Factors In Tumor Microenvironment Research Articles

Investigating Gene Expression Phenotypes and Tumor Microenvironment Factors Associated with Response to Low Dose Radiotherapy for Follicular Lymphoma

<h3>Purpose/Objective(s)</h3> Radiotherapy (RT) is the most effective single agent for the treatment of Follicular Lymphoma (FL) as shown in the FoRT trial, where the response to 4 Gy was 81%. However, as ∼20% do not respond and others develop rapid regrowth after RT, identifying molecular markers associated with response is imperative for personalized RT approaches. We sought to determine if gene expression phenotypes and tumor microenvironment factors are associated with response to low dose RT (4 Gy). <h3>Materials/Methods</h3> We retrospectively identified patients with grade 1-2 FL who had pre-treatment formalin-fixed paraffin embedded tumor tissue available prior to low dose RT and had not received systemic therapy within the prior 3 months. We measured baseline clinical factors and scored ‘responders' to RT as those with a complete or partial response by imaging or clinical criteria within 6 months, with all others defined as ‘non-responders'. We performed bulk RNA-sequencing with gene expression quantification using Salmon. Digital cytometry was performed using CIBERSORTx. We performed differential gene expression using DESeq2 with adjustment of all <i>P</i>-values for multiple hypothesis testing. All <i>P</i>-values were two-tailed and considered significant at <i>P</i><0.05. <h3>Results</h3> We identified 7 patients with 8 available tissue samples who met inclusion criteria. Median age at RT was 76.6 years (range, 57.1 – 79.3). Four were males and the majority (85.7%) had stage III-IV FL. One patient received 4 Gy in 1 fraction, all others received 2 Gy x 2. By treated site, 75% (n=6) exhibited a partial or complete response to low-dose RT and 2 progressed, representing non-responders. The non-responders had significantly higher expression of genes associated with immunosuppression (<i>CLEC4G, CCND1</i>) and tumorigenesis (<i>MAGED4, MAGED4B</i>) as compared to responders. Additionally, non-responders had significant under-expression of genes associated with tumor suppression (<i>XIST, BIK, AC009299.1</i>) and cell migration (<i>IGF2BP1, DCAF12</i>). We observed numerically larger cell fractions of CD4 T cells (median=10.1% v 6.4%, <i>P</i>=.26) and natural killer cells (median=2.7% vs 0.8%, <i>P</i>=.15) in responders whereas non-responders had higher relative abundances of follicular helper T-cells (median=11.7% v 8.6%, <i>P</i>=.15) and macrophages (median=8.5% v 5.6%, <i>P</i>=.47). <h3>Conclusion</h3> Based on the results of our pilot cohort, we observed that non-responders exhibited a gene expression phenotype characterized by immunosuppression and increased tumorigenesis. Furthermore, our results suggest immune cell abundances may differ in those that have a response to low dose RT. We aim to expand our investigation to a larger cohort of patients with the goal of informing precision approaches when using low dose RT for FL.

Muc16 depletion diminishes KRAS-induced tumorigenesis and metastasis by altering tumor microenvironment factors in pancreatic ductal adenocarcinoma.

