Jacopo Olivieri

Long-term outcome and prospective validation of NIH response criteria in 39 patients receiving imatinib for steroid-refractory chronic GVHD

Forty adults aged 28 to 73 years were entered into a prospective trial of imatinib for the treatment of steroid-refractory chronic graft-versus-host disease (SR-cGVHD). After 6 months, intention-to-treat (ITT) analysis of 39 patients who received the drug, regardless of the duration of treatment, revealed 14 partial responses (PR), 4 minor responses (MR) with relevant steroid sparing (46%) according to Couriel criteria, and 20 ≥ PR (51.3%), as per the National Institutes of Health (NIH) criteria and NIH severity score changes. The best responses were seen in the lungs, gut, and skin (35%, 50%, and 32%, respectively). After a median follow-up of 40 months, 28 patients were alive, with a 3-year overall survival (OS) and event-free survival of 72% and 46%, respectively. The 3-year OS was 94% for patients responding at 6 months and 58% for nonresponders according to NIH response, suggesting that these criteria represent a reliable tool for predicting OS after second-line treatment. Monitoring of anti-platelet-derived growth factor receptor (PDGF-R) antibodies showed a significant decrease in PDGF-R stimulatory activity in 7 responders, whereas it remained high in 4 nonresponders. This study confirms the efficacy of imatinib against SR-cGVHD and suggests that the response at 6 months significantly predicts long-term survival.

Fibrillin assemblies: extracellular determinants of tissue formation and fibrosis

The extracellular matrix (ECM) plays a key role in tissue formation, homeostasis and repair, mutations in ECM components have catastrophic consequences for organ function and therefore, for the fitness and survival of the organism. Collagen, fibrillin and elastin polymers represent the architectural scaffolds that impart specific mechanic properties to tissues and organs. Fibrillin assemblies (microfibrils) have the additional function of distributing, concentrating and modulating local transforming growth factor (TGF)-β and bone morphogenetic protein (BMP) signals that regulate a plethora of cellular activities, including ECM formation and remodeling. Fibrillins also contain binding sites for integrin receptors, which induce adaptive responses to changes in the extracellular microenvironment by reorganizing the cytoskeleton, controlling gene expression, and releasing and activating matrix-bound latent TGF-β complexes. Genetic evidence has indicated that fibrillin-1 and fibrillin-2 contribute differently to the organization and structural properties of non-collagenous architectural scaffolds, which in turn translate into discrete regulatory outcomes of locally released TGF-β and BMP signals. Additionally, the study of congenital dysfunctions of fibrillin-1 has yielded insights into the pathogenesis of acquired connective tissue disorders of the connective tissue, such as scleroderma. On the one hand, mutations that affect the structure or expression of fibrillin-1 perturb microfibril biogenesis, stimulate improper latent TGF-β activation, and give rise to the pleiotropic manifestations in Marfan syndrome (MFS). On the other hand, mutations located around the integrin-binding site of fibrillin-1 perturb cell matrix interactions, architectural matrix assembly and extracellular distribution of latent TGF-β complexes, and lead to the highly restricted fibrotic phenotype of Stiff Skin syndrome. Understanding the molecular similarities and differences between congenital and acquired forms of skin fibrosis may therefore provide new therapeutic tools to mitigate or even prevent disease progression in scleroderma and perhaps other fibrotic conditions.

Tailored therapy in an unselected population of 91 elderly patients with DLBCL prospectively evaluated using a simplified CGA.

BACKGROUND Elderly patients with diffuse large B-cell lymphoma (DLBCL) are a heterogeneous population; clinical trials have evaluated a minority of these patients. PATIENTS AND METHODS Ninety-one elderly patients with DLBCL received tailored treatment based on a comprehensive geriatric assessment (CGA). Three groups were identified: I, fit patients; II, patients with comorbidities; III, frail patients. Group I received 21-day cycles of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP-21), group II received R-CHOP-21 with liposomal doxorubicin, and group III received 21-day cycles of reduced-dose CHOP. Fifty-four patients (59%) were allocated to group I, 22 (25%) were allocated to group II, and 15 (16%) were allocated to group III. RESULTS The complete response (CR) rates were 81.5% in group I, 64% in group II, and 60% in group III. With a median follow-up of 57 months, 42 patients are alive, with 41 in continuous CR: 31 patients (57%) in group I, seven patients (32%) in group II, and four patients (20%) in group III. The 5-year overall survival, event-free survival, and disease-free survival rates in all patients were 46%, 31%, and 41%, respectively. Multivariate analysis selected group I assignment as the main significant prognostic factor for outcome. CONCLUSIONS This approach in an unselected population of elderly DLBCL patients shows that treatment tailored according to a CGA allows the evaluation of elderly patients who are currently excluded from clinical trials.

