11 results on '"Jade M. Katinas"'
Search Results
2. Supplementary Methods from Novel Pyrrolo[3,2-d]pyrimidine Compounds Target Mitochondrial and Cytosolic One-carbon Metabolism with Broad-spectrum Antitumor Efficacy
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Larry H. Matherly, Aleem Gangjee, Charles E. Dann, Zhanjun Hou, Jing Li, Joshua D. Rabinowitz, Lisa Polin, Maik Hüttemann, Seongho Kim, Juiwanna Kushner, Kathryn White, Sijana H. Dzinic, Carrie O'Connor, Adrianne Wallace-Povirk, Jenney Liu, Josephine Frühauf, Xun Bao, Changwen Ning, Arpit Doshi, Md. Junayed Nayeen, Jennifer Wong-Roushar, Jade M. Katinas, Gregory S. Ducker, Khushbu Shah, and Aamod S. Dekhne
- Abstract
Supplementary Method Descriptions Molecular modeling and computational studies Enzyme expression and purification. In vitro enzymatic assays and Ki determinations In vivo efficacy trials with MIA PaCa-2 pancreatic cancer xenografts. Cytochrome c oxidase assay Synthesis of AGF94 and 5-substituted pyrrolo[3,2-d]pyrimidine compounds
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- 2023
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3. Data from Novel Pyrrolo[3,2-d]pyrimidine Compounds Target Mitochondrial and Cytosolic One-carbon Metabolism with Broad-spectrum Antitumor Efficacy
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Larry H. Matherly, Aleem Gangjee, Charles E. Dann, Zhanjun Hou, Jing Li, Joshua D. Rabinowitz, Lisa Polin, Maik Hüttemann, Seongho Kim, Juiwanna Kushner, Kathryn White, Sijana H. Dzinic, Carrie O'Connor, Adrianne Wallace-Povirk, Jenney Liu, Josephine Frühauf, Xun Bao, Changwen Ning, Arpit Doshi, Md. Junayed Nayeen, Jennifer Wong-Roushar, Jade M. Katinas, Gregory S. Ducker, Khushbu Shah, and Aamod S. Dekhne
- Abstract
Folate-dependent one-carbon (C1) metabolism is compartmentalized into the mitochondria and cytosol and supports cell growth through nucleotide and amino acid biosynthesis. Mitochondrial C1 metabolism, including serine hydroxymethyltransferase (SHMT) 2, provides glycine, NAD(P)H, ATP, and C1 units for cytosolic biosynthetic reactions, and is implicated in the oncogenic phenotype across a wide range of cancers. Whereas multitargeted inhibitors of cytosolic C1 metabolism, such as pemetrexed, are used clinically, there are currently no anticancer drugs that specifically target mitochondrial C1 metabolism. We used molecular modeling to design novel small-molecule pyrrolo[3,2-d]pyrimidine inhibitors targeting mitochondrial C1 metabolism at SHMT2. In vitro antitumor efficacy was established with the lead compounds (AGF291, AGF320, AGF347) toward lung, colon, and pancreatic cancer cells. Intracellular targets were identified by metabolic rescue with glycine and nucleosides, and by targeted metabolomics using a stable isotope tracer, with confirmation by in vitro assays with purified enzymes. In addition to targeting SHMT2, inhibition of the cytosolic purine biosynthetic enzymes, β-glycinamide ribonucleotide formyltransferase and/or 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase, and SHMT1 was also established. AGF347 generated significant in vivo antitumor efficacy with potential for complete responses against both early-stage and upstage MIA PaCa-2 pancreatic tumor xenografts, providing compelling proof-of-concept for therapeutic targeting of SHMT2 and cytosolic C1 enzymes by this series. Our results establish structure–activity relationships and identify exciting new drug prototypes for further development as multitargeted antitumor agents.