MUC16, membrane-bound mucin, plays an oncogenic role in pancreatic ductal adenocarcinoma (PDAC). However, the pathological role of MUC16 in the PDAC progression, tumor microenvironment, and metastasis in cooperation with KrasG12D and Trp53R172H mutations remains unknown. Deletion of Muc16 with activating mutations KrasG12D/+ and Trp53R172H/+ in mice significantly decreased progression and prolonged overall survival in KrasG12D/+; Trp53R172H/+; Pdx-1-Cre; Muc16-/- (KPCM) and KrasG12D/+; Pdx-1-Cre; Muc16-/- (KCM), as compared to KrasG12D/+; Trp53R172H/+; Pdx-1-Cre (KPC) and KrasG12D/+; Pdx-1-Cre (KC) mice, respectively. Muc16 knockout pancreatic tumor (KPCM) displays decreased tumor microenvironment factors and significantly reduced incidence of liver and lung metastasis compared to KPC. Furthermore, in silico data analysis showed a positive correlation of MUC16 with activated stroma and metastasis-associated genes. KPCM mouse syngeneic cells had significantly lower metastatic and endothelial cell binding abilities than KPC cells. Similarly, KPCM organoids significantly decreased the growth rate compared to KPC organoids. Interestingly, RNA-seq data revealed that the cytoskeletal proteins Actg2, Myh11, and Pdlim3 were downregulated in KPCM tumors. Further knockdown of these genes showed reduced metastatic potential. Overall, our results demonstrate that Muc16 alters the tumor microenvironment factors during pancreatic cancer progression and metastasis by changing the expression of Actg2, Myh11, and Pdlim3 genes.

Novel Approaches in Non-Melanoma Skin Cancers-A Focus on Hedgehog Pathway in Basal Cell Carcinoma (BCC).

Basal cell carcinoma (BCC) is one of the most common neoplasms in the population. A good prognosis and mainly non-aggressive development have made it underdiagnosed and excluded from the statistics. Due to the availability of efficient surgical therapy, BCC is sometimes overlooked in the search for novel therapies. Most clinicians are unaware of its complicated pathogenesis or the availability of effective targeted therapy based on Hedgehog inhibitors (HHI) used in advanced or metastatic cases. Nevertheless, the concomitance and esthetic burden of this neoplasm are severe. As with other cancers, its pathogenesis is multifactorial and complicated with a network of dependencies. Although the tumour microenvironment (TME), genetic aberrations, and risk factors seem crucial in all skin cancers, in BCC they all have become accessible as therapeutic or prevention targets. The results of this review indicate that a central role in the development of BCC is played by the Hedgehog (Hh) signalling pathway. Two signalling molecules have been identified as the main culprits, namely Patched homologue 1 (PTCH1) and, less often, Smoothened homologue (SMO). Considering effective immunotherapy for other neoplastic growths being introduced, implementing immunotherapy in advanced BCC is pivotal and beneficial. Up to now, the US Food and Drug Administration (FDA) has approved two inhibitors of SMO for the treatment of advanced BCC. Sonidegib and vismodegib are registered based on their efficacy in clinical trials. However, despite this success, limitations might occur during the therapy, as some patients show resistance to these molecules. This review aims to summarize novel options of targeted therapies in BCC and debate the mechanisms and clinical implications of tumor resistance.

Light Activates Cdc42-Mediated Needle-Shaped Filopodia Formation via the Integration of Small GTPases.

Cdc42 has been linked to multiple human cancers and is implicated in the migration of cancer cells. Cdc42 could be activated via biochemical and biophysical factors in tumor microenvironment, the precise control of Cdc42 was essential to determine its role to cell behaviors. Needle-shaped protrusions (filopodia) could sense the extracellular biochemical cues and pave the path for cell movement, which was a key structure involved in the regulation of cancer cell motility. We used the photoactivatable Cdc42 to elucidate the breast cancer cell protrusions, the mutation of Cdc42 was to confirm the optogenetic results. We also inhibit the Cdc42, Rac or Rho respectively by the corresponding inhibitors. We identified that the activation of Cdc42 by light could greatly enhance the formation of filopodia, which was positive for the contribution of cell movement. The expression of Cdc42 active form Cdc42-Q61L in cells resulted in the longer and more filopodia while the Cdc42 inactive form Cdc42-T17N were with the shorter and less filopodia. Moreover, the inhibition of Cdc42, Rac or Rho all significantly reduced the filopodia numbers and length in the co-expression of Cdc42-Q61L, which showed that the integration of small GTPases was necessary in the formation of filopodia. Furthermore, photoactivation of Cdc42 failed to enhance the filopodia formation with the inhibition of Rac or Rho. However, with the inhibition of Cdc42, the photoactivation of Cdc42 could partially recover back the filopodia formations, which indicated that the integration of small GTPases was key for the filopodia formations. Our work highlights that light activates Cdc42 is sufficient to promote filopodia formation without the destructive structures of small GTPases, it not only points out the novel technique to determine cell structure formations but also provides the experimental basis for the efficient small GTPases-based anti-cancer strategies.