Ha-Ras stabilization mediates pro-fibrotic signals in dermal fibroblasts

BackgroundScleroderma (systemic sclerosis; SSc) is a clinically heterogeneous and often lethal acquired disorder of the connective tissue that is characterized by vascular, immune/inflammatory and fibrotic manifestations. Tissue fibrosis is the main cause of morbidity and mortality in SSc and an unmet medical challenge, mostly because of our limited understanding of the molecular factors and signalling events that trigger and sustain disease progression. Recent evidence has correlated skin fibrosis in SSc with stabilization of proto-oncogene Ha-Ras secondary to auto-antibody stimulation of reactive oxygen species production. The goal of the present study was to explore the molecular connection between Ha-Ras stabilization and collagen I production, the main read-out of fibrogenesis, in a primary dermal fibroblast culture system that replicates the early stages of disease progression in SSc.ResultsForced expression of proto-oncogene Ha-Ras in dermal fibroblasts demonstrated the promotion of an immediate collagen I up-regulation, as evidenced by enhanced activity of a collagen I-driven luciferase reporter plasmid and increased accumulation of endogenous collagen I proteins. Moreover, normal levels of Tgfβ transcripts and active transforming growth factor-beta (TGFβ) implied Ha-Ras stimulation of the canonical Smad2/3 signalling pathway independently of TGFβ production or activation. Heightened Smad2/3 signalling was furthermore correlated with greater Smad3 phosphorylation and Smad3 protein accumulation, suggesting that Ha-Ras may target both Smad2/3 activation and turnover. Additional in vitro evidence excluded a contribution of ERK1/2 signalling to improper Smad3 activity and collagen I production in cells that constitutively express Ha-Ras.ConclusionsOur study shows for the first time that constitutively elevated Ha-Ras protein levels can directly stimulate Smad2/3 signalling and collagen I accumulation independently of TGFβ neo-synthesis and activation. This finding therefore implicates the Ha-Ras pathway with the early onset of fibrosis in SSc and implicitly identifies new therapeutic targets in SSc.

Tirosin kinase inhibitors in chronic graft versus host disease: from bench to bedside.

Chronic Graft Versus Host Disease (cGVHD) is a major complication of allogeneic stem-cell transplantation (SCT). In many inflammatory fibrotic diseases, such as Systemic Scleroderma (SSc) and cGVHD with fibrotic features, an abnormal activation of transforming growth factor (TGFβ) and platelet-derived growth factor receptor (PDGF-R) pathways have been observed. Tyrosin Kinase Inhibitors (TKIs), which are currently used for treatment of patients with Chronic Myeloid Leukemia (CML), share potent antifibrotic and antiinflammatory properties, being powerful dual inhibitors of both PDGF-R and TGFβ pathways. Moreover accumulating in vitro data confirm that TKIs, interacting with the TCR and other signalling molecules, carry potent immunomodulatory effects, being involved in both T-cell and B-cell response. Translation to the clinical setting revealed that treatment with Imatinib can achieve encouraging responses in patients with autoimmune diseases and steroid-refractory cGVHD, showing a favourable toxicity profile. While the exact mechanisms leading to such efficacy are still under investigation, use of TKIs in the context of clinical trials should be promoted, aiming to evaluate the biological changes induced in vivo by TKIs and to assess the long term outcome of these patients. Second-generation TKIs, with more favourable toxicity profile are under evaluation in the same setting.

