Neuroimmune/Hematopoietic Axis with Distinct Regulation by the High-Mobility Group Box 1 in Association with Tachykinin Peptides.

Hematopoiesis is tightly regulated by the bone marrow (BM) niche. The niche is robust, allowing for the return of hematopoietic homeostasis after insults such as infection. Hematopoiesis is partly regulated by soluble factors, such as neuropeptides, substance P (SP), and neurokinin A (NK-A), which mediate hematopoietic stimulation and inhibition, respectively. SP and NK-A are derived from the Tac1 gene that is alternately spliced into four variants. The hematopoietic effects of SP and NK-A are mostly mediated via BM stroma. Array analyses with 2400 genes indicated distinct changes in SP-stimulated BM stroma. Computational analyses indicated networks of genes with hematopoietic regulation. Included among these networks is the high-mobility group box 1 gene (HMGB1), a nonhistone chromatin-associated protein. Validation studies indicated that NK-A could reverse SP-mediated HMGB1 decrease. Long-term culture-initiating cell assay, with or without NK-A receptor antagonist (NK2), showed a suppressive effect of HMGB1 on hematopoietic progenitors and increase in long-term culture-initiating cell assay cells (primitive hematopoietic cells). These effects occurred partly through NK-A. NSG mice with human hematopoietic system injected with the HMGB1 antagonist glycyrrhizin verified the in vitro effects of HMGB1. Although the effects on myeloid lineage were suppressed, the results suggested a more complex effect on the lymphoid lineage. Clonogenic assay for CFU- granulocyte-monocyte suggested that HMGB1 may be required to prevent hematopoietic stem cell exhaustion to ensure immune homeostasis. In summary, this study showed how HMGB1 is linked to SP and NK-A to protect the most primitive hematopoietic cell and also to maintain immune/hematopoietic homeostasis.

Evaluation of a developmental hierarchy for breast cancer cells to assess risk-based patient selection for targeted treatment.

This study proposes that a novel developmental hierarchy of breast cancer (BC) cells (BCCs) could predict treatment response and outcome. The continued challenge to treat BC requires stratification of BCCs into distinct subsets. This would provide insights on how BCCs evade treatment and adapt dormancy for decades. We selected three subsets, based on the relative expression of octamer-binding transcription factor 4 A (Oct4A) and then analysed each with Affymetrix gene chip. Oct4A is a stem cell gene and would separate subsets based on maturation. Data analyses and gene validation identified three membrane proteins, TMEM98, GPR64 and FAT4. BCCs from cell lines and blood from BC patients were analysed for these three membrane proteins by flow cytometry, along with known markers of cancer stem cells (CSCs), CD44, CD24 and Oct4, aldehyde dehydrogenase 1 (ALDH1) activity and telomere length. A novel working hierarchy of BCCs was established with the most immature subset as CSCs. This group was further subdivided into long- and short-term CSCs. Analyses of 20 post-treatment blood indicated that circulating CSCs and early BC progenitors may be associated with recurrence or early death. These results suggest that the novel hierarchy may predict treatment response and prognosis.

Mesenchymal Stem Cell-Derived Exosomes Stimulate Cycling Quiescence and Early Breast Cancer Dormancy in Bone Marrow.

Dormant breast cancers resurge as metastatic disease after a long dormancy period in the bone marrow, where cancer cells interact with mesenchymal stem cells (MSC). However, the nature of early interactions between breast cancer cells and MSCs in the bone marrow microenvironment that facilitate adaptation to a quiescent state remains poorly understood. Here, we report that breast cancer cells prime MSC to release exosomes containing distinct miRNA contents, such as miR-222/223, which in turn promotes quiescence in a subset of cancer cells and confers drug resistance. Building on these results, we developed a novel, nontoxic therapeutic strategy to target dormant breast cancer cells based on systemic administration of MSC loaded with antagomiR-222/223. In an immunodeficient mouse model of dormant breast cancer, this therapy sensitized breast cancer cells to carboplatin-based therapy and increased host survival. Overall, our findings illuminate the nature of the regulatory interactions between breast cancer cells and MSCs in the evolution of tumor dormancy and resurgence in the micrometastatic microenvironment of the bone marrow. Cancer Res; 76(19); 5832-44. ©2016 AACR.