Role of hypoxia in the tumor microenvironment and targeted therapy.

Tumor microenvironment (TME), which is characterized by hypoxia, widely exists in solid tumors. As a current research hotspot in the TME, hypoxia is expected to become a key element to break through the bottleneck of tumor treatment. More and more research results show that a variety of biological behaviors of tumor cells are affected by many factors in TME which are closely related to hypoxia. In order to inhibiting the immune response in TME, hypoxia plays an important role in tumor cell metabolism and anti-apoptosis. Therefore, exploring the molecular mechanism of hypoxia mediated malignant tumor behavior and therapeutic targets is expected to provide new ideas for anti-tumor therapy. In this review, we discussed the effects of hypoxia on tumor behavior and its interaction with TME from the perspectives of immune cells, cell metabolism, oxidative stress and hypoxia inducible factor (HIF), and listed the therapeutic targets or signal pathways found so far. Finally, we summarize the current therapies targeting hypoxia, such as glycolysis inhibitors, anti-angiogenesis drugs, HIF inhibitors, hypoxia-activated prodrugs, and hyperbaric medicine.

Molecular mechanisms of resistance to tyrosine kinase inhibitor in clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most common subtype of renal cell carcinoma (RCC). Loss of von Hippel-Lindau tumor suppressor gene is frequently observed in ccRCC and increases the expression of hypoxia-inducible factors and their targets, including epidermal growth factor, vascular endothelial growth factor, and platelet-derived growth factor. Tyrosine kinase inhibitors (TKIs) offer a survival benefit in metastatic renal cell carcinoma (mRCC). Recently, immune checkpoint inhibitors have been introduced in mRCC. Combination therapy with TKIs and immune checkpoint inhibitors significantly improved patient outcomes. Therefore, TKIs still play an essential role in mRCC treatment. However, the clinical utility of TKIs is compromised when primary and acquired resistance are encountered. The mechanism of resistance to TKI is not fully elucidated. Here, we comprehensively reviewed the molecular mechanisms of resistance to TKIs and a potential strategy to overcome this resistance. We outlined the involvement of angiogenesis, non-angiogenesis, epithelial-mesenchymal transition, activating bypass pathways, lysosomal sequestration, non-coding RNAs, epigenetic modifications and tumor microenvironment factors in the resistance to TKIs. Deep insight into the molecular mechanisms of resistance to TKIs will help to better understand the biology of RCC and can ultimately help in the development of more effective therapies.

A comprehensive pancancer analysis reveals the potential value of RAR-related orphan receptor C (RORC) for cancer immunotherapy.

