Endometrial carcinoma: 10 years of TCGA (the cancer genome atlas): A critical reappraisal with comments on FIGO 2023 staging.

The Cancer Genome Atlas (TCGA) Research Network described 4 molecular subgroups of endometrial carcinomas with different outcome: 1) POLE ultramutated endometrioid carcinomas which have an indolent behavior; 2) microsatellite instability hypermutated endometrioid carcinomas associated with intermediate prognosis; 3) copy-number low endometrioid carcinomas also with intermediate prognosis; and 4) copy-number high predominantly serous (non-endometrioid) but also serous-like endometrioid carcinomas, almost always carrying TP53 mutations, with poor clinical outcome. After 10 years of comprehensive analysis, it appears that the only real contribution of TCGA to the clinical management of these patients would be limited to the infrequent high-grade, early-stage endometrioid carcinomas with POLE exonuclease domain mutations, as these patients could benefit from a de-escalating treatment; knowledge about the other three subgroups has not changed significantly. The copy-number low (or non-specific genetic profile) which is the most frequent subgroup, is a mixture subgroup where investigators are currently trying to establish prognostic markers; for example, unexpected variations in a relatively small percentage of cases (i.e., CTNNB1 mutated or p53 aberrant low-grade and low-stage endometrioid carcinomas associated with unfavorable prognosis). On the other hand, TCGA has underlined that a small number of grade 3 endometrioid carcinomas, all TP53 mutated, overlap with copy-number high serous carcinomas. Recently, TCGA molecular subgroups have been integrated into the 2023 International Federation of Gynecology and Obstetrics (FIGO) staging classification which incorporates other non-anatomic parameters like histotype, tumor grade, and lymphovascular space invasion. The result is a complicated and non-intuitive classification that makes its clinical application difficult and does not facilitate correspondence with the 2009 FIGO staging.

Placental site trophoblastic tumor (PSTT): a case report and review of the literature.

Placental site trophoblastic tumor (PSTT), also known as atypical choriocarcinoma, syncytioma, chorioepitheliosis or trophoblastic pseudotumor, is a rare gestational trophoblastic disease (0.25-5% of all trophoblastic tumors) and it is composed by neoplastic proliferation of intermediate trophoblasts at placental implantation site. It consists of aggregates or sheets of large, polyhedral to round, predominantly mononucleated cells with a characteristic vascular and myometrial invasion. Main differential diagnoses are gestational choriocarcinoma (GC) and epitelioid trophoblastic tumor (ETT). We present a case of PSTT in a 25-year-old woman. Neoplastic cells showed moderate/high nuclear pleomorphism, abundant amphophilic, eosinophilic and clear cytoplasm, numerous mitotic figures (10 mitoses/10 HPF), and myometrial invasion. Other features are necrosis, vascular invasion with replacement of myometrial vessels by tumor cells and hemorrhage. The patient showed typical low serum ß-hCG levels and high serum humane placental lactogen (hPL) levels.

Simultaneous carcinomas of the breast and ovary: utility of Pax-8, WT-1, and GATA3 for distinguishing independent primary tumors from metastases.

Breast carcinomas rarely metastasize to the ovary and are even more rarely present clinically as primary ovarian tumors. However, patients with breast cancer not infrequently develop independent primary ovarian carcinomas. In these cases, distinction between independent primaries and metastatic tumors is crucial. Several comparative immunohistochemical studies have been reported, but few included significant clinicopathologic data and none investigated cases of ovarian and breast carcinomas from the same patients. In this study, we compared 18 cases of patients with bona fide independent breast and ovarian carcinomas (15 high-grade serous and 3 clear cell carcinomas), with 9 cases of patients with known mammary carcinomas (7 lobular and 2 ductal carcinomas) metastatic to the ovary. Immunohistochemical stains for Pax-8, WT-1, and GATA3 were carried out on tissue microarrays (TMA). Most primary ovarian carcinomas were larger than the metastatic tumors (P=0.001) and were diagnosed at an advanced stage. All primary ovarian tumors showed marked nuclear pleomorphism, whereas only 2 metastatic breast carcinomas had Grade 3 nuclei (P=0.000). The vast majority of ovarian metastases (7/9) showed the typical pattern of lobular breast carcinoma. Pax-8 and WT-1 expression were found in 16 of 18 (88%) and 13 of 18 (72%) primary ovarian carcinomas, respectively. In contrast, all primary ovarian carcinomas were negative for GATA3. The 2 Pax-8-negative ovarian carcinomas were also negative for WT-1. With the exception of 3 triple-negative carcinomas, all primary breast carcinomas were positive for GATA3. All metastatic breast carcinomas were positive for GATA3 and negative for Pax-8. WT-1 expression was seen in only 1 of 9 metastatic breast carcinomas (11%). Patients with ovarian metastases had worse prognosis than patients with independent breast and ovarian carcinomas (P=0.000). Pax-8, WT-1, and GATA3 immunoreactions are useful in the distinction between independent primaries and metastatic mammary carcinomas to the ovary in the light of clinicopathologic findings.

