RESUMO
In eukaryotes, the leading strand DNA is synthesized by Polε and the lagging strand by Polδ. These replicative polymerases have higher processivity when paired with the DNA clamp PCNA. While the structure of the yeast Polε catalytic domain has been determined, how Polε interacts with PCNA is unknown in any eukaryote, human or yeast. Here we report two cryo-EM structures of human Polε-PCNA-DNA complex, one in an incoming nucleotide bound state and the other in a nucleotide exchange state. The structures reveal an unexpected three-point interface between the Polε catalytic domain and PCNA, with the conserved PIP (PCNA interacting peptide)-motif, the unique P-domain, and the thumb domain each interacting with a different protomer of the PCNA trimer. We propose that the multi-point interface prevents other PIP-containing factors from recruiting to PCNA while PCNA functions with Polε. Comparison of the two states reveals that the finger domain pivots around the [4Fe-4S] cluster-containing tip of the P-domain to regulate nucleotide exchange and incoming nucleotide binding.
Assuntos
Microscopia Crioeletrônica , Antígeno Nuclear de Célula em Proliferação , Antígeno Nuclear de Célula em Proliferação/metabolismo , Antígeno Nuclear de Célula em Proliferação/química , Humanos , Holoenzimas/química , Holoenzimas/metabolismo , Modelos Moleculares , DNA Polimerase II/metabolismo , DNA Polimerase II/química , DNA Polimerase II/genética , DNA/metabolismo , DNA/química , Ligação Proteica , Domínio Catalítico , Replicação do DNA , Proteínas de Ligação a Poli-ADP-RiboseRESUMO
Patients carrying mutations in polymerase epsilon/polymerase delta have shown positive responses to immune checkpoint inhibitors. Yet, prospective trials exploring the efficacy in those with polymerase epsilon/polymerase delta mutations are still lacking. A phase II clinical trial was initiated to evaluate the efficacy of toripalimab, a humanized IgG4K monoclonal antibody to human PD-1, in patients with advanced solid tumors with unselected polymerase epsilon/polymerase delta mutations but without microsatellite instability-high. A total of 15 patients were enrolled, 14 of whom were assessed for treatment efficacy. There was a 21.4% overall response rate, with a disease control rate of 57.1%. The median overall survival and median progression-free survival were 17.9 (95% CI 13.5-not reach) months and 2.5 (95% CI 1.4-not reach) months, respectively. For patients with exonuclease domain mutations, the objective response rate was 66.7% (2/3), with a disease control rate of 66.7% (2/3). For those with non-exonuclease domain mutations, the rates were 9.1% (1/11) and 54.5% (6/11), respectively. Notably, patients with PBRM1 gene mutations exhibited a high response rate to toripalimab at 75.0% (3/4). This study showed that neither the exonuclease domain mutations nor non-exonuclease domain mutations could fully predict the efficacy of immunotherapy, urging the need for more investigations to clarify potential immune sensitization differences within polymerase epsilon/polymerase delta mutation variants.
Assuntos
Anticorpos Monoclonais Humanizados , DNA Polimerase II , Mutação , Neoplasias , Humanos , Anticorpos Monoclonais Humanizados/uso terapêutico , Feminino , Masculino , Pessoa de Meia-Idade , Idoso , Neoplasias/genética , Neoplasias/tratamento farmacológico , DNA Polimerase II/genética , DNA Polimerase III/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Adulto , Idoso de 80 Anos ou maisRESUMO
Colorectal cancer (CRC) is one of the most common malignant carcinoma worldwide. DNA polymerase epsilon 2, accessory subunit (POLE2) participates in DNA replication, repair, and cell cycle control, but its association with CRC development remains unclear. In the present study, the differentially expressed genes (DEGs) in CRC were screened from bioinformatics analysis based on GEO database. RT-qPCR was used to assess mRNA expression. CCK-8 and colony formation assays were applied for the evaluation of cell proliferation. Wound healing and transwell assays were used to detect cell migration and invasion. Protein levels were determined by Western blotting assay. We found that POLE2 was highly expressed in CRC tissues and cell lines. Inhibition of POLE2 suppressed the proliferation, migration and invasion of CRC cells. Mechanistically, Wnt/ß-catenin signaling pathway was inactivated by inhibition of POLE2. Activation of Wnt/ß-catenin pathway can reverse the function of POLE2 knockdown on CRC cells. In vivo studies demonstrated that POLE2 silencing could notably inhibit the growth of tumors, which was consistent with the results in vitro. In conclusion, we found POLE2 as a novel oncogene in CRC, providing a potential therapeutic or diagnostic target in CRC.