Background: RAR-related orphan receptor C (RORC) plays an important role in autoimmune responses and inflammation. However, its function in cancer immunity is still unclear. Its potential value in cancer immunotherapy (CIT) needs to be further studied. Methods: Expression and clinical data for 33 cancers were obtained from UCSC-Xena. The correlation between RORC expression and clinical parameters was analyzed using the limma software package to assess the prognostic value of RORC. Timer2.0 and DriverDBv3 were used to analyze the RORC mutation and methylation profiles. RORC-associated signaling pathways were identified by GSEA. The correlations of RORC expression with tumor microenvironment factors were further assessed, including immune cell infiltration (obtained by CIBERSORT) and immunomodulators (in pancancer datasets from the Tumor-Immune System Interactions and Drug Bank [TISIDB] database). In addition, the correlations of RORC with four CIT biomarkers (tumor mutational burden, microsatellite instability, programmed death ligand-1, and mismatch repair) were explored. Furthermore, three CIT cohorts (GSE67501, GSE168204, and IMvigor210) from the Gene Expression Omnibus database and a previously published study were used to determine the association between RORC expression and CIT response. Results: RORC was differentially expressed in many tumor tissues relative to normal tissues (20/33). In a small number of cancers, RORC expression was correlated with age (7/33), sex (4/33), and tumor stage (9/33). Furthermore, RORC expression showed prognostic value in many cancers, especially in kidney renal clear cell carcinoma (KIRC), brain lower grade glioma (LGG), and mesothelioma (MESO). The mutation rate of RORC in most cancer types was low, while RORC was hypermethylated or hypomethylated in multiple cancers. RORC was associated with a variety of biological processes and signal transduction pathways in various cancers. Furthermore, RORC was strongly correlated with immune cell infiltration, immunomodulators, and CIT biomarkers. However, no significant association was found between RORC and CIT response in the three CIT cohorts. Conclusion Our findings revealed the potential immunotherapeutic value of RORC for various cancers and provides preliminary evidence for the application of RORC in CIT.

High abundance of Lachnospiraceae in the human gut microbiome is related to high immunoscores in advanced colorectal cancer.

The tumor microenvironment (TME) in colorectal cancer (CRC) includes the gut microbiome, immune cells, angiogenic factors, and fibroblasts and plays a major role in cancer progression. The Immunoscore (IS) is based on tumor infiltration by immune cells that are known prognostic biomarkers for CRC. However, the interrelation between the IS, microbiome, and other TME factors in human CRC remains unclear. A cohort of 94 patients with CRC was examined at the Shiga University of Medical Science Hospital in Japan. The expression levels of CD3, CD8, CD31, and alpha-smooth muscle actin (α-SMA) in the primary tumor were evaluated by immunohistochemistry. The IS was calculated based on the results of the CD3 and CD8 staining assays. Microbiomes in patients with CRC were examined by amplicon sequencing. The expression levels of α-SMA and tumor-infiltrating lymphocytes in patients with CRC were negatively correlated (P = 0.006). A high IS was associated with high abundance of Lachnospiraceae in the microbiomes of patients with CRC. Lymphocyte infiltration into the primary tumor was marked by reduced density of cancer-associated fibroblasts and enrichment of the Lachnospiraceae family in the gut microbiome, which may influence CRC progression.

851 Cancer associated fibroblasts in different T-stage lesions of cutaneous T-cell lymphoma

The tumor microenvironment (TME) has been shown to play an important role in tumor initiation, development, and metastasis. Among all TME factors, cancer associated fibroblasts (CAF) have been suggested to play a key role. Two studies have reported that fibroblasts from cutaneous T-cell lymphoma (CTCL) lesions, mycosis fungoides (MF) and/or Sézary syndrome (SS), were hyper-activated and exerted a pro-tumorigenic activity on CTCL cells in vitro. But much regarding CAFs in development of CTCL remains to be understood.

Intratumoral Budding in Pretreatment Biopsies, among Tumor Microenvironmental Components, Can Predict Prognosis and Neoadjuvant Therapy Response in Colorectal Adenocarcinoma.

Background and Objectives: The prediction of the prognosis and effect of neoadjuvant therapy is vital for patients with advanced or unresectable colorectal carcinoma (CRC). Materials and Methods: We investigated several tumor microenvironment factors, such as intratumoral budding (ITB), desmoplastic reaction (DR), and Klintrup–Mäkinen (KM) inflammation grade, and the tumor–stroma ratio (TSR) in pretreatment biopsy samples (PBSs) collected from patients with advanced or unresectable CRC. A total of 85 patients with 74 rectal carcinomas and 11 colon cancers treated at our hospital were enrolled; 66 patients had curative surgery and 19 patients received palliative treatment. Results: High-grade ITB was associated with recurrence (p = 0.002), death (p = 0.034), and cancer-specific death (p = 0.034). Immature DR was associated with a higher grade of clinical tumor-node-metastasis stage (cTNM) (p = 0.045), cN category (p = 0.045), and cM category (p = 0.046). The KM grade and TSR were not related to any clinicopathological factors. High-grade ITB had a significant relationship with tumor regression in patients who received curative surgery (p = 0.049). Conclusions: High-grade ITB in PBSs is a potential unfavorable prognostic factor for patients with advanced CRC. Immature DR, TSR, and KM grade could not predict prognosis or therapy response in PBSs.

