Classic Galactosemia: Clinical and Computational Characterization of a Novel GALT Missense Variant (p.A303D) and a Literature Review.

Classic galactosemia is an autosomal recessive inherited liver disorder of carbohydrate metabolism caused by deficient activity of galactose-1-phosphate uridylyltransferase (GALT). While a galactose-restricted diet is lifesaving, most patients still develop long-term complications. In this study, we report on a two-week-old female patient who is a compound heterozygote for a known pathogenic variant (p.K285N) and a novel missense variant (p.A303D) in the GALT gene. Segregation analysis showed that the patient inherited the p.K285N pathogenic variant from her father and the p.A303D variant from her mother. A bioinformatics analysis to predict the impact of the p.A303D missense variant on the structure and stability of the GALT protein revealed that it may be pathogenic. Based on this finding, we performed a literature review of all GALT missense variants identified in homozygous and compound heterozygous galactosemia patients carrying the p.K285N pathogenic variant to explore their molecular effects on the clinical phenotype of the disease. Our analysis revealed that these missense variants are responsible for a wide range of molecular defects. This study expands the clinical and mutational spectrum in classic galactosemia and reinforces the importance of understanding the molecular consequences of genetic variants to incorporate genetic analysis into clinical care.

Gastric polyposis and desmoid tumours as a new familial adenomatous polyposis clinical variant associated with APC mutation at the extreme 3'-end.

Germline mutations of the APC gene, which encodes a multidomain protein of 2843 amino acid residues, cause familial adenomatous polyposis (FAP). Three FAP clinical variants are correlated with the location of APC mutations: (1) classic FAP with profuse polyposis (>1000 adenomas), associated with mutations from codon 1250 to 1424; (2) attenuated FAP (<100 adenomas), associated with mutations at APC extremities (before codon 157 and after codon 1595); (3) classic FAP with intermediate colonic polyposis (100-1000 adenomas), associated with mutations located in the remaining part of APC In an effort to decipher the clinical phenotype associated with APC C-terminal germline truncating mutations in patients with FAP, after screening APC mutations in one family whose members (n=4) developed gastric polyposis, colon oligo-polyposis and desmoid tumours, we performed a literature meta-analysis of clinically characterised patients (n=97) harbouring truncating mutations in APC C-terminus. The APC distal mutations identified in this study cluster with a phenotype characterised by colon oligo-polyposis, diffuse gastric polyposis and desmoid tumours. In conclusion, we describe a novel FAP clinical variant, which we propose to refer to as Gastric Polyposis and Desmoid FAP, that may require tailored management.

From Genetics to Histomolecular Characterization: An Insight into Colorectal Carcinogenesis in Lynch Syndrome.

Lynch syndrome is a hereditary cancer-predisposing syndrome caused by germline defects in DNA mismatch repair (MMR) genes such as MLH1, MSH2, MSH6, and PMS2. Carriers of pathogenic mutations in these genes have an increased lifetime risk of developing colorectal cancer (CRC) and other malignancies. Despite intensive surveillance, Lynch patients typically develop CRC after 10 years of follow-up, regardless of the screening interval. Recently, three different molecular models of colorectal carcinogenesis were identified in Lynch patients based on when MMR deficiency is acquired. In the first pathway, adenoma formation occurs in an MMR-proficient background, and carcinogenesis is characterized by APC and/or KRAS mutation and IGF2, NEUROG1, CDK2A, and/or CRABP1 hypermethylation. In the second pathway, deficiency in the MMR pathway is an early event arising in macroscopically normal gut surface before adenoma formation. In the third pathway, which is associated with mutations in CTNNB1 and/or TP53, the adenoma step is skipped, with fast and invasive tumor growth occurring in an MMR-deficient context. Here, we describe the association between molecular and histological features in these three routes of colorectal carcinogenesis in Lynch patients. The findings summarized in this review may guide the use of individualized surveillance guidelines based on a patient's carcinogenesis subtype.