Stromal signatures in endometrioid endometrial carcinomas.

The pattern of myometrial invasion in endometrioid endometrial carcinomas varies considerably; ie, from widely scattered glands and cell nests, often associated with a fibromyxoid stromal reaction (desmoplasia) and/or a lymphocytic infiltrate, to invasive glands with little or no stromal response. Recently, two distinct stromal signatures derived from a macrophage response (colony-stimulating factor 1, CSF1) and a fibroblastic response (desmoid-type fibromatosis, DTF) were identified in breast carcinomas and correlated with clinicopathologic features including outcome. In this study, we explored whether these stromal signatures also apply to endometrioid carcinomas and how their expression patterns correlated with morphologic changes. We studied the stromal signatures both by immunohistochemistry and in situ hybridization in 98 primary endometrioid carcinomas with (87 cases) and without (11 cases) myometrial invasion as well as in the corresponding regional lymph nodes metatases of 9 myoinvasive tumors. Desmoplasia correlated positively with the DTF expression signature. Likewise, mononuclear infiltrates were found in the stroma of tumors expressing CSF1. Twenty-four out of eighty-seven (27%) myoinvasive endometrioid carcinomas were positive for the macrophage signature and thirteen out of eighty-seven (15%) expressed the fibroblast signature. Eleven additional cases were positive for both DTF and CSF1 signatures (11/87; 13%). However, over half of the cases (39/87; 45%) and the majority of the non-myoinvasive tumors (8/11; 73%) failed to express any of the two stromal signatures. The macrophage response (CSF1) was associated with higher tumor grade, lymphovascular invasion, and PIK3CA mutations (P<0.05). There was a concordance in the expression of the CSF1 signature in the primary tumors and their corresponding lymph node metastases. This study is the first characterization of stromal signatures in endometrioid carcinomas. Our findings shed new light on the relationship between genetically different endometrioid carcinomas and various stromal responses. Preservation of the CSF1 macrophage stromal response in the metastases leds support to targeting the CSF1 pathway in endometrioid endometrial carcinomas.

Focal adhesion protein expression in human diffuse large B-cell lymphoma.

AIMS: Focal adhesions have been associated with poor prognosis in multiple cancer types, but their prognostic value in diffuse large B-cell lymphoma (DLBCL) has not been evaluated. The aim of this study was to investigate the expression patterns and the prognostic value of the focal adhesion proteins FAK, Pyk2, p130Cas and HEF1 in DLBCL. METHODS AND RESULTS: Focal adhesion protein expression was examined using immunohistochemistry in normal lymphoid tissues and in 60 DLBCL patient samples. Kaplan-Meier survival and Cox regression analysis were performed to evaluate the correlation of focal adhesion protein expression with patient prognosis. FAK, Pyk2, p130Cas and HEF1 expression was mostly found in the germinal centres of normal human lymphoid tissues. When assessed in DLBCL samples, FAK, Pyk2, p130Cas and HEF1 were highly expressed in 45%, 34%, 42% and 45% of the samples, respectively. Multivariate Cox analysis revealed that decreased FAK expression was a significant independent predictor of poorer disease outcome. CONCLUSIONS: FAK expression is an independent prognostic factor in DLBCL. Our results suggest that the addition of FAK immunostaining to the current immunohistochemical algorithms may facilitate risk stratification of DLBCL patients.

