Micrometastasis in breast cancer and other solid tumors

Hematogenous distant metastasis is the leading cause of cancer-related death in breast cancer and other solid tumors. By applying sensitive immunocytochemical and molecular assays, disseminated tumor cells (DTC) in bone marrow (BM) can be detected in 20-40% of cancer patients without any clinical or even histopathological signs of metastasis and the presence of these DTC at primary diagnosis predicts the subsequent occurrence of overt metastases in bone and other organs. cDNA-microarray analysis on primary breast carcinomas from patients with and without tumor cells in BM revealed a predominant downregulation of potential metastasis-suppressor genes in BM-positive tumors. Thus, dissemination of tumor cells appears to be an early process associated with a specific molecular signature of the primary tumor.     

Bone Metastasis: Current State of Play

Bone metastasis (BM) in cancer remains a critical issue because of its associated clinical and biological complications. Moreover, BM can alter the quality of life and survival rate of cancer patients. Growing evidence suggests that bones are a fertile ground for the development of metastasis through a “vicious circle” of bone resorption/formation and tumor growth. This review aims to outline the current major issues in the diagnosis and management of BM in the most common types of osteotropic cancers and describe the mechanisms and effects of BM. First, we discuss the incidence of BM through the following questions: Are we witnessing an increase in incidence, and are we now better equipped with modern imaging techniques? Is the advent of efficient bone resorption inhibitors affecting the bigger picture of BM management? Second, we discuss the potential effects of cancer progression and well-prescribed drugs, such as multitarget tyrosine kinase inhibitors, inhibitors of the mammalian target of rapamycin, and immune checkpoint inhibitors, on BM. Finally, we examine the duality of the effects of some therapies that may help in cancer treatment but may also contribute to further BM.     

BM micrometastases and circulating tumor cells in breast cancer patients: where have we been, where are we now and where does the future lie?

Over the past 15 years early tumor cell dissemination has been detected in patients with breast cancer using sensitive immunocytochemical and molecular assays based on the use of MAb and PCR, respectively. Clinical studies involving more than 4,000 breast cancer patients have now demonstrated that the presence of disseminated tumor cells in BM identified with immuncytochemical assays at primary diagnosis is a strong and independent prognostic factor. The published studies for the detection of disseminated tumor cells in BM fulfill the highest level of evidence as prognostic markers in primary breast cancer. In addition, various assays for the detection of circulating tumor cells in the peripheral blood have been developed recently and some studies suggest a potential clinical relevance of this parameter as a prognostic and predictive factor. Comparative analyzes indicate that the prognostic information derived from BM and blood screening seems to be complementary and not redundant. Advanced methods for molecular characterization of single tumor cells and the surrounding environment have been developed lately, and this approach allows new insights into the metastatic cascade and characterization of targets for therapeutic approaches. Taken together, these findings provide the basis for the implementation of disseminated tumor cells in BM or blood as markers for stratification and assessment of therapies in prospective clinical trials. The valuable information derived from these trials should help to improve future treatment of breast cancer patients.     

Predictive biochemical-markers for the development of brain metastases from lung cancer: Clinical evidence and future directions

BackgroundBrain metastases are a common complication of patients with lung cancer and lung cancer is one of the most common causes of brain metastases. The occurrence of brain metastases is associated with poor prognosis and high morbidity, even after intensive multimodal therapy. Therefore, identifying lung cancer patients with who are at high risk of developing brain metastases and applying effect intervention is important to reduce or delay the incidence of brain metastases. Biochemical-markers may meet an unmet need for following patients’ mechanisms of brain metastases.MethodsData for this review were identified by searches of Pubmed and Cochrane databases, and references from relevant articles using the search terms “lung cancer” and “brain metastasis”. Meeting abstracts, unpublished reports and review articles were not considered.ResultsClinical results for pathological and circulating markers including cancer molecular subtypes, miRNA, single nucleotide polymorphisms, and other markers are presented. However, these biochemical-markers are not yet established surrogate assessments for prediction of brain metastases.ConclusionsBiochemical-markers reported allowed physicians to identify which patients with lung cancer are at high risk for brain metastases. Prospective randomized clinical studies are needed to further assess the utility of these biochemical-markers.     

