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Current Opinion in Investigational... Nov 2001Researchers at the University of California at San Diego (UCSD) are developing alanosine as a potential treatment for cancer [227466], [408222]. The compound was... (Review)
Review
Researchers at the University of California at San Diego (UCSD) are developing alanosine as a potential treatment for cancer [227466], [408222]. The compound was originally under development in collaboration with Triangle, which initiated its development in 1996 [227466], but later discontinued development of the compound [406677]. As of May 2001, UCSD's ongoing clinical trials of alanosine included phase II trials for non-small cell lung cancer (NSCLC) and phase I trials for acute lymphoid leukemia (ALL), while a phase II trial for glioma at UCSD had been suspended [408222]. Alanosine is an amino acid analog originally derived from Streptomyces alanosinicus. It interferes with the de novo synthesis of adenosine in both malignant and normal cells. In cancer cells that lack methyladenosine phosphorylase (MTAP, required in the salvage pathway), alanosine should deprive such cells (but not normal cells) of de novo synthesized adenosine [277968]. In early 1997, patients were being recruited for a phase II pilot efficacy trial of alanosine as a treatment for glioma and NSCLC, since a significant number of these tumor types lack MTAP and, it was hoped, would therefore be sensitive to alanosine [239280], [248260]. Phase I and II trials were completed in the 1980s by the NCI before they were discontinued because alanosine caused toxicity typically associated with chemotherapy, and did not produce significant response rates in common tumors such as breast or colon cancers. Researchers at UCSD found that some types of cancer lack MTAP, which was responsible for alanosine's previous clinical failure [227466]; phase II trials were being carried out at the university in 1997 [269338]. Triangle obtained an option for a worldwide license from the Regents of the University of California that expired in September 1998 (but had an option to extend the period for a further one year) [277968].
Topics: Alanine; Animals; Antibiotics, Antineoplastic; Humans
PubMed: 11763167
DOI: No ID Found -
Angewandte Chemie (International Ed. in... Mar 2020The formation of a N-N bond is a unique biochemical transformation, and nature employs diverse biosynthetic strategies to activate nitrogen for bond formation. Among...
The formation of a N-N bond is a unique biochemical transformation, and nature employs diverse biosynthetic strategies to activate nitrogen for bond formation. Among molecules that contain a N-N bond, biosynthetic routes to diazeniumdiolates remain enigmatic. We here report the biosynthetic pathway for the diazeniumdiolate-containing amino acid l-alanosine. Our work reveals that the two nitrogen atoms in the diazeniumdiolate of l-alanosine arise from glutamic acid and aspartic acid, and we clarify the early steps of the biosynthetic pathway by using both in vitro and in vivo approaches. Our work demonstrates a peptidyl-carrier-protein-based mechanism for activation of the precursor l-diaminopropionate, and we also show that nitric oxide can participate in non-enzymatic diazeniumdiolate formation. Furthermore, we demonstrate that the gene alnA, which encodes a fusion protein with an N-terminal cupin domain and a C-terminal AraC-like DNA-binding domain, is required for alanosine biosynthesis.
Topics: Alanine; Aspartic Acid; Glutamic Acid; Molecular Structure; Multigene Family; Streptomyces
PubMed: 31823464
DOI: 10.1002/anie.201913458 -
Chembiochem : a European Journal of... Apr 2020N-Nitroso-containing natural products are bioactive metabolites with antibacterial and anticancer properties. In particular, compounds containing the diazeniumdiolate...
N-Nitroso-containing natural products are bioactive metabolites with antibacterial and anticancer properties. In particular, compounds containing the diazeniumdiolate (N-nitrosohydroxylamine) group display a wide range of bioactivities ranging from cytotoxicity to metal chelation. Despite the importance of this structural motif, knowledge of its biosynthesis is limited. Herein we describe the discovery of a biosynthetic gene cluster in Streptomyces alanosinicus ATCC 15710 responsible for producing the diazeniumdiolate natural product l-alanosine. Gene disruption and stable isotope feeding experiments identified essential biosynthetic genes and revealed the source of the N-nitroso group. Additional biochemical characterization of the biosynthetic enzymes revealed that the non-proteinogenic amino acid l-2,3-diaminopropionic acid (l-Dap) is synthesized and loaded onto a free-standing peptidyl carrier protein (PCP) domain in l-alanosine biosynthesis, which we propose may be a mechanism of handling unstable intermediates generated en route to the diazeniumdiolate. These discoveries will facilitate efforts to determine the biochemistry of diazeniumdiolate formation.
