Checkpoint blockade can reverse T-cell exhaustion and promote antitumor responses. Although blocking the PD-1 pathway has been successful in Hodgkin lymphoma, response rates have been modest in ...B-cell non-Hodgkin lymphoma (NHL). Coblockade of checkpoint receptors may therefore be necessary to optimize antitumor T-cell responses. Here, characterization of coinhibitory receptor expression in intratumoral T cells from different NHL types identified TIGIT and PD-1 as frequently expressed coinhibitory receptors. Tumors from NHL patients were enriched in CD8
and CD4
T effector memory cells that displayed high coexpression of TIGIT and PD-1, and coexpression of these checkpoint receptors identified T cells with reduced production of IFNγ, TNFα, and IL2. The suppressed cytokine production could be improved upon
culture in the absence of ligands. Whereas PD-L1 was expressed by macrophages, the TIGIT ligands CD155 and CD112 were expressed by lymphoma cells in 39% and 50% of DLBCL cases and in some mantle cell lymphoma cases, as well as by endothelium and follicular dendritic cells in all NHLs investigated. Collectively, our results show that TIGIT and PD-1 mark dysfunctional T cells and suggest that TIGIT and PD-1 coblockade should be further explored to elicit potent antitumor responses in patients with NHL.
Tumor-infiltrating regulatory T cells (Tregs) contribute to an immunosuppressive tumor microenvironment. Despite extensive studies, the prognostic impact of tumor-infiltrating Tregs in B-cell ...non-Hodgkin lymphomas (B-NHLs) remains unclear. Emerging studies suggest substantial heterogeneity in the phenotypes and suppressive capacities of Tregs, emphasizing the importance of understanding Treg diversity and the need for additional markers to identify highly suppressive Tregs. Here, we applied single-cell RNA sequencing and T-cell receptor sequencing combined with high-dimensional cytometry to decipher the heterogeneity of intratumoral Tregs in diffuse large B-cell lymphoma and follicular lymphoma (FL), compared with that in nonmalignant tonsillar tissue. We identified 3 distinct transcriptional states of Tregs: resting, activated, and unconventional LAG3+FOXP3- Tregs. Activated Tregs were enriched in B-NHL tumors, coexpressed several checkpoint receptors, and had stronger immunosuppressive activity compared with resting Tregs. In FL, activated Tregs were found in closer proximity to CD4+ and CD8+ T cells than other cell types. Furthermore, we used a computational approach to develop unique gene signature matrices, which were used to enumerate each Treg subset in cohorts with bulk gene expression data. In 2 independent FL cohorts, activated Tregs was the major subset, and high abundance was associated with adverse outcome. This study demonstrates that Tregs infiltrating B-NHL tumors are transcriptionally and functionally diverse. Highly immunosuppressive activated Tregs were enriched in tumor tissue but absent in the peripheral blood. Our data suggest that a deeper understanding of Treg heterogeneity in B-NHL could open new paths for rational drug design, facilitating selective targeting to improve antitumor immunity.
CHK1 is an important regulator of the cell cycle and DNA damage response, and its altered expression has been identified in various tumors. Chk1 inhibitors are currently being evaluated as ...monotherapy and as potentiators of chemotherapy in clinical settings. However, to our knowledge, no previous study has investigated either the activation status or the therapeutic potential of CHK1 targeting in vulvar cancer. Therefore, we examined the expression status of activated CHK1 forms pCHK1Ser345, pCHK1Ser317, pCHK1Ser296, and pCHK1Ser280 in 294 vulvar squamous cell carcinomas (VSCC) using immunohistochemistry and analyzed their relationships with various clinicopathological variables and clinical outcome. To aid translation of preclinical studies, we also assessed cell sensitivity to the Chk1 inhibition in two vulvar cancer cell lines. Compared to the levels of pCHK1Ser345, pCHK1Ser317, pCHK1Ser296, and pCHK1Ser280 in normal vulvar squamous epithelium, high nuclear pCHK1Ser345 expression was found in 57% of vulvar carcinomas, whereas low nuclear pCHK1Ser317, pCHK1Ser296, and pCHK1Ser280 expressions were observed in 58%, 64%, and 40% of the cases, respectively. Low levels of pCHK1Ser317 and pCHK1Ser280 in the nucleus correlated significantly with advanced tumor behaviors and aggressive features. None of pCHK1Ser345, pCHK1Ser317, pCHK1Ser296, and pCHK1Ser280 forms were identified as prognostic factors. In vitro inhibition of CHK1 by small molecular inhibitors or siRNA reduced viability by inducing DNA damage and apoptosis of vulvar cancer cell lines. In summary, we conclude that cellular functions regulated by CHK1 are phosphorylation/localization‐dependent and deregulation of CHK1 function occurs in VSCC and might contribute to tumorigenesis. Targeting CHK1 might represent as a useful antitumor strategy for the subgroup of VSCC harboring p53 mutations.