The role of adjuvant clodronate therapy for prevention of bone metastasis in breast cancer

Bisphosphonates are effective against increased bone resorption because they inhibit osteoclast activity. The use of these drugs is well established for the treatment of metastatic breast and other cancers; they reduce skeletal complications, hypercalcemia, bone pain, and metastatic progression and they can improve the overall survival and quality of life. Preclinical observations and early clinical data indicate that early bisphosphonate treatment reduces the incidence and the extent of newly developed metastases in breast cancer. There is considerable interest in determining whether bisphosphonate treatment is to prevent the incidence of bone metastases and associated complications. To date three randomized, controlled clinical trials have examined the effect of long-term use of clodronate (1600 mg/d po.) on the incidence of bone metastases, other metastases, the survival of patients, and the side effects of the study drug as well. All the trials have observed significant reduction of the occurrence of bone metastases, although this reduction was significant only during the medication period. One of the trials mentioned have shown an unexpected reduction in non-osseous metastases, and two of them have revealed significant improvements in the death rates. These promising results need further evaluation by large clinical trials with longer treatment periods to establish the clinical role of adjuvant bisphosphonate treatment of primary breast cancer.     

Detection and molecular characterisation of disseminated tumour cells: Implications for anti-cancer therapy

Haematogenous distant metastasis is the leading cause of cancer-related death in solid tumours. By applying sensitive immunocytochemical and molecular assays, disseminated tumour cells (DTC) in bone marrow (BM) can be detected in 20–40% of cancer patients without any clinical or even histopathological signs of metastasis, and the presence of these DTC at primary diagnosis predicts the subsequent occurrence of overt metastases in bone and other organs. The detection and characterisation of DTC in BM may lead to a better understanding of the biology initiating metastatic spread in cancer patients and will eventually contribute to the development of more effective strategies to eliminate DTC. In this review, we will therefore discuss the detection and characterisation of DTC in the light of new therapeutic strategies targeting tumour-associated molecules and signalling pathways.     

Femoral Bone Marrow Aspiration in Live Mice

Serial sampling of the cellular composition of bone marrow (BM) is a routine procedure critical to clinical hematology. This protocol describes a detailed step-by-step technical procedure for an analogous procedure in live mice which allows for serial characterization of cells present in the BM. This procedure facilitates studies aimed to detect the presence of exogenously administered cells within the BM of mice as would be done in xenograft studies for instance. Moreover, this procedure allows for the retrieval and characterization of cells enriched in the BM such as hematopoietic stem and progenitor cells (HSPCs) without sacrifice of mice. Given that the cellular composition of peripheral blood is not necessarily reflective of proportions and types of stem and progenitor cells present in the marrow, procedures which provide access to this compartment without requiring termination of the mice are very helpful. The use of femoral bone marrow aspiration is illustrated here for cytological analysis of marrow cells, flow cytometric characterization of the hematopoietic stem/progenitor compartment, and culture of sorted HSPCs obtained by femoral BM aspiration compared with conventional marrow harvest.     

A novel rich source of human mesenchymal stem cells from the debris of bone marrow samples

The debris from human bone marrow (BM) samples is generally filtered out and discarded prior to isolation of mesenchymal stem cells (MSCs). The purpose of this study is to develop a method to harvest MSCs from the debris and investigate their biological characteristics compared with the marrow counterparts. The BM tissue fragments were digested with collagenase and this treatment yielded mononuclear cells half to those from the corresponding filtered BM. The frequencies of colony-forming unit-fibroblast in these two cell populations were not significantly different. MSCs of two origins exhibited similar morphological and phenotypic features. Fluorescent dye-dilution assay showed that they grew at comparable rates both in the primary and passaging cultures. Further, they could be induced into osteoblasts, chondroblasts and adipocytes, as revealed by histological and molecular examinations. Thus, BM tissue fragments may serve as a new source of MSCs in the settings of bench experiments and clinical trials.     