Consensus recommendations for improvement of unmet clinical needs—the example of chronic graft-versus-host disease: a systematic review and meta-analysis

BACKGROUND Consensus recommendations are used to improve the methodology of research about rare disorders, but their uptake is unknown. We studied the uptake of consensus recommendations in steroid-refractory chronic graft-versus-host disease (SR-cGVHD). Although in 2006 the National Institutes of Health (NIH) cGVHD consensus project produced recommendations for clinical trials, guidelines have emphasised the scarcity of valuable evidence for all tested interventions. METHODS We searched Medline (PubMed) between Jan 1, 1998, and Oct 1, 2013, for non-randomised studies of systemic treatment for SR-cGVHD. To measure adherence to NIH recommendations, we applied a 61 item checklist derived from the NIH consensus document. We did a meta-analysis to measure pooled effect size for overall response rate (ORR) and meta-regression analyses to measure the effect of deviations from NIH recommendations on pooled effect size. FINDINGS We included 82 studies related to nine interventions. Conformity to NIH recommendations was evenly low across the analysed timeframe (1998-2013), and did not change significantly after publication of NIH recommendations. The pooled effect size for ORR for systemic treatment of SR-cGVHD was 0.66 (95% CI 0.62-0.70). Increased adherence to NIH recommendations in a score of items defining correct response assessment was associated with a significant reduction in ORR (-4.2%, 95% CI -6.6 to -1.9; p=0.001). We recorded no significant association between ORR and sets of items related to correct diagnostic definition of SR-cGVHD (change in ORR -3.1%, 95% CI -7.7 to 1.5), specification of primary intervention (0, -3.8 to 3.6), or concomitant treatments (-1.6%, -5.4 to 2.3). The score of items defining correct response assessment increased after publication of NIH recommendations. INTERPRETATION Our findings show evidence of bias in the reported efficacy of treatment of SR-cGVHD. The overall effect of NIH recommendations in scientific literature is scarce; however, NIH recommendations improved assessment of response, possibly reducing the overestimation bias. Better implementation of NIH recommendations might reduce false expectations about new interventions, and thus prevent clinical studies with ineffective treatments. FUNDING None.

Modern Management of Anthracycline‐Induced Cardiotoxicity in Lymphoma Patients: Low Occurrence of Cardiotoxicity with Comprehensive Assessment and Tailored Substitution by Nonpegylated Liposomal Doxorubicin

BACKGROUND Anthracyclines (AC) are still undeniable drugs in lymphoma treatment, despite occasionally causing cardiotoxicity. Liposomal AC may reduce cardiotoxicity while retaining clinical efficacy; also, biomarker monitoring during chemotherapy allows early detection of cardiac damage, enabling strategies to prevent left ventricular ejection fraction (LVEF) deterioration. MATERIALS AND METHODS We conducted a prospective observational trial in a real-life population of lymphoma patients, combining advanced echocardiography and biomarkers (Troponin I [TnI]) for early detection of cardiotoxicity; we applied a prespecified policy to minimize cardiotoxicity, selecting patients with higher baseline risk to replace doxorubicin with nonpegylated liposomal doxorubicin (NPLD) and starting cardioprotective treatment when subclinical cardiotoxicity was detected. RESULTS Ninety-nine patients received ≥1 cycle of chemotherapy (39 with NPLD): 38 (NPLD = 34) were older than 65 years. At baseline, the NPLD subgroup had more cardiovascular risk factors and comorbidities than the doxorubicin subgroup. After treatment, echocardiographic parameters did not worsen in the NPLD subgroup; significant LVEF reduction occurred in two patients treated with doxorubicin. Over treatment course, TnI rises increased linearly in the doxorubicin subgroup but modestly in the NPLD subgroup. At doxorubicin doses >200 mg/m2 the difference was statistically significant, with more TnI rises in the doxorubicin subgroup. NPLD-treated patients did not experience higher rates of grade 3-4 adverse events. Within the diffuse large B-cell lymphomas category, we observed similar rates of complete and overall responses between doxorubicin- and NPLD-treated patients. CONCLUSION A comprehensive strategy to prevent, detect, and treat cardiotoxicity allows an optimal management of the lymphoma with low incidence of cardiac complications. The Oncologist 2017;22:422-431 IMPLICATIONS FOR PRACTICE: Despite the recent advances of targeted therapy in cancer, old cytotoxic drugs such as anthracyclines (AC) still play a fundamental role in the treatment of many lymphoma patients. We tested and validated in a real-life setting a personalized approach to prevent, detect, and treat AC-induced cardiotoxicity; biomarker monitoring was accomplished by Troponin I measurements before and after chemotherapy infusions, allowing detection of early subclinical cardiotoxicity, which was preemptively treated with cardio-protectants (beta blockers and angiotensin-converting-enzyme inhibitors). A telemedicine system allowed interdisciplinary management of the patients with an expert cardiologist. Furthermore, tailored use of liposomal AC following a prespecified policy appeared to prevent the excess cardiotoxicity expected in high-risk patients.