In this manuscript, we investigated 1) the expression status of phosphorylated and activated pCHK1Ser345, pCHK1Ser317, pCHK1Ser296, and pCHK1Ser280 forms in 294 vulvar squamous cell carcinomas (VSCC) and their relationship with various clinicopathological variables and clinical outcome; 2) the effects of in Vitro inhibition of CHK1 by small molecules of siRNA in vulvar cancer cell lines. Our results show that low levels of pCHK1Ser317 and pCHK1Ser280 in the nucleus correlate significantly with advanced tumor behaviors and aggressive features. Furthermore, in vitro targeting of CHK1 leads to induction of DNA damage and apoptosis in vulvar cancer cell lines harboring p53 mutation.
Abstract
CHK
1 is an important regulator of the cell cycle and
DNA
damage response, and its altered expression has been identified in various tumors. Chk1 inhibitors are currently being evaluated as ...monotherapy and as potentiators of chemotherapy in clinical settings. However, to our knowledge, no previous study has investigated either the activation status or the therapeutic potential of
CHK
1 targeting in vulvar cancer. Therefore, we examined the expression status of activated
CHK
1 forms
pCHK
1
Ser345
,
pCHK
1
Ser317
,
pCHK
1
Ser296
, and
pCHK
1
Ser280
in 294 vulvar squamous cell carcinomas (
VSCC
) using immunohistochemistry and analyzed their relationships with various clinicopathological variables and clinical outcome. To aid translation of preclinical studies, we also assessed cell sensitivity to the Chk1 inhibition in two vulvar cancer cell lines. Compared to the levels of
pCHK
1
Ser345
,
pCHK
1
Ser317
,
pCHK
1
Ser296
, and
pCHK
1
Ser280
in normal vulvar squamous epithelium, high nuclear
pCHK
1
Ser345
expression was found in 57% of vulvar carcinomas, whereas low nuclear
pCHK
1
Ser317
,
pCHK
1
Ser296
, and
pCHK
1
Ser280
expressions were observed in 58%, 64%, and 40% of the cases, respectively. Low levels of
pCHK
1
Ser317
and
pCHK
1
Ser280
in the nucleus correlated significantly with advanced tumor behaviors and aggressive features. None of
pCHK
1
Ser345
,
pCHK
1
Ser317
,
pCHK
1
Ser296
, and
pCHK
1
Ser280
forms were identified as prognostic factors.
In vitro
inhibition of
CHK
1 by small molecular inhibitors or si
RNA
reduced viability by inducing
DNA
damage and apoptosis of vulvar cancer cell lines. In summary, we conclude that cellular functions regulated by
CHK
1 are phosphorylation/localization‐dependent and deregulation of
CHK
1 function occurs in
VSCC
and might contribute to tumorigenesis. Targeting
CHK
1 might represent as a useful antitumor strategy for the subgroup of VSCC harboring p53 mutations.
