Kurzfassung: Peptides are highly selective and efficacious for the treatment of cardiovascular and other diseases. However, it is currently not possible to administer peptides for cardiac-targeting therapy via a noninvasive procedure, thus representing scientific and technological challenges. We demonstrate that inhalation of small ( 〈 50 nm in diameter) biocompatible and biodegradable calcium phosphate nanoparticles (CaPs) allows for rapid translocation of CaPs from the pulmonary tree to the bloodstream and to the myocardium, where their cargo is quickly released. Treatment of a rodent model of diabetic cardiomyopathy by inhalation of CaPs loaded with a therapeutic mimetic peptide that we previously demonstrated to improve myocardial contraction resulted in restoration of cardiac function. Translation to a porcine large animal model provides evidence that inhalation of a peptide-loaded CaP formulation is an effective method of targeted administration to the heart. Together, these results demonstrate that inhalation of biocompatible tailored peptide nanocarriers represents a pioneering approach for the pharmacological treatment of heart failure.

Kurzfassung: Primary Myelofibrosis (PMF) is a Philadelphia-negative myeloproliferative neoplasm characterized by the presence of hyperplastic/dysplastic megakaryocytes and the development of myelofibrosis and osteosclerosis. This disruption of the bone marrow (BM) structure results from a deregulated release of growth factors by malignant hematopoietic cells, particularly monocytes and megakaryocytes, that activate BM stromal cells (Desterke-C, Mediators Inflamm, 2015). Therefore, it remains of primary importance to identify novel therapeutic strategies to restore BM microenvironment homeostasis in PMF patients. In order to better characterize the molecular mechanisms underlying PMF pathogenesis, we have recently compared the gene expression profile (GEP) of CD34+ hematopoietic progenitor cells (HPCs) from PMF patients and healthy donors, and we observed the upregulation of the transcription factor MAF (v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog) (Norfo-R, Blood, 2014). With the aim to investigate the role of MAF in PMF pathogenesis and to reproduce the condition observed in PMF CD34+ cells, we studied the effects of MAF overexpression on HPCs differentiation. Human cord blood-derived CD34+ cells were transduced with a retroviral vector carrying the coding sequences for the two MAF transcript variants (LMAFVAR1IDN and LMAFVAR2IDN). The megakaryocyte lineage-committed CD41+ population was increased in LMAFVAR1IDN (38.2±2.3%) and LMAFVAR2IDN (39.4±2.3%) samples compared with the empty vector LXIDN (16.1±2.7%) at day 3 after NGFR+ cells purification. Values are reported as mean ± SEM from 3 independent experiments, p 〈 .05. Moreover, MAF overexpression determined a remarkable increase of the CD14+ monocyte population in LMAFVAR1IDN (45.5±5.7%) and LMAFVAR2IDN (43.1±8.7%) compared to LXIDN control (8.0±0.4%) at t0 post-purification. Data represent mean ± SEM from 3 independent experiments, p 〈 .05. Next, we analyzed the GEP of MAF-overexpressing CD34+ cells and observed the upregulation of genes involved in megakaryopoiesis, such as MERTK and RCAN1, and in monocyte/macrophage differentiation (e.g. EGR2 and KLF4). In particular, GEP revealed the upregulation of genes related to processes deregulated in PMF, such as fibrosis (i.e. ENPP2, ADAM9) and inflammation (i.e. TNF, CCL2 and CCL3). Noteworthy, several upregulated transcripts encoded for secreted proteins. Based on their role in inflammation and fibrosis, we selected a set of secreted molecules (IL8, MMP9, CCL2, SPP1, LGALS3 and PLAUR) and assessed them by quantitative enzyme-linked immunoassay (ELISA) in the culture supernatants from LMAFVAR1IDN-, LMAFVAR2IDN- and LXIDN-transduced cells. The levels in culture supernatants of all these molecules were increased in MAF-overexpressing samples compared to control. In order to correlate the expression of these molecules with MAF upregulation