Topics: Alanine; Azo Compounds; Bacterial Proteins; Biosynthetic Pathways; Molecular Structure; Multigene Family; Streptomyces
PubMed: 31643127
DOI: 10.1002/cbic.201900565 -
Journal of the American Chemical Society Aug 2023l-Alanosine is a diazeniumdiolate (-nitrosohydroxylamine) antibiotic that inhibits MTAP-deficient tumor cells by blocking adenine biosynthesis. Previous work revealed...
l-Alanosine is a diazeniumdiolate (-nitrosohydroxylamine) antibiotic that inhibits MTAP-deficient tumor cells by blocking adenine biosynthesis. Previous work revealed the early steps in the biosynthesis of l-alanosine. In the present study, we used genome mining to discover two new l-alanosine-producing strains that lack the spartate-itrouccinate pathway genes found in the original l-alanosine producer. Instead, nitrate is reduced with a unique set of nitrate-nitrite reductases. These enzymes are typically used as part of the nitrogen cycle for denitrification or assimilation, and our report here shows how enzymes from the nitrogen cycle can be repurposed for the biosynthesis of specialized metabolites. The widespread distribution of nitric-oxide-producing reductases also indicates a potential for the discovery of new nitric-oxide-derived natural products.
Topics: Nitric Oxide; Nitrates; Oxidoreductases; Nitrite Reductases; Nitrate Reductases
PubMed: 37478476
DOI: 10.1021/jacs.3c04447 -
Advances in Pharmacology and... 1984
Review
Topics: Alanine; Animals; Antibiotics, Antineoplastic; Antiviral Agents; Bacteria; Chemical Phenomena; Chemistry; Drug Evaluation; Drug Resistance; Humans; Immunosuppressive Agents; Kinetics; Stereoisomerism
PubMed: 6398969
DOI: 10.1016/s1054-3589(08)60265-3 -
Biomedicines Mar 2022Glioblastoma (GBM) is a lethal brain cancer exhibiting high levels of drug resistance, a feature partially imparted by tumor cell stemness. Recent work shows that...
Glioblastoma (GBM) is a lethal brain cancer exhibiting high levels of drug resistance, a feature partially imparted by tumor cell stemness. Recent work shows that homozygous deletion, a genetic alteration occurring in about half of all GBMs, promotes stemness in GBM cells. Exploiting MTAP loss-conferred deficiency in purine salvage, we demonstrate that purine blockade via treatment with L-Alanosine (ALA), an inhibitor of de novo purine synthesis, attenuates stemness of -deficient GBM cells. This ALA-induced reduction in stemness is mediated in part by compromised mitochondrial function, highlighted by ALA-induced elimination of mitochondrial spare respiratory capacity. Notably, these effects of ALA are apparent even when the treatment was transient and with a low dose. Finally, in agreement with diminished stemness and compromised mitochondrial function, we show that ALA sensitizes GBM cells to temozolomide (TMZ) in vitro and in an orthotopic GBM model. Collectively, these results identify purine supply as an essential component in maintaining mitochondrial function in GBM cells and highlight a critical role of mitochondrial function in sustaining GBM stemness. We propose that purine synthesis inhibition can be beneficial in combination with the standard of care for -deficient GBMs, and that it may be feasible to achieve this benefit without inflicting major toxicity.
PubMed: 35453502
DOI: 10.3390/biomedicines10040751 -
Cancer Research Apr 19762-Amino-3-(hydroxynitrosoamino)propionic acid (alanosine), at a concentration as low as 2.7 muM, completely inhibits the incorporation of hypoxanthine into adenosine...