Assuntos
Proliferação de Células , Neoplasias Colorretais , Via de Sinalização Wnt , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Camundongos , Animais , Movimento Celular , Linhagem Celular Tumoral , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Camundongos Nus , Inativação Gênica , DNA Polimerase II/genética , DNA Polimerase II/metabolismoRESUMO
An organism's genomic DNA must be accurately duplicated during each cell cycle. DNA synthesis is catalysed by DNA polymerase enzymes, which extend nucleotide polymers in a 5' to 3' direction. This inherent directionality necessitates that one strand is synthesised forwards (leading), while the other is synthesised backwards discontinuously (lagging) to couple synthesis to the unwinding of duplex DNA. Eukaryotic cells possess many diverse polymerases that coordinate to replicate DNA, with the three main replicative polymerases being Pol α, Pol δ and Pol ε. Studies conducted in yeasts and human cells utilising mutant polymerases that incorporate molecular signatures into nascent DNA implicate Pol ε in leading strand synthesis and Pol α and Pol δ in lagging strand replication. Recent structural insights have revealed how the spatial organization of these enzymes around the core helicase facilitates their strand-specific roles. However, various challenging situations during replication require flexibility in the usage of these enzymes, such as during replication initiation or encounters with replication-blocking adducts. This review summarises the roles of the replicative polymerases in bulk DNA replication and explores their flexible and dynamic deployment to complete genome replication. We also examine how polymerase usage patterns can inform our understanding of global replication dynamics by revealing replication fork directionality to identify regions of replication initiation and termination.
Assuntos
Replicação do DNA , Humanos , DNA/metabolismo , DNA/biossíntese , DNA Polimerase Dirigida por DNA/metabolismo , Animais , DNA Polimerase II/metabolismo , Eucariotos/enzimologia , Eucariotos/genética , DNA Polimerase III/metabolismo , Células Eucarióticas/metabolismo , Células Eucarióticas/enzimologia , DNA Polimerase I/metabolismoRESUMO
The molecular classification of endometrial carcinoma defines four main groups: polymeraseÉ(PolE) gene mutated, microsatellite unstable (MSI), p53 abnormal tumors and tumors with no specific molecular profile (NSMP). This classification provides significant insights into the prognosis and therapeutic decisions. Each group exhibits unique genetic profiles identified through immunohistochemistry and molecular diagnostics, enabling personalized treatment. The identification of these molecular signatures necessitates precise analytical methods, selected based on the local circumstances at each site. The approach to molecular classification highlights the critical role of pathology in the diagnosis and emphasizes the necessity of collaboration between the clinic and pathology.
Assuntos
Neoplasias do Endométrio , Instabilidade de Microssatélites , Humanos , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/classificação , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/diagnóstico , Feminino , Mutação , Imuno-Histoquímica , Biomarcadores Tumorais/genética , Proteína Supressora de Tumor p53/genética , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genéticaRESUMO
Among the 4 molecular subgroups of endometrial carcinoma, the p53 abnormal (copy number high) subgroup has the worst prognosis; however, the histologic characteristics of this subgroup are not well established. Also, it is not well established whether low-grade tumors can belong to the p53 abnormal molecular subgroup and if so, what is the prognostic significance of the p53-mutated molecular subgroup in low-grade tumors. In the current study, we included 146 p53-mutated endometrial carcinomas and performed molecular subgrouping either based on a combination of immunohistochemical studies for p53 and MMR protein expression and POLE mutation testing (81 cases) or based on array-based and sequencing-based technologies (65 cases). We excluded cases that belonged to the POLE mutant or MSI molecular subgroups and only studied p53 abnormal (molecular subgroup) endometrial carcinomas (125 cases). In 71 cases, the molecular subgroup was determined by a combination of immunohistochemical studies and POLE mutation testing, and in 54 cases by array-based and sequencing-based methods. We reviewed 1 to 2 representative digital slides from each case and recorded the morphologic characteristics as well as clinical, treatment, and survival follow-up data. Overall, 47 cases were classified as endometrioid carcinoma, 55 serous carcinoma, and 23 other histotypes. Eight cases were FIGO 1, 21 were FIGO 2, and 91 were FIGO 3. A significant proportion of the cases (24.2%) were histologically classified as low-grade (FIGO 1 or 2) endometrioid carcinoma. There was no morphologic characteristic that showed prognostic implication. There was no significant difference in survival among different histotypes (P=0.60). There was no significant difference in survival among low-grade endometrioid (FIGO 1 or 2) versus high-grade (FIGO 3) tumors (P=0.98). Early-stage (stage I), low-grade tumors showed no significant survival advantage over early-stage, high-grade tumors (P=0.16) and this was more evident in FIGO 2 tumors. Although not statistically significant, the FIGO 2 tumors showed a trend toward worse survival than FIGO 3 tumors. Among the cases with available treatment data, more patients with early-stage high-grade tumors received adjuvant treatment, compared to patients with early-stage low-grade tumors, possibly explaining this trend (P=0.03). In conclusion, the findings of our study suggest that low-grade p53 abnormal endometrioid endometrial carcinomas (especially FIGO 2 tumors) have an aggressive course, with a prognosis similar to high-grade tumors. Furthermore, our study suggests that patients who had early-stage low-grade p53 abnormal disease might have been undertreated because of the "low-grade" histotype.