A large de novo 9p21.3 deletion in a girl affected by astrocytoma and multiple melanoma.

BACKGROUND: Association of melanoma, neural system tumors and germ line mutations at the 9p21 region in the CDKN2A, CDKN2B and CDKN2BAS genes has been reported in a small number of families worldwide and described as a discrete syndrome in melanoma families registered as a rare disease, the melanoma-astrocytoma syndrome. CASE PRESENTATION: We here studied two young patients developing melanoma after radiotherapy for astrocytoma, both reporting lack of family history for melanoma or neural system tumors at genetic counselling. Patient A is a girl treated for anaplastic astrocytoma at 10 years and for multiple melanomas on the scalp associated to dysplastic nevi two years later. Her monozygotic twin sister carried dysplastic nevi and a slow growing, untreated cerebral lesion. Direct sequencing analysis showed no alterations in melanoma susceptibility genes including CDKN2A, CDK4, MC1R and MITF or in TP53. By microsatellite analysis, multiplex ligation-dependent probe amplification, and array comparative genomic hybridization a deletion including the CDKN2A, CDKN2B and CDKN2BAS gene cluster was detected in both twin sisters, encompassing a large region at 9p21.3 and occurring de novo after the loss of one paternal allele.Patient B is a boy of 7 years when treated for astrocytoma then developing melanoma associated to congenital nevi on the head 10 years later: sequencing and multiplex ligation-dependent probe amplification revealed a normal profile of the CDKN2A/CDKN2B/CDKN2BAS region. Array comparative genomic hybridization confirmed the absence of deletions at 9p21.3 and failed to reveal known pathogenic copy number variations. CONCLUSIONS: By comparison with the other germ line deletions at the CDKN2A, CDKN2B and CDKN2BAS gene cluster reported in melanoma susceptible families, the deletion detected in the two sisters is peculiar for its de novo origin and for its extension, as it represents the largest constitutive deletion at 9p21.3 region identified so far.In addition, the two studied cases add to other evidence indicating association of melanoma with exposure to ionizing radiation and with second neoplasm after childhood cancer. Melanoma should be considered in the monitoring of pigmented lesions in young cancer patients.

Interstitial 22q13 deletions not involving SHANK3 gene: a new contiguous gene syndrome.

Phelan-McDermid syndrome (22q13.3 deletion syndrome) is a contiguous gene disorder resulting from the deletion of the distal long arm of chromosome 22. SHANK3, a gene within the minimal critical region, is a candidate gene for the major neurological features of this syndrome. We report clinical and molecular data from a study of nine patients with overlapping interstitial deletions in 22q13 not involving SHANK3. All of these deletions overlap with the largest, but not with the smallest deletion associated with Phelan-McDermid syndrome. The deletion sizes and breakpoints varied considerably among our patients, with the largest deletion spanning 6.9 Mb and the smallest deletion spanning 2.7 Mb. Eight out of nine patients had a de novo deletion, while in one patient the origin of deletion was unknown. These patients shared clinical features common to Phelan-McDermid syndrome: developmental delay (11/12), speech delay (11/12), hypotonia (9/12), and feeding difficulties (7/12). Moreover, the majority of patients (8/12) exhibited macrocephaly. In the minimal deleted region, we identified two candidate genes, SULT4A1 and PARVB (associated with the PTEN pathway), which could be associated in our cohort with neurological features and macrocephaly/hypotonia, respectively. This study suggests that the haploinsufficiency of genes in the 22q13 region beside SHANK3 contributes to cognitive and speech development, and that these genes are involved in the phenotype associated with the larger Phelan-McDermid syndrome 22q13 deletions. Moreover, because the deletions in our patients do not involve the SHANK3 gene, we posit the existence of a new contiguous gene syndrome proximal to the smallest terminal deletions in the 22q13 region.

The novel SMYD3 inhibitor EM127 impairs DNA repair response to chemotherapy-induced DNA damage and reverses cancer chemoresistance.

