ALBERT

Description: Introduction: Acute Myeloblastic Leukemia (AML) is the most frequent acute leukemia in the adults and its incidence increases with age. There are few studies about the demography and outcomes of AML patients in Chile and the only report belongs to a public hospital from 2000. We discuss the results of patients treated in our institution with AML non promyelocytic. Patients and Methods: Retrospective analysis of the epidemiologic, clinical and laboratory characteristics of diagnosis (cytology and flow cytometry) and treatment of AML non promyelocytic patients between 2010-2014. Statistical analysis of the data was performed using SPSS Statistics v21 software. Results: 63 patients were diagnosed with AML non M3, 52 males (66%), with a median age of 55.4 years (range: 16 - 89). Diagnosis laboratory tests (mean values and ranges) were: WBC 45.989/mm3 (range: 700 - 405.000); hemoglobin 9,1 g/dl (range: 5,2 - 14,1); platelets 75.548/mm3 (range: 10.000 - 454.000); peripheral blood blasts 38% (range 0 - 100); bone marrow blasts 74% (range 25 - 100%). The cytogenetic risk groups were: favorable (n=5, 8%), intermediate (n=33, 52%), adverse (n=8, 13%) and unknown (n=17, 27%). Of all the patients, 75% (n=47) received induction chemotherapy (CT) and 25% (n=16) palliative care. The mean age of the group with cytogenetic analysis was 51.2 years and only 8.6% did not receive consolidation CT. On the other hand, the group of patients with unknown cytogenetics had a mean age of 68 years and 57% did not receive consolidation CT. The mean survival of the CT group was 27.3 month (range: 0 - 53). By contrast, the mean survival in the palliative care group was 1 month (range: 0 - 6). The mean follow up in all patients was 13 months, (range: 1 - 55) and 17 months (range: 1 - 54) in the group that received CT. 87% (n=41) of patients with CT had febrile neutropenia with respiratory and intestinal focus most commonly identified. The induction mortality was 4,2% (n=2). Complete cytologic remission was achieved in 70% (n=33). The 3-year relapse free survival (RFS) and overall survival (OS) in the CT group were 25% and 31%, respectively. The multivariate survival analysis using Cox’s regression demonstrated that the variables that had significant impact in RFS and OS were: age at diagnosis (

Description: Background: Deletions of the 9p21 chromosomal region which encompasses CDKN2A, a gene encoding both p16INK4a and p14ARF, are frequent in childhood acute lymphoblastic leukemia (ALL). Their prognostic relevance is controversial. Indeed, small retrospective series of patients were analysed so far, which usually mixed B- and T-lineage ALL. Moreover, the prognosis could vary according to the extent of the deletion since the inactivation of contiguous genes such as CDKN2B encoding p15INK4b or MTAP encoding methylthioadenosine phosphorylase, may influence chemosensitivity. Methods: 9p21 deletions were retrospectivelly studied in 227 children (aged 2 mo to 16 yrs) with B-lineage ALL, using a real-time PCR assay for CDKN2B, CDKN2A (e1b and e3), and MTAP gene dosage, and loss of heterozygosity analysis (Bertin et al., 2003). CDKN2A and CDKN2B inactivation by promoter methylation was also investigated. TEL-AML1 fusion was screened in 182 (80%) of these patients. All patients were enrolled in EORTC trials 58 881 or 58 951. Median follow-up was 6 years; total number of events was 55 and the overall 6-year EFS rate was 74%. Results: CDKN2A bi- and mono-allelic inactivation was found in 38 (17%), and 31 (14%) B-lineage ALL respectively. Patients with a bi-allelic inactivation did not differ from undeleted ones regarding age, sex, immunophenotype and response to prephase treatment, but tended to have a higher WBC count at diagnosis and were more often allocated to the NCI high risk group (42% vs 27%). Genetic defects, including TEL-AML1, were independent of 9p21 alteration with exception of hyperdiploidy (〉50 chromosomes) which was less frequent in patients with CDKN2A inactivation (5% vs 34%). The 6-year EFS rates of patients with bi-allelic, mono allelic and no inactivation did not differ significantly (68%, 80%, and 75% respectively). Bi-allelic CDKN2B inactivation by either deletion or promoter methylation was found in 36 (16%) patients and the 6-year EFS rate of these patients was similar to that of undeleted patients (71% vs 74%). MTAP bi-allelic inactivation was found in 24 (11%) of patients. Although MTAP deficient cells are in vitro very sensitive to methotrexate, the 6-year EFS rate (70% vs 75%) did not differ from that of MTAP positive patients even when the analysis was restricted to CDKN2A inactivated ALL. Conclusion: Although bi-alleic CDKN2A inactivation was more often associated with bad prognostic parameters in B-lineage ALL, it failed to significantly influence the outcome of the patients. We did not observe either any influence of the co-inactivation of CDKN2B and MTAP on the outcome. Considering that our study is the largest so far using molecular analysis of the 9p21 region, we assume that any bad prognosis associated with 9p21 alteration, if present, should be weak in B-lineage ALL, and would probably require larger cohorts of patients to be ascertained.