Promoter hypermethylation contributes to TIMP3 down-regulation in high stage endometrioid endometrial carcinomas.

AIMS: Expression of tissue inhibitor of metalloproteinases-3 (TIMP-3) has been found to be decreased in several types of cancer by promoter gene hypermethylation. However, little is known regarding the silencing effect of TIMP3 promoter hypermethylation on gene and protein expression in endometrial carcinomas and its prognostic significance. METHODS AND RESULTS: TIMP3 promoter hypermethylation and gene copy number variations were evaluated using a methylation-specific multiplex ligation-dependent probe amplification approach in 60 cases of endometrioid endometrial carcinomas. TIMP3 expression was also evaluated at the transcript and protein levels. Loss of TIMP-3 protein expression was found in 44 (73%) of 60 carcinomas. Promoter hypermethylation was identified in 25% (15 of 60); was more frequent in stages II-IV (55%, six of 11) than in stage I (18%, nine of 49; P = 0.021); and was found more commonly in tumours with deep myometrial invasion. MLH1 and TIMP3 promoters were hypermethylated simultaneously in the same group of tumours (P < 0.001). A correlation between TIMP3 methylation and microsatellite instability (MSI) was found (P = 0.005). TIMP3 copy number changes were frequently a loss (35%), whereas a gain was detected in only 5%. CONCLUSIONS: TIMP3 promoter hypermethylation was associated with high stage endometrioid endometrial tumours with extrauterine spread. Nevertheless, promoter hypermethylation and loss of heterozygosity are not the only mechanisms for TIMP3 inactivation.

MYC protein expression and genetic alterations have prognostic impact in patients with diffuse large B-cell lymphoma treated with immunochemotherapy.

MYC alterations influence the survival of patients with diffuse large B-cell lymphoma. Most studies have focused on MYC translocations but there is little information regarding the impact of numerical alterations and protein expression. We analyzed the genetic alterations and protein expression of MYC, BCL2, BCL6, and MALT1 in 219 cases of diffuse large B-cell lymphoma. MYC rearrangement occurred as the sole abnormality (MYC single-hit) in 3% of cases, MYC and concurrent BCL2 and/or BCL6 rearrangements (MYC double/triple-hit) in 4%, MYC amplifications in 2% and MYC gains in 19%. MYC single-hit, MYC double/triple-hit and MYC amplifications, but not MYC gains or other gene rearrangements, were associated with unfavorable progression-free survival and overall survival. MYC protein expression, evaluated using computerized image analysis, captured the unfavorable prognosis of MYC translocations/amplifications and identified an additional subset of patients without gene alterations but with similar poor prognosis. Patients with tumors expressing both MYC/BCL2 had the worst prognosis, whereas those with double-negative tumors had the best outcome. High MYC expression was associated with shorter overall survival irrespectively of the International Prognostic Index and BCL2 expression. In conclusion, MYC protein expression identifies a subset of diffuse large B-cell lymphoma with very poor prognosis independently of gene alterations and other prognostic parameters.

ROR2 is a novel prognostic biomarker and a potential therapeutic target in leiomyosarcoma and gastrointestinal stromal tumour.

Soft-tissue sarcomas are a group of malignant tumours whose clinical management is complicated by morphological heterogeneity, inadequate molecular markers and limited therapeutic options. Receptor tyrosine kinases (RTKs) have been shown to play important roles in cancer, both as therapeutic targets and as prognostic biomarkers. An initial screen of gene expression data for 48 RTKs in 148 sarcomas showed that ROR2 was expressed in a subset of leiomyosarcoma (LMS), gastrointestinal stromal tumour (GIST) and desmoid-type fibromatosis (DTF). This was further confirmed by immunohistochemistry (IHC) on 573 tissue samples from 59 sarcoma tumour types. Here we provide evidence that ROR2 expression plays a role in the invasive abilities of LMS and GIST cells in vitro. We also show that knockdown of ROR2 significantly reduces tumour mass in vivo using a xenotransplantation model of LMS. Lastly, we show that ROR2 expression, as measured by IHC, predicts poor clinical outcome in patients with LMS and GIST, although it was not independent of other clinico-pathological features in a multivariate analysis, and that ROR2 expression is maintained between primary tumours and their metastases. Together, these results show that ROR2 is a useful prognostic indicator in the clinical management of these soft-tissue sarcomas and may represent a novel therapeutic target.

