The SF3B1 inhibitor spliceostatin A (SSA) elicits apoptosis in chronic lymphocytic leukemia cells through downregulation of Mcl-1

The pro-survival Bcl-2 family member Mcl-1 is expressed in chronic lymphocytic leukemia (CLL), with high expression correlated with progressive disease. The spliceosome inhibitor spliceostatin A (SSA), is known to regulate Mcl-1 and so here we assessed the ability of SSA to elicit apoptosis in CLL. SSA induced apoptosis of CLL cells at low nanomolar concentrations in a dose- and time-dependent manner, but independently of SF3B1 mutational status, IGHV status and CD38 or ZAP70 expression. However, normal B and T cells were less sensitive than CLL cells (P=0.006 and P<0.001, respectively). SSA altered the splicing of anti-apoptotic MCL-1L to MCL-1s in CLL cells coincident with induction of apoptosis. Overexpression studies in Ramos cells suggested Mcl-1 was important for SSA-induced killing since its expression inversely correlated with apoptosis (P=0.001). IL4 and CD40L, present in patient lymph nodes, are known to protect tumor cells from apoptosis and significantly inhibited SSA, ABT-263 and ABT-199 induced killing following administration to CLL cells (P=0.008). However, by combining SSA with the Bcl-2/Bcl-xL antagonists ABT-263 or ABT-199, we were able to overcome this pro-survival effect. We conclude that SSA combined with Bcl-2/Bcl-xL antagonists may have therapeutic utility for CL

Genetics and prognostication in splenic marginal zone lymphoma: revelations from deep sequencing

PURPOSE: Mounting evidence supports the clinical significance of gene mutations and immunogenetic features in common mature B-cell malignancies.EXPERIMENTAL DESIGN: We undertook a detailed characterization of the genetic background of splenic marginal zone lymphoma (SMZL), using targeted re-sequencing and explored potential clinical implications in a multinational cohort of 175 SMZL patients.RESULTS: We identified recurrent mutations in TP53 (16%), KLF2 (12%), NOTCH2 (10%), TNFAIP3 (7%), MLL2 (11%), MYD88 (7%) and ARID1A (6%), all genes known to be targeted by somatic mutation in SMZL. KLF2 mutations were early, clonal events, enriched in patients with del(7q) and IGHV1-2*04 B-cell receptor immunoglobulins, and were associated with a short median time-to-first-treatment (0.12 vs. 1.11 yrs; P=0.01). In multivariate analysis mutations in NOTCH2 (HR 2.12, 95%CI 1.02-4.4, P=0.044) and 100% germline IGHV gene identity (HR 2.19, 95%CI 1.05-4.55, P=0.036) were independent markers of short time-to-first-treatment, while TP53 mutations were an independent marker of short overall survival (HR 2.36, 95% CI 1.08-5.2, P=0.03).CONCLUSIONS: We identify key associations between gene mutations and clinical outcome, demonstrating for the first time that NOTCH2 and TP53 gene mutations are independent markers of reduced treatment-free and overall survival, respectively.<br/

Determination and assessment of the invariant position of the X-ray beam focal spot with the flattening filter generated by the TrueBeam accelerator

Zastosowanie małych pól promieniowania w radioterapii stereotaktycznej pozwala na precyzyjne zdeponowanie dawki w obszarze zmiany nowotworowej przy jednoczesnym maksymalnym oszczędzeniu zdrowych tkanek. Zastosowanie obrazowania (IGRT) podczas procesu radioterapii pozwala na jej osiągnięcie, pod warunkiem zachowania wymaganego poziomu zgodność położenia izocentrum obrazowania i promieniowania. Położenie ogniska (focal spot) wiązki promieniowania X jest jednym z parametrów, które mogą mieć wpływ zarówno na położenie izocentrum promieniowania, jak i symetrię wiązki, a także szerokość półcieni – szczególnie dla małych pól stosowanych w radioterapii stereotaktycznej. Na podstawie pracy Chojnowskiego (J Appl Clin Med Phys. 2017;18(5):175-183) opracowano i wdrożono metodę badania położenia ogniska wiązki promieniowania X względem osi obrotu kolimatora. Rejestrowano obrazy portalowe generowane przez wiązki wysokoenergetycznego promieniowania X z filtrem spłaszczającym dla dwóch aparatów TrueBeam firmy Varian zainstalowanych w Zakładzie Radioterapii I w Centrum Onkologii – Instytucie im. Marii Skłodowskiej-Curie w Warszawie. Obrazy uzyskane dla pól kształtowanych przez szczęki kolimatora głównego i przez kolimator wielolistkowy analizowano z wykorzystaniem programu ImageJ. Dla obu akceleratorów odległość między ogniskiem wiązki a osią obrotu kolimatora mierzona w płaszczyźnie lateralnej nie przekraczała 0,2 mm, a w ponad 90% przypadków mieściła się w granicach 0,1 mm. Odległości mierzone w płaszczyźnie strzałkowej były większe i sięgały 0,76 mm, przy czym dla jednego z akceleratorów nie przekraczały 0,4 mm.The use of small fields of radiation in stereotactic radiotherapy allows for precise depositing of the dose in the area of neoplastic lesions with the simultaneous maximum saving of healthy tissues. The use of imaging (IGRT) during the radiotherapy process allows it to be achieved, provided that the required level compliance of the imaging and radiation isocenter is maintained. The focal spot position of the X-ray beam is one of the parameters that can affect both the position of the radiation isocentre and the symmetry of the beam, as well as the width of penumbra – especially for small fields used in stereotactic radiotherapy. Based on the Chojnowski paper (J Appl Clin Med Phys. 2017;18(5):175-183), a method for examining the location of the focal spot X-ray beam relative to the collimator rotation axis was developed and implemented. Portal images generated by high-energy X-ray beams with a flattening filter for two True- Beam Varian accelerators installed in The Maria Sklodowska – Curie Memorial Cancer Center and Institute of Oncology in Warsaw were registered. The images obtained for the fields shaped by the jaws of the main collimator and the multi-leaf collimator were analyzed using the ImageJ program. For both accelerators, the distance between the beam focal spot and the collimator rotation axis measured in the crossplane direction did not exceed 0.2 mm, and in more than 90% of cases it was within 0.1 mm. The distances measured in the inplane direction were larger and for one accelerator reached even 0.76 mm, whereas for the one the range of results was smaller and did not exceed 0.4 mm