Description: INTRODUCTION: The increased survival of hematological patients has required to widen the care of these patients, with emphasis in factors related to quality of life and late mortality. Among them, osteoporosis (OP) is a fundamental problem. Patients undergoing hematopoietic cell transplantation (HCT) are at great risk of OP, mainly due to prolonged exposure to chemotherapy, immunosuppresants and the hypogonadism frequently associated to these treatments. Despite this, there is lack of strong evidence on this matter and HCT guidelines are not clear on this problem. OBJECTIVES: To evaluate parameters related to bone health in patients undergoing HCT. To measure the frequency and severity of the alterations in this group of patients. PATIENTS AND METHODS: Observational and retrospective analysis of patients undergoing HCT at the Catholic University Hematology-Oncology Department. All patients undergoing HCT were recommended to have determination of 25-OH vitamin D, PTHi, Calcium, Phosphorus and bone densitometry (DXA). We included all patients with the evaluation before HCT (preHCT) and 1 year after HCT (postHCT). RESULTS: We obtained data from 20 patients preHCT and 22 patients postHCT. Main diagnosis were acute myeloid leukemia (n=13; 32%), multiple myeloma (n=11; 26%), acute lymphoblastic leukemia (n=9; 21%) and Hodgkin's Lymphoma (n=5; 12%). Mean age was 40 years (range: 17-67) and 67% were males. In the preHCT group, the median 25-OH vitamin D levels were 13,6 ng/ml (range: 4,9-26,7 ng/ml) and 100% of the patients were in the insuficiency range leveles (

Description: Introduction: Patients undergoing hematopoietic cell transplantation (HCT) have an increased risk of Clostridium difficile infection (CD) and multiple risk factors have been found. Literature reports CD infection in nearly 20% of transplanted patients. No information about this infection in HCT patients has been reported in Chile. Patients and Methods: We performed a retrospective analysis of 250 patients undergoing HCT at the Catholic University Hospital in Santiago, Chile, between 2000 and the first semester of 2013, including allogeneic (allo) HCT and autologous (auto) HCT patients. Statistical analysis of the data was conducted using SPSS Statistics v21. Results: Of the 250 transplanted patients studied, 59% (n=147) were allo-HCT and 41% (n=103) were auto-HCT. The mean age was 39 years old (range: 15-69), with a male predominance (151 patients; 60%). The main indication for HCT was acute leukemia (n=104; 42%) followed by multiple myeloma (n=36; 14%) and lymphoma (n=49; 20%). 93% of patients received myeloablative (MA) conditioning regimen, and all of them received proton pump inhibitors and prophylactic antibiotics during the previous months of the HCT. Of the 250 patients studied, 192 (77%) had at least one episode of diarrhea that required study, among them 13% (n=25) were documented as positive for CD (toxin assay or PCR test), the mean age of this group was 36 years old (range: 18-62), also with a male predominance (15 patients; 60%). All of the infected patients had mild to moderate diseases and there were no deaths attributed to it. 80% (n=20) of the infected patients underwent allo-HCT and 20% (n=5) auto-HCT. In the allo-HCT group, 53% had acute lymphoblastic leukemia (ALL), 6% acute myeloid leukemia (AML), 24% chronic myeloid leukemia (CML) and 12% other causes. In the auto-HCT group, 40% were transplanted due to multiple myeloma, 20% amyloidosis, 20% germinal cancer and 20% acute myeloid leukemia. No patient required total central parenteral nutrition previous to the infection. During the 3 months before HCT, 84% (n=21) of the infected patients used antibiotics including cephalosporins, carbapenem, aminoglicosides and vamcomycin. The overall incidence of CD infection in the first week, month and year after transplant, was 4.0%, 6.4% and 10%, respectively, with a median time frame from transplantation to infection diagnosis of 20 days. In auto-HCT, 7 days, 30 days and 1 year CD incidence was 1.9, 2.9 and 4.9%, respectively. In allo-HCT, 7 days, 30 days and 1 year CD incidence was 5.4, 8.8 and 13.6%, respectively. There was no significant statistical difference in overall survival (OS) between the infected and non-infected patients one year after the transplant (OS 67.6% for CD negative vs. 72.0% for CD positive, p=0.61). Conclusions: In our institution CD infection in patients undergoing HCT had a similar incidence to other reports. Most of cases occur before the first week after HCT (40% of the cases), and the incidence of the infection remained stable during the first year after the procedure. We identified the type of transplant (allo-HCT 3 times higher risk than auto-HCT) and disease (ALL 3 times higher risk than AML) as risk factors for CD infection. Disclosures No relevant conflicts of interest to declare.