Abstract 6133: Intratumoral budding in pretreatment biopsies can predict prognosis and neoadjuvant therapy response among the tumor microenvironmental components in colorectal adenocarcinoma

Abstract Purpose: Neoadjuvant therapy has been used as one of a treatment option in locally advanced rectal cancer and unresectable colorectal cancers (CRCC) with oligo-metastasis. Considering that radical specimen has been deformed due to this treatment, predicting prognosis is more complex. We evaluated several tumor microenvironmental factors in the pretreatment biopsies; those were known as prognostic factors in radical specimen. Methods: From 2010 to 2021, 85 patients with neoadjuvant therapy and surgery for CRCC were enrolled in Uijeongbu St. Mary’s hospital. Intratumoral budding (ITB), tumor stromal ratio, presence of immature stroma and Klintruo-Makinen grade for inflammatory cells were evaluated in pretreatment biopsies by three pathologists, independently. Results: Only high grade ITB (OS: p = 0.030, CSS: p = 0.005, DFS: p &amp;lt; 0.001) was poor survival factor in univariate analysis. In addition, high grade ITB was related with pathological stage (p = 0.017), lymphatic invasion (p = 0.010) and chemoresistance in non-metastatic CRCC group (p = 0.046). Presence of immature fibrosis was related with clinical stage (p = 0.045) and pathological stage (p = 0.033). In multivariate analysis, high grade ITB was independent poor survival factor (OS: p = 0.031, CSS: p = 0.009, DFS: p = 0.001). Conclusion: Our study revealed that high grade ITB in pretreatment biopsied was poor survival factor in CRCC with neoadjuvant treatment. In our observation, we recommend that ITB result should be included in the pathology report when considering neoadjuvant treatment. Citation Format: Kwangil Yim, Won Mo Jang, Yosep Chong, OK Ran Shin, Kyung Jin Seo. Intratumoral budding in pretreatment biopsies can predict prognosis and neoadjuvant therapy response among the tumor microenvironmental components in colorectal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6133.

Abstract 6268: Discovery and characterization of the CD96 antibody GSK6097608, a high-affinity, antagonistic anti-CD96 antibody for cancer immunotherapy

