Validation of a clinicopathological and gene expression profile model to identify patients with cutaneous melanoma where sentinel lymph node biopsy is unnecessary.

BACKGROUND: In patients with cutaneous melanoma, sentinel lymph node biopsy (SLNB) serves as an important technique to asses disease stage and to guide adjuvant systemic therapy. A model using clinicopathologic and gene expression variables (CP-GEP; Merlin Assay) has recently been introduced to identify patients that may safely forgo SLNB. Herein we present data from an independent validation cohort of the CP-GEP model in Swedish patients. METHODS: Archival histological material (primary melanoma tissue) from a prospectively collected cohort of 421 consecutive patients with pT1-T4 melanoma undergoing SLNB between 2006 and 2014 was analyzed using the CP-GEP model. CP-GEP combines Breslow thickness and patient age with the expression levels of eight genes from the primary melanoma. Stratification is based on their risk for nodal metastasis: CP-GEP Low Risk or CP-GEP High Risk. RESULTS: The SLNB positivity rate was 13%. Of 421 primary melanomas, the CP-GEP model identified 86 patients as having a low risk for nodal metastasis. In patients with pT1-2 melanomas, the SLNB reduction rate was 35.4% (95% CI: 29.4-41.8) with a negative predictive value (NPV) of 96.5% (95% CI: 90.0-99.3). Among patients with pT1-3 melanomas, CP-GEP suggested a SLNB reduction rate of 24.0% (95% CI: 19.7-28.8) and a NPV of 96.5% (95% CI: 90.1-99.3). Only one of 118 pT3 tumors was classified as CP-GEP Low Risk, and all pT4 tumors were classified as being high risk for nodal metastasis. CONCLUSION: This study demonstrates that CP-GEP can identify patients with a low risk for nodal metastasis. Patients with pT1-2 melanomas have the highest clinical benefit from using the test, where 35% of the patients could forgo a SLNB procedure.

The CD34+ Cell Dose Matters in Hematopoietic Stem Cell Transplantation with Peripheral Blood Stem Cells from Sibling Donors.

The effect of CD34+ cell dose in allogeneic hematopoietic stem cell transplantation (HSCT) on overall survival (OS) and incidence of acute and chronic graft-versus-host disease (GvHD) has not been established and few studies have been performed. Our single center analysis included 189 patients with hematological malignancies who received peripheral blood stem cell (PBSC) grafts from sibling donors. Myeloablative conditioning was used in 88 cases and 101 received reduced intensity conditioning. The median CD34+ cell dose was 5.6 × 106/kg (0.6-17.0). In the multivariate analysis, a CD34 cell dose of 6-7 × 106/kg was associated with better OS and lower transplant-related mortality (TRM), while a dose of <5 × 106/kg led to increased relapse and reduced chronic GVHD (cGVHD). A high CD34 cell-dose (>6.5 × 106/kg) correlated with less acute GVHD (aGVHD) II-IV. We conclude that the CD34 cell dose has an impact on the outcome of HSCT from sibling donor PBSCs.

A platform for phenotypic discovery of therapeutic antibodies and targets applied on Chronic Lymphocytic Leukemia.

Development of antibody drugs against novel targets and pathways offers great opportunities to improve current cancer treatment. We here describe a phenotypic discovery platform enabling efficient identification of therapeutic antibody-target combinations. The platform utilizes primary patient cells throughout the discovery process and includes methods for differential phage display cell panning, high-throughput cell-based specificity screening, phenotypic in vitro screening, target deconvolution, and confirmatory in vivo screening. In this study the platform was applied on cancer cells from patients with Chronic Lymphocytic Leukemia resulting in discovery of antibodies with improved cytotoxicity in vitro compared to the standard of care, the CD20-specific monoclonal antibody rituximab. Isolated antibodies were found to target six different receptors on Chronic Lymphocytic Leukemia cells; CD21, CD23, CD32, CD72, CD200, and HLA-DR of which CD32, CD200, and HLA-DR appeared as the most potent targets for antibody-based cytotoxicity treatment. Enhanced antibody efficacy was confirmed in vivo using a patient-derived xenograft model.

Change of apheresis device decreased the incidence of severe acute graft-versus-host disease among patients after allogeneic stem cell transplantation with sibling donors.

