Fragment size and level of cell-free DNA provide prognostic information in patients with advanced pancreatic cancer.

BACKGROUND: It was recently demonstrated that the size of cell-free DNA (cfDNA) fragments that originates from tumor cells are shorter than cfDNA fragments that originates from non-malignant cells. We investigated whether cfDNA fragment size and cfDNA levels might have prognostic value in patients with advanced pancreatic cancer. METHODS: Blood samples were obtained from patients with advanced pancreatic cancer, before (n = 61) initiation of chemotherapy and after the first cycle of chemotherapy (n = 39). Samples were separated with density centrifugation and plasma DNA was isolated. Mode cfDNA fragment size and cfDNA levels were then determined using a 2100 Bioanalyzer. A cohort of partially age-matched healthy volunteers (n = 28) constituted the control group. RESULTS: Both a pre-treatment cfDNA fragment size of ≤ 167 bp (mode) and high pre-treatment cfDNA levels were associated with shorter progression-free survival (PFS) (p = 0.002 and p < 0.001, respectively) and overall survival (OS) (p = 0.001 and p = 0.001, respectively). Furthermore, multivariable Cox regression analyses demonstrated that pre-treatment cfDNA levels could independently predict prognosis for both PFS (HR = 3.049, p = 0.005) and OS (HR = 2.236, p = 0.028). CONCLUSION: This study demonstrates that cfDNA fragment size and cfDNA levels can be used to predict disease outcome in patients with advanced pancreatic cancer. The described approach, using a rapid, economic and simple test to reveal prognostic information, has potential for future treatment stratification and monitoring.

Coculture of native human acute myelogenous leukemia blasts with fibroblasts and osteoblasts results in an increase of vascular endothelial growth factor levels.

OBJECTIVES: Angiogenesis seems important in the development of acute myelogenous leukemia (AML). Proangiogenic vascular endothelial growth factor (VEGF) is constitutively secreted by the AML blasts for a subset of patients, but it can also be released by non-leukemic bone marrow cells. METHODS: VEGF levels were determined after coculture of native human AML blasts with fibroblast lines, osteoblastic sarcoma cell lines, normal bone marrow stromal cells and normal osteoblasts. Cultures were prepared with leukemic and non-leukemic cells separated by a semipermeable membrane or in direct contact. RESULTS: The non-leukemic cells usually showed higher spontaneous VEGF release than AML cell populations. Coculture of AML blasts with HFL1 fibroblasts caused a supra-additive increase of VEGF levels when the cell populations were cultured separately, and the increase was also observed when cells were cultured in direct contact. An increase was also observed when AML blasts were cultured with osteoblastic sarcoma cells, normal bone marrow stromal cells and normal osteoblasts. Coculture had divergent effects on VEGF mRNA levels both for leukemic and non-leukemic cells, but increased mRNA levels were commonly observed especially for the non-leukemic cells. Cytokine inhibition experiments suggest that IL1 is important for the VEGF-increasing crosstalk, whereas the mechanisms are probably heterogeneous for coculture with osteoblasts. CONCLUSION: The bi-directional crosstalk via local cytokine networks between AML blasts and non-leukemic cells results in increased local VEGF levels, an observation suggesting that VEGF-targeting antiangiogenic therapy should be considered as a general therapeutic strategy in AML.

Osteoblasts increase proliferation and release of pro-angiogenic interleukin 8 by native human acute myelogenous leukemia blasts.

BACKGROUND AND OBJECTIVES: Interactions between acute myelogenous leukemia (AML) blasts and non-leukemic cells in the bone marrow seem to be important for both disease development and susceptibility to chemotherapy. Recent studies have focused on the endothelial cells, but other non-leukemic cells may also be involved. In the present study we investigated how osteoblasts affect native human AML blasts. DESIGN AND METHODS: AML cells were derived from a large group of consecutive patients. The AML blasts and osteoblastic sarcoma cell lines (Cal72, SJSA-1) were incubated together in different chambers separated by a semipermeable membrane. We investigated effects of co-culture on proliferation, apoptosis and cytokine release. RESULTS: The cross-talk between these two cell populations, achieved via release of soluble mediators, resulted in increased AML blast proliferation, including increased proliferation of clonogenic progenitors, but did not affect spontaneous in vitro apoptosis. Both interleukin (IL) 1-b and granulocyte-macrophage colony-stimulating factor were involved in this growth-enhancing cross-talk, and normal osteoblasts could also increase the AML blast proliferation. Furthermore, co-culture of AML blasts with osteoblastic sarcoma cells as well as normal osteoblasts increased the levels of the pro-angiogenic mediator IL8. INTERPRETATION AND CONCLUSIONS: Our in vitro results suggest that the release of soluble mediators by osteoblasts supports leukemic hematopoiesis through two major mechanisms: (i) direct enhancement of AML blast proliferation; and (ii) enhanced angiogenesis caused by increased IL8 levels.