Abstract The overall therapeutic benefit of blocking the first generation of immune checkpoint pathways (PD-1 and CTLA-4) has been demonstrated across multiple tumor types, yielding long term protection in some patients. However, most patients do not respond to these single-agent approaches. Thus, collaborative strategies that seek to engage novel pathways and cell types may provide therapeutic options for patients wherein pre-existing host and tumor microenvironment factors do not favor current immunotherapeutic agents or in tumors where adaptive resistance has occurred. In recent years, the CD226 (DNAX Accessory Molecule-1 [DNAM-1]) axis has emerged as a relevant regulatory node in for natural killer (NK) and T cells - particularly in the context of tumor immunology. Similar to the competitive interplay between CTLA-4/CD28 and B7 (CD80/86), inhibitory receptors within the axis (e.g., CD96 [TACTILE]) effectively compete with the co-stimulatory receptor CD226 for binding to shared ligands (e.g., CD155), thereby impairing the initiation and/or promotion of ongoing antitumor immune responses. Indeed, genetic or monoclonal antibody (mAb)-based co-inhibition of CD96 with other immune checkpoints has proven efficacious in several nonclinical tumor models. CD96 has been shown to impact both T cell and NK cell function, offering a level of versatility as a target for cancer immunotherapy. For example, in the setting of anti-PD-1 neoadjuvant treatment, significantly improved survival of pancreatic ductal adenocarcinoma (PDAC) tumor-bearing mice was observed following enhancement of NK cell activity by CD96 antibody treatment. Equally, pronounced effects on primary tumor growth following anti-PD-1, -TIGIT, and -CD96 mAb treatment in a colorectal carcinoma tumor model (CT26) was found to be dependent on CD8+ T cells. GSK6097608 is a clinical-stage fully-human immunoglobulin G1 (IgG1)κ mAb that targets the inhibitory immune receptor CD96. GSK6097608 was identified, in part, for its ability prevent and disrupt CD96:CD155 interactions, thereby promoting T and NK cell function. GSK6097608 demonstrated concentration-dependent rescue of human immune cell activity following exposure to plate-bound recombinant CD155 (cognate receptor/ligand); an effect that was improved with concomitant TIGIT blockade. Notably, functional activity was found to be dependent on intact Fc-FcγR co-engagement, as potentiation of primary human T and NK cell function was lost when GSK6097608 was grafted on an Fc-attenuated backbone. Here, we describe some of the biophysical and functional characteristics that support the rationale for clinical evaluation of GSK6097608. Citation Format: Yan Degenhardt, Jun Guan, Peter Morley, David Jones, Michael Conner, Stephen Eastman, Wei Wang, Andrew Sanderson, Anand Ravindran, Julie Krueger, Iris Roth, James Smothers, Jeremy D. Waight. Discovery and characterization of the CD96 antibody GSK6097608, a high-affinity, antagonistic anti-CD96 antibody for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6268.

Precision Medicine in Cholangiocarcinoma: Past, Present, and Future.

Cholangiocarcinoma (CCA), or biliary tract cancer, has a poor prognosis. The median survival time among patients with CCA is under 2 years from diagnosis, and the global 5-year survival rate is only 10%. First-line therapy with chemotherapeutic agents, gemcitabine plus cisplatin, has traditionally been used to treat unresectable advanced CCA. In recent years, precision medicine has become a mainstream cancer treatment due to innovative next-generation sequencing technology. Several genetic alterations, including mutations, gene fusions, and copy number variations, have been found in CCA. In this review, we summarized the current understanding of genetic profiling in CCA and targeted therapy in CCA. Owing to the high heterogeneity of CCA, tumor microenvironmental factors, and the complexity of tumor biology, only pemigatinib, infigratinib, ivosidenib, larotrbctinib, and entrectinib are currently approved for the treatment of CCA patients with fibroblast growth factor receptor 2 gene (FGFR2) fusion, isocitrate dehydrogenase gene (IDH1) mutation, and neurotrophin receptor tyrosine kinase gene (NRTK) fusion, respectively. Additional targeted therapies, including other FGFR2 inhibitors, PI3K/AKT/mTOR inhibitors, and BRAF-directed targeted therapy, have been discussed for the management of CCA, and immune checkpoint inhibitors, particularly pembrolizumab, can be administered to patients with high microsatellite instability tumors. There is a further need for improvement in precision medicine therapies in the treatment of CCA and discuss the approved and potential targeted therapies for CCA.

Pan-cancer analysis of altered glycosyltransferases confers poor clinical outcomes.

Pan-cancer analysis of altered glycosyltransferases confers poor clinical outcomes.

Targeting PARP11 to avert immunosuppression and improve CAR T therapy in solid tumors.