Is there a role for antibody testing in the diagnosis of invasive candidiasis?

During the last decades, the use of antibody tests for the diagnosis of invasive mycoses has declined as a consequence of the general belief that they are insensitive and non-specific. However, there is a clear evidence that antibodies can be detected in highly immunodeficient patients (such as bone marrow transplant recipients), and that those antibodies are useful for the diagnosis. Antibody tests are currently in use as diagnostic tools for some primary mycoses, such as the endemic mycoses, aspergilloma, allergic bronchopulmonary aspergilosis and sporothrichosis. For invasive candidiasis, diagnostic methods must differentiate Candida colonization of mucous membranes or superficial infection from tissue invasion by this microorganism. Substantial progress has been made in diagnosis of invasive candidiasis with the development of a variety of methods for the detection of antibodies and antigens. However, no single test has found widespread clinical use and there is a consensus that diagnosis based on a single specimen lacks sensitivity. It is necessary to test sequential samples taken while the patient is at greatest risk for developing invasive candidiasis to optimize the diagnosis. Results obtained from a panel of diagnostic tests in association with clinical aspects will likely be the most useful strategy for early diagnosis and therapy.     

NBI及AFE在喉癌早期诊断中的应用

研究表明,喉癌的早期诊断、及时治疗不仅可以提高治愈率,而且也减少了患者的手术创伤和经济负担。积极开展喉癌的早期诊断研究具有重要的临床和社会意义。发现早期喉癌常规方法主要有电子喉镜、纤维喉镜、颈部CT及MRI检查,但并不能明显有效提高早期诊断率。而窄带成像(narrow band imaging,NBI)及自体荧光内镜(autofluorescence endoscopy AFE)是近几年用于喉癌早期诊断的两种新颖的内镜技术。NBI是一种通过变窄光波的波长,使粘膜上皮内乳头样毛细血管袢及粘膜下静脉的结构形成鲜明的对比,从而提高组织表面细微结构的对比度,便于发现病灶。而AFE技术是一种利用自发荧光聚集于病变组织的某个区域产生的差异强度,来区别正常组织与肿瘤性病变,从而用于肿瘤的早期诊断及识别癌前病变。因此,对NBI及AFE的进一步研究及认识对喉癌早期诊断提供非常重要的临床应用价值。     

Beta-CTX and ICTP act as indicators of skeletal metastasis status in male patients with non-small cell lung cancer

Bone metastasis is common in lung cancer patients and associated with reduced quality of life and reduced overall and median survival, so the early detection of bone metastasis and monitoring of its status is very important for clinicians. Serum bone-specific alkaline phosphatase (BAP), osteocalcin (OC), beta isomer of C-terminal telopeptide of type I collagen (beta-CTX) and cross-linked C-terminal telopeptide of type I collagen (ICTP) were compared with regard to their usefulness as indicators of bone metastasis in lung cancer. The serum concentrations of the 4 markers were measured by commercially available tests in 96 male patients with non-small cell lung cancer and 30 male patients with other pulmonary diseases. The levels of both a-CTX and ICTP were significantly higher in 61 lung cancer patients with bone metastases than in 35 lung cancer patients without bone metastases (both p<0.001), and significantly correlated with the extent of bone disease. Although ICTP had a better sensitivity and accuracy than beta-CTX (75.4% vs 65.6% and 72.9% vs 68.8%, respectively), they had a similar area under the receiver operating characteristic curve (0.85 vs 0.83). These results support the use of beta-CTX and ICTP as an adjunct tool for the diagnosis and screening of bone metastasis in lung cancer.     