Description: Recent studies have demonstrated that innate immune cells, i.e. neutrophils and monocytes, provide the initiating stimulus for venous thrombus development. Gas6 was found to promote inflammation by inducing interactions between the endothelium and innate immune cells. In addition, we recently showed that Gas6 was involved in venous thrombosis by inducing tissue factor expression in the endothelium. Since Gas6 is expressed in monocytes, we hypothesize that Gas6 may be involved in monocyte recruitment during venous thrombosis. Venous thrombosis was induced in the inferior vena cava of wild type (WT) and Gas6 deficient (-/-) mice using 5% FeCl3. Using ultrasonography, we found that global monocyte depletion by clodronate resulted in the formation of smaller thrombi. Selective depletion of the pro-inflammatory (Ly6Chi) monocyte subset, using an anti-CCR2 antibody, also induced the formation of smaller clots. In addition, MOMA-2 (monocyte-macrophage marker) staining showed a reduced number of monocytes in thrombi from Gas6-/- mice. More importantly, immunofluorescent staining revealed that fewer Ly6Chi monocytes were recruited to the thrombi of Gas6-/- mice compared to WT. However, Ly6Clow (patrolling) monocytes were equivalently recruited between Gas6-/- and WT mice. In vitro, mRNA expression of CCR2 was increased by thrombin in WT but not in Gas6-/- monocytes. The mRNA and protein expression of the CCR2 ligand, CCL2, was also increased by thrombin in WT but not in Gas6-/- endothelial cells. CCL2 secretion, as demonstrate by ELISA, was induced by thrombin treatment in WT but not in Gas6-/- endothelial cells. Conditioned media from WT or Gas6-/- endothelial treated by thrombin was used for monocyte migration experiments. The conditioned media from WT endothelial cells treated with thrombin increased migration of WT monocytes compared to media from untreated or Gas6-/- endothelial cells. Our results demonstrate an important role for Ly6Chi monocytes in thrombus formation and that Gas6 is specifically involved in the recruitment of these monocytes through the expression of CCL2 and CCR2. Disclosures No relevant conflicts of interest to declare.

Description: Introduction Autologous hematopoietic transplantation (autoHT) is an effective treatment in myeloma and lymphoma patients. To perform it, an adequate harvest of stem cells is required. Collection of stem cells can be challenging in some patients, because they do not achieve satisfactory CD34 cell counts with GCSF + - chemotherapy. Plerixafor is a potent drug that promotes the release of stem cells from medullar niche to peripheral blood and allows satisfactory harvests but at a high cost. Different criteria of use have been applied in several countries to select patients eligible for treatment with plerixafor, with a high variability of prescription modalities. Even more, there are difficulties in precise the optimal schedule and dosification mainly by differences in the number defined as optimal collection, because some centers use the minimum tolerated collection cellularity of 2.0 x106 CD34/ kg and others use more stringent cells recounts. Development of our own algorithm for stem cell harvest is mandatory. Methodology We performed a retrospective analysis of the harvests performed in patients treated with autoHT in our institution since 1993. In 2013 plerixafor was available in our country and was included in our transplantation program. Satisfactory harvest was defined with total cellularity obtained of 2.5 x106 CD34/ kg. Results During the last 26 years, 273 patients with myeloma and lymphoma were treated with autoHT. Before 2013, all patients (n=112) were mobilized with GCSF +- chemotherapy. In this period, 75% of patients had successful harvests, requiring more than 2 apheresis procedures in 30% of them. Median cellularity obtained was 2.7 x106 CD34/ kg (range 1.2-15). Since 2013, 161 patients were treated with autoHT and we found that 80% of patients obtained satisfactory harvests with 1 apheresis and satisfactory harvests were obtained in 97% using plerixafor in 50% of them (p 〈 0.001). Median cellularity obtained was 4.8 x106 CD34/ kg (range 2-14) (p 〈 0.001). In multivariate analysis, after 2013 the only factor that predicted the need for plerixafor was the presence of less than 30,000 CD34 / ul at the day of the apheresis (OR 0,3 p