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- 2023
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4. Supplementary Data from Novel Pyrrolo[3,2-d]pyrimidine Compounds Target Mitochondrial and Cytosolic One-carbon Metabolism with Broad-spectrum Antitumor Efficacy
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Larry H. Matherly, Aleem Gangjee, Charles E. Dann, Zhanjun Hou, Jing Li, Joshua D. Rabinowitz, Lisa Polin, Maik Hüttemann, Seongho Kim, Juiwanna Kushner, Kathryn White, Sijana H. Dzinic, Carrie O'Connor, Adrianne Wallace-Povirk, Jenney Liu, Josephine Frühauf, Xun Bao, Changwen Ning, Arpit Doshi, Md. Junayed Nayeen, Jennifer Wong-Roushar, Jade M. Katinas, Gregory S. Ducker, Khushbu Shah, and Aamod S. Dekhne
- Abstract
Supplementary Figure S1 shows in vitro antiproliferative activity and targeted pathways of previously reported AICARFTase and GARFTase inhibitors. Supplementary Figure S2 shows docking of AGF291, AGF320, and AGF347 (along with 5-formyl-THF) in human SHMT2 and rabbit SHMT1. Supplementary Figure S3 shows plasma membrane folate transporter expression levels in H460, HCT116, and MIA PaCa-2 human tumor cell lines as compared to those in the IGROV-1 epithelial ovarian cancer cell line. Supplementary Figure S4 shows in vitro antiproliferative activity and targeted pathways of AGF291, AGF320, and AGF347 along with previously reported GARFTase inhibitor AGF94 in H460, HCT116, and MIA PaCa-2 cell lines. Supplementary Figure S5 shows targeted metabolomics data on total serine, serine isotope labeling patterns, total GAR, total AICAR, total adenine nucleotides, and total dTTP along with isotope labeling patterns of H460, HCT116, and MIA PaCa-2 cell lines treated with AGF291, AGF320, and AGF347 not shown in main text Figure 4. Supplementary Figure S6 shows a Western blot confirming knockdown of SHMT2 in H460 SHMT2 KD cell line and knockout of SHMT2 in HCT116 SHMT2 KO cell line. Supplementary Figure S7 shows in vivo efficacies of AGF347 and gemcitabine towards MIA PaCa-2 early and late stage tumor xenograft models where mouse serum folate levels were not depleted to approximate those found in humans. Supplementary Figure S8 shows a cytochrome c oxidase assay on tumors harvested from the metabolomics arm of the late stage in vivo AGF347 trial. Table S1 shows docking scores of novel compounds in human SHMT2 and rabbit SHMT1 (corresponding to Supplementary Figure S2). Table S2 shows quantitative data from in vivo early- and late-stage trials of AGF347 and gemcitabine against MIA PaCa-2 tumor xenografts in NCR SCID mice.
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- 2023
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5. Correction: Novel Pyrrolo[3,2-d]Pyrimidine Compounds Target Mitochondrial and Cytosolic One-Carbon Metabolism with Broad-spectrum Antitumor Efficacy
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Aamod S. Dekhne, Khushbu Shah, Gregory S. Ducker, Jade M. Katinas, Jennifer Wong-Roushar, Md. Junayed Nayeen, Arpit Doshi, Changwen Ning, Xun Bao, Josephine Frühauf, Jenney Liu, Adrianne Wallace-Povirk, Carrie O'Connor, Sijana H. Dzinic, Kathryn White, Juiwanna Kushner, Seongho Kim, Maik Hüttemann, Lisa Polin, Joshua D. Rabinowitz, Jing Li, Zhanjun Hou, Charles E. Dann, Aleem Gangjee, and Larry H. Matherly
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Cancer Research ,Oncology - Published
- 2023
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6. Novel pyrrolo[3,2-d]pyrimidine compounds target mitochondrial and cytosolic one-carbon metabolism with broad-spectrum antitumor efficacy
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Lisa Polin, Khushbu Shah, Zhanjun Hou, Arpit Doshi, Josephine Frühauf, Md. Junayed Nayeen, Kathryn White, Gregory S. Ducker, Jade M. Katinas, Seongho Kim, Larry H. Matherly, Changwen Ning, Adrianne Wallace-Povirk, Jing Li, Juiwanna Kushner, Maik Hüttemann, Jennifer Wong-Roushar, Carrie O'Connor, Jenney Liu, Xun Bao, Charles E. Dann, Sijana H. Dzinic, Aleem Gangjee, Joshua D. Rabinowitz, and Aamod Dekhne
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0301 basic medicine ,Purine ,Cancer Research ,Ribonucleotide ,Antineoplastic Agents ,CHO Cells ,Mice, SCID ,Mitochondrion ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,Inhibitory Concentration 50 ,0302 clinical medicine ,Cricetulus ,Cytosol ,Cell Line, Tumor ,Cricetinae ,Animals ,Metabolomics ,Pyrroles ,Amino acid synthesis ,chemistry.chemical_classification ,Xenograft Model Antitumor Assays ,Carbon ,Biosynthetic Pathways ,Mitochondria ,030104 developmental biology ,Enzyme ,Pyrimidines ,Oncology ,chemistry ,Biochemistry ,Purines ,030220 oncology & carcinogenesis ,Serine hydroxymethyltransferase ,Female ,NAD+ kinase - Abstract