Assuntos
Carcinoma Endometrioide , Neoplasias do Endométrio , Imuno-Histoquímica , Mutação , Proteína Supressora de Tumor p53 , Humanos , Feminino , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/diagnóstico , Neoplasias do Endométrio/mortalidade , Proteína Supressora de Tumor p53/genética , Carcinoma Endometrioide/patologia , Carcinoma Endometrioide/genética , Carcinoma Endometrioide/diagnóstico , Carcinoma Endometrioide/mortalidade , Prognóstico , Pessoa de Meia-Idade , Idoso , Cistadenocarcinoma Seroso/patologia , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/diagnóstico , Cistadenocarcinoma Seroso/mortalidade , Idoso de 80 Anos ou mais , Adulto , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genéticaRESUMO
The proliferating cell nuclear antigen (PCNA) clamp encircles DNA to hold DNA polymerases (Pols) to DNA for processivity. The Ctf18-RFC PCNA loader, a replication factor C (RFC) variant, is specific to the leading-strand Pol (Polε). We reveal here the underlying mechanism of Ctf18-RFC specificity to Polε using cryo-electron microscopy and biochemical studies. We found that both Ctf18-RFC and Polε contain specific structural features that direct PCNA loading onto DNA. Unlike other clamp loaders, Ctf18-RFC has a disordered ATPase associated with a diverse cellular activities (AAA+) motor that requires Polε to bind and stabilize it for efficient PCNA loading. In addition, Ctf18-RFC can pry prebound Polε off of DNA, then load PCNA onto DNA and transfer the PCNA-DNA back to Polε. These elements in both Ctf18-RFC and Polε provide specificity in loading PCNA onto DNA for Polε.
Assuntos
Replicação do DNA , Antígeno Nuclear de Célula em Proliferação , Proteína de Replicação C , Humanos , ATPases Associadas a Diversas Atividades Celulares/metabolismo , ATPases Associadas a Diversas Atividades Celulares/química , Microscopia Crioeletrônica , DNA/química , DNA/metabolismo , DNA Polimerase II/metabolismo , DNA Polimerase II/química , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/química , Proteínas Nucleares , Antígeno Nuclear de Célula em Proliferação/metabolismo , Antígeno Nuclear de Célula em Proliferação/química , Ligação Proteica , Proteína de Replicação C/metabolismo , Proteína de Replicação C/química , Domínios ProteicosRESUMO
Genetic sequencing has revolutionized immunotherapy in colorectal cancer (CRC). Recent clinical trials have revealed a positive response to immunotherapy-based systemic therapies in CRC patient subgroups with microsatellite instability (MSI)-High or DNA polymerase epsilon (POLE) mutation. However, the unsatisfactory response rates was the major limitation in real-world practice of the precision immunotherapy in CRC. Adding photodynamic therapy (PDT) to systemic immunotherapy has showed synergetic anti-tumor effect by modulating tumor microenvironment, while the eligible patient's subgroups which would benefit from this combination remained equivocal. Here we reported a synchronous colorectal cancer patient with MSI-High and POLE mutation who had accelerated response in less than 2 cycles (42 days) of immunotherapy-based systemic therapies after tumor-directed PDT and has remained progression-free by far. This case enlightened the synergetic effect of PDT in immunotherapy-treated CRC patients, with the MSI and POLE-mutation status as predictors of survival benefits.