Evasion of antitumor immunity and resistance to therapies in solid tumors are aided by an immunosuppressive tumor microenvironment (TME). We found that TME factors, such as regulatory T cells and adenosine, downregulated type I interferon receptor IFNAR1 on CD8+ cytotoxic T lymphocytes (CTLs). These events relied upon poly-ADP ribose polymerase-11 (PARP11), which was induced in intratumoral CTLs and acted as a key regulator of the immunosuppressive TME. Ablation of PARP11 prevented loss of IFNAR1, increased CTL tumoricidal activity and inhibited tumor growth in an IFNAR1-dependent manner. Accordingly, genetic or pharmacologic inactivation of PARP11 augmented the therapeutic benefits of chimeric antigen receptor T cells. Chimeric antigen receptor CTLs engineered to inactivate PARP11 demonstrated a superior efficacy against solid tumors. These findings highlight the role of PARP11 in the immunosuppressive TME and provide a proof of principle for targeting this pathway to optimize immune therapies.

High throughput screen for the improvement of inducible promoters for tumor microenvironment cues

Cancer immunotherapies are highly potent and are gaining wide clinical usage. However, severe side effects require focusing effector immune cell activities on the tumor microenvironment (TME). We recently developed a chimeric antigen receptor tumor-induced vector (CARTIV), a synthetic promoter activated by TME factors. To improve CARTIV functions including background, activation levels, and synergism, we screened a library of promoters with variations in key positions. Here, we present a screening method involving turning ON/OFF stimulating TNFα and IFNγ cytokines, followed by sequential cell sorting. Sequencing of enriched promoters identified seventeen candidates, which were cloned and whose activities were then validated, leading to the identification of two CARTIVs with lower background and higher induction. We further combined a third hypoxia element with the two-factor CARTIV, demonstrating additional modular improvement. Our study presents a method of fine-tuning synthetic promoters for desired immunotherapy needs.

Differential immune landscapes in appendicular versus axial skeleton.

Roughly 400,000 people in the U.S. are living with bone metastases, the vast majority occurring in the spine. Metastases to the spine result in fractures, pain, paralysis, and significant health care costs. This predilection for cancer to metastasize to the bone is seen across most cancer histologies, with the greatest incidence seen in prostate, breast, and lung cancer. The molecular process involved in this predilection for axial versus appendicular skeleton is not fully understood, although it is likely that a combination of tumor and local micro-environmental factors plays a role. Immune cells are an important constituent of the bone marrow microenvironment and many of these cells have been shown to play a significant role in tumor growth and progression in soft tissue and bone disease. With this in mind, we sought to examine the differences in immune landscape between axial and appendicular bones in the normal noncancerous setting in order to obtain an understanding of these landscapes. To accomplish this, we utilized mass cytometry by time-of-flight (CyTOF) to examine differences in the immune cell landscapes between the long bone and vertebral body bone marrow from patient clinical samples and C57BL/6J mice. We demonstrate significant differences between immune populations in both murine and human marrow with a predominance of myeloid progenitor cells in the spine. Additionally, cytokine analysis revealed differences in concentrations favoring a more myeloid enriched population of cells in the vertebral body bone marrow. These differences could have clinical implications with respect to the distribution and permissive growth of bone metastases.

Role of the Bone Marrow Microenvironment in Drug Resistance of Hematological Malignances.

The unique features of the tumor microenvironment (TME) govern the biological properties of many cancers, including hematological malignancies. TME factors can trigger an invasion and protect against drug cytotoxicity by inhibiting apoptosis and activating specific signaling pathways (e.g. NF-ΚB). TME remodeling is facilitated due to the high self-renewal ability of the bone marrow. Progressing tumor cells can alter some extracellular matrix (ECM) components which act as a barrier to drug penetration in the TME. The initial progression of the cell cycle is controlled by the MAPK pathway (Raf/MEK/ERK) and Hippo pathway, while the final phase is regulated by the PI3K/Akt /mTOR and WNT pathways. This review summarizes the main signaling pathways involved in drug resistance (DR) and some mechanisms by which DR can occur in the bone marrow. The relationship between autophagy, endoplasmic reticulum stress, and cellular signaling pathways in DR and apoptosis is covered in the TME.

Regulation of Epithelial-Mesenchymal Transition of A549 Cells by Prostaglandin D2.