Folate-dependent one-carbon (C1) metabolism is compartmentalized into the mitochondria and cytosol and supports cell growth through nucleotide and amino acid biosynthesis. Mitochondrial C1 metabolism, including serine hydroxymethyltransferase (SHMT) 2, provides glycine, NAD(P)H, ATP, and C1 units for cytosolic biosynthetic reactions, and is implicated in the oncogenic phenotype across a wide range of cancers. Whereas multitargeted inhibitors of cytosolic C1 metabolism, such as pemetrexed, are used clinically, there are currently no anticancer drugs that specifically target mitochondrial C1 metabolism. We used molecular modeling to design novel small-molecule pyrrolo[3,2-d]pyrimidine inhibitors targeting mitochondrial C1 metabolism at SHMT2. In vitro antitumor efficacy was established with the lead compounds (AGF291, AGF320, AGF347) toward lung, colon, and pancreatic cancer cells. Intracellular targets were identified by metabolic rescue with glycine and nucleosides, and by targeted metabolomics using a stable isotope tracer, with confirmation by in vitro assays with purified enzymes. In addition to targeting SHMT2, inhibition of the cytosolic purine biosynthetic enzymes, β-glycinamide ribonucleotide formyltransferase and/or 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase, and SHMT1 was also established. AGF347 generated significant in vivo antitumor efficacy with potential for complete responses against both early-stage and upstage MIA PaCa-2 pancreatic tumor xenografts, providing compelling proof-of-concept for therapeutic targeting of SHMT2 and cytosolic C1 enzymes by this series. Our results establish structure–activity relationships and identify exciting new drug prototypes for further development as multitargeted antitumor agents.
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- 2019
7. Abstract 2348: Targeting mitochondrial and cytosolic one-carbon metabolism in epithelial ovarian cancer via folate receptor alpha
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Jennifer Wong-Roushar, Aamod Dekhne, Charles E. Dann, Lisa Polin, Xun Bao, Seongho Kim, Jade M. Katinas, Adrianne Wallace-Povirk, Jose Cardiel Nunez, Md. Junayed Nayeen, Khushbu Shah, Zhanjun Hou, Aleem Gangjee, Carrie O'Connor, Jing Li, and Larry Matherly
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Folate Receptor Alpha ,Cancer Research ,One-carbon metabolism ,Cytosol ,Oncology ,Chemistry ,Cancer research ,Epithelial ovarian cancer - Abstract
Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. Though most patients initially respond to platinum-based therapy, the likelihood of disease reoccurrence is nearly 100%. Thus, new tumor-selective therapies for EOC are urgently needed. One such treatment option involves targeting tumors via folate receptor α (FRα), which is overexpressed in up to 90% of EOCs and shows increasing expression with higher stage and grade of disease. Our laboratory discovered novel 5-substituted pyrrolo[3,2-d]pyrimidine analogs (AGF347, AGF359, AGF362 and AGF363) that inhibit mitochondrial one-carbon (C1) metabolism at serine hydroxymethyltransferase (SHMT) 2, with secondary inhibition at cytosolic enzyme targets including those in de novo purine biosynthesis. Potent inhibition was seen with several FRα-expressing EOC tumor cells. Inhibitory potencies were in order, AGF347 > AGF359 > AGF362 > AGF363. Drug effects were substantially reduced with excess folic acid (FA), confirming FRα-mediated drug uptake. Toward cisplatin resistant SKOV3, TOV112D and A2780 EOC cells, inhibition in the nanomolar range was detected with all compounds. Targeted metabolomics, using L-[2,3,3-2H]serine as a tracer in wild-type or SHMT2 knockdown SKOV3 cells, confirmed all compounds inhibited cytosolic and mitochondrial C1-metabolism (at SHMT2). Apoptosis was detected for all compounds by Annexin V/PI, with partial rescue of apoptosis observed upon addition of glutathione (GSH). Glutathione pools (GSH and total GSH+GSSG) were significantly perturbed by drug treatment with all inhibitors in SKOV3 cells, comparable to GSH levels observed in SHMT2 KD cells. In vivo efficacy studies with SKOV3 xenografts treated with either AGF347 or cisplatin in SCID mice showed cisplatin resistance, while AGF347 demonstrated efficacy and delay in disease progression with a median tumor growth delay of 10 days, with the longest delay being 15 days. Our studies describe a series of novel inhibitors targeting mitochondrial and cytosolic C1-metabolism, selectively delivered via FRα, which show direct cytotoxic effects against cisplatin resistant EOC in vitro and in vivo, and display additional mechanisms of cytotoxicity mediated through glycine depletion. Citation Format: Adrianne C. Wallace-Povirk, Carrie O'Connor, Xun Bao, Jade Katinas, Jennifer Wong-Roushar, Aamod Dekhne, Zhanjun Hou, Md. Junayed Nayeen, Khushbu Shah, Jose Cardiel Nunez, Jing Li, Seongho Kim, Lisa Polin, Charles E. Dann, Aleem Gangjee, Larry H. Matherly. Targeting mitochondrial and cytosolic one-carbon metabolism in epithelial ovarian cancer via folate receptor alpha [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2348.
