T2-FLAIR mismatch sign, an imaging biomarker for CDKN2A-intact in non-enhancing astrocytoma, IDH-mutant.

INTRODUCTION: The WHO classification of central nervous system tumors (5th edition) classified astrocytoma, IDH-mutant accompanied with CDKN2A/B homozygous deletion as WHO grade 4. Loss of immunohistochemical (IHC) staining for methylthioadenosine phosphorylase (MTAP) was developed as a surrogate marker for CDKN2A-HD. Identification of imaging biomarkers for CDKN2A status is of immense clinical relevance. In this study, we explored the association between radiological characteristics of non-enhancing astrocytoma, IDH-mutant to the CDKN2A/B status. METHODS: Thirty-one cases of astrocytoma, IDH-mutant with MTAP results by IHC were included in this study. The status of CDKN2A was diagnosed by IHC staining for MTAP in all cases, which was further confirmed by comprehensive genomic analysis in 12 cases. The T2-FLAIR mismatch sign, cystic component, calcification, and intratumoral microbleeding were evaluated. The relationship between the radiological features and molecular pathological diagnosis was analyzed. RESULTS: Twenty-six cases were identified as CDKN2A-intact while 5 cases were CDKN2A-HD. The presence of > 33% and > 50% T2-FLAIR mismatch was observed in 23 cases (74.2%) and 14 cases (45.2%), respectively, and was associated with CDKN2A-intact astrocytoma (p = 0.0001, 0.0482). None of the astrocytoma, IDH-mutant with CDKN2A-HD showed T2-FLAIR mismatch sign. Cystic component, calcification, and intratumoral microbleeding were not associated with CDKN2A status. CONCLUSION: In patients with non-enhancing astrocytoma, IDH-mutant, the T2-FLAIR mismatch sign is a potential imaging biomarker for the CDKN2A-intact subtype. This imaging biomarker may enable preoperative prediction of CDKN2A status among astrocytoma, IDH-mutant.

Prevalence of S-methyl-5'-thioadenosine Phosphorylase (MTAP) Deficiency in Human Cancer: A Tissue Microarray Study on 13,067 Tumors From 149 Different Tumor Types.

Loss of S-methyl-5'-thioadenosine phosphorylase (MTAP) expression is a common event in cancer leading to a critical vulnerability of cancer cells towards anti-cancer drugs. Homozygous MTAP deletions result in a complete expression loss that can be detected by immunohistochemistry (IHC). In this study, a tissue microarray containing 17,078 samples from 149 different tumor entities was analyzed by IHC, and complete MTAP loss was validated by fluorescence in situ hybridization. MTAP loss was observed in 83 of 149 tumor categories, including neuroendocrine neoplasms (up to 80%), Hodgkin lymphoma (50.0%), mesothelioma (32.0% to 36.8%), gastro-intestinal adenocarcinoma (4.0% to 40.5%), urothelial neoplasms (10.5% to 36.7%), squamous cell carcinomas (up to 38%), and various types of sarcomas (up to 20%) and non-Hodgkin lymphomas (up to 14%). Homozygous MTAP deletion was found in 90% to 100% of cases with MTAP expression loss in most tumor categories. However, neuroendocrine tumors, Hodgkin lymphomas, and other lymphomas lacked MTAP deletions. MTAP deficiency was significantly linked to unfavorable tumor phenotype in selected tumor entities and the presence of PD-L1 expression on tumor cells, absence of PD-L1 expression on immune cells, and a low density of CD8 + lymphocytes. In summary, MTAP deficiency can occur in various tumor entities and is linked to unfavorable tumor phenotype and noninflamed tumor microenvironment, but is not always related to deletions. MTAP IHC is of considerable diagnostic value for the detection of neoplastic transformation in multiple different applications.

Spatial Landscape of Malignant Pleural and Peritoneal Mesothelioma Tumor Immune Microenvironments.

