DISCOVERY OF A SERIES OF SMALL MOLECULE SHMT1/2 INHIBITORS
• Synthetized compounds exert potency in SHMT1/2 biochemical assay as well as in
cellular assay (measured by the C13 serine to glycine conversion)
• Tested compounds inhibit cancer cell growth
• Reduced viability could be rescued with formate
SHMT1: SEL302-01612
SHMT2: SEL302-01612
SHMT1: SEL302-00332
SHMT2: SEL302-00332
SHMT1: SEL302-00621
SHMT2: SEL302-00621
Discovery of novel SHMT small molecule inhibitors for cancer treatment
Anna Bartosik, Paweł Guzik, Marta Sowińska, Karolina Gluza, Marcin Król, Anna Wróbel, Agnieszka Dreas, Faustyna Iwańska, Magdalena Zastawna, Urszula Kulesza, Nicolas Boutard, David Schultz,
Justyna Wujkowska Karolina Pyziak, Agnieszka Sroka-Porada, Agnieszka Przybyłowicz, Agnieszka Adamus, Magdalena Sieprawska-Lupa, Przemysław Golik,
Piotr Kowalczyk, Krzysztof Brzózka, Tomasz Rzymski, Mateusz Nowak
Selvita S.A. Krakow, Poland www.selvita.com
Glucose
3-P-GlyceratePHGDH
THF 5,10-CH2-THF 10-formyl-THF
MTHFD1
Formate
DNA pyrimidine methylation
MTHFD1
Redox Proteins
Pyruvate
Serine Glycine
THF 5,10-CH2-THF 10-formyl-THF
Serine Glycine
SHMT2
Formate
MTHFD2/2L MTHFD1L
Redox ATP
SHMT1
Mitochondria
Cytoplasm
Breast (TCGA)
Enrichment Score (ES) 0.76
Normalized Enrichment Score (NES)
2.20
Nominal p-value 0.001
Enrichment Score (ES) 0.57
Normalized Enrichment Score (NES)
1.86
Nominal p-value 0.005
CRC (TCGA) NSCLC (TCGA)
Enrichment Score (ES) 0.54
Normalized Enrichment Score (NES)
1.71
Nominal p-value 0.03
INTRODUCTION
Over-activation of the serine synthesis pathway, upregulation of SHMT2 has beendescribed in over 20% of solid tumors (e.g. breast, lung, colorectal cancers). Such cancercells are highly dependent on serine. Serine hydroxymethyltransferase (SHMT isoform 1and 2) plays a key role in a so-called one-carbon pathway, a group of biochemicalreactions involved in amino acid metabolism. SHMT catalyzes the conversion of serine toglycine and also plays a role in the folate (vitamin B9) cycle. Antagonists of folatemetabolism or antifolates are an established chemotherapy in certain cancers. Folateantagonism disrupts cell division, DNA/RNA synthesis and protein synthesis. Pemetrexed(for non-small cell lung carcinoma, mesothelioma) and methotrexate (for autoimmuneconditions like rheumatoid arthritis and certain cancers) are two well established andeffective antifolates. The main drawback with antifolates in cancer treatment is thedevelopment of resistance. In this study we report development of a series of smallmolecule SHMT1/2 inhibitors. Synthetized compounds exert potency in SHMT1/2biochemical assay as well as in cellular assay (measured by the C13 serine to glycineconversion) in a low nanomolar range. Therapeutic effect of the compounds wasinvestigated in the panel of cancer cell lines with different genetic background as well aswith different SHMT2 levels. We identified several cell lines in which tested compoundsinhibited cancer cells growth with nM GI50 values. Taken together, presented datasupports our rationale for using SHMT1/2 inhibitors as a novel and interesting promisingapproach for the cancer therapy.