Summary Leiomyosarcoma (LMS) is the most common uterine sarcoma. Although the disease is relatively rare, it is responsible for considerable mortality due to frequent metastasis and chemoresistance. ...The molecular events related to LMS metastasis are unknown to date. The present study compared the global gene expression patterns of primary uterine LMSs and LMS metastases. Gene expression profiles of 13 primary and 15 metastatic uterine LMSs were analyzed using the HumanRef-8 BeadChip from Illumina. Differentially expressed candidate genes were validated using quantitative real-time polymerase chain reaction (PCR) and immunohistochemistry. To identify differently expressed genes between primary and metastatic tumors, we performed one-way analysis of variance with Benjamini-Hochberg correction. This led to identification of 203 unique probes that were significantly differentially expressed in the 2 tumor groups by greater than 1.58-fold with P < .01, of which 94 and 109 were overexpressed in primary and metastatic LMSs, respectively. Genes overexpressed in primary uterine LMSs included OSTN, NLGN4X , NLGN1, SLITRK4 , MASP1 , XRN2 , ASS1 , RORB, HRASLS , and TSPAN7 . Genes overexpressed in LMS metastases included TNNT1 , FOLR3 , TDO2 , CRYM , GJA1 , TSPAN10 , THBS1 , SGK1 , SHMT1 , EGR2 , and AGT . Quantitative real-time PCR confirmed significant anatomical site-related differences in FOLR3 , OSTN , and NLGN4X levels; and immunohistochemistry showed significant differences in TDO2 expression. Gene expression profiling differentiates primary uterine LMSs from LMS metastases. The molecular signatures unique to primary and metastatic LMSs may aid in understanding tumor progression in this cancer and in providing a molecular basis for prognostic studies and therapeutic target discovery.
The purpose of this study was to determine the expression and potential clinical role of epithelial‐to‐mesenchymal transition (EMT)‐related factors in malignant ovarian germ cell tumors (MOGCT). ...Protein expression of E‐cadherin, N‐cadherin, P‐cadherin, Zeb1, HMGA2, and vimentin by immunohistochemistry was analyzed in 42 MOGCT from patients treated in Norway during the period 1981–2001. Expression was analyzed for association with clinicopathologic parameters. E‐cadherin (p = 0.016) and HMGA2 (p = 0.002) expression was significantly higher in immature teratomas and yolk sac tumors compared with dysgerminomas. Vimentin (p < 0.001) and Zeb1 (p = 0.029) staining was significantly higher in immature teratomas compared with yolk sac tumors and dysgerminomas, whereas no significant differences were observed for N‐cadherin and P‐cadherin. EMT‐associated markers were not significantly related to clinicopathologic parameters including age, tumor diameter, and FIGO stage. In conclusion, based on this limited series, EMT‐associated markers are not associated with clinical parameters in MOGCT, in contrast to ovarian carcinoma. EMT‐related proteins are differentially expressed among various MOGCT subtypes, suggesting differences in biological characteristics associated with invasion and metastasis.
Cyclin B1-CDK1 complex plays an important role in the regulation of cell cycle. Activation of Cyclin B1 and CDK1 and the formation of the complex in G2/M are under multiple regulations involving many ...regulators such as isoforms of 14-3-3 and CDC25 and Wee1. Abnormal expression of Cyclin B1 and CDK1 has been detected in various tumors. However, to our knowledge no previous study has investigated Cyclin B1 and CDK1 in vulvar cancer. Therefore, we evaluated the statuses of CDK1Tyr15, pCDK1Thr161, Cyclin B1 (total) and pCyclin B1Ser126 in 297 cases of vulvar squamous cell carcinomas by immunohistochemistry. Statistical analyses were performed to explore their clinicopathological and prognostic values. In at least 25% of tumor cases high expression of CDK1Tyr15, pCDK1Thr161, Cyclin B1 (total) and pCyclin B1Ser126 was observed, compared to the low levels in normal vulvar squamous epithelium. Elevated levels of CDK1Tyr15, pCDK1Thr161, Cyclin B1 (total) and pCyclin B1Ser126 were correlated with advanced tumor behaviors and aggressive features. Although CDK1Tyr15, pCDK1Thr161, Cyclin B1 (total) and pCyclin B1Ser126 could not be identified as prognostic factors, combinations of (pCDK1Thr161 C+N + 14-3-3σN), (pCDK1Thr161 C+N + 14-3-3ηC), (pCDK1Thr161 C+N + Wee1C) and (pCDK1Thr161 C+N + 14-3-3σN + 14-3-3ηC + Wee1C) were correlated with disease-specific survival (p = 0.036, p = 0.029, p = 0.042 and p = 0.007, respectively) in univariate analysis. The independent prognostic significance of (pCDK1Thr161 C+N + 14-3-3σN + 14-3-3ηC + Wee1C) was confirmed by multivariate analysis. In conclusion, CDK1Tyr15, pCDK1Thr161, Cyclin B1 (total) and pCyclin B1Ser126 may be involved in progression of vulvar squamous cell carcinoma. The combination of pCDK1Thr161, 14-3-3σ, 14-3-3η and Wee1 was a statistically independent prognostic factor.
