one estrogen receptor agonist (levonorgestrel), one serine/threonine protein kinase inhibitor (MK-2206) and other small molecules. Key genes from network and PPI analysis allowed us to identify potential drugs for ICC. The identification of novel gene expression profiles and related drug screening may accelerate the identification of potential novel drug therapies for ICC.