SSP/1C highSSP/1C highSSP/1C high
SSP POSITIVE SOLID TUMORS ARE C-MYC POSITIVE AND
SHOW INDUCTION OF AMINOACID INTEGRATED STRESS
RESPONSE (ISR)
PSAT1
CONCLUSIONS
Gene Set Enrichment Analysis for the class of SSP/1C high solid tumors
SSP and 1C pathways in cancer
Project supported by the European Regional Development Funds under the Measure 1.1.1. Operational Program - Smart Growth (POIR.01.01.01-00-0673/15-00)
SHMT INHIBITORS DECREASE VIABILITY OF BREAST AND LUNG
CANCER CELLS BY THE INHIBITION OF 1C PATHWAY
FORMATE SENSITIZES CELL LINES DEFICIENT IN GLYCINE
TRANSPORT TO SHMT1/2 INHIBITORS
• Cancer specific SSP and 1C proteins are attractive targets for pharmacological inhibition in oncology
• SHMT1/2 inhibitors are a novel approach for cancer therapy
• Inhibition of SHMT by small molecules effectively blocks influx of 1C units form serine in cancer cells
• Anticancer effects of compounds are SHMT-dependent as revealed by rescue experiments
• Defective glycine import as well as glycine starvation could be a promising therapeutic strategy for SHMT2
inhibitors in DLBCL
References:
1. Yang and Vousden. Serine and one-carbon metabolism in cancer. Nat. Rev. Can. 2016
2. Jain et al. Metabolite profiling identifies a key role for glycine in rapid cancer cell proliferation. Science. 2012
3. Labuschagne et al. Serine, but not glycine, supports one-carbon metabolism and proliferation of cancer cells. Cell Reports. 2014
4. Newman and Maddocks. One-carbon metabolism in cancer. British Journal of Cancer. 2017.
TOP TABLE
COMPOUND SLV# SEL302-01612 SEL302-00687 SEL302-00745 SEL302-00753
IC50 [µM] SHMT2 0.02 0.03 0.05 0.04
IC50 [µM] SHMT1 0.35 tbd tbd tbd
AD
ME/
DM
PK
Metabolic Stability(mic. mouse) (1h)
0% 80.4% 80.7% 45.8%
Metabolic StabilityMouse S9 fraction (phase I and II)
0% 86.9% tbd 55.6%
hERG Channel Inhibition [uM] 17 tbd tbd >100
CEL
L A
SSA
YS
Ser-Gly Flux InhBreast cancer cell line
IC50 [uM] 0.03 0.34 tbd tbd
Viability 72hBreast cancer cell line
GI50 [uM](efficiency)
0.2 4.1 2.3 2.5 1.1 1.8
Viability 72hLung cancer cell line
GI50 [uM](efficiency)
0.3 >10 >10 9.7 5.5 6.9
Results from combination study indicate that glycine starvation (e.g. in a form of low glycine diet)
can be used to render similar dependency in cancer cell without glycine transport deficiency.
Failure of formate to rescue growth in the DLBCL cell lines is most likely due to a
requirement for both glycine and 1C units made by SHMT in these cells. In the
absence of glycine, formate can enhance the cytotoxicity of SHMT inhibition, by
driving residual SHMT enzymatic function in the glycine-consuming direction.
Human SHMT inhibitors reveal defective glycine import as a targetable metabolic vulnerability of diffuse large B-cell lymphoma. Proc NatlAcad Sci U S A. 2017 Oct 24;114(43):11404-11409.
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SEL302-00332
biochemical assay SHMT1 IC50 (µM) SHMT2 IC50 (µM)
SEL302-01612 0.01 0.02
SEL302-00332 0.22 0.01
SEL302-00621 0.08 0.16
serine flux IC50 (µM)
SEL302-01612 0.03
SEL302-00332 0.05
SEL302-00621 0.4
SEL302-01612 SEL302-00621
SEL302-01612
SEL302-00332
SEL302-00621
FULL MEDIUM 1 mM FORMATE
10X GLYCINE 10x GLYCINE + 1 mM FORMATE
SEL302-01612Breast cancer
cell line (-FA)
Breast cancer
cell line (+FA)
Lung cancer
cell line (-FA)
Lung cancer
cell line (+FA)
GI50 (µM) 0.23 > 10 0.35 > 10LC (µM) 2 > 10 3 > 10
SEL302-00332Breast cancer
cell line (-FA)
Breast cancer
cell line (+FA)
Lung cancer
cell line (-FA)
Lung cancer
cell line (+FA)
GI50 (µM) 0.9 > 10 0.6 > 10LC (µM) > 10 > 10 7 > 10
SEL302-00621Breast cancer
cell line (-FA)
Breast cancer
cell line (+FA)
Lung cancer
cell line (-FA)
Lung cancer
cell line (+FA)
GI50 (µM) 0.89 > 10 3.2 > 10LC (µM) 2.6 > 10 > 10 > 10
Combination study was performed using DLBCL cell line with defective glycine import.