Mark Wroblewski

Axl, a prognostic and therapeutic target in acute myeloid leukemia mediates paracrine crosstalk of leukemia cells with bone marrow stroma

Acute myeloid leukemia (AML) represents a clonal disease of hematopoietic progenitors characterized by acquired heterogenous genetic changes that alter normal mechanisms of proliferation, self-renewal, and differentiation.(1) Although 40% to 45% of patients younger than 65 years of age can be cured with current therapies, only 10% of older patients reach long-term survival.(1) Because only very few novel AML drugs were approved in the past 2 decades, there is an urgent need to identify novel targets and therapeutic strategies to treat underserved AML patients. We report here that Axl, a member of the Tyro3, Axl, Mer receptor tyrosine kinase family,(2-4) represents an independent prognostic marker and therapeutic target in AML. AML cells induce expression and secretion of the Axl ligand growth arrest-specific gene 6 (Gas6) by bone marrow-derived stromal cells (BMDSCs). Gas6 in turn mediates proliferation, survival, and chemoresistance of Axl-expressing AML cells. This Gas6-Axl paracrine axis between AML cells and BMDSCs establishes a chemoprotective tumor cell niche that can be abrogated by Axl-targeting approaches. Axl inhibition is active in FLT3-mutated and FLT3 wild-type AML, improves clinically relevant end points, and its efficacy depends on presence of Gas6 and Axl. Axl inhibition alone or in combination with chemotherapy might represent a novel therapeutic avenue for AML.

Mast cells decrease efficacy of anti-angiogenic therapy by secreting matrix-degrading granzyme B

Resistance towards VEGF-centered anti-angiogenic therapy still represents a substantial clinical challenge. We report here that mast cells alter the proliferative and organizational state of endothelial cells which reduces the efficacy of anti-angiogenic therapy. Consequently, absence of mast cells sensitizes tumor vessels for anti-angiogenic therapy in different tumor models. Mechanistically, anti-angiogenic therapy only initially reduces tumor vessel proliferation, however, this treatment effect was abrogated over time as a result of mast cell-mediated restimulation of angiogenesis. We show that mast cells secrete increased amounts of granzyme b upon therapy, which mobilizes pro-angiogenic laminin- and vitronectin-bound FGF-1 and GM-CSF from the tumor matrix. In addition, mast cells also diminish efficacy of anti-angiogenic therapy by secretion of FGF-2. These pro-angiogenic factors act beside the targeted VEGFA–VEGFR2-axis and reinduce endothelial cell proliferation and angiogenesis despite the presence of anti-angiogenic therapy. Importantly, inhibition of mast cell degranulation with cromolyn is able to improve efficacy of anti-angiogenic therapy. Thus, concomitant mast cell-targeting might lead to improved efficacy of anti-angiogenic therapy.Resistance towards VEGF-centered anti-angiogenic therapy is an important clinical challenge. Here, the authors show that mast cells mediate resistance to anti-angiogenetic inhibitors by altering the proliferative and organizational state of endothelial cells through mobilization of FGF-1 and GM-CSF from the tumor matrix and secretion of FGF-2.

Blockade of myeloid-derived suppressor cell expansion with all-trans retinoic acid increases the efficacy of anti-angiogenic therapy

Intrinsic and adaptive resistance hampers the success of antiangiogenic therapies (AAT), especially in breast cancer where this treatment modality has proven largely ineffective. Therefore, novel strategies to improve the efficacy of AAT are warranted. Solid tumors such as breast cancer are characterized by a high infiltration of myeloid-derived suppressor cells (MDSC), which are key drivers of resistance to AAT. Therefore, we hypothesized that all-trans retinoic acid (ATRA), which induces differentiation of MDSC into mature cells, could improve the therapeutic effect of AAT. ATRA increased the efficacy of anti-VEGFR2 antibodies alone and in combination with chemotherapy in preclinical breast cancer models. ATRA reverted the anti-VEGFR2-induced accumulation of intratumoral MDSC, alleviated hypoxia, and counteracted the disorganization of tumor microvessels. Mechanistic studies indicate that ATRA treatment blocked the AAT-induced expansion of MDSC secreting high levels of vessel-destabilizing S100A8. Thus, concomitant treatment with ATRA holds the potential to improve AAT in breast cancer and possibly other tumor types.Significance: Increasing the therapeutic efficiency of antiangiogenic drugs by reducing resistance-conferring myeloid-derived suppressor cells might improve breast cancer treatment.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/12/3220/F1.large.jpg Cancer Res; 78(12); 3220-32. ©2018 AACR.