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- 2021
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8. Inhibitory Effects of Unique Sulfonamides on Leishmania tarentolae and Potential Pathway of Inhibition
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Jade M. Katinas
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biology ,Chemistry ,Sulfonamide (medicine) ,Leishmania tarentolae ,medicine ,Leishmaniasis ,Inhibitory postsynaptic potential ,Leishmania ,biology.organism_classification ,medicine.disease ,Microbiology ,medicine.drug - Published
- 2018
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9. Abstract 2992: Cellular pharmacodynamics of mitochondrial one-carbon metabolism-targeting 5-substituted pyrrolo[3,2-d]pyrimidine antifolates
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Aamod Dekhne, Khushbu Shah, Gregory S. Ducker, Md. Junayed Nayeen, Jade M. Katinas, Jennifer Wong, Arpit Doshi, Xun Bao, Hasini Kalpage, Jenney Liu, Seongho Kim, Adrianne Wallace-Povirk, Changwen Ning, Carrie O'Connor, Zhanjun Hou, Lisa Polin, Jing Li, Maik Hüttemann, Joshua D. Rabinowitz, Charles E. Dann, Aleem Gangjee, and Larry Matherly
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Cancer Research ,Oncology - Abstract
Folate-dependent one-carbon metabolism (1CM) is compartmentalized in the mitochondria and cytosol and generates a number of metabolites critical to tumor propagation. Folates are taken up by the plasma membrane facilitative transporters, reduced folate carrier (RFC; major tissue transporter) and proton-coupled folate transporter (PCFT; narrow physiological niche, but commonly expressed in solid tumors), and then transported into mitochondria via the mitochondrial folate transporter (MFT; SLC25A32). Although drug-targeting of cytosolic 1CM remains a clinical mainstay for a variety of cancers, development of clinically-useful agents targeting mitochondrial 1CM remains elusive. Of particular pharmacological interest is the mitochondrial 1CM enzyme, serine hydroxymethyltransferase2 (SHMT2). SHMT2 expression correlates with the oncogenic phenotype in a host of different cancers and, overall, SHMT2 is the fifth-most differentially expressed metabolic enzyme in cancer versus normal tissues. Despite its unequivocal oncogenic importance and therapeutic potential, there are no clinically relevant inhibitors of SHMT2. In this study, we characterized cellular pharmacodynamics of novel 5-substituted pyrrolo[3,2-d]pyrimidine antifolates (AGF291, AGF320, and AGF347) which show in vitroantitumor efficacy toward H460 lung, HCT-116 colon, and MIA PaCa-2 pancreatic cancer cells. Inhibition of mitochondrial SHMT2 and cytosolic 1CM at the purine nucleotide biosynthesis enzymes glycinamide ribonucleotide formyltransferase (GARFTase) and/or 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase (AICARFTase) by this series was established by in vitro targeted metabolomics in H460, HCT-116, and MIA PaCa-2 cells and in vitro cell-free assays with purified enzymes. By depleting SHMT2-derived formate, these compounds potentiate their own direct inhibition of GARFTase and AICARFTase. Depletion of adenine nucleotide pools in vitro by all compounds led to inhibition of mTOR signaling to S6K1 in HCT116 cells. Subcellular fractionation of MIA PaCa-2 and MFT-null and human MFT-transfected glyB Chinese hamster ovary cells confirmed synthesis of polyglutamyl forms of AGF347 in both cytosol and mitochondria with mitochondrial uptake of AGF347 in part mediated by MFT. Treatment by all compounds decreased the cellular GSH/GSSG ratio, indicating depleted ability to combat oxidative stress. In vivo, AGF347 demonstrated potent antitumor efficacy against MIA PaCa-2 xenografts in SCID mice (n=5) with median tumor growth delay (T-C) in 4 mice >38 days and 1 of 5 tumor-free survivors (cured). In vivo metabolomics on tumor xenografts confirmed inhibition of serine catabolism and purine biosynthesis. Collectively, our studies establish the exceptional therapeutic potential of inhibitors dual-targeting mitochondrial and cytosolic 1CM. Citation Format: Aamod Dekhne, Khushbu Shah, Gregory S. Ducker, Md. Junayed Nayeen, Jade M. Katinas, Jennifer Wong, Arpit Doshi, Xun Bao, Hasini Kalpage, Jenney Liu, Seongho Kim, Adrianne Wallace-Povirk, Changwen Ning, Carrie O'Connor, Zhanjun Hou, Lisa Polin, Jing Li, Maik Hüttemann, Joshua D. Rabinowitz, Charles E. Dann, Aleem Gangjee, Larry Matherly. Cellular pharmacodynamics of mitochondrial one-carbon metabolism-targeting 5-substituted pyrrolo[3,2-d]pyrimidine antifolates [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2992.