2-Amino-3-(hydroxynitrosoamino)propionic acid (alanosine), at a concentration as low as 2.7 muM, completely inhibits the incorporation of hypoxanthine into adenosine triphosphate by cultured Novikoff rat hepatoma cells. Alanosine inhibits the first step in the conversion of inosine monophosphate to adenosine monophosphate because inosine monophosphate, but not adenylosuccinate, accumulates in treated cells. However, the alanosine inhibition is not prevented by aspartic acid, even at a concentration of 1 mM. Alanosine treatment results in the inhibition of cell division, DNA synthesis, RNA and protein synthesis (in this order), and a depletion of the cells of adenosine triphosphate. Some of the cells accumulate in late G2 or M, but the remainder become arrested in other stages of the cell cycle. All effects are due to the inhibition of adenosine monophosphate synthesis and the consequent depletion of the adenosine triphosphate pool since they are completely prevented or reversed by addition of adenine, but not hypoxanthine, to the medium. Pyrimidine nucleotide synthesis is not significantly inhibited by alanosine, since the uridine triphosphate pool is not affected and uridine fails to reverse the cytotoxicity of alanosine. Alanosine also inhibits the transport of aspartic acid, but has a much lower affinity for this transport system than aspartic acid.
Topics: Adenine; Adenosine Monophosphate; Adenosine Triphosphate; Alanine; Aspartic Acid; Carcinoma, Hepatocellular; Cell Division; Cell Line; Cells, Cultured; DNA, Neoplasm; Depression, Chemical; Glucose; Hypoxanthines; Inosine Nucleotides; Liver Neoplasms; Mengovirus; Neoplasm Proteins; Neoplasms, Experimental; Nitroso Compounds; Purine Nucleotides; Pyrimidine Nucleotides; RNA, Neoplasm; Succinates; Uracil Nucleotides; Uridine
PubMed: 177207
DOI: No ID Found -
Biochemical Pharmacology Dec 1968
Topics: Acetates; Adenine; Amino Acids; Anti-Bacterial Agents; Carbon Isotopes; Chromatography, Paper; Escherichia coli; Ligases; Nucleosides; Penicillium; Propionates; Streptomyces; Succinates
PubMed: 4888630
DOI: 10.1016/0006-2952(68)90144-5 -
Cancer Research Dec 1980The conjugate of L-alanosine [L-2-amino-3-(N-hydroxyN-nitrosamino)propionic acid] and 5-amino-4-imidazolecarboxylic acid ribonucleotide has been synthesized in good...
Identification of the antimetabolite of L-alanosine, L-alanosyl-5-amino-4-imidazolecarboxylic acid ribonucleotide, in tumors and assessment of its inhibition of adenylosuccinate synthetase.