Immunotherapies have demonstrated limited clinical efficacy in malignant mesothelioma treatment. We conducted multiplex immunofluorescence analyses on tissue microarrays (n = 3) from patients with malignant pleural mesothelioma (MPM, n = 88) and malignant peritoneal mesothelioma (MPeM, n = 25). Our study aimed to elucidate spatial distributions of key immune cell populations and their association with lymphocyte activation gene 3 (LAG3), BRCA1-associated protein 1 (BAP1), neurofibromatosis type 2 (NF2), and methylthioadenosine phosphorylase (MTAP), with MTAP serving as a cyclin-dependent kinase inhibitor 2A/2B (CDKN2A/B) surrogate marker. Additionally, we examined the relationship between the spatial distribution of major immune cell types and prognosis and clinical characteristics of patients with malignant mesothelioma. We observed a higher degree of interaction between immune cells and tumor cells in MPM compared with MPeM. Notably, within MPM tumors, we detected a significantly increased interaction between tumor cells and CD8+ T cells in tumors with low BAP1 expression compared with those with high BAP1 expression. To support the broader research community, we have developed The Human Spatial Atlas of Malignant Mesothelioma, containing hematoxylin and eosin and multiplex immunofluorescence images with corresponding metadata. SIGNIFICANCE: Considering the limited therapeutic options available to patients with malignant mesothelioma, there is substantial translational potential in understanding the correlation between the spatial architecture of the malignant mesothelioma tumor immune microenvironment and tumor biology. Our investigation reveals critical cell-cell interactions that may influence the immune response against malignant mesothelioma tumors, potentially contributing to the differential behaviors observed in MPM and MPeM. These findings represent a valuable resource for the malignant mesothelioma cancer research community.

Enzymatic Transglycosylation Features in Synthesis of 8-Aza-7-Deazapurine Fleximer Nucleosides by Recombinant E. coli PNP: Synthesis and Structure Determination of Minor Products.

Enzymatic transglycosylation of the fleximer base 4-(4-aminopyridine-3-yl)-1H-pyrazole using recombinant E. coli purine nucleoside phosphorylase (PNP) resulted in the formation of "non-typical" minor products of the reaction. In addition to "typical" N1-pyrazole nucleosides, a 4-imino-pyridinium riboside and a N1-pyridinium-N1-pyrazole bis-ribose derivative were formed. N1-Pyrazole 2'-deoxyribonucleosides and a N1-pyridinium-N1-pyrazole bis-2'-deoxyriboside were formed. But 4-imino-pyridinium deoxyriboside was not formed in the reaction mixture. The role of thermodynamic parameters of key intermediates in the formation of reaction products was elucidated. To determine the mechanism of binding and activation of heterocyclic substrates in the E. coli PNP active site, molecular modeling of the fleximer base and reaction products in the enzyme active site was carried out. As for N1-pyridinium riboside, there are two possible locations for it in the PNP active site. The presence of a relatively large space in the area of amino acid residues Phe159, Val178, and Asp204 allows the ribose residue to fit into that space, and the heterocyclic base can occupy a position that is suitable for subsequent glycosylation. Perhaps it is this "upside down" arrangement that promotes secondary glycosylation and the formation of minor bis-riboside products.

Mechanistic safety assessment via multi-omic characterisation of systemic pathway perturbations following in vivo MAT2A inhibition.

The tumour suppressor p16/CDKN2A and the metabolic gene, methyl-thio-adenosine phosphorylase (MTAP), are frequently co-deleted in some of the most aggressive and currently untreatable cancers. Cells with MTAP deletion are vulnerable to inhibition of the metabolic enzyme, methionine-adenosyl transferase 2A (MAT2A), and the protein arginine methyl transferase (PRMT5). This synthetic lethality has paved the way for the rapid development of drugs targeting the MAT2A/PRMT5 axis. MAT2A and its liver- and pancreas-specific isoform, MAT1A, generate the universal methyl donor S-adenosylmethionine (SAM) from ATP and methionine. Given the pleiotropic role SAM plays in methylation of diverse substrates, characterising the extent of SAM depletion and downstream perturbations following MAT2A/MAT1A inhibition (MATi) is critical for safety assessment. We have assessed in vivo target engagement and the resultant systemic phenotype using multi-omic tools to characterise response to a MAT2A inhibitor (AZ'9567). We observed significant SAM depletion and extensive methionine accumulation in the plasma, liver, brain and heart of treated rats, providing the first assessment of both global SAM depletion and evidence of hepatic MAT1A target engagement. An integrative analysis of multi-omic data from liver tissue identified broad perturbations in pathways covering one-carbon metabolism, trans-sulfuration and lipid metabolism. We infer that these pathway-wide perturbations represent adaptive responses to SAM depletion and confer a risk of oxidative stress, hepatic steatosis and an associated disturbance in plasma and cellular lipid homeostasis. The alterations also explain the dramatic increase in plasma and tissue methionine, which could be used as a safety and PD biomarker going forward to the clinic.