Malignant rhabdoid tumors of the vulva versus epithelioid sarcomas: a clinicopathologic, immunohistochemical, and molecular genetics study.

It has been suggested that most, if not all, extrarenal rhabdoid tumors of the vulva represent "proximal-type" epithelioid sarcomas. To better understand rhabdoid tumors of the vulva, we studied the clinicopathologic, immunohistochemical (IHC), and molecular features of 8 of these tumors and 13 extragenital epithelioid sarcomas. IHC analysis for cytokeratin AE1/AE3, EMA, S100, CD34, ERG, smooth muscle actin, desmin, and SMARCB1 (INI1) was performed. Ultrastructural study was done in one vulvar rhabdoid tumor. Next-generation sequencing of the SMARCB1 gene was performed in all cases. The 8 vulvar tumors occurred in adult women (mean age, 49 years). They were poorly differentiated neoplasms with a rhabdoid morphology. The ultrastructural study showed large amounts of intermediate filaments (10 nm). All cases had loss of expression of INI1 and were negative for CD34 and ERG. One case showed 2 SMARCB1 mutations: c.592C>T in exon 5 and c.782delG in exon 6. Follow-up revealed that 4 patients died of disease, 1 was alive with disease, and 3 were alive without evidence of disease. Epithelioid sarcomas occurred in young adults (mean age, 41 years), mostly men. Seven tumors arose in the distal extremities and the other 6 had a proximal location. They showed the characteristic "granulomatous" arrangement of the neoplastic cells. The recurrent tumors were more proximal and often showed a rhabdoid morphology. All cases had loss of expression of INI1. CD34 and ERG were expressed by 8 (62%) and 5 (38%) tumors, respectively. No SMARCB1 mutations were encountered. Follow-up revealed that 5 patients died of disease, 1 was alive with disease, and 7 were alive without evidence of disease. Based on their different morphology and biological behavior, we conclude that rhabdoid tumors of the vulva and epithelioid sarcomas are different diseases with distinct clinicopathologic features. Undifferentiated vulvar tumors with rhabdoid morphology should be classified as malignant rhabdoid tumors, rather than "proximal-type" epithelioid sarcomas.

Ovarian high-grade serous carcinoma with transitional-like (SET) morphology: a homologous recombination-deficient tumor.

Thirteen years ago, we pointed out that ovarian transitional cell carcinomas (TCCs) and conventional high-grade serous carcinomas (HGSCs) had similar genetic alterations and clinical behavior. Consequently, ovarian TCC is now classified as a morphologic variant of HGSC. Defective homologous recombination, resulting from genetic or epigenetic inactivation of DNA damage repair genes, such as BRCA1/2, occurs in approximately 50% of the HGSCs. Although BRCA mutations have been associated with HGSCs with solid, pseudoendometrioid or transitional (SET) features, little is known about the role of non-BRCA homologous recombinationrepair (HRR) genes and the HRR status in these tumors. Using two commercially available assays (Myriad Genetics MyChoice CDx Plus test and SOPHiA Dx Homologous Recombination Deficiency Solution), we study mutations of BRCA1/2 and non-BRCA HRR genes (ATM, BARD1, BRIP1, CDK12, CHEK1/2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L), and the HRR status in 19 HGSCs with SET features and 20 HGSCs with classic morphology. We also studied, as control cases, 5 endometrioid carcinomas, 1 clear cell carcinoma, 2 low-grade serous carcinomas, and 1 malignant Brenner tumor. Seven HGSCs with SET features (7/19; 37%) showed BRCA mutations (4 BRCA1, 2 BRCA2, and 1 BRCA1/2). Mutations in non-BRCA HRR genes were found in ATM (1/15; 7%), BARD1 (1/15; 7%), and BRIP1 (1/19; 5%). Most HGSCs with SET features (17/19; 90%) were considered to be homologous recombination-deficient tumors. Three HGSCs with classic morphology (3/20; 15%) showed BRCA2 mutations. Mutations in non-BRCA HRR genes were found in CDK12 (2/14; 14%), FANCL (1/14; 7%), RAD51B (1/14; 7%), and RAD54L (1/14; 7%). Eleven HGSCs with classical morphology (11/20; 55%) were considered to be homologous recombination deficient. In contrast, all ovarian carcinoma control cases (5 endometrioid carcinomas, 1 clear cell carcinoma, 2 low-grade serous carcinomas, and 1 malignant Brenner tumor) were homologous recombination proficient and did not have BRCA mutations. Our results show that the majority of HGSCs with SET features are homologous recombination-deficient tumors independently of the BRCA status and highlight the importance of the HRR tumor testing, especially in BRCA wild-type tumors. Recognition of transitional cell variant of HGSCs may help to identify patients most likely to benefit from PARP inhibitors.