Assuntos
Neoplasias Colorretais , DNA Polimerase II , Imunoterapia , Instabilidade de Microssatélites , Mutação , Fotoquimioterapia , Proteínas de Ligação a Poli-ADP-Ribose , Humanos , Neoplasias Colorretais/terapia , Neoplasias Colorretais/genética , Fotoquimioterapia/métodos , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Imunoterapia/métodos , Terapia Combinada , Masculino , Resultado do Tratamento , Neoplasias Primárias Múltiplas/terapia , Neoplasias Primárias Múltiplas/genética , Pessoa de Meia-Idade , FemininoAssuntos
Neoplasias Colorretais , DNA Polimerase III , DNA Polimerase II , Inibidores de Checkpoint Imunológico , Proteínas de Ligação a Poli-ADP-Ribose , Humanos , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Inibidores de Checkpoint Imunológico/uso terapêutico , DNA Polimerase II/genética , DNA Polimerase II/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/genética , DNA Polimerase III/genética , DNA Polimerase III/metabolismo , MutaçãoRESUMO
The clinical success of PARP1/2 inhibitors (PARPi) prompts the expansion of their applicability beyond homologous recombination deficiency. Here, we demonstrate that the loss of the accessory subunits of DNA polymerase epsilon, POLE3 and POLE4, sensitizes cells to PARPi. We show that the sensitivity of POLE4 knockouts is not due to compromised response to DNA damage or homologous recombination deficiency. Instead, POLE4 loss affects replication speed leading to the accumulation of single-stranded DNA gaps behind replication forks upon PARPi treatment, due to impaired post-replicative repair. POLE4 knockouts elicit elevated replication stress signaling involving ATR and DNA-PK. We find POLE4 to act parallel to BRCA1 in inducing sensitivity to PARPi and counteracts acquired resistance associated with restoration of homologous recombination. Altogether, our findings establish POLE4 as a promising target to improve PARPi driven therapies and hamper acquired PARPi resistance.
Assuntos
Proteína BRCA1 , DNA Polimerase II , Replicação do DNA , Inibidores de Poli(ADP-Ribose) Polimerases , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Humanos , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , DNA Polimerase II/metabolismo , DNA Polimerase II/genética , Replicação do DNA/efeitos dos fármacos , Dano ao DNA , Linhagem Celular Tumoral , Recombinação Homóloga/genética , Recombinação Homóloga/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genéticaRESUMO
BACKGROUND: POLE and POLD1 proofreading deficiency (POLE/D1pd) define a rare subtype of ultramutated metastatic colorectal cancer (mCRC; over 100 mut/Mb). Disease-specific data about the activity and efficacy of immune checkpoint inhibitors (ICIs) in POLE/D1pd mCRC are lacking and it is unknown whether outcomes may be different from mismatch repair-deficient (dMMR)/microsatellite instability-high (MSI-H) mCRCs treated with ICIs. PATIENTS AND METHODS: In this global study, we collected 27 patients with mCRC harboring POLE/D1 mutations leading to proofreading deficiency and treated with anti-programmed cell death-ligand 1 alone +/- anti-cytotoxic T-lymphocyte antigen-4 agents. We collected clinicopathological and genomic characteristics, response, and survival outcomes after ICIs of POLE/D1pd mCRC and compared them with a cohort of 610 dMMR/MSI-H mCRC patients treated with ICIs. Further genomic analyses were carried out in an independent cohort of 7241 CRCs to define POLE and POLD1pd molecular profiles and mutational signatures. RESULTS: POLE/D1pd was associated with younger age, male sex, fewer RAS/BRAF driver mutations, and predominance of right-sided colon cancers. Patients with POLE/D1pd mCRC showed a significantly higher overall response rate (ORR) compared to dMMR/MSI-H mCRC (89% versus 54%; P = 0.01). After a median follow-up of 24.9 months (interquartile range: 11.3-43.0 months), patients with POLE/D1pd showed a significantly superior progression-free survival (PFS) compared to dMMR/MSI-H mCRC [hazard ratio (HR) = 0.24, 95% confidence interval (CI) 0.08-0.74, P = 0.01] and superior overall survival (OS) (HR = 0.38, 95% CI 0.12-1.18, P = 0.09). In multivariable analyses including the type of DNA repair defect, POLE/D1pd was associated with significantly improved PFS (HR = 0.17, 95% CI 0.04-0.69, P = 0.013) and OS (HR = 0.24, 95% CI 0.06-0.98, P = 0.047). Molecular profiling showed that POLE/D1pd tumors have higher tumor mutational burden (TMB). Responses were observed in both subtypes and were associated with the intensity of POLE/D1pd signature. CONCLUSIONS: Patients with POLE/D1pd mCRC showed more favorable outcomes compared to dMMR/MSI-H mCRC to treatment with ICIs in terms of tumor response and survival.