BACKGROUND: SMYD3 has been found implicated in cancer progression. Its overexpression correlates with cancer growth and invasion, especially in gastrointestinal tumors. SMYD3 transactivates multiple oncogenic mechanisms, favoring cancer development. Moreover, it was recently shown that SMYD3 is required for DNA restoration by promoting homologous recombination (HR) repair. METHODS: In cellulo and in vivo models were employed to investigate the role of SMYD3 in cancer chemoresistance. Analyses of SMYD3-KO cells, drug-resistant cancer cell lines, patients' residual gastric or rectal tumors that were resected after neoadjuvant therapy and mice models were performed. In addition, the novel SMYD3 covalent inhibitor EM127 was used to evaluate the impact of manipulating SMYD3 activity on the sensitization of cancer cell lines, tumorspheres and cancer murine models to chemotherapeutics (CHTs). RESULTS: Here we report that SMYD3 mediates cancer cell sensitivity to CHTs. Indeed, cancer cells lacking SMYD3 functions showed increased responsiveness to CHTs, while restoring its expression promoted chemoresistance. Specifically, SMYD3 is essential for the repair of CHT-induced double-strand breaks as it methylates the upstream sensor ATM and allows HR cascade propagation through CHK2 and p53 phosphorylation, thereby promoting cancer cell survival. SMYD3 inhibition with the novel compound EM127 showed a synergistic effect with CHTs in colorectal, gastric, and breast cancer cells, tumorspheres, and preclinical colorectal cancer models. CONCLUSIONS: Overall, our results show that targeting SMYD3 may be an effective therapeutic strategy to overcome chemoresistance.

Uncoupling p38α nuclear and cytoplasmic functions and identification of two p38α phosphorylation sites on ß-catenin: implications for the Wnt signaling pathway in CRC models.

BACKGROUND: Activation of the Wnt pathway has been linked to colorectal cancer (CRC). Previous reports suggest that Wnt3a can activate p38. Besides, p38α feeds into the canonical Wnt/ß-catenin pathway by inhibiting GSK3ß through phosphorylation. Recently, we identified p38α as a new druggable member of ß-catenin chromatin-associated kinase complexes in CRC. METHODS: The functional relationship between p38α and ß-catenin was characterized in CRC cells, patient-derived CRC stem cells, patient-derived tumor intestinal organoids, and in vivo models (C57BL/6-APCMin/+ mice). The role of p38α in ß-catenin transcriptional activity was assessed by pharmacological inhibition with ralimetinib. RESULTS: We used the GSK3ß inhibitor TWS-119, which promotes the activation of Wnt signaling, to uncouple p38α nuclear/cytoplasmatic functions in the Wnt pathway. Upon GSK3ß inhibition, nuclear p38α phosphorylates ß-catenin at residues S111 and T112, allowing its binding to promoter regions of Wnt target genes and the activation of a transcriptional program implicated in cancer progression. If p38α is pharmacologically inhibited in addition to GSK3ß, ß-catenin is prevented from promoting target gene transcription, which is expected to impair carcinogenesis. CONCLUSIONS: p38α seems to play a dual role as a member of the ß-catenin destruction complex and as a ß-catenin chromatin-associated kinase in CRC. This finding may help elucidate mechanisms contributing to human colon tumor pathogenesis and devise new strategies for personalized CRC treatment.

The chromatin remodeling factors EP300 and TRRAP are novel SMYD3 interactors involved in the emerging 'nonmutational epigenetic reprogramming' cancer hallmark.