Description: Introduction Steroids refractory (SR) acute (a) or chronic (c) graft versus host disease (GVHD) carries morbidity and lethality. Treatment algorithms varies widely among institutions meaning that the most effective treatment is not yet established. Extracorporeal photopheresis (ExP) and Ruxolitinib have shown in retrospective studies a hopeful efficacy in the treatment of this disease. Moreover, prolonged corticoid or immunosuppressant drugs are associated with opportunistic infection and mortality. Consequently our GVHD algorithm consider 7 days of high dose steroids before moving on to the second line treatments mentioned above to avoid prolonged infectious risk. Because of quick availability and reported benefit, without adding more immunosuppression, we have prefered ExP over other treatments. However this apparent advantages are contrasted with its great cost, as each session exceeds $US 2000. Ruxolitinib in the other hand, is less expensive, but since it is only administered orally, doubts regarding its absorption frequently appear in situations of intestinal GVHD. Hence we used Ruxolitinib as third line therapy after ExP. To know real outcomes of our patients we analyzed all patients with SR-GVHD of our adult hematopoietic transplantation program. Methodology Since 1993 we prospectively performed a registry of all transplanted patients. For this analysis we reviewed all allogeneic transplantated patients and data of those who developed SR-GVHD was extracted. In 2013, ExP was available in our center and was included in our algorithm. SR-GVHD was defined as patients that did not respond despite 5-7 of high dose steroids. As some patients suffer from aGVHD and cGVHD we analyse them separately for global response purpose. Considering that some events can affect more than one organ, when less than 3 organs were affected, we also analyse responses to each one of them separately. When 3 or more organs were affected we consider it as multisystemic affectation Results Until 2013 150 allogeneic transplants were performed; of those, 14 patients (9%) developed SR-GVHD, with 90% mortality. Unsuccessful treatmens included ATG, high doses of methylprednisolone, etanercept, and cyclosporine adjustment. Since 2013 we performed 135 allogeneic transplants and 20 patients (14,8%) developed SR-GVHD. Twelve patients presented with SR-aGVHD, 5 patients with SR-cGVHD and 4 patients with acute and chronic SR-GVHD in different time. For analysis of global response, then, we consider 16 SR-aGVHD patients and 9 SR-cGVHD patients (patients characteristics are shown in table 1). All 16 SR-aGVHD patients were treated with ExP with a median of 5,3 sessions. Among SR-aGVHD patients 37% responded to ExP, all being complete responses. When including ruxolitinib addition, final responses reached 56,3%. Mortality was high in this group (56,3% died, 66% of them because of aGVHD). In patients with no response to ExP and Ruxolitinib, mortality rate was 85,7%. Most affected organs were small intestine and skin, affecting 43,7% of patients each. Responses to therapy by organ affectation are shown in table 2. In SR-cGVHD, ExP was used in 7/9 patients, and 85,7% responded. In this case most patients only achieved partial response (71,4%) and ruxolitinib did not improved response any further. Among SR-cGVHD, skin affectation was the most common, 66% of SR-cGVHD patients presented with skin affectation either exclusively or combine with other organ affectation. Conclusions SR-GVHD remains a challenging disease, and no standard treatment exist. Beyond the high mortality rate, cost associated with treatment, procedures and hospitalization makes this patients even more difficult to manage. Therefore it is imperative that every center analyse their own data and treatment pathways to optimize both, GVHD treatment results and hospital resources. As a matter of fact, after reviewing our own data, we switched to a combined modality of ExP and Ruxolitinib for affectation other than skin, in SR- aGVHD. We hope this new algorithm overcomes, specially, the bad results of monotherapy in intestinal SR-aGVHD seen in our center, as these patients are exposed to high risk of malnutrition, dehydration, prolonged hospitalization and death. In SR-cGVHD we found a consistent tendency of partial responses with ExP. Keywords: Steroids refractory graft versus host disease, Extracorporeal Photopheresis, Ruxolitinib. Disclosures No relevant conflicts of interest to declare.