Assuntos
Neoplasias Colorretais , DNA Polimerase III , DNA Polimerase II , Inibidores de Checkpoint Imunológico , Mutação , Proteínas de Ligação a Poli-ADP-Ribose , Humanos , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Masculino , Feminino , Inibidores de Checkpoint Imunológico/uso terapêutico , Pessoa de Meia-Idade , Idoso , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , DNA Polimerase III/genética , Adulto , Instabilidade de Microssatélites , Idoso de 80 Anos ou mais , Reparo de Erro de Pareamento de DNARESUMO
INTRODUCTION: The study aimed to validate the Betella algorithm, focusing on molecular analyses exclusively for endometrial cancer patients, where molecular classification alters risk assessment based on ESGO/ESTRO/ESP 2020 guidelines. MATERIALS AND METHODS: Conducted between March 2021 and March 2023, the retrospective research involved endometrial cancer patients undergoing surgery and comprehensive molecular analyses. These included p53 and mismatch repair proteins immunohistochemistry, as well as DNA sequencing for POLE exonuclease domain. We applied the Betella algorithm to our population and evaluated the proportion of patients in which the molecular analysis changed the risk class attribution. RESULTS: Out of 102 patients, 97 % obtained complete molecular analyses. The cohort exhibited varying molecular classifications: 10.1 % as POLE ultra-mutated, 30.3 % as mismatch repair deficient, 11.1 % as p53 abnormal, and 48.5 % as non-specified molecular classification. Multiple classifiers were present in 3 % of cases. Integrating molecular classification into risk group calculation led to risk group migration in 11.1 % of patients: 7 moved to lower risk classes due to POLE mutations, while 4 shifted to higher risk due to p53 alterations. Applying the Betella algorithm, we can spare the POLE sequencing in 65 cases (65.7 %) and p53 immunochemistry in 17 cases (17.2 %). CONCLUSION: In conclusion, we externally validated the Betella algorithm in our population. The application of this new proposed algorithm enables assignment of the proper risk class and, consequently, the appropriate indication for adjuvant treatment, allowing for the rationalization of the resources that can be allocated otherwise, not only for the benefit of settings with low resources, but of all settings in general.
Assuntos
Algoritmos , DNA Polimerase II , Neoplasias do Endométrio , Proteína Supressora de Tumor p53 , Humanos , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Feminino , Estudos Retrospectivos , Pessoa de Meia-Idade , Idoso , Proteína Supressora de Tumor p53/genética , DNA Polimerase II/genética , Mutação , Imuno-Histoquímica , Proteínas de Ligação a Poli-ADP-Ribose/genética , Medição de Risco/métodos , Reparo de Erro de Pareamento de DNA , Idoso de 80 Anos ou mais , Adulto , Análise de Sequência de DNA/métodosRESUMO
The advent of PARP inhibitors (PARPis) has profoundly changed the treatment landscape of BRCA1/BRCA2-mutated cancers. Despite this, the development of resistance to these compounds has become a major challenge. Hence, a detailed understanding of the mechanisms underlying PARPi sensitivity is crucially needed. Here, we show that loss of the POLE3-POLE4 subunits of DNA polymerase epsilon (Polε) strongly sensitizes cancer cells to PARPis in a Polε level-independent manner. Loss of POLE3-POLE4 is not associated with defective RAD51 foci formation, excluding a major defect in homologous recombination. On the contrary, treatment with PARPis triggers replicative gap accumulation in POLE3-POLE4 knockout (KO) cells in a PRIMPOL-dependent manner. In addition to this, the loss of POLE3-POLE4 further sensitizes BRCA1-silenced cells to PARPis. Importantly, the knockdown of 53BP1 does not rescue PARPi sensitivity in POLE3-POLE4 KO cells, bypassing a common PARPi resistance mechanism and outlining a potential strategy to sensitize cancer cells to PARPis.