Immunomodulatory Effects of Tyrosine Kinase Inhibitors (TKIs) in Vitro and in Vivo Study.

Pathogenesis of chronic graft-versus-host disease (cGVHD) is incompletely defined, involving donor-derived CD4 and CD8-positive T lymphocytes as well as B cells. Standard treatment is lacking for steroid-dependent/refractory cases; therefore, the potential usefulness of tyrosine kinase inhibitors (TKIs) has been suggested, based on their potent antifibrotic effect. However, TKIs seem to have pleiotropic activity. We sought to evaluate the in vitro and in vivo impact of different TKIs on lymphocyte phenotype and function. Peripheral blood mononuclear cells (PBMCs) from healthy donors were cultured in the presence of increasing concentrations of nilotinib, imatinib, dasatinib, and ponatinib; in parallel, 44 PBMC samples from 15 patients with steroid-dependent/refractory cGVHD treated with nilotinib in the setting of a phase I/II trial were analyzed at baseline, after 90, and after 180 days of therapy. Flow cytometry was performed after labeling lymphocytes with a panel of monoclonal antibodies (CD3, CD4, CD16, CD56, CD25, CD19, CD45RA, FoxP3, CD127, and 7-amino actinomycin D). Cytokine production was assessed in supernatants of purified CD3+ T cells and in plasma samples from nilotinib-treated patients. Main T lymphocyte subpopulations were not significantly affected by therapeutic concentrations of TKIs in vitro, whereas proinflammatory cytokine (in particular, IL-2, IFN-γ, tumor necrosis factor-α, and IL-10) and IL-17 production showed a sharp decline. Frequency of T regulatory, B, and natural killer (NK) cells decreased progressively in presence of therapeutic concentrations of all TKIs tested in vitro, except for nilotinib, which showed little effect on these subsets. Of note, naive T regulatory cell (Treg) subset accumulated after exposure to TKIs. Results obtained in vivo on nilotinib-treated patients were largely comparable, both on lymphocyte subset kinetics and on cytokine production by CD3-positive cells. This study underlines the anti-inflammatory and immunomodulatory effects of TKIs and supports their potential usefulness as treatment for patients with steroid-dependent/refractory cGVHD. In addition, both in vitro and in vivo data point out that compared with other TKIs, nilotinib could better preserve the integrity of some important regulatory subsets, such as Treg and NK cells.

Reversal of poor graft function with iron-chelating therapy after allogeneic transplantation for severe aplastic anemia