in PMF, we assessed them in plasma samples of PMF patients (n=30) and healthy donors (n=10). All the selected molecules displayed higher plasma levels in PMF patients than in healthy controls (Figure 1). Since the increase of SPP1 and LGALS3 has never been described before in PMF, we investigated which cell type could be responsible for SPP1 and LGALS3 production. Real Time qRTPCR performed on healthy donors peripheral blood-derived cells showed that the highest SPP1 expression levels were observed in CD34+ cells-derived megakaryocytes and granulocytes, whereas LGALS3 was expressed at higher levels in monocytes compared to the other cell types. Because the increased proliferation of fibroblasts is involved in BM fibrosis and is a PMF feature, we investigated the effects of LGALS3 and SPP1 on normal human primary dermal fibroblasts. The treatment with recombinant human LGALS3 (1000 ng/ml) or SPP1 (500 ng/ml) caused an increase in fibroblasts cell count of 20.9±2.7% (p 〈 .001) and 14.9±4.1% (p 〈 .01) compared to the untreated control, respectively. Values are reported as mean ± SEM from 7 independent experiments. Finally, we observed that increased SPP1 plasma levels in PMF patients correlate with a more severe fibrosis and lower overall survival (p 〈 .05). In conclusion, our data unveil the involvement of MAF in PMF pathogenesis through the induction of a pro-fibrotic and pro-inflammatory phenotype in the malignant myeloid differentiated progeny. Disclosures Vannucchi: Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau.

Kurzfassung: Introduction Malignancy can be heralded by unprovoked venous thromboembolism (VTE) but also by arterial thrombosis. To date it is unknown whether this association is present also in myeloproliferative neoplasms (MPN), in which arterial thrombosis is more frequent that venous thrombosis and solid tumors are reported with an increased frequency. The MPN-K nested case-control study addressed the impact of cytoreductive drugs on the risk of developing second cancer in MPN patients (Barbui T et al, Leukemia 2019); here we re-examined the study database to evaluate the frequency and type of vascular complications in MPN patients with second cancer excluding leukemia and to establish whether arterial and venous thrombosis during follow-up after diagnosis of MPN could predict the occurrence of a second cancer. Patients and methods Cases were patients with second cancer diagnosed concurrently or subsequent to the diagnosis of MPN. Controls were MPN patients without second cancer. For each case with second cancer, up to 3 cancer-free controls were matched by each center for sex, age at MPN diagnosis, date of MPN diagnosis, and MPN disease duration. Each set consisting of one case and their matched-controls had a similar observational period (from MPN diagnosis until the index date of diagnosis for the second cancer). The study included 647 cases with second cancer (carcinoma, non-melanoma-skin cancers, hematological secondary cancer and melanoma). The most frequent category was carcinoma (n=426, 65.8%). Cases were comparable with the 1,234 matched controls for demographics, type of MPN, and exposure to potential confounders such as mutational profile, abnormal karyotype and cardiovascular risk factors. The thrombotic events of interest were ischemic stroke, transient ischemic attacks, acute coronary syndromes, peripheral arterial thrombosis, deep venous thrombosis (including thrombosis of cerebral and splanchnic veins) and pulmonary embolism. Thrombosis had to be concurrent with or in the 2 years before MPN diagnosis or occurring after MPN diagnosis. The cumulative incidence of either arterial or venous thrombosis from MPN diagnosis was estimated by the Kaplan-Meier method and was compared between cases and controls using the log-rank test. A conditional logistic regression model estimated the Odds Ratio (OR) with 95% Confidence Interval (CI) of second cancer associated with the occurrence of thrombosis before/at diagnosis of MPN and during follow-up. Other