Metabolomics of the Tumor Microenvironment in Pediatric Acute Lymphoblastic Leukemia

The tumor microenvironment is emerging as an important therapeutic target. Most studies, however, are focused on the protein components, and relatively little is known of how the microenvironmental metabolome might influence tumor survival. In this study, we examined the metabolic profiles of paired bone marrow (BM) and peripheral blood (PB) samples from 10 children with acute lymphoblastic leukemia (ALL). BM and PB samples from the same patient were collected at the time of diagnosis and after 29 days of induction therapy, at which point all patients were in remission. We employed two analytical platforms, high-resolution magnetic resonance spectroscopy and gas chromatography-mass spectrometry, to identify and quantify 102 metabolites in the BM and PB. Standard ALL therapy, which includes l-asparaginase, completely removed circulating asparagine, but not glutamine. Statistical analyses of metabolite correlations and network reconstructions showed that the untreated BM microenvironment was characterized by a significant network-level signature: a cluster of highly correlated lipids and metabolites involved in lipid metabolism (p<0.006). In contrast, the strongest correlations in the BM upon remission were observed among amino acid metabolites and derivatives (p<9.2×10^-10). This study provides evidence that metabolic characterization of the cancer niche could generate new hypotheses for the development of cancer therapies.     

Moving cancer diagnostics from bench to bedside

To improve treatment and reduce the mortality from cancer, a key task is to detect the disease as early as possible. To achieve this, many new technologies have been developed for biomarker discovery and validation. This review provides an overview of omics technologies in biomarker discovery and cancer detection, and highlights recent applications and future trends in cancer diagnostics. Although the present omic methods are not ready for immediate clinical use as diagnostic tools, it can be envisaged that simple, fast, robust, portable and cost-effective clinical diagnosis systems could be available in near future, for home and bedside use.     

Long-term survival after adoptive bone marrow T cell therapy of advanced metastasized breast cancer: follow-up analysis of a clinical pilot trial

Background

The bone marrow (BM) of breast cancer patients harbors tumor-reactive memory T cells (TCs) with therapeutic potential. We recently described the immunologic effects of adoptive transfer of ex vivo restimulated tumor-reactive memory TCs from the BM of 12 metastasized breast cancer patients in a clinical phase-I study. In this trial, adoptive T cell transfer resulted in the occurrence of circulating tumor antigen-reactive type-1 TCs. We here describe the long-term clinical outcome and its correlation with tumor-specific cellular immune response in 16 metastasized breast cancer patients, including 12 included in the original study.

Methods

Sixteen metastatic breast cancer patients with preexisting tumor-reactive BM memory TCs were included into the study. The study protocol involved one transfusion of TCs which were reactivated in vitro with autologous dendritic cells pulsed with lysates of MCF-7 breast cancer cells as source of tumor antigens. The presence of tumor-reactive memory TCs was analyzed by IFN-γ ELISpot assays.

Results

Tumor-reactive memory TCs in the peripheral blood were induced de novo in 7/16 patients (44 %) after adoptive TC transfer. These patients were considered immunologic responders to the therapy. Positive adoptive immunotherapy (ADI) response was observed significantly more often in patients without bone metastases (p = 0.0051), in patients with high levels of tumor-reactive BM TCs prior to therapy (p = 0.036) and correlated significantly with the estimated numbers of transferred tumor-reactive TCs (p = 0.0021). After the treatment, we observed an overall median survival of 33.8 months in the total cohort with three patients alive at last follow-up and more than 7 years after ADI. Numbers of transferred tumor-reactive TCs correlated significantly with the overall survival of patients (p = 0.017). Patients with an immunologic response to ADI in the peripheral blood had a significantly longer median survival than nonresponders (median survival 58.6 vs. 13.6 months; p = 0.009).

Conclusion

In metastasized breast cancer patients, adoptive transfer of BM TCs can induce the presence of tumor antigen-reactive type-1 TCs in the peripheral blood. Patients with immunologic response after ADI show a significantly longer overall survival. Patients with bone metastases significantly less frequently respond to the treatment and, therefore, might not be optimal candidates for ADI. Although the present study does not yet prove the therapeutic effect of ADI, these findings shed light on the relation between immune response and cancer prognosis and suggest that transfer of reactivated BM TCs might bear therapeutic potential.     