Assuntos
Inibidores de Poli(ADP-Ribose) Polimerases , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Humanos , Replicação do DNA/efeitos dos fármacos , Linhagem Celular Tumoral , Proteína BRCA1/metabolismo , Proteína BRCA1/genética , DNA Polimerase II/metabolismo , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/metabolismo , Rad51 Recombinase/metabolismoRESUMO
INTRODUCTION: Pathogenic mutations in POLE/POLD1 lead to decreased fidelity of DNA replication, resulting in a high tumor mutational burden (TMB-H), defined as TMB ≥â 10 mut/Mb, independent of deficient mismatch repair (dMMR) and microsatellite instability high (MSI-H) status. METHODS: De-identified records of patients with colorectal cancer (CRC) profiled with the Tempus xT assay (DNA-seq of 595-648 genes at 500×) were identified from the Tempus Database. RESULTS: Among 9136 CRC samples profiled, the frequency of POLE/POLD1 genomic alterations was 2.4% (nâ =â 217). Copy number loss was the most common genomic alteration (64%, nâ =â 138) of POLE/POLD1, followed by copy number amplifications (18%, nâ =â 40) and short variant mutations (18%, nâ =â 39). The POLE/POLD1 mutated group presented with a higher frequency of TMB-H phenotype relative to wild type (WT; 22% vs. 9%, P <â .001), with a median TMB of 127 mut/Mb in the TMB-H POLE/POLD1 subset. The TMB showed a dramatic contrast between POLE/POLD1 short variant mutations as compared to the group with copy number alterations, with a TMB of 159 mut/Mb vs 15 mut/Mb, respectively. Thus, the short variant mutations represented the so-called ultra-hypermutated phenotype. The POLE/POLD1 mutated group, as compared to WT, exhibited a higher rate of coexisting mutations, including APC, ALK, ATM, BRCA2, and RET mutations. CONCLUSION: Patients with POLE/POLD1 mutations exhibited significant differences across immunological markers (ie, TMB, MMR, and MSI-H) and molecular co-alterations. Those with short variant mutations represented 18% of the POLE/POLD1 cohort and 0.4% of the total cohort examined. This group of patients had a median TMB of 159 mut/Mb (range 34-488), representing the ultra-hypermutated phenotype. This group of patients is important to identify given the potential for exceptional response to immune checkpoint inhibitors.
Assuntos
Neoplasias Colorretais , DNA Polimerase III , DNA Polimerase II , Mutação , Proteínas de Ligação a Poli-ADP-Ribose , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Masculino , Feminino , DNA Polimerase III/genética , Idoso , Pessoa de Meia-Idade , Idoso de 80 Anos ou mais , Adulto , Instabilidade de Microssatélites , Genômica/métodos , Variações do Número de Cópias de DNA , Biomarcadores Tumorais/genéticaAssuntos
Ciclina B1 , DNA Polimerase II , Neoplasias do Endométrio , Imuno-Histoquímica , Proteínas de Ligação a Poli-ADP-Ribose , Proteína Supressora de Tumor p53 , Humanos , Feminino , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/genética , Ciclina B1/genética , Ciclina B1/metabolismo , DNA Polimerase II/genética , DNA Polimerase II/metabolismo , Mutação , Pessoa de Meia-Idade , Idoso , AdultoRESUMO
PURPOSE: Molecular classification of endometrial cancer (EC) has become a promising information to tailor preoperatively the surgical treatment. We aimed to evaluate the rate of lymph node metastases (LNM) in patients with EC according to molecular profile. METHODS: A systematic review and meta-analysis were performed according to PRISMA guidelines by searching in two major electronic databases (PubMed and Scopus), including original articles reporting lymph node metastases according to the molecular classification of EC as categorized in the ESGO-ESMO-ESP guidelines. RESULTS: Fifteen studies enrolling 3056 patients were included. Pooled prevalence LNM when considering only patients undergoing lymph node assessment was 4% for POLE-mutated (95%CI: 0-12%), 22% for no specific molecular profile (95% CI: 9-39%), 23% for Mismatch repair-deficiency (95%CI: 10-40%) and 31% for p53-abnormal (95%CI: 24-39%). CONCLUSIONS: The presence of LNM seems to be influenced by molecular classification. P53-abnormal group presents the highest rate of nodal involvement, and POLE-mutated the lowest.