Is reduction of tumor burden sufficient for the 21st century?

Currently, animal models are used to test the efficacy of tumor treatment. A significant reduction of tumor mass is lauded as great improvement. As we begin the 21st century, one wonders if this is sufficient and acceptable for cancer treatment. Although the presence of cancer stem cell (CSCs) is not a new phenomenon, their role in the initiation of the tumor for clinical resurgence is mostly ignored when testing drugs. The current treatment then poses a major limitation to aggressively target the cells most responsible for tumor initiation and resurgence. The review does not trivialize the problem since it is acknowledged that the tumors and cells within the tissue microenvironment would interact through complex mechanisms. It is quite possible that the interaction by CSCs and the microenvironment will vary, depending on the tissue, e.g., bone marrow versus brain. Research studies are needed to investigate if CSCs from the same organ differ after migrating to other tissues. If so, this will pose an economic dilemma for targeted drug development. It will not be feasible to develop drugs for each organ. Besides, the cost, there could be problems to effectively deliver the drugs to all organs, problems to assess drug distribution to particular tissues and toxicity for specific drugs. If multiple drugs are required to eradicate CSCs in different tissues, there is a problem of possible untoward effect for the simultaneous delivery of multiple drugs to a single cancer patient. As new drugs are developed, the investigators will need to pay attention for dedifferentiation of non-CSCs to CSCs. The metabolic pathways will have to be given equal attention as the stem cells genes since their pathways might show major differences rather than the stem cells genes, which are shared by the normal stem cells.

Hierarchy of breast cancer cells: key to reverse dormancy for therapeutic intervention.

An understanding of how cancer cells adapt dormancy would allow for targeted treatment. The current literature suggests that the cancer stem cells might be the major cells with the ability to become quiescent and to resist current drug treatment. The properties of cancer stem cells and healthy stem cells are functionally similar, thereby posing a challenge to target the dormant cells. The bone marrow is particularly a challenge because the dormant breast cancer cells are close to the endosteum, which is also home to the endogenous hematopoietic stem cells. Here we discuss how research studies could bring an understanding of the cellular and molecular interactions between the cancer stem cells and cells within the bone marrow microenvironment. This will allow for intervention to reverse dormancy for targeted treatment. The treatment will require studies within the normal organ functions to ensure treatment without toxicity.

Treg/Th17 polarization by distinct subsets of breast cancer cells is dictated by the interaction with mesenchymal stem cells.

Breast cancer (BC) cells (BCCs) exist within a hierarchy beginning with cancer stem cells (CSCs). Unsorted BCCs interact with mesenchymal stem cells (MSCs) to induce regulatory T cells (Tregs). This study investigated how distinct BCC subsets interacted with MSCs to polarize T-cell response, Tregs versus T helper 17 (Th17). This study tested BC initiating cells (CSCs) and the relatively more mature early and late BC progenitors. CSCs interacted with the highest avidity to MSCs. This interaction required CXCR4 and connexin 43 (Cx43)-dependant gap junctional intercellular communication (GJIC). This interaction induced Treg whereas interactions between MSCs and the progenitors induced Th17 response. The increases in Treg and Th17 depended on MSCs but not CTLA-4, which was increased in the presence of MSCs. Studies with BM stroma (fibroblasts) and MSCs from the same donors, indicated specific effects of MSCs. In total, MSC-CSC interaction required CXCR4 for GJIC. This led to increased Tregs and TGFß, and decreased Th17. In contrast, late and early BCCs showed reduced formation of GJIC, decreased Treg and increased Th17 and IL-17. These findings have significance to the methods by which CSCs evade the immune response. The findings could provide methods of intervention to reverse immune-mediated protection and support of BC.