covariates were patient age, cardiovascular risk factors, the JAK2V617F mutation, and treatment during follow-up. Results Approximately 20% of either MPN cases or controls had thrombosis before MPN or at diagnosis (19.8% vs. 21.1%, respectively, p=0.462). After a median observation time from diagnosis of MPN to an index date of 4.5 years (interquartile range 1.5-8.2) in cases and 3.7 years (interquartile range 1.5-7.5) in controls, cases showed a percentage of thrombosis higher than in controls (75/647, 11.6% vs. 100/1234, 8.1%, respectively, p=0.013). Approximately one-third of thrombosis preceding cancer occurred in the 12 months before the diagnosis of second cancer (22/75, 29.3%). The excess of thrombosis in cases was due to a higher frequency of arterial thrombosis (6.2% vs. 3.7%, p=0.015), whereas no significant difference was found for venous thrombosis (5.4% vs. 4.3%, p=0.277). While the cumulative incidence of venous thrombosis over time was similar among cases and controls (p=0.864), the cumulative incidence of arterial thrombosis was higher in cases with second cancer (p=0.006) (Figure 1). The excess of arterial thrombosis after MPN diagnosis was limited to cases with carcinoma (6.8% vs 3.9%, p=0.027). In a multivariable model, arterial thrombosis during the follow-up was confirmed to be an independent predictor factor for carcinoma, with an odds ratio of 1.97 (95%CI 1.14-3.41, p=0.015). Conclusions. These findings reveal an association of arterial thrombosis with subsequent second cancer (namely carcinoma) in MPN patients. A possible biological plausibility for this link may be related to an underlying common pathogenic mechanism such as an aberrant inflammatory response consistently found in MPN. This observation may have practical implications and suggests careful clinical surveillance for early diagnosis of second cancer in MPN patients with arterial thrombosis during the follow-up. Disclosures Palandri: Novartis: Consultancy, Honoraria. Iurlo:Novartis: Other: Speaker Honoraria; Incyte: Other: Speaker Honoraria; Pfizer: Other: Speaker Honoraria. Bonifacio:Incyte: Honoraria; Novartis: Honoraria; Amgen: Honoraria; Pfizer: Honoraria; BMS: Honoraria. Rumi:novartis: Honoraria, Research Funding. Elli:Novartis: Membership on an entity's Board of Directors or advisory committees. Lunghi:Pfizer: Honoraria; Novartis: Honoraria; Incyte: Honoraria. Benevolo:Novartis Pharmaceuticals: Consultancy. McMullin:Italopharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Speakers Bureau; Daiko Sanyo: Membership on an entity's Board of Directors or advisory committees. Griesshammer:Novartis: Consultancy, Honoraria, Speakers Bureau. Vannucchi:CTI: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Membership on an entity's Board of Directors or advisory committees; Italfarmaco: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene Corporation: Membership on an entity's Board of Directors or advisory committees. Rambaldi:Pfizer: Consultancy, Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau.

Kurzfassung: Introduction: Autologous stem cell transplantation (ASCT) has proven effective in Multiple Myeloma (MM) patients (pts) aged 〈 65. However in several studies ASCT has proven safe and effective also in selected MM pts aged 〉 65. With the increase of treatment options, several geriatric assessment tools have been proposed to help physicians in selecting treatment of appropriate intensity. However specific criteria for evaluating ASCT eligibility in elderly MM pts have been seldom investigated.Patients and Methods: From January 2013 to December 2017 131 consecutive newly diagnosed symptomatic MM pts aged 65-75 (M/F: 66/65) were considered for ASCT at our center according to physician's clinical judgement. The variables included in Charlson Comorbidity Index (CCI), Hematopoietic cell transplantation comorbidity index (HCT-CI), and International Myeloma Working Group (IMWG) frailty score were not considered for selection but were retrieved for this study and pts were classified accordingly. Of