Description: Introduction. The use of cryopreservation of hematopoietic precursors for autologous or allogeneic transplantation with DMSO is a procedure commonly used in many countries, but it is expensive and is not exempt of risks, since serious adverse events have been described. Our group recently published a comparative study between two centers and showed that avoiding DMSO cryopreservation favors better transplantation tolerance, significantly shorter hospitalizations, fewer episodes of febrile neutropenia and mucositis. In this report we show the financial analysis of both modalities of CRYO and Non CRYO preserved transplants. Methodology Database of the adult hematopoietic transplant program of our institution was retrospectively analyzed. Since Non CRYO modality was initiated in our institution on 2015, we compared 3 years before and after that date, assigning two groups (CRYO and Non CRYO). Costs data of mobilization, apheresis, hospitalization, freezing and blood banking were analyzed. Results Between 2013 and 2018, 90 autologous hematopoietic transplants were performed, 41 CRYO and 49 Non CRYO. The average cost of mobilization therapy with filgrastim and the use of plerixafor did not differ between groups (p = 0.26). The number of apheresis needed to achieve a satisfactory count was higher in CRYO (median 1.5 vs 1.2 p = 0.001) with 55% higher cost on average. The cost of transplant hospitalization including antibiotics costs, blood banking costs, analgesics requirements and nutritional parenteral support was lower in Non CRYO (minimum cost USD 8000, maximum USD 160000 average value USD 20.000) than in CRYO (minimum cost USD 11.000, maximum USD 200.000, average value USD 32000) (p = 0.001). Conclusions In addition to promoting better patient tolerance, Non CRYO modality in our country has lower cost. This information has important relevance for health systems of developing countries and can promote better access to transplant. Disclosures No relevant conflicts of interest to declare.

Description: A goal in precision medicine is to use patient-derived material to predict disease course and intervention outcomes. Here, we use mechanistic observations in a preclinical animal model to design an ex vivo platform that recreates genetic susceptibility to T-cell–mediated damage. Intestinal graft-versus-host disease (GVHD) is a life-threatening complication of allogeneic hematopoietic cell transplantation. We found that intestinal GVHD in mice deficient in Atg16L1, an autophagy gene that is polymorphic in humans, is reversed by inhibiting necroptosis. We further show that cocultured allogeneic T cells kill Atg16L1-mutant intestinal organoids from mice, which was associated with an aberrant epithelial interferon signature. Using this information, we demonstrate that pharmacologically inhibiting necroptosis or interferon signaling protects human organoids derived from individuals harboring a common ATG16L1 variant from allogeneic T-cell attack. Our study provides a roadmap for applying findings in animal models to individualized therapy that targets affected tissues.

Description: Venous thromboembolism (VTE) is a leading cause of morbidity and mortality. Gas6-/- mice are protected against VTE suggesting an important role for Gas6 in the pathophysiology of this disorder. VCAM-1 is a key adhesion molecule, expressed on the endothelium, implicated in the pathophysiology of venous thrombosis. We previously demonstrated that Gas6 induced the phosphorylation of Forkhead box O1 (FoxO-1) through the activation of the PI3K/Akt signaling pathway. Thus, we hypothesize that Gas6 promotes thrombin-induced VCAM-1 expression through the regulation of FoxO-1 in endothelial cells. Thrombin induces VCAM-1 mRNA and protein expression in WT but not in Gas6-/- endothelial cells. Silencing FoxO-1, using siRNA, enhances VCAM-1 expression both in WT and in Gas6-/- endothelial cells. Thrombin induces FoxO-1 phosphorylation and nuclear exclusion in WT but not in Gas6-/- endothelial cells. Inhibition of FoxO-1 phosphorylation by overexpression of a phosphorylation resistant FoxO-1 mutant (Tm-FoxO-1) decreases VCAM-1 expression, both in WT and Gas6-/- endothelial cells. Inhibition of Akt by LY294002 prevents FoxO-1 phosphorylation and VCAM-1 expression. Finally, thrombin-induced VCAM-1 expression in WT endothelial cells was associated with an increase in bone marrow mononuclear cell (BM-MC) adhesion. BM-MC adhesion was further increased when endothelial cells were transfected with a FoxO-1 siRNA and decreased when transfected with the Tm-FoxO-1 plasmid. Thus, we demonstrate that Gas6 regulates VCAM-1 expression through phosphorylation and nuclear exclusion of FoxO-1, thereby promoting BM-MC adhesion. These data suggest that the Gas6/FoxO-1 signaling axis plays an important role in VCAM-1 expression during thrombosis. Disclosures No relevant conflicts of interest to declare.