Psychiatric medications: adverse cutaneous drug reactions.

Psychiatric medications are among the most widely prescribed medications in the United States. Adverse cutaneous drug reactions are associated with psychiatric medications in approximately 2% to 5% of the individuals for whom they are prescribed. Although most adverse cutaneous drug reactions associated with psychotropic medications are benign and easily treated, some can be disfiguring or life-threatening, particularly those associated with the mood stabilizers. Adverse cutaneous drug reactions associated with antidepressants, antipsychotics, and mood stabilizers are reviewed, and important issues that are of concern for the dermatologist who must consider when and how to safely discontinue a psychotropic medication in their patients are presented.

Delivery of Functional Anti-miR-9 by Mesenchymal Stem Cell-derived Exosomes to Glioblastoma Multiforme Cells Conferred Chemosensitivity.

Glioblastoma multiforme (GBM), the most common and lethal tumor of the adult brain, generally shows chemo- and radioresistance. MicroRNAs (miRs) regulate physiological processes, such as resistance of GBM cells to temozolomide (TMZ). Although miRs are attractive targets for cancer therapeutics, the effectiveness of this approach requires targeted delivery. Mesenchymal stem cells (MSCs) can migrate to the sites of cancers, including GBM. We report on an increase in miR-9 in TMZ-resistant GBM cells. miR-9 was involved in the expression of the drug efflux transporter, P-glycoprotein. To block miR-9, methods were developed with Cy5-tagged anti-miR-9. Dye-transfer studies indicated intracellular communication between GBM cells and MSCs. This occurred by gap junctional intercellular communication and the release of microvesicles. In both cases, anti-miR-9 was transferred from MSCs to GBM cells. However, the major form of transfer occurred with the microvesicles. The delivery of anti-miR-9 to the resistant GBM cells reversed the expression of the multidrug transporter and sensitized the GBM cells to TMZ, as shown by increased cell death and caspase activity. The data showed a potential role for MSCs in the functional delivery of synthetic anti-miR-9 to reverse the chemoresistance of GBM cells.Molecular Therapy-Nucleic Acids (2013) 2, e126; doi:10.1038/mtna.2013.60; published online 1 October 2013.

Exogenous CXCL12 activates protein kinase C to phosphorylate connexin 43 for gap junctional intercellular communication among confluent breast cancer cells.

Despite ongoing attempts to improve the overall breast cancer (BC) survival rate, BC cells' (BCCs) predilection for metastasizing to the bone marrow has enabled BCCs to not only remain dormant, but also evade detection. BCCs are able to acquire quiescence by establishing gap junctional intercellular communication (GJIC) with the stroma through the assembly of connexins (Cxs). The chemoattractant CXCL12 also appears to play a role in GJIC based on its tendency to decrease when GJIC is formed between BCCs and bone marrow stroma. This study investigates the role CXCL12 has on Cx43 expression and PKC-mediated Cx43 phosphorylation. Cx43 gene reporter assays revealed that as the BCCs come in contact with each other and establish GJIC, there is an inverse relationship between CXCL12 level and Cx43 expression. Immunoblot analyses confirmed this relationship at the level of protein, showing decreased Cx43 and reduced Cx43 phosphorylation at higher CXCL12 concentrations. However, real-time PCR studies revealed little change in Cx43 mRNA levels, despite stimulation with different concentrations of CXCL12, indicating CXCL12's effect on Cx43 is post-translational, through phosphorylation. Immunoblot analyses and functional dye exchange studies showed activation of PKC by exogenous CXCL12 in the phosphorylation, which in turn, increased intercellular communication. These findings elucidate the importance of considering the microenvironment's role in micrometastasis in clinical studies pertaining to prospective breast cancer treatment.

Can breast cancer stem cells evade the immune system?