SMDY3 is a histone-lysine N-methyltransferase involved in several oncogenic processes and is believed to play a major role in various cancer hallmarks. Recently, we identified ATM, BRCA2, CHK2, MTOR, BLM, MET, AMPK, and p130 as direct SMYD3 interactors by taking advantage of a library of rare tripeptides, which we first tested for their in vitro binding affinity to SMYD3 and then used as in silico probes to systematically search the human proteome. Here, we used this innovative approach to identify further SMYD3-interacting proteins involved in crucial cancer pathways and found that the chromatin remodeling factors EP300 and TRRAP interact directly with SMYD3, thus linking SMYD3 to the emerging 'nonmutational epigenetic reprogramming' cancer hallmark. Of note, we validated these interactions in gastrointestinal cancer cell lines, including HCT-116 cells, which harbor a C-terminal truncating mutation in EP300, suggesting that EP300 binds to SMYD3 via its N-terminal region. While additional studies are required to ascertain the functional mechanisms underlying these interactions and their significance, the identification of two novel SMYD3 interactors involved in epigenetic cancer hallmark pathways adds important pieces to the puzzle of how SMYD3 exerts its oncogenic role.

SMYD3 Modulates AMPK-mTOR Signaling Balance in Cancer Cell Response to DNA Damage.

Cells respond to DNA damage by activating a complex array of signaling networks, which include the AMPK and mTOR pathways. After DNA double-strand breakage, ATM, a core component of the DNA repair system, activates the AMPK-TSC2 pathway, leading to the inhibition of the mTOR cascade. Recently, we showed that both AMPK and mTOR interact with SMYD3, a methyltransferase involved in DNA damage response. In this study, through extensive molecular characterization of gastrointestinal and breast cancer cells, we found that SMYD3 is part of a multiprotein complex that is involved in DNA damage response and also comprises AMPK and mTOR. In particular, upon exposure to the double-strand break-inducing agent neocarzinostatin, SMYD3 pharmacological inhibition suppressed AMPK cascade activation and thereby promoted the mTOR pathway, which reveals the central role played by SMYD3 in the modulation of AMPK-mTOR signaling balance during cancer cell response to DNA double-strand breaks. Moreover, we found that SMYD3 can methylate AMPK at the evolutionarily conserved residues Lys411 and Lys424. Overall, our data revealed that SMYD3 can act as a bridge between the AMPK and mTOR pathways upon neocarzinostatin-induced DNA damage in gastrointestinal and breast cancer cells.

SMYD3 Modulates the HGF/MET Signaling Pathway in Gastric Cancer.

Gastric cancer (GC) is the third most deadly cancer worldwide. Considerable efforts have been made to find targetable drivers in order to improve patient outcomes. MET is one of the most important factors involved in GC initiation and progression as it plays a major role in GC invasiveness and is related to cancer stemness. Unfortunately, treatment strategies targeting MET are still limited, with a proportion of patients responding to therapy but later developing resistance. Here, we showed that MET is a molecular partner of the SMYD3 methyltransferase in GC cells. Moreover, we found that SMYD3 pharmacological inhibition affects the HGF/MET downstream signaling pathway. Extensive cellular analyses in GC models indicated that EM127, a novel active site-selective covalent SMYD3 inhibitor, can be used as part of a synergistic approach with MET inhibitors in order to enhance the targeting of the HGF/MET pathway. Importantly, our data were confirmed in a 3D GC cell culture system, which was used as a surrogate to evaluate stemness characteristics. Our findings identify SMYD3 as a promising therapeutic target to impair the HGF/MET pathway for the treatment of GC.

Tumor Testing and Genetic Analysis to Identify Lynch Syndrome Patients in an Italian Colorectal Cancer Cohort.

Lynch syndrome (LS) is an inherited cancer susceptibility syndrome caused by germline mutations in a DNA mismatch repair (MMR) gene or in the EPCAM gene. LS is associated with an increased lifetime risk of colorectal cancer (CRC) and other malignancies. The screening algorithm for LS patient selection is based on the identification of CRC specimens that have MMR loss/high microsatellite instability (MSI-H) and are wild-type for BRAFV600. Here, we sought to clinically and molecularly characterize patients with these features. From 2017 to 2023, 841 CRC patients were evaluated for MSI and BRAFV600E mutation status, 100 of which showed MSI-H. Of these, 70 were wild-type for BRAFV600. Among these 70 patients, 30 were genetically tested for germline variants in hereditary cancer predisposition syndrome genes. This analysis showed that 19 of these 30 patients (63.3%) harbored a germline pathogenic or likely pathogenic variant in MMR genes, 2 (6.7%) harbored a variant of unknown significance (VUS) in MMR genes, 3 (10%) harbored a VUS in other cancer-related genes, and 6 (20%) were negative to genetic testing. These findings highlight the importance of personalized medicine for tailored genetic counseling, management, and surveillance of families with LS and other hereditary cancer syndromes.