Graft failure (GF) after allogeneic stem cell transplantation (SCT) is a serious complication with high mortality. Graft rejection and poor graft function (PGF) are common causes of GF and they can be distinguished by chimerism analysis: in rejection, chimerism status is mixed, with the presence of recipient cells, whereas full donor chimerism is present in PGF. [1] PGF is defined as severe cytopenia of at least two cell lines (Hb510 g/dl, neutrophil count51 10/ml, platelet count530 10/ml) for at least two consecutive weeks beyond d + 14 posttransplant, with transfusion requirements, associated with hypoplastic-aplastic BM, in the presence of complete donor chimerism and in the absence of severe GVHD and relapse. [2] About 5–27% of transplanted patients will develop PGF; [2,3] several factors have been shown to influence its incidence, such as donor type, HLA matching, ABO incompatibility, cell dose, stem cell source, graft-versus-host disease (GVHD), viral infections, intensity of conditioning regimen, and the use of myelotoxic agents, such as ganciclovir. [4,5] The pathogenesis of PGF is probably multifactorial and includes immunologic issues as well as abnormalities in the bone marrow (BM) microenvironment, number of progenitor cells, and type of the underlying disease. [3] We report of a 27-year-old woman presenting to our clinic in September 2013 because of severe pancytopenia, following an episode of acute cryptogenetic hepatitis in March 2013. She had concomitant autoimmune hypothyroidism treated with L-thyroxine. She had hyporegenerative anemia (Hb 6.6 g/dl, reticulocyte count 2.7% reticulocyte index 0.6), severe thrombocytopenia (Platelets 3 10/l) and moderate neutropenia (840/mmc). Findings of BM aspirate and biopsy (marrow cellularity55%, presence of a lymphoplasmocytic infiltrate 15–20%) were consistent with aplastic anemia (AA). BM colony-forming unit assays showed absent growth for all three hematopoietic lineages. Serologic analyses for viral (HAV, HBV, HCV) and autoimmune (antinuclear, anti-ENA, anti-DNA antibodies) diseases resulted negative. Physical examination was notable for oral petechiae and several bruises at lower limbs. While waiting for availability of horse anti-lymphocyte globulin (h-ALG), she was started on Cyclosporine (150 mg bid) but, after 1 month, she had no hematologic improvement. H-ALG (ATGAM) 40 mg/kg/d was then administered for 4 days. Due to high transfusion requirements (6.4 RBC units/ month) and laboratory evidence of iron overload (serum ferritin 1787 ng/ml [normal value 20–200 ng/ml], transferrin saturation 87.6%) deferasirox 750 mg bid was started in December 2013 (72 days before transplant). Follow-up CBC counts did not show response to hALG (Table I). Due to lack of a matched related donor, she was referred for allogeneic transplantation from matched unrelated donor (UD). Search on the International Bone Marrow Donor Registry (IBMDR) identified a 10/12 male donor with major AB0 incompatibility. The BM was preferred over the peripheral source; the delivered graft contained 2.8 10/kg total nucleated cells; the donor was A Rh-positive, while the patient 0 Rh-positive, with a titre anti-A of 1:64. Therefore an erythrodepletion was planned before reinfusion. She underwent immunosuppressive conditioning with TBI (200 cGy), Cyclophosphamide (300 mg/m) and Fludarabine (30 mg/m). [6] GVHD prophylaxis included antithymocyte globulin 7.5 mg/kg (3.75 mg/kg on days 4 and 3), Cyclosporine 2 mg/kg starting on day 3, Methotrexate 15 mg on d + 2, and 13 mg on days +4, +7. Due to insufficient recovery of mononuclear cells after erythrodepletion of the marrow graft, the whole graft content was reinfused at day 0, after reducing the anti-A titre (to 1:4) with plasma-exchange. [7] She achieved full donor chimerism at d + 14; hematologic recovery was slightly delayed (neutrophils

Predicting failure of hematopoietic stem cell mobilization before it starts: the Predicted Poor Mobilizer (pPM) score

Predicting mobilization failure before it starts may enable patient-tailored strategies. Although consensus criteria for predicted PM (pPM) are available, their predictive performance has never been measured on real data. We retrospectively collected and analyzed 1318 mobilization procedures performed for MM and lymphoma patients in the plerixafor era. In our sample, 180/1318 (13.7%) were PM. The score resulting from published pPM criteria had sufficient performance for predicting PM, as measured by AUC (0.67, 95%CI: 0.63–0.72). We developed a new prediction model from multivariate analysis whose score (pPM-score) resulted in better AUC (0.80, 95%CI: 0.76–0.84, p < 0001). pPM-score included as risk factors: increasing age, diagnosis of NHL, positive bone marrow biopsy or cytopenias before mobilization, previous mobilization failure, priming strategy with G-CSF alone, or without upfront plerixafor. A simplified version of pPM-score was categorized using a cut-off to maximize positive likelihood ratio (15.7, 95%CI: 9.9–24.8); specificity was 98% (95%CI: 97–98.7%), sensitivity 31.7% (95%CI: 24.9–39%); positive predictive value in our sample was 71.3% (95%CI: 60–80.8%). Simplified pPM-score can “rule in” patients at very high risk for PM before starting mobilization, allowing changes in clinical management, such as choice of alternative priming strategies, to avoid highly likely mobilization failure.