CSF1 expression in nongynecological leiomyosarcoma is associated with increased tumor angiogenesis.

Leiomyosarcoma (LMS) is a malignant tumor of smooth muscle cells for which few effective therapies exist. A subset of LMS cases express macrophage colony-stimulating factor (CSF1) and the resultant tumor-associated macrophage (TAM) infiltration predicts poor clinical outcome. Further, TAMs have been shown to increase tumor angiogenesis. Here, we analyzed 149 LMS cases by immunohistochemistry for vascular marker CD34 and show that high microvessel density (MVD) in nongynecological LMS cases significantly predicts poor patient outcome. The majority of high MVD cases were also CSF1-positive, and when combining high MVD with CSF1 expression, an even stronger prognostic correlation with patient outcome was obtained. Gene expression profiling revealed that MVD has a stronger correlation with CSF1 expression than with expression of vascular endothelial growth factor isoforms, which have traditionally been used as markers of angiogenesis and as anti-angiogenic therapeutic targets. Finally, patterns of CSF1 expression and TAM recruitment remained consistent between primary tumors and their metastases, and between primary tumors and those grown as xenografts in mice, highlighting the stability of these features to the biology of LMS tumors. Together, these findings suggest an important role for CSF1 and the resulting TAM infiltration in the pathological neovascularization of LMS tumors and provide a rationale for CSF1-targeted therapies in LMS.

Identification, molecular characterization, clinical prognosis, and therapeutic targeting of human bladder tumor-initiating cells.

Major clinical issues in bladder cancer include the identification of prediction markers and novel therapeutic targets for invasive bladder cancer. In the current study, we describe the isolation and characterization of a tumor-initiating cell (T-IC) subpopulation in primary human bladder cancer, based on the expression of markers similar to that of normal bladder basal cells (Lineage-CD44(+)CK5(+)CK20(-)). The bladder T-IC subpopulation was defined functionally by its enriched ability to induce xenograft tumors in vivo that recapitulated the heterogeneity of the original tumor. Further, molecular analysis of more than 300 bladder cancer specimens revealed heterogeneity among activated oncogenic pathways in T-IC (e.g., 80% Gli1, 45% Stat3, 10% Bmi-1, and 5% beta-catenin). Despite this molecular heterogeneity, we identified a unique bladder T-IC gene signature by gene chip analysis. This T-IC gene signature, which effectively distinguishes muscle-invasive bladder cancer with worse clinical prognosis from non-muscle-invasive (superficial) cancer, has significant clinical value. It also can predict the progression of a subset of recurring non-muscle-invasive cancers. Finally, we found that CD47, a protein that provides an inhibitory signal for macrophage phagocytosis, is highly expressed in bladder T-ICs compared with the rest of the tumor. Blockade of CD47 by a mAb resulted in macrophage engulfment of bladder cancer cells in vitro. In summary, we have identified a T-IC subpopulation with potential prognostic and therapeutic value for invasive bladder cancer.

