Acute lymphoblastic leukaemia (ALL) has a marked propensity to metastasize to the central nervous system (CNS). In contrast to brain metastases from solid tumours, metastases of ALL seldom involve the parenchyma but are isolated to the leptomeninges, which is an infrequent site for carcinomatous invasion. Although metastasis to the CNS occurs across all subtypes of ALL, a unifying mechanism for invasion has not yet been determined. Here we show that ALL cells in the circulation are unable to breach the blood-brain barrier in mice; instead, they migrate into the CNS along vessels that pass directly between vertebral or calvarial bone marrow and the subarachnoid space. The basement membrane of these bridging vessels is enriched in laminin, which is known to coordinate pathfinding of neuronal progenitor cells in the CNS. The laminin receptor α6 integrin is expressed in most cases of ALL. We found that α6 integrin-laminin interactions mediated the migration of ALL cells towards the cerebrospinal fluid in vitro. Mice with ALL xenografts were treated with either a PI3Kδ inhibitor, which decreased α6 integrin expression on ALL cells, or specific α6 integrin-neutralizing antibodies and showed significant reductions in ALL transit along bridging vessels, blast counts in the cerebrospinal fluid and CNS disease symptoms despite minimally decreased bone marrow disease burden. Our data suggest that α6 integrin expression, which is common in ALL, allows cells to use neural migratory pathways to invade the CNS.

• Stromal OPN anchors leukemia cells in prodormancy BM niches.• Inhibiting this interaction leads dormant cells to proliferate, sensitizing them to chemotherapy.Malignant cells may evade death from cytotoxic agents if they are in a dormant state. The host microenvironment plays important roles in cancer progression, but how niches might control cancer cell dormancy is little understood. Here we show that osteopontin (OPN), an extracellular matrix molecule secreted by osteoblasts, can function to anchor leukemic blasts in anatomic locations supporting tumor dormancy. We demonstrate that acute lymphoblastic leukemia (ALL) cells specifically adhere to OPN in vitro and secrete OPN when localized to the endosteal niche in vivo. Using intravital microscopy to perform imaging studies of the calvarial bone marrow (BM) of xenografted mice, we show that OPN is highly expressed adjacent to dormant tumor cells within the marrow. Inhibition of the OPN-signaling axis significantly increases the leukemic cell Ki-67 proliferative index and leads to a twofold increase in tumor burden in treated mice. Moreover, using cell-cycle-dependent Ara-C chemotherapy to produce minimal residual disease (MRD) in leukemic mice, we show that OPN neutralization synergizes with Ara-C to reduce detectable BM MRD. Taken together, these data suggest that ALL interacts with extracellular OPN within the malignant BM, and that this interaction induces cell cycle exit in leukemic blasts, protecting them from cytotoxic chemotherapy. (Blood. 2013;121(24):4821-4831) IntroductionAcute lymphoblastic leukemia (ALL) in adults initially responds well to induction chemotherapy, with greater than 80% of patients attaining a complete remission (CR). Unfortunately, most initial CRs are short lived, and overall survival rate is only 30% to 40% for adults who are diagnosed before age 60 years.1 Although outcomes in the pediatric population are better, a significant number of patients still experience relapsed or refractory disease.2 Relapses in both populations are believed to be the outgrowth of minimal residual disease (MRD) that is not completely eliminated by chemotherapy. Indeed, it has been demonstrated that patients with the lowest levels of detectable MRD at CR have the best prognosis and least likelihood of relapse.2 Strategies to overcome resistance and reduce MRD may therefore have the potential to increase overall survival duration.Antiapoptotic signals from the host tissue microenvironment are increasingly recognized as important mechanisms of malignant cell survival against chemotherapy. Our previous work using the Nalm-6 model of ALL has shown that the bone marrow (BM) microenvironment plays a critical role in disease spread and in the dysregulation of normal hematopoiesis that occurs during leukemic growth. 3,4 To metastasize and outcompete native BM cells, leukemic cells co-opt normal signaling mechanisms within hematopoietic stem cell (HSC) niches. At least 2 distinct HSC niches, one perivascular and one endosteal (or bony), exist in the BM. 5 In t...

Breast cancer metastatic relapse can occur years after therapy, indicating that disseminated breast cancer cells (BCCs) have a prolonged dormant phase before becoming proliferative. A major site of disease dissemination and relapse is bone, although the critical signals that allow circulating BCCs to identify bone microvasculature, enter tissue, and tether to the microenvironment are poorly understood. Using real-time in vivo microscopy of bone marrow (BM) in a breast cancer xenograft model, we show that dormant and proliferating BCCs occupy distinct areas, with dormant BCCs predominantly found in E-selectin- and stromal cell-derived factor 1 (SDF-1)-rich perisinusoidal vascular regions. We use highly specific inhibitors of E-selectin and C-X-C chemokine receptor type 4 (CXCR4) (SDF-1 receptor) to demonstrate that E-selectin and SDF-1 orchestrate opposing roles in BCC trafficking. Whereas E-selectin interactions are critical for allowing BCC entry into the BM, the SDF-1/CXCR4 interaction anchors BCCs to the microenvironment, and its inhibition induces mobilization of dormant micrometastases into circulation. Homing studies with primary BCCs also demonstrate that E-selectin regulates their entry into bone through the sinusoidal niche, and immunohistochemical staining of patient BMs shows dormant micrometastatic disease adjacent to SDF-1(+) vasculature. These findings shed light on how BCCs traffic within the host, and suggest that simultaneous blockade of CXCR4 and E-selectin in patients could molecularly excise dormant micrometastases from the protective BM environment, preventing their emergence as relapsed disease.

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Visual, block-based programming environments present an alternative way of teaching programming to novices and have proven successful in classrooms and informal learning settings. However, few studies have been able to attribute this success to specific features of the environment. In this study, we isolate the most fundamental feature of these environments, the block interface, and compare it directly to its textual counterpart. We present analysis from a study of two groups of novice programmers, one assigned to each interface, as they completed a simple programming activity. We found that while the interface did not seem to affect users' attitudes or perceived difficulty, students using the block interface spent less time off task and completed more of the activity's goals in less time.