Assuntos
Neoplasias do Endométrio , Metástase Linfática , Humanos , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Feminino , Proteína Supressora de Tumor p53/genética , Mutação , Proteínas de Ligação a Poli-ADP-Ribose/genética , DNA Polimerase II/genética , Linfonodos/patologia , Biomarcadores Tumorais/genéticaRESUMO
OBJECTIVE: The Cancer Genome Atlas (TCGA) project identified favorable prognosis regarding the ultra-mutated endometrial cancer (EC) subtype linked to polymerase epsilon gene (POLE) mutations. This study investigated POLE mutations in EC of Indian patients. METHODS: This retrospective analytical study was conducted between January 2016 and January 2023 at the Government Medical College, Kozhikode, and the MVR Cancer Center, Kozhikode, Kerala. Sanger sequencing of POLE gene exons 9 and 13 in 151 EC patients was carried out to analyze the relationship between mutations and epidemiological factors, clinicopathologic features, and treatment outcomes. RESULTS: Among 151 cases enrolled, 39 were unique POLE-mutated cases. Significant associations were high-grade tumors, myometrial invasion >50%, and Lymph-vascular space invasion (LVSI). The median follow-up was 40 months (95% confidence interval [CI], 34-46). A lower mean disease-specific survival (DSS) of 51.7 months (95% CI, 43.7-59.6) was noted in the POLE-mutated group compared with 72.11 months (95% CI, 67.60-76.62) for the POLE wild-type. A statistically significant hazard ratio (HR) of 2.683 for DSS in the POLE-mutated group was noted. In advanced stages (FIGO stages II-IV), a nine-fold HR for DSS and overall survival (OS) compared with POLE wild-type was identified. After controlling for treatment effects using Cox proportional HR, advanced-stage POLE-mutated tumors had a significantly higher HR of 8.67 for DSS compared with POLE-wild-type tumors of the same stage. CONCLUSION: This study identified a unique set of POLE mutations in Indian EC patients associated with poor prognosis, which were particularly pronounced in advanced stages. Advanced stage of presentation, type of POLE mutations, and possibly ethnicity are predictors of adverse outcomes in POLE-mutated EC. The present study highlights ethnicity as a determinant of phenotypic expression of genetic change.
Assuntos
DNA Polimerase II , Neoplasias do Endométrio , Mutação , Proteínas de Ligação a Poli-ADP-Ribose , Humanos , Feminino , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/mortalidade , Índia , DNA Polimerase II/genética , Pessoa de Meia-Idade , Estudos Retrospectivos , Prognóstico , Proteínas de Ligação a Poli-ADP-Ribose/genética , Idoso , Adulto , Prevalência , Estadiamento de NeoplasiasRESUMO
OBJECTIVES: The characterization of DNA polymerase epsilon (POLE) mutations has transformed the classification of endometrial endometrioid carcinomas (EECs), highlighting the need for efficient identification methods. This study aims to examine the relationship between distinct morphologic features-namely, squamous morules and squamous differentiation (SD), as well as ß-catenin expression-and the POLE mutation status in endometrial cancer (EC). METHODS: Our study included 35 POLE-mutated (POLEmut) EC cases and 395 non-POLEmut EEC cases. RESULTS: Notably, we observed no presence of morules in POLEmut cases, while SD was identified in 20% of instances. Conversely, morules and SD were identified in 12.7% and 26.1% of non-POLEmut EC cases, respectively, with morules consistently linked to a POLE wild-type status. The nuclear ß-catenin expression is typically absent in tumors with wild-type POLE (wt-POLE) status. CONCLUSIONS: Our findings suggest that the presence of either morules or nuclear ß-catenin expression in EEC could practically rule out the presence of POLE mutations. These morphologic and immunohistochemical features can be used as preliminary screening tools for POLE mutations, offering significant savings in time and resources and potentially enhancing clinical decision-making and patient management strategies. However, further validation in larger, multi-institutional studies is required to fully understand the implications of these findings on clinical practice.