The conjugate of L-alanosine [L-2-amino-3-(N-hydroxyN-nitrosamino)propionic acid] and 5-amino-4-imidazolecarboxylic acid ribonucleotide has been synthesized in good yield by enzymatic means, using partially purified chicken liver 5-amino-4-imidazole-N-succinocarboxamide ribonucleotide synthetase (EC 6.3.2.6). The chromatographic behavior of this molecule was characterized, as was its ability to inhibit adenylosuccinate synthetase, an enzyme long considered to be the locus of action of the drug. The Ki of-L-alanosyl-5-amino-4-imidazolecarboxylic acid ribonucleotide versus a partially purified adenylosuccinate synthetase frm the L5178y/AR leukemia of C57BL X DBA/2 F1 (hereafter called BD2F1) mice was 0.228 microM, whereas the Ki of L-alanosine was 57.23 mM. Administration of 50 microCi of DL-[1-14C]alanosine along with unlabeled L-alanosine (500 mg/kg) to BD2F1 mice bearing s.c. nodules of Leukemia L5178Y/AR resulted in the accumulation in tumors of a material with properties compatible with those of L-alanosyl-5-amino-4-imidazolecarboxylic acid ribonucleotide. It coeluted with L-alanosyl-5-amino-r-imidazolecarboxylic acid ribonucleotide in the high-resolution chromatographic system used, was Bratton-Marshall positive, and inhibited adenylosuccinate synthetase strongly. In tumor nodules 2 hr after dosage, the concentration of this compound approximated 70 microM. Under the same circumstances, the intratumoral concentration of L-alanosine was found to be 440 microM. At this concentration, the antibiotic itself exerts only a marginal inhibition of leukemic adenylosuccinate synthetase. In ancillary studies, it was shown for the first time in vivo that the parenteral administration of L-alanosine reduces the specific activity of intratumoral adenylosuccinate synthetase by 70% and depresses the synthesis of DNA to an equivalent or greater extent; adenine but not hypoxanthine (both at 250 mg/kg) was able to reverse the latter inhibition. No effect on purine salvage enzymes was exerted by L-alanosine. Viewed in concert, these experiments establish that the adduct of L-alanosine with 5-amino-4-imidazolecarboxylic acid is formed by neoplastic cells in vivo and that this anabolite is most probably responsible for the inhibition of adneylosuccinate synthetase and, in turn, for the diminished synthesis of DNA seen after a therapeutic dose of L-alanosine.
Topics: Adenylosuccinate Synthase; Alanine; Animals; Antibiotics, Antineoplastic; Biotransformation; DNA, Neoplasm; Ligases; Mice; Neoplasms, Experimental; Nitrosamines; Purines; Ribonucleotides
PubMed: 7438071
DOI: No ID Found -
Biochemical Pharmacology Aug 2003The methylthioadenosine phosphorylase (MTAP) gene gained considerable interest as therapeutic target for tumors with the 9p21 deletion. This gene maps to 9p21 and loss...
The methylthioadenosine phosphorylase (MTAP) gene gained considerable interest as therapeutic target for tumors with the 9p21 deletion. This gene maps to 9p21 and loss of this chromosomal region in tumors offers an unique opportunity for chemoselective treatment, since MTAP is an important salvage enzyme for the formation of adenine that is needed for DNA synthesis. L-Alanosine, an antibiotic from Streptomyces alanosinicus, blocks the common de novo purine biosynthesis pathway and, thereby, inhibits tumor cells with MTAP deficiency. Normal cells escape the detrimental effects of L-alanosine due to their proficiency in the MTAP salvage pathway. The present analysis was undertaken to gain insights into the molecular architecture of tumor cells that determines the response to L-alanosine apart from the MTAP gene. Analysis of cell doubling times and IC(50) values for L-alanosine showed that slowly growing cell lines were more resistant to L-alanosine than rapidly growing ones. Mining the database of the National Cancer Institute (N.C.I.), for the mRNA expression of 9706 genes in 60 cell lines by means of Kendall's tau-test, false discovery rate calculation, and hierarchical cluster analysis pointed to 11 genes or expressed sequence tags whose mRNA expression correlated with the IC(50) values for L-alanosine. Furthermore, we tested L-alanosine for cross-resistance in multidrug-resistant cell lines which overexpress selectively either the P-glycoprotein/MDR1 (CEM/ADR5000), MRP1 (HL-60/AR), or BCRP (MDA-MB-231-BCRP) genes. None of the multidrug-resistant cell lines was cross-resistant to L-alanosine indicating that L-alanosine may be suitable to treat multidrug-resistant, refractory tumors in the clinic. Finally, the IC(50) values for L-alanosine of the 60 cell lines were correlated to the p53 mutational status and expression of p53 downstream genes. We found that p53 mutated cell lines were more resistant to L-alanosine than p53 wild type cell lines.
Topics: Alanine; Antibiotics, Antineoplastic; Cell Division; Cluster Analysis; Drug Resistance, Multiple; Gene Expression Profiling; HL-60 Cells; Humans; Tumor Suppressor Protein p53
PubMed: 12906926
DOI: 10.1016/s0006-2952(03)00341-1