Understanding the Genetic Landscape of Pancreatic Ductal Adenocarcinoma to Support Personalized Medicine: A Systematic Review.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most fatal malignancies worldwide. While population-wide screening recommendations for PDAC in asymptomatic individuals are not achievable due to its relatively low incidence, pancreatic cancer surveillance programs are recommended for patients with germline causative variants in PDAC susceptibility genes or a strong family history. In this study, we sought to determine the prevalence and significance of germline alterations in major genes (ATM, BRCA1, BRCA2, CDKN2A, EPCAM, MLH1, MSH2, MSH6, PALB2, PMS2, STK11, TP53) involved in PDAC susceptibility. We performed a systematic review of PubMed publications reporting germline variants identified in these genes in PDAC patients. Overall, the retrieved articles included 1493 PDAC patients. A high proportion of these patients (n = 1225/1493, 82%) were found to harbor alterations in genes (ATM, BRCA1, BRCA2, PALB2) involved in the homologous recombination repair (HRR) pathway. Specifically, the remaining PDAC patients were reported to carry alterations in genes playing a role in other cancer pathways (CDKN2A, STK11, TP53; n = 181/1493, 12.1%) or in the mismatch repair (MMR) pathway (MLH1, MSH2, MSH6, PMS2; n = 87/1493, 5.8%). Our findings highlight the importance of germline genetic characterization in PDAC patients for better personalized targeted therapies, clinical management, and surveillance.

A Low Glycemic Index Mediterranean Diet Combined with Aerobic Physical Activity Rearranges the Gut Microbiota Signature in NAFLD Patients.

Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease, and its prevalence worldwide is increasing. Several studies support the pathophysiological role of the gut-liver axis, where specific signal pathways are finely tuned by intestinal microbiota both in the onset and progression of NAFLD. In the present study, we investigate the impact of different lifestyle interventions on the gut microbiota composition in 109 NAFLD patients randomly allocated to six lifestyle intervention groups: Low Glycemic Index Mediterranean Diet (LGIMD), aerobic activity program (ATFIS_1), combined activity program (ATFIS_2), LGIMD plus ATFIS_1 or ATFIS2 and Control Diet based on CREA-AN (INRAN). The relative abundances of microbial taxa at all taxonomic levels were explored in all the intervention groups and used to cluster samples based on a statistical approach, relying both on the discriminant analysis of principal components (DAPCs) and on a linear regression model. Our analyses reveal important differences when physical activity and the Mediterranean diet are merged as treatment and allow us to identify the most statistically significant taxa linked with liver protection. These findings agree with the decreased 'controlled attenuation parameter' (CAP) detected in the LGIMD-ATFIS_1 group, measured using FibroScan®. In conclusion, our study demonstrates the synergistic effect of lifestyle interventions (diet and/or physical activity programs) on the gut microbiota composition in NAFLD patients.

Identification and Somatic Characterization of the Germline PTEN Promoter Variant rs34149102 in a Family with Gastrointestinal and Breast Tumors.