Prognostic significance of FOXL2 mutation and mRNA expression in adult and juvenile granulosa cell tumors of the ovary.

Recently, mutation of the FOXL2 gene has been consistently identified in adult granulosa cell tumors of the ovary. The purpose of this study is to investigate whether the FOXL2 mutation and mRNA expression have a role in the pathogenesis of juvenile and adult granulosa cell tumors and influence tumor progression. Thirty-four adult granulosa cell tumors and 20 juvenile granulosa cell tumors were examined for the presence of the FOXL2 (C402G) mutation. Expression levels were studied by quantitative PCR and immunohistochemistry. We found that FOXL2 (C402G) mutation was present in 19/27 (70%) of the adult type tumors but in none of the juvenile granulosa cell tumors (0/18). No correlation was encountered between the presence of FOXL2 mutation and various clinicopathologic parameters except for the presence of a different sex-cord component, which was more frequently found in the subgroup of wild-type adult granulosa cell tumors than in the mutated tumors. Patients with tumors harboring the FOXL2 (C402G) mutation had a worse disease-free survival than those with the wild-type gene. Expression levels of FOXL2 mRNA had an impact on disease-free survival in both adult and juvenile granulosa cell tumors. We also found that the mutated tumors had a higher immunohistochemical expression of the FOXL2 protein, and there was a linear correlation between mRNA and immunohistochemical FOXL2 expression in both adult and juvenile granulosa cell tumors. Patients with juvenile granulosa cell tumors and higher FOXL2 protein expression had worse overall survival and disease-free survival than those with negative or weakly immunoreactive tumors. Our data suggest that FOXL2 mutation and mRNA expression are of prognostic importance in both adult and juvenile granulosa cell tumors.

Gene expression analysis identifies two groups of ovarian high-grade serous carcinomas with different prognosis.

Gene expression profiling is an important tool to evaluate genetic heterogeneity in carcinomas and is useful to develop expression-based classifications for many types of cancer, as well as markers of disease outcome. In this study, we have investigated the expression profile of 22 genes involved in the PI3K-AKT pathway in 26 high-grade ovarian carcinomas (19 serous and 7 clear cell carcinomas). Unsupervised hierarchical clustering divided high-grade ovarian carcinomas into three groups. Although all clear cell carcinomas clustered in one group, high-grade serous carcinomas were segregated into two separate groups with different prognosis (P=0.05). High expression of CASP3, XIAP (X-linked inhibitor of apoptosis) , NFKB1, FAS, and GSK3B mRNAs identified high-grade serous carcinomas with better prognosis. In multivariate analysis, these cluster groups were of prognostic significance independent of age, tumor size, and tumor stage (P=0.008). To validate the mRNA expression data, we studied the immunohistochemical expression of caspase-3 and XIAP on a tissue microarray. Immunoreaction for caspase-3 was concordant with the results obtained by mRNA expression analysis (Spearman r=0.762, P=0.000). Caspase-3 was exclusively expressed by the macrophages. Furthermore, co-expression of caspase-3 and XIAP identified high-grade serous carcinomas with different prognosis (P=0.03). Our results suggest that there are different biological subtypes of high-grade serous carcinomas.

Uterine leiomyosarcomas: tumor size, mitotic index, and biomarkers Ki67, and Bcl-2 identify two groups with different prognosis.