The evidence seems to be growing in favor of the stem cell theory of cancer with the emergence of studies demonstrating the parallel mechanisms of self renewing pathways in stem cells and particular subsets of cancer cells. The finding of leukemia stem cells and subsequently breast cancer stem cells (BCSC) further supports the concept. The importance of these findings lends itself to the selfrenewal properties of stem cells in addition to the survival characteristics of stem cells, mechanisms that will have to be overcome when creating treatment modalities. In particular, research has shown that stem cells and a specific type of stem cells, mesenchymal stem cells (MSC), have special drug effluxing properties and some interactions with particular cells of the immune system that may serve major roles in immunosuppresion and overall tumor cell survival. Furthermore, the recent discovery of epithelial to mesenchymal transition (EMT) has laid out a possible mechanism for tumor cells to lose particular phenotypic epithelial cell markers and gain phenotypic mesenchymal cell markers. This process is implicated in metastasis in addition to overall tumor survival and evasion of the immune system. This review examines the current understanding of how tumor cells evade the immune system, but will first explore stem cells, cancer stem cells, normal immune interaction with tumor cells, and EMT.

Gap junction-mediated import of microRNA from bone marrow stromal cells can elicit cell cycle quiescence in breast cancer cells.

Bone marrow (BM) metastasis of breast cancer (BC) can recur even decades after initial diagnosis and treatment, implying the long-term survival of disseminated cancer cells in a dormant state. Here we investigated the role of microRNAs (miRNA) transmitted from BM stroma to BC cells via gap junctions and exosomes in tumor cell quiescence. MDA-MB-231 and T47D BC cells arrest in G(0) phase of the cell cycle when cocultured with BM stroma. Analyses of miRNA expression profiles identified numerous miRNAs implicated in cell proliferation including miR-127, -197, -222, and -223 targeting CXCL12. Subsequently, we showed that these CXCL12-specific miRNAs are transported from BM stroma to BC cells via gap junctions, leading to reduced CXCL12 levels and decreased proliferation. Stroma-derived exosomes containing miRNAs also contributed to BC cell quiescence, although to a lesser degree than miRNAs transmitted via gap junctions. This study shows that the transfer of miRNAs from BM stroma to BC cells might play a role in the dormancy of BM metastases.

A selective phosphatase of regenerating liver phosphatase inhibitor suppresses tumor cell anchorage-independent growth by a novel mechanism involving p130Cas cleavage.

The phosphatase of regenerating liver (PRL) family, a unique class of oncogenic phosphatases, consists of three members: PRL-1, PRL-2, and PRL-3. Aberrant overexpression of PRL-3 has been found in multiple solid tumor types. Ectopic expression of PRLs in cells induces transformation, increases mobility and invasiveness, and forms experimental metastases in mice. We have now shown that small interfering RNA-mediated depletion of PRL expression in cancer cells results in the down-regulation of p130Cas phosphorylation and expression and prevents tumor cell anchorage-independent growth in soft agar. We have also identified a small molecule, 7-amino-2-phenyl-5H-thieno[3,2-c]pyridin-4-one (thienopyridone), which potently and selectively inhibits all three PRLs but not other phosphatases in vitro. The thienopyridone showed significant inhibition of tumor cell anchorage-independent growth in soft agar, induction of the p130Cas cleavage, and anoikis, a type of apoptosis that can be induced by anticancer agents via disruption of cell-matrix interaction. Unlike etoposide, thienopyridone-induced p130Cas cleavage and apoptosis were not associated with increased levels of p53 and phospho-p53 (Ser(15)), a hallmark of genotoxic drug-induced p53 pathway activation. This is the first report of a potent selective PRL inhibitor that suppresses tumor cell three-dimensional growth by a novel mechanism involving p130Cas cleavage. This study reveals a new insight into the role of PRL-3 in priming tumor progression and shows that PRL may represent an attractive target for therapeutic intervention in cancer.