Genetic variants located in non-coding regions can affect processes that regulate protein expression, functionally contributing to human disease. Germline heterozygous mutations in the non-coding region of the PTEN gene have been previously identified in patients with PTEN hamartoma tumor syndrome (PHTS) diagnosed with breast, thyroid, and/or endometrial cancer. In this study, we report a PTEN promoter variant (rs34149102 A allele) that was identified by direct sequencing in an Italian family with a history of gastroesophageal junction (GEJ) adenocarcinoma and breast cancer. In order to investigate the putative functional role of the rs34149102 A allele variant, we evaluated the status of PTEN alterations at the somatic level. We found that PTEN protein expression was absent in the GEJ adenocarcinoma tissue of the index case. Moreover, we detected the occurrence of copy number loss involving the PTEN rs34149102 major C allele in tumor tissue, revealing that the second allele was somatically inactivated. This variant is located within an active regulatory region of the PTEN core promoter, and in silico analysis suggests that it may affect the binding of the nuclear transcription factor MAZ and hence PTEN expression. Overall, these results reveal the functional role of the PTEN promoter rs34149102 A allele variant in the modulation of PTEN protein expression and highlight its contribution to hereditary cancer risk.

c-MYC Protein Stability Is Sustained by MAPKs in Colorectal Cancer.

c-MYC is one of the most important factors involved in colorectal cancer (CRC) initiation and progression; indeed, it is found to be upregulated in up to 80% of sporadic cases. During colorectal carcinogenesis, c-MYC is maintained upregulated through ß-catenin-mediated transcriptional activation and ERK-mediated post-translational stabilization. Our data demonstrate that p38α, a kinase involved in CRC metabolism and survival, contributes to c-Myc protein stability. Moreover, we show that p38α, like ERK, stabilizes c-MYC protein levels by preventing its ubiquitination. Of note, we found that p38α phosphorylates c-MYC and interacts with it both in vitro and in cellulo. Extensive molecular analyses in the cellular and in vivo models revealed that the p38α kinase inhibitors, SB202190 and ralimetinib, affect c-MYC protein levels. Ralimetinib also exhibited a synthetic lethality effect when used in combination with the MEK1 inhibitor trametinib. Overall, our findings identify p38α as a promising therapeutic target, acting directly on c-MYC, with potential implications for countering c-MYC-mediated CRC proliferation, metastatic dissemination, and chemoresistance.

Investigation of modifier genes within copy number variations in Rett syndrome.

MECP2 mutations are responsible for two different phenotypes in females, classical Rett syndrome and the milder Zappella variant (Z-RTT). We investigated whether copy number variants (CNVs) may modulate the phenotype by comparison of array-CGH data from two discordant pairs of sisters and four additional discordant pairs of unrelated girls matched by mutation type. We also searched for potential MeCP2 targets within CNVs by chromatin immunopreceipitation microarray (ChIP-chip) analysis. We did not identify one major common gene/region, suggesting that modifiers may be complex and variable between cases. However, we detected CNVs correlating with disease severity that contain candidate modifiers. CROCC (1p36.13) is a potential MeCP2 target, in which a duplication in a Z-RTT and a deletion in a classic patient were observed. CROCC encodes a structural component of ciliary motility that is required for correct brain development. CFHR1 and CFHR3, on 1q31.3, may be involved in the regulation of complement during synapse elimination, and were found to be deleted in a Z-RTT but duplicated in two classic patients. The duplication of 10q11.22, present in two Z-RTT patients, includes GPRIN2, a regulator of neurite outgrowth and PPYR1, involved in energy homeostasis. Functional analyses are necessary to confirm candidates and to define targets for future therapies.

p53 Arg72Pro and MDM2 309 SNPs in hereditary retinoblastoma.

The tumor suppressor p53 and its negative regulator MDM2 have crucial roles in a variety of cellular functions such as the control of the cell cycle, senescence, genome stability and apoptosis, and are frequently deregulated in carcinogenesis. Previous studies have highlighted the contribution of the common functional polymorphisms p53 p.Arg72Pro and MDM2 309SNP to the risk of both common cancers and Li-Fraumeni syndrome. Their possible role in retinoblastoma has recently been addressed by Castéra et al, who however only studied the MDM2 309SNP. Here, for the first time, we analyzed both single nucleotide polymorphisms (SNPs) in a case-control study of 111 Italian hereditary retinoblastoma patients. We found a significant association of the p53 Pro/Pro genotype with the disease (odds ratio=3.58, P=0.002). The MDM2 309SNP showed a weak negative association of allele G that deserves further investigation. These findings further support the hypothesis that genetic variability of the p53 pathway contributes to the individual susceptibility to retinoblastoma, as shown for Li-Fraumeni syndrome and a variety of non-hereditary cancers.