Bone marrow biopsy (BMB). II. Bone marrow biopsy in myeloproliferative disorders

The myeloproliferative disorders (MPD) are a domain in which the bone marrow biopsy (BMB) greatly proved its utility. We have studied the histology of the bone marrow (BM) in all the four entities of MPD: chronic myeloid leukemia (CML) with its subtype, chronic megakaryocytic granulocytic myelosis (CMGM), polycythemia vera (PV), hemorrhagic thrombocythemia (HT) and myeloid metaplasia with myelofibrosis (MMM). The work presents in short some of the clinical and hematologic characters of MPD with special stress upon the histologic modifications of BM, either specific or common to all MPD entities, underlying also the criteria for differential diagnosis.     

Mesenchymal stem cells in hematopoietic stem cell transplantation

Mesenchymal stromal/stem cells (MSC) of bone marrow (BM) origin not only provide the supportive microenvironmental niche for hematopoietic stem cells (HSC) but are capable of differentiating into various cell types of mesenchymal origin, such as bone, fat and cartilage. In vitro and in vivo data suggest that MSC have low inherent immunogenicity, modulate/suppress immunologic responses through interactions with immune cells, and home to damaged tissues to participate in regeneration processes through their diverse biologic properties. MSC derived from BM are being evaluated for a wide range of clinical applications, including disorders as diverse as myocardial infarction and newly diagnosed diabetes mellitus type 1. However, their use in HSC transplantation, either for enhancement of hematopoietic engraftment or for treatment/prevention of graft-versus-host disease, is far ahead of other indications. Ease of isolation and ex vivo expansion of MSC, combined with their intriguing immunomodulatory properties and their impressive record of safety in a wide variety of clinical trials, make these cells promising candidates for further investigation.     

Angiogenesis and the role of bone marrow endothelial cells in haematological malignancies

Increased microvessel density (MVD) has been observed in the bone marrow (BM) of patients with multiple myeloma (MM), acute lymphoblastic leukaemia, acute myeloid leukaemia, and myelodysplastic and myeloproliferative syndrome. The MVD is the net result of cumulative phases of angiogenesis and angio-regression and is as such not an indicator of the ongoing angiogenesis at the time of biopsy. There is, therefore, a need for additional methods that allow the estimation of ongoing angiogenesis. Double immunostainings for CD34 and Ki-67 can be used on paraffin-embedded tissue to determine the endothelial proliferation fraction. The BM endothelial cells, as a component of the BM stroma, have a close interaction with the malignant cells. In MM, for example, they are involved in the specific homing and are a source of paracrine growth factors. Targeting the BM microvessels will not only influence the nutrient and oxygen supply, but will in addition reduce the growth stimuli provided by the EC.     

Mesenchymal stromal cells from amniotic fluid are less prone to senescence compared to those obtained from bone marrow: An in vitro study

Mesenchymal stromal cells (MSCs) are considered to be an excellent source in regenerative medicine. They contain several cell subtypes, including multipotent stem cells. MSCs are of particular interest as they are currently being tested using cell and gene therapies for a number of human diseases. They represent a rare population in tissues; for this reason, they require, before being transplanted, an in vitro amplification. This process may induce replicative senescence, thus affecting differentiation and proliferative capacities. Increasing evidence suggests that MSCs from fetal tissues are significantly more plastic and grow faster than MSCs from bone marrow. Here, we compare amniotic fluid mesenchymal stromal cells (AF‐MSCs) and bone marrow mesenchymal stromal cells (BM‐MSCs) in terms of cell proliferation, surface markers, multidifferentiation potential, senescence, and DNA repair capacity. Our study shows that AF‐MSCs are less prone to senescence with respect to BM‐MSCs. Moreover, both cell models activate the same repair system after DNA damage, but AF‐MSCs are able to return to the basal condition more efficiently with respect to BM‐MSCs. Indeed, AF‐MSCs are better able to cope with genotoxic stress that may occur either during in vitro cultivation or following transplantation in patients. Our findings suggest that AF‐MSCs may represent a valid alternative to BM‐MSCs in regenerative medicine, and, of great relevance, the investigation of the mechanisms involved in DNA repair capacity of both AF‐MSCs and BM‐MSCs may pave the way to their rational use in the medical field.