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- 2019
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10. Abstract 811: Multi-targeted inhibitors of mitochondrial one-carbon metabolism and cytosolic de novo purine synthesis enzymes as anti-tumor agents
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Aleem Gangjee, Junayed Nayeen, Aamod Dekhne, Changwon Ning, Zhanjun Hou, Larry H. Matherly, Carrie O'Connor, Khushbu Shah, Jennifer Wong, Charles E. Dann, and Jade M. Katinas
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De novo synthesis ,chemistry.chemical_classification ,Cancer Research ,Pyrimidine analogue ,Ribonucleotide ,Oncology ,Chemistry ,Cell growth ,Serine hydroxymethyltransferase ,Glycine ,Cancer cell ,Nucleotide ,Molecular biology - Abstract
Targeting cellular one-carbon (1C) metabolism in cancer cells with pyrrolo[3,2-d]pyrimidine analogs provides in vivo antitumor efficacy. We discovered a first-in-class series of 5-substituted pyrrolo[3,2-d]pyrimidine analogs (AGF291, AGF320, AGF347) with inhibition of serine hydroxymethyltransferase 2 (SHMT2), a key enzyme in mitochondrial 1C metabolism and a reported oncodriver. In addition, AGF291, AGF320 and AGF347 inhibited de novo purine nucleotide biosynthesis at glycinamide ribonucleotide formyltransferase (GARFTase) and 5-aminoimidazole-4-carboxamide (AICA) ribonucleotide formyltransferase (AICARFTase), and serine hydroxymethyltransferase 1 (SHMT1). In vivo studies demonstrated excellent antitumor efficacy for AGF347 against MIA PaCa-2 pancreatic adenocarcinoma xenografts in SCID mice with a median tumor growth delay (T-C) of >38 days for 4 mice, and 1 of 5 mice tumor-free 122 days post-treatment. AGF359, a new analog in this series, was a potent inhibitor of KB human tumor cell proliferation in vitro. AGF359 inhibition of KB human tumor cells was reversed with glycine and adenosine, establishing mitochondrial 1C metabolism and de novo purine biosynthesis as the targeted pathways; AICA plus glycine was incompletely protective, implicating AICARFTase as a direct cellular target. AGF359, like AGF291, AGF320, and AGF347, inhibited purified human SHMT2 (Ki = 0.399 ± 0.174 µM), SHMT1 (Ki = 0.70 ± 0.088 µM) and AICARFTase (Ki = 8.86 ± 2.83 µM). There was no inhibition of 5,10-methylene tetrahydrofolate dehydrogenase 2 (MTHFD2). Further structural modifications of these multi-targeted agents afforded pyrrolo[3,2-d]pyrimidine analogs AGF307 and AGF312. Cell-based glycine/nucleotide rescue experiments in KB tumor cells established that, as with AGF359, AGF307 and AGF312 were dual inhibitors of the mitochondrial 1C metabolism (AGF307) and de novo purine biosynthesis (AGF307, AGF312). These compounds are currently in further preclinical evaluation as a prelude to possible clinical development as antitumor agents. Citation Format: Md Junayed Nayeen, Khushbu Shah, Aamod Dekhne, Changwon Ning, Carrie O'Connor, Jade M. Katinas, Jennifer Wong, Zhanjun Hou, Charles E. Dann III, Larry H. Matherly, Aleem Gangjee. Multi-targeted inhibitors of mitochondrial one-carbon metabolism and cytosolic de novo purine synthesis enzymes as anti-tumor agents [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 811.