Assuntos
Carcinoma Endometrioide , DNA Polimerase II , Neoplasias do Endométrio , Mutação , Proteínas de Ligação a Poli-ADP-Ribose , beta Catenina , Humanos , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/diagnóstico , beta Catenina/genética , beta Catenina/metabolismo , Feminino , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Carcinoma Endometrioide/genética , Carcinoma Endometrioide/patologia , Carcinoma Endometrioide/diagnóstico , Pessoa de Meia-Idade , Biomarcadores Tumorais/genética , Idoso , Análise Custo-Benefício , AdultoRESUMO
INTRODUCTION: Patients with polymerase epsilon (POLE) mutation (POLEmut) subtype, MMR-deficient (MMR-d) subtype as classified by The Cancer Genome Atlas (TCGA), and a high tumor mutation burden (TMB-high) potentially benefit from immunotherapy. However, characteristics of the cytological morphology within these populations remain unknown. METHODS: DNA extracted from formalin-fixed paraffin-embedded tissues was subjected to next-generation sequencing analysis. Genomic mutations related to gynecological cancers, TMB, and microsatellite instability were analyzed and were placed in four TCGA classification types. The following morphological cytological investigations were conducted on endometrial cancer using a liquid-based preparation method, prior to the commencement of initial treatment: (i) cytological backgrounds; (ii) differences between each count of neutrophils and lymphocytes as described below. RESULTS: Insignificant differences in the cytological background patterns of TCGA groups and TMB status were found. Although there was no significant difference in neutrophil count (p = 0.955) in the TCGA groups, POLEmut and MMR-d had significantly higher lymphocyte counts than no specific molecular profile (NSMP) (p = 0.019 and 0.037, respectively); furthermore, p53mut also tended to be significant (p = 0.064). Lymphocyte counts in TMB-high were also significantly greater than TMB-low (p = 0.002). POLEmut showed a positive correlation between TMB levels and lymphocyte counts. For predicting patients with POLEmut plus MMR-d, lymphocyte counts demonstrated a superior diagnostic accuracy of area under the curve (AUC) (0.70, 95% CI: 0.57-0.84), with a cutoff value of 26 high-power field. CONCLUSION: Lymphocyte count using liquid-based cytology for patients with endometrial cancer may predict POLEmut plus MMR-d of TCGA groups and TMB-high in those who can benefit from immunotherapy.
Assuntos
Biomarcadores Tumorais , DNA Polimerase II , Neoplasias do Endométrio , Endométrio , Imunoterapia , Mutação , Humanos , Feminino , Neoplasias do Endométrio/patologia , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/terapia , Pessoa de Meia-Idade , Imunoterapia/métodos , Idoso , Biomarcadores Tumorais/genética , Endométrio/patologia , Endométrio/imunologia , DNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/genética , Neutrófilos/patologia , Adulto , Contagem de Linfócitos/métodos , Instabilidade de Microssatélites , Valor Preditivo dos Testes , Idoso de 80 Anos ou mais , Seleção de Pacientes , Análise Mutacional de DNA , Linfócitos/patologia , Tomada de Decisão Clínica , CitologiaRESUMO
Recycling of parental histones is an important step in epigenetic inheritance. During DNA replication, DNA polymerase epsilon subunit DPB3/DPB4 and DNA replication helicase subunit MCM2 are involved in the transfer of parental histones to the leading and lagging strands, respectively. Single Dpb3 deletion (dpb3Δ) or Mcm2 mutation (mcm2-3A), which each disrupts one parental histone transfer pathway, leads to the other's predominance. However, the biological impact of the two histone transfer pathways on chromatin structure and DNA repair remains elusive. In this study, we used budding yeast Saccharomyces cerevisiae to determine the genetic and epigenetic outcomes from disruption of parental histone H3-H4 tetramer transfer. We found that a dpb3Δ mcm2-3A double mutant did not exhibit the asymmetric parental histone patterns caused by a single dpb3Δ or mcm2-3A mutation, suggesting that the processes by which parental histones are transferred to the leading and lagging strands are independent. Surprisingly, the frequency of homologous recombination was significantly lower in dpb3Δ, mcm2-3A and dpb3Δ mcm2-3A mutants, likely due to the elevated levels of free histones detected in the mutant cells. Together, these findings indicate that proper transfer of parental histones during DNA replication is essential for maintaining chromatin structure and that lower homologous recombination activity due to parental histone transfer defects is detrimental to cells.