Concordance between CDKN2A homozygous deletion and MTAP immunohistochemical loss in fluoroedenite-induced pleural mesothelioma: An immunohistochemical and molecular study on a single-institution series.

Fluoroedenite-induced pleural mesothelioma (FE-induced-PM) is a rare and small subset of PM that shares with its asbestos-induced counterpart the same aggressive biological behavior and poor prognosis, but that differs from it from a pathogenetic point of view as it is associated with exposure to fluoroedenite, a carcinogenic agent that shows similarities with tremolite amphibolic asbestos fibers. Although it has been demonstrated that asbestos-induced PMs frequently harbor CDKN2A homozygous deletion and that the immunohistochemical loss of MTAP may represent a cheap and reliable surrogate marker for this molecular alteration, little is known about the molecular landscape and the reliability of MTAP immunohistochemistry in this peculiar subset of PM. The study herein presented investigated the prevalence of CDKN2A homozygous deletion and its concordance with MTAP immunohistochemical status on a cohort of 10 cases of FE-induced-PM from patients with environmental exposure to FE fibers, who were residents in the small town of Biancavilla (Sicily, Italy) or nearby areas. CDKN2A homozygous deletions were found in 3 out of 10 cases (30%) and all these cases showed concomitant cytoplasmic loss of MTAP with a concordance rate of 100%. Despite the relatively low number of cases included in our series, MTAP immunohistochemistry seemed to represent a reliable immunohistochemical surrogate marker of CDKNA homozygous deletion even in this subset of PMs.

Loss of p16 Immunoexpression and Deletions of CDKN2A in the Progression of Extramammary Paget Disease: An Immunohistochemical and Genetic Study of 24 Invasive/Metastatic Cases.

ABSTRACT: Information regarding the genetic alterations in extramammary Paget disease (EMPD) is scarce. This study investigated the significance of CDKN2A and MTAP alterations in EMPD progression using immunohistochemistry and panel DNA sequencing. In total, 24 invasive/metastatic EMPD cases were included in this study. The immunoexpression of p16 and MTAP in the primary in situ, primary invasive, and metastatic tumor components was evaluated. Panel DNA sequencing was performed for metastatic tumor components in 5 of the 24 cases. Immunoexpression of p16 in the in situ tumor component was at least partially preserved in all 19 tested cases (100%). By contrast, the invasive tumor component was diffusely or partially lost in 18 (81.8%) of 22 tested cases. Regarding the foci of lymph node metastasis, 13 (81.2%) of the 16 patients showed a significant loss of p16 expression. Loss of MTAP immunoexpression was observed less frequently compared with the loss of p16 expression. CDKN2A homozygous deletions were confirmed in all 5 tested cases by sequencing, whereas MTAP deletions were detected in only 2 cases. In conclusion, p16 expression loss and CDKN2A deletions can be frequently seen in invasive/metastatic cases of EMPD.

Loss of methylthioadenosine phosphorylase immunoreactivity correlates with poor prognosis and elevated uptake of 11C-methionine in IDH-mutant astrocytoma.