Despite significant advances in diagnostic and operative techniques, lung cancer remains one of the most lethal malignancies worldwide. Since prostaglandins such as prostaglandin D2 (PGD2) is involved in various pathophysiological process, including inflammation and tumorigenesis, this study aims to investigate the role of PGD2 during the process of epithelial-mesenchymal transition (EMT) in A549 cells. A549 cells were stimulated with PGD2 and expression of EMT markers was analyzed by immunoblotting and immunofluorescence. EMT-related gene, Slug expression was evaluated using quantitative real-time polymerase chain reaction (qPCR). Migration and invasion abilities of A549 cells were determined in chemotaxis and Matrigel invasion assays, respectively. We also inhibited the TGF/Smad signaling pathway using a receptor inhibitor or silencing of TGF-β1 and TGFβ type I receptor (TGFβRI), and protein expressionwas assessed by immunoblotting and immunofluorescence. Here, we found that stimulation of A549 cells with PGD2 resulted in morphological changes into a mesenchymal-like phenotype under low serum conditions. Stimulation of A549 cells with PGD2 resulted in a significant reduction in proliferation, whereas invasion and migration were enhanced. The expression of E-cadherin was markedly downregulated, while Vimentin expression was upregulated after treatment of A549 cells with PGD2. Slug expression was markedly upregulated by stimulating A549 cells with PGD2, and stimulation of A549 cells with PGD2 significantly enhanced TGF-β1 expression, and silencing of TGF-β1 significantly blocked PGD2-induced EMT and Smad2 phosphorylation. In addition, PGD2-induced Smad2 phosphorylation and EMT were significantly abrogated by either pharmacological inhibition or silencing of TGFβRI. PGD2-induced expression of Slug and EMT were significantly augmented in low nutrient and low serum conditions. Finally, the subsequent culture of mesenchymal type of A549 cells under normal culture conditions reverted the cell's phenotype to an epithelial type. Given these results, we suggest that tumor microenvironmental factors such as PGD2, nutrition, and growth factors could be possible therapeutic targets for treating metastatic cancers.

Novel antiangiogenic therapy targeting biglycan using tumor endothelial cell-specific liposomal siRNA delivery system.

Tumor blood vessels play important roles in tumor progression and metastasis. Targeting tumor endothelial cells (TECs) is one of the strategies for cancer therapy. We previously reported that biglycan, a small leucine‐rich proteoglycan, is highly expressed in TECs. TECs utilize biglycan in an autocrine manner for migration and angiogenesis. Furthermore, TEC‐derived biglycan stimulates tumor cell migration in a paracrine manner leading to tumor cell intravasation and metastasis. In this study, we explored the therapeutic effect of biglycan inhibition in the TECs of renal cell carcinoma using an in vivo siRNA delivery system known as a multifunctional envelope‐type nanodevice (MEND), which contains a unique pH‐sensitive cationic lipid. To specifically deliver MEND into TECs, we incorporated cyclo(Arg–Gly–Asp–d–Phe–Lys) (cRGD) into MEND because αVβ3 integrin, a receptor for cRGD, is selective and highly expressed in TECs. We developed RGD‐MEND‐encapsulating siRNA against biglycan. First, we confirmed that MEND was delivered into OS‐RC‐2 tumor‐derived TECs and induced in vitro RNAi‐mediated gene silencing. MEND was then injected intravenously into OS‐RC‐2 tumor‐bearing mice. Flow cytometry analysis demonstrated that MEND was specifically delivered into TECs. Quantitative RT‐PCR indicated that biglycan was knocked down by biglycan siRNA‐containing MEND. Finally, we analyzed the therapeutic effect of biglycan silencing by MEND in TECs. Tumor growth was inhibited by biglycan siRNA‐containing MEND. Tumor microenvironmental factors such as fibrosis were also normalized using biglycan inhibition in TECs. Biglycan in TECs can be a novel target for cancer treatment.