BET-inhibition by JQ1 promotes proliferation and self-renewal capacity of hematopoietic stem cells

Although inhibitors of bromodomain and extra terminal domain (BET) proteins show promising clinical activity in different hematologic malignancies, a systematic analysis of the consequences of pharmacological BET inhibition on healthy hematopoietic (stem) cells is urgently needed. We found that JQ1 treatment decreases the numbers of pre-, immature and mature B cells while numbers of early pro-B cells remain constant. In addition, JQ1 treatment increases apoptosis in T cells, all together leading to reduced cellularity in thymus, bone marrow and spleen. Furthermore, JQ1 induces proliferation of long-term hematopoietic stem cells, thereby increasing stem cell numbers. Due to increased numbers, JQ1-treated hematopoietic stem cells engrafted better after stem cell transplantation and repopulated the hematopoietic system significantly faster after sublethal myeloablation. As quantity and functionality of hematopoietic stem cells determine the duration of life-threatening myelosuppression, BET inhibition might benefit patients in myelosuppressive conditions.

Abstract 3253: Mast cell-derived granzyme b contributes to resistance against anti-angiogenic therapy

Significance: Targeted therapies have revolutionized the treatment of cancer. However, efficacy of anti-angiogenic therapies is limited due to significant resistance. Recent studies showed that the tumor microenvironment is involved in resistance towards targeted anti-angiogenic treatment. Based on the correlation of mast cell (MC) density with tumor growth and angiogenesis we put forward the hypothesis that MC might be implicated in anti-angiogenic therapy resistance. Methods: C57BL/6J, NSG or MC-deficient KitW-sh (Wsh) mice were subcutaneously injected with 5×105 (Panc02 and EL4) or 1×106 (TD2) cells +/- bone marrow derived MC. Tumors were treated with 20 mg/kg anti-VEGFR2 antibody (DC101) or 25 mg/kg cromoglicic acid (Cromo). BrdU was injected 12 h before sacrifice. Results: We show that MC alter the proliferative and organizational state of endothelial cells (EC). MC dose-dependently induced EC-proliferation (158 ± 12%; *p In WT mice, AAT only initially reduced the proliferation of tumor vessels by 60% (*p Microarray analysis of tumor-resident MC unraveled increased expression levels of ECM-degrading granzyme b (gzmb) in response to therapy. MC-specific knock down of gzmb rendered tumors more susceptible for AAT and lowered the levels of alternative, pro-angiogenic mediators beside the VEGF-VEGFR2-axis in the tumor microenvironment. Conclusions: Our results indicate that tumor-resident MC interfere with AAT. We provide evidence that MC-derived gzmb liberates ECM-bound pro-angiogenic factors besides the targeted VEGF-VEGFR2 axis, thereby fine-tuning vessel maturation and proliferation, which ultimately decreases therapeutic efficacy. Importantly, knock down of gzmb and pharmacological inhibition of MC degranulation improved the therapeutic response towards AAT. Citation Format: Mark A. Wroblewski, Raimund Bauer, Miguel Cubas Cordova, Florian Udonta, Isabel Ben Batalla, Victoria Gensch, Stefanie Sawall, Jonas S. Waizenegger, Julian Pardo Jimeno, Klaus Pantel, Carsten Bokemeyer, Sonja Loges. Mast cell-derived granzyme b contributes to resistance against anti-angiogenic therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3253.

Abstract 1161: COX-2 blockade improves efficacy of VEGF-targeting drugs

Efficacy of anti-angiogenic drugs is hampered by hypoxia-induced resistance. Because cyclooxygenase-2 (Cox-2) is upregulated in hypoxic conditions we analyzed mRNA expression levels of cyclooxygenase-1 (Cox-1) and Cox-2 in GFP+ FACS-sorted tumor cells from 4T1 tumors after treatment with anti-VEGFR2 antibodies (DC101) or with sunitinib. Cox-2 but not Cox-1 mRNA was upregulated by 2.3-fold upon anti-angiogenic treatment. In addition we found 5.2-fold increased prostaglandin E2 levels in 4T1 tumors after anti-angiogenic therapy. We hypothesized that concomitant blockade of Cox-2 could increase efficacy of anti-angiogenic agents. Therefore we treated 4T1 tumor-bearing mice with sunitinib or DC101 alone and in combination with acetylsalicyclic acid (ASA). We found that single treatment with ASA or angiogenesis inhibitors inhibited tumor growth and that combined inhibition of Cox-2 and VEGF(R) signaling exerted additive therapeutic efficacy (n=5; 1142±84 (ASS); 1148±78 (Sunitnib) vs. 63±5 mg (combination); p In addition, Cox-2 and PGE2 can promote tumor angiogenesis. This alternative pro-angiogenic pathway would be enhanced by increased Cox-2 expression and PGE2 levels and could contribute to resistance against anti-angiogenic treatments. In line with this hypothesis the MVD was decreased 4T1 tumors treated with combined Cox-2 and VEGF blockade compared to the respective monotherapy (n=7; 31±2.4 (Sunitinib); 28.58±0.83 (ASS) vs. 9.67±1.71 (combination); p Citation Format: Isabel Ben Batalla, Miguel Cubas-Cordova, Florian Udonta, Mark Wroblewski, Stefanie Sawall, Victoria Gensch, Klaus Pantel, Carsten Bokemeyer, Sonja Loges. COX-2 blockade improves efficacy of VEGF-targeting drugs. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1161. doi:10.1158/1538-7445.AM2014-1161