PURPOSE: The proximate localization of MTAP, which encodes methylthioadenosine phosphorylase, and CDKN2A/B on Chromosome 9q21 has allowed the loss of MTAP expression as a surrogate for homozygous deletion of CDKN2A/B. This study aimed to determine whether MTAP status correlates with clinical outcomes and 11C-methionine uptake in astrocytomas with IDH mutations. METHODS: We conducted immunohistochemistry for MTAP in 30 patients with astrocytoma, IDH-mutant who underwent 11C-methionine positron emission tomography scans prior to surgical resection. The tumor-to-normal (T/N) ratio of 11C-methionine uptake was calculated using the mean standardized uptake value (SUV) for tumor and normal brain tissues. Cox regression analysis was used for multivariate survival analysis. RESULTS: Among IDH-mutant astrocytomas, 26.7% (8/30) exhibited the loss of cytoplasmic MTAP expression, whereas 73.3% (22/30) tumors retained MTAP expression. The median progression-free survival (PFS) was significantly shorter in patients with MTAP loss than those with MTAP retention (1.88 years vs. 6.80 years, p = 0.003). The median overall survival (OS) was also shorter in patients with MTAP loss than in MTAP-retaining counterparts (5.23 years vs. 10.69 years, p = 0.019). Multivariate analysis identified MTAP status (hazard ratio (HR), 0.081) and extent of resection (HR, 0.104) as independent prognostic factors for PFS. Astrocytomas lacking cytoplasmic MTAP expression showed a significantly higher median T/N ratio for 11C-methionine uptake than tumors retaining MTAP (2.12 vs. 1.65, p = 0.012). CONCLUSION: Our study revealed that the loss of MTAP expression correlates with poor prognosis and an elevated T/N ratio of 11C-methionine uptake in astrocytoma, IDH-mutant.

[Methylthioadenosine phosphorylase and p16 as surrogate diagnostic markers for CDKN2A homozygous deletion in brain tumors].

Objective: To examine whether immunohistochemistry of methylthioadenosine phosphorylase (MTAP) and p16 could be used to predict the CDKN2A status in various brain tumors. Methods: A total of 118 cases of IDH-mutant astrocytomas, 16 IDH-wildtype glioblastoma, 17 polymorphic xanthoastrocytoma (PXA) and 20 meningiomas diagnosed at Xuanwu Hospital, Capital Medical University, Beijing, China from November 2017 to October 2023 were collected and analyzed. The CDKN2A status was detected by using fluorescence in situ hybridization or next-generation sequencing. Expression of MTAP and p16 proteins was detected with immunohistochemistry. The association of loss of MTAP/p16 expression with CDKN2A homozygous/heterozygous deletion was examined. Results: Among the 118 cases of IDH-mutant astrocytoma, 13 cases showed homozygous deletion of CDKN2A. All of them had no expression of MTAP while 9 cases had no expression of p16. Among the 16 cases of IDH wild-type glioblastoma, 6 cases showed homozygous deletion of CDKN2A. All 6 cases had no expression of MTAP, while 3 of these cases had no expression of p16 expression. Among the 17 PXA cases, 4 cases showed homozygous deletion of CDKN2A, and the expression of MTAP and p16 was also absent in these 4 cases. Among the 20 cases of meningiomas, 4 cases showed homozygous deletion of CDKN2A. Their expression of MTAP and p16 was also absent. Among the four types of brain tumors, MTAP was significantly correlated with CDKN2A homozygous deletion (P<0.05), with a sensitivity of 100%. However, it was only significantly correlated with the loss of heterozygosity (LOH) of CDKN2A in astrocytomas (P<0.001). P16 was associated with CDKN2A homozygous deletion in IDH-mutant astrocytoma and PXA (P<0.001), but not with the LOH of CDKN2A. Its sensitivity and specificity were lower than that of MTAP. Conclusions: MTAP could serve as a predictive surrogate for CDKN2A homozygous deletion in adult IDH-mutant astrocytoma, PXA, adult IDH-wildtype glioblastoma and meningioma. However, p16 could only be used in the first two tumor types, and its specificity and sensitivity are lower than that of MTAP.

Loss of MTAP Expression by Immunohistochemistry Is a Surrogate Marker for Homozygous 9p21.3 Deletion in Urothelial Carcinoma.

