Although immune checkpoint inhibitors improve median overall survival in patients with metastatic urothelial cancer (mUC), only a minority of patients benefit from it. Early blood-based response ...biomarkers may provide a reliable way to assess response weeks before imaging is available, enabling an early switch to other therapies. We conducted an exploratory study aimed at the identification of early markers of response to anti-PD-1 in patients with mUC. Whole blood RNA sequencing and phenotyping of peripheral blood mononuclear cells were performed on samples of 26 patients obtained before and after 2 to 6 weeks of anti-PD-1. Between baseline and on-treatment samples of patients with clinical benefit, 51 differentially expressed genes (DEGs) were identified, of which 37 were upregulated during treatment. Among the upregulated genes was PDCD1, the gene encoding PD-1. STRING network analysis revealed a cluster of five interconnected DEGs which were all involved in DNA replication or cell cycle regulation. We hypothesized that the upregulation of DNA replication/cell cycle genes is a result of T cell proliferation and we were able to detect an increase in Ki-67+ CD8+ T cells in patients with clinical benefit (median increase: 1.65%, range −0.63 to 7.06%, p = 0.012). In patients without clinical benefit, no DEGs were identified and no increase in Ki-67+ CD8+ T cells was observed. In conclusion, whole blood transcriptome profiling identified early changes in DNA replication and cell cycle regulation genes as markers of clinical benefit to anti-PD-1 in patients with urothelial cancer. Although promising, our findings require further validation before implementation in the clinic.
Lipid metabolism is altered in several cancer settings leading to different ratios of intermediates. Ovarian cancer is the most lethal gynecological malignancy. Cancer cells disperse in the abdominal ...space and ascites occurs. T cells obtained from ascites are unable to proliferate after an antigenic stimulus. The proliferation of ascites-derived T cells can be restored after culturing the cells for ten days in normal culture medium. No pathway aberrancies were detected. The acellular fraction of ascites can inhibit the proliferation of autologous as well as allogeneic peripheral blood lymphocytes, indicating the presence of soluble factors that interfere with T cell functionality. Therefore, we analyzed 109 lipid mediators and found differentially regulated lipids in suppressive ascitic fluid compared to normal abdominal fluid. Our study indicates the presence of lipid intermediates in ascites of ovarian cancer patients, which coincidences with T cell dysfunctionality. Since the immune system in the abdominal cavity is compromised, this may explain the high seeding efficiency of disseminated tumor cells. Further research is needed to fully understand the correlation between the various lipids and T cell proliferation, which could lead to new treatment options.
Eradication of cancer stem cells to abrogate tumor growth is a new treatment modality. However, like normal cells cancer cells show plasticity. Differentiated tumor stem cells can acquire stem cell ...properties when they gain access to the stem cell niche. This indicates that eradicating of stem cells (emptying of the niche) alone will not lead to eradication of the tumor. Treatment should be directed to cancer stem cells and more mature cancer cells.
Ascites is a prominent feature of ovarian cancer and could serve as liquid biopsy to assess the immune status of patients. Tumor-infiltrating T lymphocytes are correlated with improved survival in ...ovarian cancer. To investigate whether immune cells in ascites are associated with patient outcome, we analyzed the amount of dendritic cell (DC) and T cell subsets in ascites from ovarian cancer patients diagnosed with high-grade serous cancer (HGSC). Ascites was collected from 62 HGSC patients prior to chemotherapy. Clinicopathological, histological and follow-up data from patients were collected. Ascites-derived immune cells were isolated using density-gradient centrifugation. The presence of myeloid DCs (BDCA-1
, BDCA-3
, CD16
), pDCs (CD123
BDCA-2
), and T cells (CD4
, CD8
) was analyzed using flow cytometry. Complete cytoreduction, response to primary treatment and chemosensitivity were associated with improved patient outcome. In contrast, immune cells in ascites did not significantly correlate with patient survival. However, we observed a trend toward improved outcome for patients having low percentages of CD4
T cells. Furthermore, we assessed the expression of co-stimulatory and co-inhibitory molecules on T cells and non-immune cells in 10 ascites samples. PD-1 was expressed by 30% of ascites-derived T cells and PD-L1 by 50% of non-immune cells. However, the percentage of DC and T cell subsets in ascites was not directly correlated to the survival of HGSC patients.
Radium-223 improves overall survival (OS) in men with bone metastatic castration-resistant prostate cancer (mCRPC). While the exact mechanism behind this survival benefit remains unclear, ...radium-induced immunological mechanisms might contribute to the OS advantage. We performed a comprehensive evaluation of the immunological changes in mCRPC patients by phenotyping the peripheral blood mononuclear cells (PBMCs) during radium-223 therapy.