BACKGROUND: Prognostic factors for uterine leiomyosarcomas are not well established. Although most tumors are associated with poor prognosis even when apparently confined to the uterus (stage I), some cases that exhibited morphologic features of malignancy had prolonged survival. METHODS: Using tissue microarrays of 84 uterine leiomyosarcomas, we investigated conventional clinico-pathologic parameters, including International Federation of Gynecology and Obstetrics (FIGO) stage, together with expression of Ki67, p53, p16, and Bcl-2, attempting to distinguish leiomyosarcomas with different prognosis. The rate of CD163 immunoreactive tumor macrophages was also investigated. RESULTS: Tumor size and mitotic index were significant prognostic factors by univariate (p=0.018 and p=0.003, respectively) and multivariate (p=0.006 and p=0.001) analyses. Of the biomarkers investigated, only Ki67 immunoreaction was significant by univariate analysis and was associated with adverse prognosis (p=0.01). However, combination of tumor size, mitotic index, Ki67, and Bcl-2 worked even better. Using these 4 parameters, unsupervised hierarchical clustering identified 2 groups of tumors with different prognosis (p=0.001): group 1 consisted mostly of smaller leiomyosarcomas (<10cm) with mitotic index <20 MF/10 HPF, negative Ki67, and positive or negative Bcl-2 immunostaining. These tumors were associated with better prognosis. In contrast, group 2 leiomyosarcomas which were mostly≥10cm in diameter had higher mitotic index (≥ 20 MF/10 HPF), and were positive for Ki67 and negative for Bcl-2 had worse prognosis. Also, the number of CD163-macrophages was greater in group 2 than group 1 (p=0.007). CONCLUSIONS: Tumor size and mitotic index are morphologic predictors of malignancy in uterine leiomyosarcomas. Combination of tumor size, mitotic index, Ki67, and Bcl-2 protein expression allows distinguishing 2 groups of leiomyosarcomas with different survival. Leiomyosarcomas associated with poor outcome had a higher number of CD163 stromal macrophages.

Variations in stromal signatures in breast and colorectal cancer metastases.

The tumour microenvironment (TME) plays an important role in tumour survival and growth, but little is known about the degree of preservation between different stromal response patterns found in primary tumours and their metastases. We have previously identified gene expression profiles for two distinct stromal signatures in breast carcinoma of fibroblast (aka DTF) and macrophage (aka CSF1) response and found them to be correlated with clinicopathological features, including outcome. In this study, we compare the DTF fibroblast and CSF1 macrophage stromal response patterns in primary breast and colorectal cancers to their matched lymph node metastases. In both breast and colorectal cancer, there was a significant positive correlation between the CSF1 macrophage signature in the primary tumours and the matched lymph node metastases, as assessed by immunohistochemical markers. No such correlation was observed for the DTF fibroblast signature. A similar result was seen in independent analysis of two published gene expression microarray datasets. The variations of these stromal reaction patterns from the primary to the metastasis shed light on the relationship between the neoplastic cells and the non-neoplastic cells in the TME. The preservation of the CSF1 macrophage response pattern in metastases lends support to targeting the CSF1 pathway in cancer.

Atypical Endometrial Hyperplasia, Low-grade: "Much ADO About Nothing".

Atypical endometrial hyperplasia (AEH) is considered a precursor of endometrioid carcinoma. The 2020 World Health Organization (WHO) classification divides endometrial hyperplasia into 2 categories: hyperplasia without atypia and atypical hyperplasia/endometrioid intraepithelial neoplasia (EIN); however, this classification does not consider the degree of nuclear atypia. We graded nuclear atypia for estimating the risk of finding carcinoma at hysterectomy. Also, we investigated genes involved in endometrial carcinogenesis including mismatch repair (MMR) genes and ARID1A, PIK3CA, PTEN, KRAS, and CTNNB1. We reviewed 79 biopsies of AEH from 79 patients who underwent hysterectomy within a 1-year interval. Intraobserver and interobserver agreement of grading nuclear atypia and the relationship between the grade of nuclear atypia at biopsy and the findings at hysterectomy were evaluated. Immunohistochemistry for MMR status was performed in all cases and targeted sequencing in 11. Using low-grade versus high-grade nuclear atypia, κ values ranged from 0.74 to 0.91 (89% to 96%) and from 0.72 to 0.81 (87% to 91%) for the intraobserver and the interobserver agreement, respectively. The degree of nuclear atypia at biopsy was highly predictive of the findings at hysterectomy (P=1.6×10-15). Of 53 patients with low-grade AEH, none had carcinoma at hysterectomy, whereas 6 (6/26; 23%) with high-grade AEH in the biopsy also had high-grade AEH in the uterus and 16 (16/26; 61%) had FIGO grade 1 carcinoma. MMR deficiency was found in 3 of the 79 patients. None of the genes showed a mutational load significantly associated with the degree of nuclear atypia. In summary, our data show high reproducibility within and between observers for the diagnosis of low-grade and high-grade AEH. Most cases of AEH had low-grade nuclear atypia and neither high-grade AEH nor carcinoma was encountered in the corresponding hysterectomy specimens.