Homozygous deletion of the chromosomal region 9p21.3 is common in urothelial carcinoma (UC) and leads to loss of several genes, including CDKN2A and MTAP, resulting in loss of MTAP protein expression. Here, we aimed to explore the diagnostic potential of MTAP immunohistochemistry (IHC) as a surrogate marker for homozygous 9p21.3 deletion (9p21 homozygous deletion [HD]) in UC. MTAP status was determined by IHC on 27 UC tissue specimens with known 9p21.3 status as defined by fluorescence in situ hybridization in matched cytological specimens, by IHC and fluorescence in situ hybridization on a tissue microarray (TMA) containing 359 UC at different stages, and by IHC on 729 consecutive UC from routine practice. Moreover, we analyzed a longitudinal series of matched specimens from 38 patients with MTAP-negative recurrent UC. MTAP loss by IHC was found in all 17 patients with 9p21 HD and in 2/8 cases without 9p21 HD. In the TMA, MTAP loss was more common in metastases (53%) than in muscle-invasive (33%) and non-muscle-invasive UC (29%) (P = .03). In the consecutive series, 164/729 (22%) cases showed loss of MTAP expression. In 41 of these 164 cases (25%), loss of MTAP expression was heterogenous. We also discovered loss of MTAP expression in flat urothelium adjacent to MTAP-negative low-grade UC, suggesting true flat low-grade neoplasia that could not be diagnosed by morphology alone. Longitudinal analysis of recurrences showed persistent negative MTAP status over time in 37/38 (97%) patients. MTAP IHC can serve as a surrogate marker for 9p21 HD in UC and as a diagnostic tool to differentiate reactive urothelium from urothelial neoplasia. It also provides a unique opportunity to study clinicopathological associations and the heterogeneity of 9p21 HD across the whole spectrum of UC manifestations.

Polyamine Inhibitor SAM486A Augments Cytarabine Cytotoxicity in Methylthioadenosine Phosphorylase-deficient Leukemia Cells.

BACKGROUND/AIM: Methionine metabolism contributes to supplying sulfur-containing amino acids, controlling the methyl group transfer reaction, and producing polyamines in cancer cells. Polyamines play important roles in various cellular functions. Methylthioadenosine phosphorylase (MTAP), the key enzyme of the methionine salvage pathway, is reported to be deficient in 15-62% of cases of hematological malignancies. MTAP-deficient cancer cells accumulate polyamines, resulting in enhanced cell proliferation. The aim of this study was to investigate the combined effects of the polyamine synthesis inhibitor SAM486A and the anticancer antimetabolite cytarabine in MTAP-deficient leukemic cells in vitro. MATERIALS AND METHODS: The leukemia cell line U937 and the subline, U937/MTAP(-), in which MTAP was knocked down by shRNA, were used. The experiments were performed in media supplemented with 20% methionine (low methionine), which was the minimum concentration for maintaining cellular viability. RESULTS: The knockdown efficiency test confirmed a 70% suppression of the expression of the MTAP gene in U937/MTAP(-) cells. Even in the media with low methionine, the intracellular methionine concentration was not reduced in U937/MTAP(-) cells, suggesting that the minimum supply of methionine was sufficient to maintain intracellular levels of methionine. Both U937/MTAP(+) and U937/MTAP(-) cells were comparably sensitive to anticancer drugs (cytarabine, methotrexate, clofarabine and 6-thioguanine). The combination of SAM486A and cytarabine was demonstrated to have synergistic cytotoxicity in U937/MTAP(-) cells with regard to cell growth inhibition and apoptosis induction, but not in U937/MTAP(+) cells. Mechanistically, SAM486A altered the intracellular polyamine concentrations and reduced the antiapoptotic proteins. CONCLUSION: Methionine metabolism and polyamine synthesis can be attractive therapeutic targets in leukemia.

Methylthioadenosine Phosphorylase Genomic Loss in Advanced Gastrointestinal Cancers.