APC Splicing Mutations Leading to In-Frame Exon 12 or Exon 13 Skipping Are Rare Events in FAP Pathogenesis and Define the Clinical Outcome.

Familial adenomatous polyposis (FAP) is caused by germline mutations in the tumor suppressor gene APC. To date, nearly 2000 APC mutations have been described in FAP, most of which are predicted to result in truncated protein products. Mutations leading to aberrant APC splicing have rarely been reported. Here, we characterized a novel germline heterozygous splice donor site mutation in APC exon 12 (NM_000038.5: c.1621_1626+7del) leading to exon 12 skipping in an Italian family with the attenuated FAP (AFAP) phenotype. Moreover, we performed a literature meta-analysis of APC splicing mutations. We found that 119 unique APC splicing mutations, including the one described here, have been reported in FAP patients, 69 of which have been characterized at the mRNA level. Among these, only a small proportion (9/69) results in an in-frame protein, with four mutations causing skipping of exon 12 or 13 with loss of armadillo repeat 2 (ARM2) and 3 (ARM3), and five mutations leading to skipping of exon 5, 7, 8, or (partially) 9 with loss of regions not encompassing known functional domains. The APC splicing mutations causing skipping of exon 12 or 13 considered in this study cluster with the AFAP phenotype and reveal a potential molecular mechanism of pathogenesis in FAP disease.

Pharmacological targeting of the novel ß-catenin chromatin-associated kinase p38α in colorectal cancer stem cell tumorspheres and organoids.

The prognosis of locally advanced colorectal cancer (CRC) is currently unsatisfactory. This is mainly due to drug resistance, recurrence, and subsequent metastatic dissemination, which are sustained by the cancer stem cell (CSC) population. The main driver of the CSC gene expression program is Wnt signaling, and previous reports indicate that Wnt3a can activate p38 MAPK. Besides, p38 was shown to feed into the canonical Wnt/ß-catenin pathway. Here we show that patient-derived locally advanced CRC stem cells (CRC-SCs) are characterized by increased expression of p38α and are "addicted" to its kinase activity. Of note, we found that stage III CRC patients with high p38α levels display reduced disease-free and progression-free survival. Extensive molecular analysis in patient-derived CRC-SC tumorspheres and APCMin/+ mice intestinal organoids revealed that p38α acts as a ß-catenin chromatin-associated kinase required for the regulation of a signaling platform involved in tumor proliferation, metastatic dissemination, and chemoresistance in these CRC model systems. In particular, the p38α kinase inhibitor ralimetinib, which has already entered clinical trials, promoted sensitization of patient-derived CRC-SCs to chemotherapeutic agents commonly used for CRC treatment and showed a synthetic lethality effect when used in combination with the MEK1 inhibitor trametinib. Taken together, these results suggest that p38α may be targeted in CSCs to devise new personalized CRC treatment strategies.

Targeting SMYD3 to Sensitize Homologous Recombination-Proficient Tumors to PARP-Mediated Synthetic Lethality.

SMYD3 is frequently overexpressed in a wide variety of cancers. Indeed, its inactivation reduces tumor growth in preclinical in vivo animal models. However, extensive characterization in vitro failed to clarify SMYD3 function in cancer cells, although confirming its importance in carcinogenesis. Taking advantage of a SMYD3 mutant variant identified in a high-risk breast cancer family, here we show that SMYD3 phosphorylation by ATM enables the formation of a multiprotein complex including ATM, SMYD3, CHK2, and BRCA2, which is required for the final loading of RAD51 at DNA double-strand break sites and completion of homologous recombination (HR). Remarkably, SMYD3 pharmacological inhibition sensitizes HR-proficient cancer cells to PARP inhibitors, thereby extending the potential of the synthetic lethality approach in human tumors.