Objectives: To analyze the clinical role of hormone receptors in a large uterine sarcomas series with long-term follow-up.
Methods: Protein expression of estrogen receptor (ER) and progesterone ...receptor (PR) by immunohistochemistry was studied in tissue microarrays from 294 patients diagnosed with uterine sarcoma in Norway from 1970 to 2000 and analyzed for an association with clinicopathologic parameters and outcome.
Results: ER and PR were detected in 136 of 291 and 184 of 291 tumors (three noninformative cases each), respectively. Expression was unrelated to histology, patient age, tumor diameter, the degree of atypia, the presence of necrosis or vascular invasion, or mitotic counts. ER and PR expression was unrelated to survival in the analysis of the entire cohort. When survival analysis was confined to stage I leiomyosarcoma (n = 147), higher PR score was significantly related to longer overall survival (OS) (P = .042). Clinicopathologic prognosticators in this group were age (P = .041), tumor diameter (P = .001), and mitotic count (P = .007), with a trend for atypia (P = .087). In Cox multivariate analysis, PR score (P = .019), tumor diameter (P = .013), and mitotic count (P = .002) were independent prognosticators of OS.
Conclusions: Hormone receptor expression is not informative of outcome in the analysis of uterine sarcomas of all stages and histologic types. PR expression identifies patients with longer survival in stage I leiomyosarcoma.
Cyclin B1-CDK1 complex plays an important role in the regulation of cell cycle. Activation of Cyclin B1 and CDK1 and the formation of the complex in G2/M are under multiple regulations involving many ...regulators such as isoforms of 14-3-3 and CDC25 and Wee1. Abnormal expression of Cyclin B1 and CDK1 has been detected in various tumors. However, to our knowledge no previous study has investigated Cyclin B1 and CDK1 in vulvar cancer. Therefore, we evaluated the statuses of CDK1.sup.Tyr15, pCDK1.sup.Thr161, Cyclin B1 (total) and pCyclin B1.sup.Ser126 in 297 cases of vulvar squamous cell carcinomas by immunohistochemistry. Statistical analyses were performed to explore their clinicopathological and prognostic values. In at least 25% of tumor cases high expression of CDK1.sup.Tyr15, pCDK1.sup.Thr161, Cyclin B1 (total) and pCyclin B1.sup.Ser126 was observed, compared to the low levels in normal vulvar squamous epithelium. Elevated levels of CDK1.sup.Tyr15, pCDK1.sup.Thr161, Cyclin B1 (total) and pCyclin B1.sup.Ser126 were correlated with advanced tumor behaviors and aggressive features. Although CDK1.sup.Tyr15, pCDK1.sup.Thr161, Cyclin B1 (total) and pCyclin B1.sup.Ser126 could not be identified as prognostic factors, combinations of (pCDK1.sup.Thr161 C+N + 14-3-3sigma.sup.N ), (pCDK1.sup.Thr161 C+N + 14-3-3eta.sup.C ), (pCDK1.sup.Thr161 C+N + Wee1.sup.C) and (pCDK1.sup.Thr161 C+N + 14-3-3sigma.sup.N + 14-3-3eta.sup.C + Wee1.sup.C) were correlated with disease-specific survival (p = 0.036, p = 0.029, p = 0.042 and p = 0.007, respectively) in univariate analysis. The independent prognostic significance of (pCDK1.sup.Thr161 C+N + 14-3-3sigma.sup.N + 14-3-3eta.sup.C + Wee1.sup.C) was confirmed by multivariate analysis. In conclusion, CDK1.sup.Tyr15, pCDK1.sup.Thr161, Cyclin B1 (total) and pCyclin B1.sup.Ser126 may be involved in progression of vulvar squamous cell carcinoma. The combination of pCDK1.sup.Thr161, 14-3-3sigma, 14-3-3eta and Wee1 was a statistically independent prognostic factor.
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