BACKGROUND: One of the most common sporadic homozygous deletions in cancers is 9p21 loss, which includes the genes methylthioadenosine phosphorylase (MTAP), CDKN2A, and CDKN2B, and has been correlated with worsened outcomes and immunotherapy resistance. MTAP-loss is a developing drug target through synthetic lethality with MAT2A and PMRT5 inhibitors. The purpose of this study is to investigate the prevalence and genomic landscape of MTAP-loss in advanced gastrointestinal (GI) tumors and investigate its role as a prognostic biomarker. MATERIALS AND METHODS: We performed next-generation sequencing and comparative genomic and clinical analysis on an extensive cohort of 64 860 tumors comprising 5 GI cancers. We compared the clinical outcomes of patients with GI cancer harboring MTAP-loss and MTAP-intact tumors in a retrospective study. RESULTS: The prevalence of MTAP-loss in GI cancers is 8.30%. MTAP-loss was most prevalent in pancreatic ductal adenocarcinoma (PDAC) at 21.7% and least in colorectal carcinoma (CRC) at 1.1%. MTAP-loss tumors were more prevalent in East Asian patients with PDAC (4.4% vs 3.2%, P = .005) or intrahepatic cholangiocarcinoma (IHCC; 6.4% vs 4.3%, P = .036). Significant differences in the prevalence of potentially targetable genomic alterations (ATM, BRAF, BRCA2, ERBB2, IDH1, PIK3CA, and PTEN) were observed in MTAP-loss tumors and varied according to tumor type. MTAP-loss PDAC, IHCC, and CRC had a lower prevalence of microsatellite instability or elevated tumor mutational burden. Positive PD-L1 tumor cell expression was less frequent among MTAP-loss versus MTAP-intact IHCC tumors (23.2% vs 31.2%, P = .017). CONCLUSION: In GI cancers, MTAP-loss occurs as part of 9p21 loss and has an overall prevalence of 8%. MTAP-loss occurs in 22% of PDAC, 15% of IHCC, 8.7% of gastroesophageal adenocarcinoma, 2.4% of hepatocellular carcinoma, and 1.1% of CRC and is not mutually exclusive with other targetable mutations.

Combined inhibition of MTAP and MAT2a mimics synthetic lethality in tumor models via PRMT5 inhibition.

Homozygous 5'-methylthioadenosine phosphorylase (MTAP) deletions occur in approximately 15% of human cancers. Co-deletion of MTAP and methionine adenosyltransferase 2 alpha (MAT2a) induces a synthetic lethal phenotype involving protein arginine methyltransferase 5 (PRMT5) inhibition. MAT2a inhibitors are now in clinical trials for genotypic MTAP-/- cancers, however the MTAP-/- genotype represents fewer than 2% of human colorectal cancers (CRCs), limiting the utility of MAT2a inhibitors in these and other MTAP+/+ cancers. Methylthio-DADMe-immucillin-A (MTDIA) is a picomolar transition state analog inhibitor of MTAP that renders cells enzymatically MTAP-deficient to induce the MTAP-/- phenotype. Here, we demonstrate that MTDIA and MAT2a inhibitor AG-270 combination therapy mimics synthetic lethality in MTAP+/+ CRC cell lines with similar effects in mouse xenografts and without adverse histology on normal tissues. Combination treatment is synergistic with a 104-fold increase in drug potency for inhibition of CRC cell growth in culture. Combined MTDIA and AG-270 decreases S-adenosyl-L-methionine and increases 5'-methylthioadenosine in cells. The increased intracellular methylthioadenosine:S-adenosyl-L-methionine ratio inhibits PRMT5 activity, leading to cellular arrest and apoptotic cell death by causing MDM4 alternative splicing and p53 activation. Combination MTDIA and AG-270 treatment differs from direct inhibition of PRMT5 by GSK3326595 by avoiding toxicity caused by cell death in the normal gut epithelium induced by the PRMT5 inhibitor. The combination of MTAP and MAT2a inhibitors expands this synthetic lethal approach to include MTAP+/+ cancers, especially the remaining 98% of CRCs without the MTAP-/- genotype.

SERS analysis of cancer cell-secreted purines reveals a unique paracrine crosstalk in MTAP-deficient tumors.