Immunoglobulin G fragment C receptor polymorphisms and KRAS mutations: are they useful biomarkers of clinical outcome in advanced colorectal cancer treated with anti-EGFR-based therapy?

KRAS mutations have been identified as a strong predictor of resistance to anti-epidermal growth factor receptor (EGFR) therapies. Besides inhibiting the EGFR pathway, anti-EGFR monoclonal antibodies may exert antitumor effects through antibody-dependent cell-mediated cytotoxicity (ADCC). Through this mechanism, the antibody fragment C portion (Fcγ) interacts with Fc receptors (FcγRs) expressed by immune effectors cells. We investigated the association of FcγR polymorphisms and KRAS mutation with the clinical outcome of 104 refractory metastatic colorectal cancer (mCRC) patients treated with anti-EGFR antibodies. FcγRIIa-H131R and FcγRIIIa-V158F polymorphisms were analyzed in genomic DNA using a 48.48 dynamic array on the BioMark system (Fluidigm, South Sanfrancisco, CA, USA). Tumor tissues from 96 cases were screened for KRAS mutations. KRAS mutation was associated with a lower response rate (RR) (P = 0.035) and a shorter progression-free survival (PFS) (3 vs 7 months; P = 0.36). FcγRIIa-H131R and FcγRIIIa-V158F polymorphisms did not show statistically significant associations with response, PFS, or KRAS status. In the logistic regression analysis, KRAS status (P = 0.04) and skin toxicity (P = 0.03) were associated with RR. By multivariate analysis, the clinical risk classification (P = 0.006) and skin toxicity (P < 0.0001) were found to be independent risk factors for PFS. In conclusion, the FcγRIIa and FcγRIIIa polymorphisms are not useful as molecular markers for clinical outcome in mCRC patients. To date, the EORTC (European Organization for Research and Treatment of Cancer Classification), skin toxicity, and KRAS status are the only reliable biomarkers to identify patients that would benefit from anti-EGFR therapy.

Expression profiling of 22 genes involved in the PI3K-AKT pathway identifies two subgroups of high-grade endometrial carcinomas with different molecular alterations.

Previously, we showed that PIK3CA and p53 alterations in uterine endometrial carcinomas correlate with poor prognosis. However, the contribution of phosphatidylinositol 3-kinase (PI3K) -AKT deregulation to endometrial carcinogenesis is not completely understood. The purpose of this study was to analyze alterations of this pathway in endometrial carcinomas and correlate them with the most common genetic abnormalities. Expression profiling of 22 genes involved in PI3K-AKT signaling pathway was analyzed in 38 endometrial carcinomas using TaqMan low-density array (TLDA) analysis. The gene expression pattern was analyzed by hierarchical clustering analysis. Unsupervised clustering divided the high-grade endometrial carcinomas into two clusters. One cluster identified tumors with alterations in the PI3K-AKT signaling pathway (exon 20 PIK3CA mutations and/or PTEN mutations 9/15; 60%), and p16 protein overexpression (8/13; 62%). Almost all non-endometrioid adenocarcinomas (serous and clear cell adenocarcinomas) were segregated into this cluster. In contrast, the other cluster identified tumors with p53 alterations (6/6; 100%), p16 protein overexpression (5/5; 100%), and exon 9 PIK3CA mutations (2/6; 33%). Exon 20 PIK3CA and PTEN mutations were not found in this subgroup. Low-grade endometrial carcinomas clustered in a third subgroup characterized by high frequency of PTEN mutations (10/17; 59%) and microsatellite instability (6/17; 35%). Our results show that gene expression profile differences in the PI3K-AKT signaling pathway identify two subgroups of high-grade endometrial carcinomas with different molecular alterations (PI3K-AKT pathway vs p53 alterations) that may have distinct roles in endometrial carcinogenesis. Identification of these subgroups can provide insight into the biology of these tumors and may facilitate the development of future treatments.