note, ADL and IADL were referred to pts status prior the occurrence of MM related symptoms. The impact of age and the predictive role of the above mentioned scores on progression-free survival (PFS) of pts selected or not for ASCT was analyzed. Pts characteristics are shown in Table 1. Results: Of 131 pts, 85 (65%) were judged transplant eligible (ASCT ITT arm) by the clinician and received bortezomib-based induction (VTD 94%, VD 5%, VCD 1%); 72 of them (85%) actually underwent ASCT (70% single, 30% double) with melphalan conditioning 200 mg/sqm in 68% of cases. The 46 pts considered ineligible to ASCT received a less intensive first line treatment (89% VMP, 4% Daratumumab-VMP, 2% MP, 5% steroids/palliation). Complete remission (CR) after the first ASCT was higher in the ASCT (ITT) vs the NO ASCT group (43,5% vs 26%, respectively; p 0.048), whereas ORR and ³VGPR rates were comparable (83% vs 74% and 76,4% vs 61%, respectively). Transplant related mortality (TRM) was 0%. 2 year death rate was 13% in the ASCT (ITT) arm and was due to PD in 91% of cases (the latter 9% concerns a cardiac arrest secondary to aspiration pneumonia in a pt in VGPR 3 months after first ASCT). After a median follow-up of 27 months, PFS was 35,6 in the ASCT (ITT) group vs 19,9 months in NO ASCT pts, respectively; p 0,013, HR 0,42 (95% CI: 0,25-0,71). HCT-CI was ≥2 in 87% of pts overall. PFS was better in pts with HCT-CI 0-1 compared to HCT≥2 (NR vs 27,3 months; p 0,025, HR 0,45, 95% CI 0,23-0,91) and also in pts with HCT-CI ³2 undergoing ASCT (ITT) than in NO ASCT pts: median 34 vs 19,9 months, p 0,008 (HR 0,5, 95%CI 0,30-0,84). CCI was 0-3 in 78% of pts. Their outcome according was similar to pts with CCI 〉 3 but ASCT performed better than NO ASCT also in the few (median PFS 51,4 vs 16,9 months; p 0,04, HR 0,37, 95%CI 0,15-0,95). IMWG frailty score was more useful. No pts classified as FRAIL had been considered eligible to ASCT by clinical decision with a significantly worse outcome compared to FIT and UNFIT pts (median PFS: 7,9 months vs 32,9 and 29,6; FIT vs FRAIL: p 〈 0.0001, HR 0,02, 95%CI 0,004-0,008; UNFIT vs FRAIL: p 〈 0,001, HR 0,03 95%CI 0,007-0,12). Unlike HCT-CI and CCI, the distribution of patients according to IMWG score was more balanced both overall and between the ASCT (ITT) and NO ASCT groups, particularly for UNFIT pts. However on the whole no outcome differences were observed between FIT and UNFIT pts. PFS was better in FIT & UNFIT pts in the ASCT (ITT) group than in the NO ASCT group: median 35,6 months vs 25,8 months; p 0,04, HR 0,54, 95%CI 0,31-0,97. However, in the ASCT (ITT) group, the age group 65-69 years fared better than pts ³70 years (51,5 vs 27,7 months, p 0,0037; HR 0.34, 95% CI 0.17-0.7). Indeed, in IMWG UNFIT patients aged ³70, the PFS of the ASCT (ITT) group was comparable to NO ASCT group (18 vs 27 months, p 0,33) whereas in UNFIT pts aged 65-69, PFS was superior in pts treated more intensively by physician choice: 43,3 months in ASCT (ITT) arm vs 18,4 months in NO ASCT (p 0.01, HR 0.03; 95%CI 0.003-0.24). Conclusion: In an unselected series of elderly MM pts aged 65-75 and undergoing ASCT according to clinical judgement the outcome of ASCT pts was better than those of NO ASCT. CCI and HCT-CI score proved of little help for better identifying the best candidates to ASCT. Conversely the IMWG frailty score would be of help in identifying the category of UNFIT pts aged 70-75 whose outcome with ASCT selected by clinical judgement was no better than with less intensive treatments. Table 1: Disclosures Belotti: Amgen: Other: Advisory Board; Celgene: Other: Advisory Board. Cattaneo:GILEAD: Other: Advisory Board. Rossi:TEVA: Other: ADVISORY BOARD; CELGENE: Other: ADVISORY BOARD; SANOFI: Other: ADVISORY BOARD; MUNDIPHARMA: Honoraria; ABBVIE: Other: ADVISORY BOARD; JANNSEN: Other; PFIZER: Other: ADVISORY BOARD; ROCHE: Other: Advisory Board; NOVARTIS: Honoraria; SANDOZ: Honoraria; GILEAD: Other: ADVISORY BOARD; BMS: Honoraria; JAZZ: Other: ADVISORY BOARD; AMGEN: Other: ADVISORY BOARD.