The tumor microenvironment (TME) is a dynamic pseudoorgan that shapes the development and progression of cancers. It is a complex ecosystem shaped by interactions between tumor and stromal cells. Although the traditional focus has been on the paracrine communication mediated by protein messengers, recent attention has turned to the metabolic secretome in tumors. Metabolic enzymes, together with exchanged substrates and products, have emerged as potential biomarkers and therapeutic targets. However, traditional techniques for profiling secreted metabolites in complex cellular contexts are limited. Surface-enhanced Raman scattering (SERS) has emerged as a promising alternative due to its nontargeted nature and simplicity of operation. Although SERS has demonstrated its potential for detecting metabolites in biological settings, its application in deciphering metabolic interactions within multicellular systems like the TME remains underexplored. In this study, we introduce a SERS-based strategy to investigate the secreted purine metabolites of tumor cells lacking methylthioadenosine phosphorylase (MTAP), a common genetic event associated with poor prognosis in various cancers. Our SERS analysis reveals that MTAP-deficient cancer cells selectively produce methylthioadenosine (MTA), which is taken up and metabolized by fibroblasts. Fibroblasts exposed to MTA exhibit: i) molecular reprogramming compatible with cancer aggressiveness, ii) a significant production of purine derivatives that could be readily recycled by cancer cells, and iii) the capacity to secrete purine derivatives that induce macrophage polarization. Our study supports the potential of SERS for cancer metabolism research and reveals an unprecedented paracrine crosstalk that explains TME reprogramming in MTAP-deleted cancers.

Enzyme-mediated depletion of methylthioadenosine restores T cell function in MTAP-deficient tumors and reverses immunotherapy resistance.

Chromosomal region 9p21 containing tumor suppressors CDKN2A/B and methylthioadenosine phosphorylase (MTAP) is one of the most frequent genetic deletions in cancer. 9p21 loss is correlated with reduced tumor-infiltrating lymphocytes (TILs) and resistance to immune checkpoint inhibitor (ICI) therapy. Previously thought to be caused by CDKN2A/B loss, we now show that it is loss of MTAP that leads to poor outcomes on ICI therapy and reduced TIL density. MTAP loss causes accumulation of methylthioadenosine (MTA) both intracellularly and extracellularly and profoundly impairs T cell function via the inhibition of protein arginine methyltransferase 5 (PRMT5) and by adenosine receptor agonism. Administration of MTA-depleting enzymes reverses this immunosuppressive effect, increasing TILs and drastically impairing tumor growth and importantly, synergizes well with ICI therapy. As several studies have shown ICI resistance in 9p21/MTAP null/low patients, we propose that MTA degrading therapeutics may have substantial therapeutic benefit in these patients by enhancing ICI effectiveness.

Neurologic Status of Patients with Purine Nucleoside Phosphorylase Deficiency Before and After Hematopoetic Stem Cell Transplantation.

BACKGROUND: Purine nucleoside phosphorylase (PNP) deficiency is a rare autosomal recessive combined immunodeficiency. The phenotype is profound T cell deficiency with variable B and NK cell functions and results in recurrent and persistent infections that typically begin in the first year of life. Neurologic findings occur in approximately two-thirds of patients. The mechanism of neurologic abnormalities is unclear. Hematopoietic stem cell transplantation (HSCT) is the only curative treatment for PNP deficiency. METHODS: We report here six patients from five unrelated families with PNP deficiency treated in two centers in Turkey. We evaluated the neurological status of patients and compared to post-transplantation period if available. Then, we performed PubMed, Google Scholar, and Researchgate searches using the terms "PNP" and "hematopoietic stem cell transplantation" to find all reported cases of PNP transplantation and compared to our cohort. RESULTS: Six patients were treated in two centers in Turkey. One patient died from post-transplant complications. The other four patients underwent successful HSCT with good immune reconstitution after transplantation (follow-up 21-48 months) and good neurological outcomes. The other patient with a new mutation is still waiting for a matching HLA donor. DISCUSSION: In PNP deficiency, clinical manifestations are variable, and this disease should be considered in the presence of many different clinical findings. Despite the comorbidities that occurred before transplantation, HSCT currently appears to be the only treatment option for this disease. HSCT not only cures immunologic disorders, but probably also improves or at least stabilizes the neurologic status of patients.