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- 2019
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11. Abstract 4794: Targeting mitochondrial and cytosolic one carbon metabolism of pancreatic adenocarcinoma via the proton-coupled folate transporter with novel 5-substituted pyrrolo[3,2-d]pyrimidine analogs
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Jennifer Wong, Adrianne Wallace-Povirk, Jade M. Katinas, Larry H. Matherly, Zhanjun Hou, Jing Li, Xun Bao, Charles E. Dann, Josephine Frühauf, Carrie O'Connor, Changwen Ning, Aleem Gangjee, Md. Junayed Nayeen, and Aamod Dekhne
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Purine ,chemistry.chemical_classification ,Cancer Research ,Metabolism ,Mitochondrion ,Molecular biology ,chemistry.chemical_compound ,Cytosol ,Enzyme ,Oncology ,chemistry ,Cell culture ,Serine hydroxymethyltransferase ,Purine metabolism - Abstract
Pancreatic cancer (PaC) represents the 4th leading cause of cancer-related deaths in the US with a mortality rate of 99%. The 5-year overall survival rate for PaC is currently 8%. One-carbon (C1) metabolism is frequently altered in cancer. For PaC, TCGA data sets show that elevated expression of key enzymes involved in cytosolic [5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase (AICARFTase) and serine hydroxymethyltransferase (SHMT)1] and mitochondrial [SHMT2 and methylene tetrahydrofolate dehydrogenase 2 (MTHFD2)] C1 metabolism is associated with poor survival. Antifolate therapeutics disrupt cytosolic C1 pathways required for syntheses of thymidylate, purines, and certain amino acids, and are a mainstay for therapy of several cancers. Antifolate uptake into tumors and tissues involves the reduced folate carrier, the major tissue folate transporter, and the proton-coupled folate transporter (PCFT), which shows a more limited tissue distribution but is widely expressed in human solid tumors and is active only at acidic pHs characterizing the tumor microenvironment. We discovered novel 5-substituted pyrrolo[3,2-d]pyrimidine analogs (AGF347, AGF359) with PCFT transport that potently inhibited proliferation of PaC cell lines (AsPC-1, BxPC-3, CFPAC-1, HPAC, MIA PaCa-2 and PANC-1), of which HPAC (KRAS mutant) and BxPC-3 (KRAS wild-type) cells were most sensitive. The PaC cell lines all expressed PCFT transcripts and proteins that were active for PCFT transport with 3H-AGF347 at acid pH. When HPAC cells were incubated with 3H-AGF347 over 48 h, drug accumulated in both cytosol and mitochondria. 3H-AGF347 was extensively metabolized to polyglutamates. Treatment of PaC cells with AGF347 and AGF359 inhibited proliferation by inducing glycine and adenosine auxotrophy that was rescued by excess glycine and adenosine. This implied that both mitochondria and cytosolic C1 metabolism was inhibited. Inhibition of mitochondrial SHMT2 and cytosolic SHMT1, glycinamide ribonucleotide formyltransferase and/or AICARFTase was confirmed by in vitro targeted metabolomics and assays with purified enzymes. Tumor cell killing was confirmed (with HPAC and BxPC-3) by colony-forming assays with AGF347 and AGF359 and drug-induced apoptosis with AGF347 was demonstrated (with HPAC) by annexin V-PI staining and flow cytometry. AGF347 and AGF359 depleted purine nucleotides and inhibited mTOR signaling via S6K1 at least in part (for BxPC-3) via activation of AMPK, likely due to elevated ZMP accompanying suppression of AICARFTase. Collectively, our studies identify first-in-class inhibitors and establish the considerable therapeutic potential of dual-targeting mitochondrial and cytosolic C1 metabolism in PaC independent of KRAS mutation status and reflecting cellular uptake by PCFT. Citation Format: Changwen Ning, Aamod Dekhne, Md. Junayed Nayeen, Jade M. Katinas, Jennifer Wong, Josephine Frühauf, Xun Bao, Carrie O’Connor, Adrianne Wallace-Povirk, Jing Li, Charles E. Dann, Aleem Gangjee, Larry H. Matherly, Zhanjun Hou. Targeting mitochondrial and cytosolic one carbon metabolism of pancreatic adenocarcinoma via the proton-coupled folate transporter with novel 5-substituted pyrrolo[3,2-d]pyrimidine analogs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4794.
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- 2019
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