In this prospective, single-arm, exploratory study, PBMCs of 30 mCRPC patients were collected before, during, and after treatment with radium-223. Lymphocyte and monocyte counts were analyzed to get insight into general immune cell trends. Next, we analyzed changes in T cell subsets, myeloid-derived suppressor cells (MDSCs), and immune checkpoint expression using linear regression models. Per subset, the 6-month change (% of baseline) was determined. Bootstrapped 95% confidence intervals were used to measure the degree of uncertainty of our findings.
We observed a substantial decrease in absolute lymphocyte counts (-0.12 * 10^9 cells/L per injection, 95% CI: -0.143 - -0.102). Simultaneously, an increase was observed in the proportion of T cells that expressed costimulatory (ICOS) or inhibitory (TIM-3, PD-L1, and PD-1) checkpoint molecules. Moreover, the fraction of two immunosuppressive subsets - the regulatory T cells and the monocytic MDSCs - increased throughout treatment. These findings were not more pronounced in patients with an alkaline phosphatase response during therapy.
Immune cell subsets in patients with mCRPC changed during radium-223 therapy, which warrants further research into the possible immunological consequences of these changes.
Ovarian cancer is one of the most fatal tumors in women. Due to a lack of symptoms and adequate screening methods, patients are diagnosed at advanced stages with extensive tumor burden (Jelovac and ...Armstrong, 2011). Interestingly, ovarian cancer metastasis is generally found within the peritoneal cavity rather than other tissues (Lengyel, 2010; Tan
, 2006 ). The reason behind this tissue tropism of the peritoneal cavity remains elusive. A prominent feature of this selectivity is ascites, the accumulation of fluid within the peritoneal cavity, containing, amongst others, immune cells, tumor cells and various soluble factors that can be involved in the progression of ovarian cancer ( Kipps
, 2013 ). The protocol described here is used to isolate mononuclear cells from ascites to study the functionality of the immune system within the peritoneal cavity.
The identification of growth and differentiation pathways that are responsible for the proliferation and survival of cancer stem cells (CSCs) has opened avenues for the discovery of novel therapeutic ...targets. In the initial phase of an anticancer immune response, T cells specific for tumor-associated antigens develop in patients and, at least under selected circumstances, are able to eliminate malignant cells. However, it remains unknown whether CSC-specific T cells are also operational. We found naturally occurring multifunctional CD4
+
and CD8
+
T cells specific for the stem cell marker OCT4 among the peripheral blood mononuclear cells (PBMCs) of both healthy individuals and ovarian cancer patients. Moreover, lymphocytes isolated from the ascites of patients affected by ovarian malignancies also contained OCT4-specific T cells. OCT4-reactive CD4
+
T cells did not produce interferon γ (IFNγ) and IFNγ-inducible protein 10 (IP-10) but were capable of proliferation upon stimulation with dendritic cells (DCs) loaded with an OCT4-derived peptide or OCT4 mRNA. OCT4-reactive CD8
+
cells did not proliferate in response to a similar challenge, yet produced IP-10 as well as sufficient amounts of IFNγ to induce IP-10 . Furthermore, CD8
+
cytotoxic T cells were able to release their lysosomal components, as indicated by the mobilization of CD107a. These results demonstrate the existence of anti-CSC specific T cells in ovarian cancer patients.
Dendritic cells (DCs) play a key role in orchestrating adaptive immune responses. In human blood, three distinct subsets exist: plasmacytoid DCs (pDCs) and BDCA3+ and CD1c+ myeloid DCs. In addition, ...a DC-like CD16+ monocyte has been reported. Although RNA-expression profiles have been previously compared, protein expression data may provide a different picture. Here, we exploited label-free quantitative mass spectrometry to compare and identify differences in primary human DC subset proteins. Moreover, we integrated these proteomic data with existing mRNA data to derive robust cell-specific expression signatures with more than 400 differentially expressed proteins between subsets, forming a solid basis for investigation of subset-specific functions. We illustrated this by extracting subset identification markers and by demonstrating that pDCs lack caspase-1 and only express low levels of other inflammasome-related proteins. In accordance, pDCs were incapable of interleukin (IL)-1β secretion in response to ATP.
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•We present a comprehensive quantitative proteome comparison of primary human DC subsets•Proteome comparison reveals many expression differences between DC subsets•We provide a resource to derive markers and examine subset functional specialization•pDCs lack caspase-1 and have a decreased inflammasome response
Worah et al. present a comprehensive quantitative proteomic comparison of four human blood-derived DC-like subsets. Through integration of proteomic and transcriptomic data, the authors derive expression signatures for each subset that provide a resource for study of subset functional specialization.
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