Efficient synthesis of 2'-deoxyguanosine in one-pot cascade by employing an engineered purine nucleoside phosphorylase from Brevibacterium acetylicum.

2'-deoxyguanosine is a key medicinal intermediate that could be used to synthesize anti-cancer drug and biomarker in type 2 diabetes. In this study, an enzymatic cascade using thymidine phosphorylase from Escherichia coli (EcTP) and purine nucleoside phosphorylase from Brevibacterium acetylicum (BaPNP) in a one-pot whole cell catalysis was proposed for the efficient synthesis of 2'-deoxyguanosine. BaPNP was semi-rationally designed to improve its activity, yielding the best triple variant BaPNP-Mu3 (E57A/T189S/L243I), with a 5.6-fold higher production of 2'-deoxyguanosine than that of wild-type BaPNP (BaPNP-Mu0). Molecular dynamics simulation revealed that the engineering of BaPNP-Mu3 resulted in a larger and more flexible substrate entrance channel, which might contribute to its catalytic efficiency. Furthermore, by coordinating the expression of BaPNP-Mu3 and EcTP, a robust whole cell catalyst W05 was created, capable of producing 14.8 mM 2'-deoxyguanosine (74.0% conversion rate) with a high time-space yield (1.32 g/L/h) and therefore being very competitive for industrial applications.

Purine Nucleoside Phosphorylase Deficient Severe Combined Immunodeficiencies: A Case Report and Systematic Review (1975-2022).

Purine nucleoside phosphorylase deficient severe combined immunodeficiency (PNP SCID) is one of the rare autosomal recessive primary immunodeficiency disease, and the data on epidemiology and outcome are limited. We report the successful management of a child with PNP SCID and present a systematic literature review of published case reports, case series, and cohort studies on PNP SCID listed in PubMed, Web of Science, and Scopus from 1975 until March 2022. Forty-one articles were included from the 2432 articles retrieved and included 100 PNP SCID patients worldwide. Most patients presented with recurrent infections, hypogammaglobulinaemia, autoimmune manifestations, and neurological deficits. There were six reported cases of associated malignancies, mainly lymphomas. Twenty-two patients had undergone allogeneic hematopoietic stem cell transplantation with full donor chimerism seen mainly in those receiving matched sibling donors and/or conditioning chemotherapy before the transplant. This research provides a contemporary, comprehensive overview on clinical manifestations, epidemiology, genotype mutations, and transplant outcome of PNP SCID. These data highlight the importance of screening for PNP SCID in cases presented with recurrent infections, hypogammaglobulinaemia, and neurological deficits.

Phosphoinositide and redox dysregulation by the anticancer methylthioadenosine phosphorylase transition state inhibitor.

Methylthio-DADMe-immucillin-A (MTDIA) is an 86 picomolar inhibitor of 5'-methylthioadenosine phosphorylase (MTAP) with potent and specific anti-cancer efficacy. MTAP salvages S-adenosylmethionine (SAM) from 5'-methylthioadenosine (MTA), a toxic metabolite produced during polyamine biosynthesis. Changes in MTAP expression are implicated in cancer growth and development, making MTAP an appealing target for anti-cancer therapeutics. Since SAM is involved in lipid metabolism, we hypothesised that MTDIA alters the lipidomes of MTDIA-treated cells. To identify these effects, we analysed the lipid profiles of MTDIA-treated Saccharomyces cerevisiae using ultra-high resolution accurate mass spectrometry (UHRAMS). MTAP inhibition by MTDIA, and knockout of the Meu1 gene that encodes for MTAP in yeast, caused global lipidomic changes and differential abundance of lipids involved in cell signaling. The phosphoinositide kinase/phosphatase signaling network was specifically impaired upon MTDIA treatment, and was independently validated and further characterised via altered localization of proteins integral to this network. Functional consequences of dysregulated lipid metabolism included a decrease in reactive oxygen species (ROS) levels induced by MTDIA that was contemporaneous with changes in immunological response factors (nitric oxide, tumour necrosis factor-alpha and interleukin-10) in mammalian cells. These results indicate that lipid homeostasis alterations and concomitant downstream effects may be associated with MTDIA mechanistic efficacy.