BACKGROUND: The composition of the graft used for allogeneic hematopoietic stem cell transplantation (HSCT) is important for the treatment outcome. Different apheresis devices may yield significant differences in peripheral blood stem cell graft cellular composition. We compared stem cell grafts produced by Cobe Spectra (Cobe) and Spectra Optia (Optia) with use of the mononuclear cell (MNC) protocol, and evaluated clinical outcome parameters such as graft-versus-host disease (GvHD), transplant-related mortality (TRM), relapse, and overall survival. STUDY DESIGN AND METHODS: During 5 years, 31 Cobe Spectra and 40 Spectra Optia grafts were analyzed for CD34, CD3, CD4, CD8, CD19, and CD56 cell content. Clinical outcome parameters were correlated and compared between the two patient groups using different apheresis devices. RESULTS: Optia grafts contained fewer lymphocytes compared to Cobe (p < 0.001). Optia grafts had a significantly lower incidence of acute GvHD Grades II through IV (Cobe 45% vs. Optia 23%; p = 0.039) and TRM (16% vs. 2.5%; p < 0.05) but higher chronic GvHD (32% vs. 67%; p = 0.005) compared to Cobe grafts. Finally, the multivariate analysis showed a significant correlation among the different apheresis devices and both acute GvHD II through IV and severe chronic GvHD. The multivariate analysis also showed a significant correlation between the CD3+ cell dose and the incidence of severe acute GvHD. CONCLUSION: Optia-obtained grafts yielded a lower acute GvHD Grades II-IV and TRM risk, but had no impact on relapse or overall survival in this study. Understanding and further improvement of peripheral blood stem cell (PBSC) apheresis techniques may be used in the future to personalize HSCT by, for example, fine-tuning the GvHD incidence.

Patients' experiences of different care settings and a new life situation after allogeneic haematopoietic stem cell transplantation.

Over the past 20 years, considerable healthcare resources have shifted from an inpatient to an outpatient setting. To be in an outpatient setting or at home after allogeneic haematopoietic stem cell transplantation (allo-HSCT) has been shown to be medically safe and beneficial to the patient. In this study we describe patients' experiences of different care settings (hospital or home) and a new life situation during the acute post-transplant phase after HSCT. Semi-structured interviews were conducted with 15 patients (six women and nine men) 29-120 days after HSCT. An inductive qualitative content analysis was performed to analyse the data. The analysis resulted in four categories: To be in a safe place, To have a supportive network, My way of taking control, and My uncertain return to normality. The findings showed that patients undergoing HSCT felt medically safe regardless of the care setting. The importance of a supportive network (i.e. the healthcare team, family and friends) was evident for all patients. Both emotional and problem-focused strategies were used to cope with an uncertain future. Being at home had some positive advantages, including freedom, having the potential for more physical activity, and being with family members. The study highlights some key areas thought to provide more personalised care after HSCT.

Assessment of TREC, KREC and telomere length in long-term survivors after allogeneic HSCT: the role of GvHD and graft source and evidence for telomere homeostasis in young recipients.

Reconstitution of the adaptive immune system following allogeneic hematopoietic stem cell transplantation is crucial for beneficial outcome and is affected by several factors, such as GvHD and graft source. The impact of these factors on immune reconstitution has been thoroughly investigated during the early phase after transplantation. However, little is known about their long-term effect. Similarly, leukocyte telomere length (TL) shortening has been reported shortly after transplantation. Nevertheless, whether TL shortening continues in long-term aspect is still unsettled. Here, we assessed T-cell receptor excision circle (TREC), kappa deleting recombination excision circle (KREC) and leukocyte TL in recipients and donors several years post transplantation (median 17 years). Our analysis showed that, recipients who received bone marrow (BM) as the graft source have higher levels of both TREC and KREC. Also, chronic GvHD affected TREC levels and TL but not KREC levels. Finally, we show that recipient's TL was longer than respective donors in a group of young age recipients with high KREC levels. Our results suggest that BM can be beneficial for long-term adaptive immune recovery. We also present supporting evidence for recipient telomere homeostasis, especially in young age recipients, rather than telomere shortening.

Family members' life situation and experiences of different caring organisations during allogeneic haematopoietic stem cells transplantation-A qualitative study.

The aim of this study was to describe family members' life situation and experiences of care in two different care settings, the patient's home or in hospital during the acute post-transplantation phase after allogeneic haematopoietic stem cell transplantation (HSCT). Data were collected through semi-structured interviews with 14 family members (seven women and seven men). An inductive qualitative content analysis was used to analyse the data. The majority of the family members' (n = 10) had experiences from home care. The findings show the family members' voice of the uncertainty in different ways, related with the unknown prognosis of the HSCT, presented as Being me being us in an uncertain time. The data are classified into; To meet a caring organisation, To be in different care settings, To be a family member and To have a caring relationship. Positive experiences such as freedom and security from home care were identified. The competence and support from the healthcare professionals was profound. Different strategies such as adjusting, having hope and live in the present used to balance to live in an uncertain time. The healthcare professionals need to identify psychosocial problems, and integrate the psychosocial support for the family to alleviate or decrease anxiety during HSCT, regardless of the care setting.

Functionality testing of stem cell grafts to predict infectious complications after allogeneic hematopoietic stem cell transplantation.

BACKGROUND AND OBJECTIVES: Allogeneic hematopoietic stem cell transplantation (HSCT) is a routine clinical procedure performed to treat patients with haematological malignancies, primary immune deficiencies or metabolic disorders. Infections during lymphopenia after allogeneic HSCT are associated with high mortality and morbidity. Typical infectious agents are Epstein-Barr virus, cytomegalovirus, herpes simplex virus, varicella-zoster virus and fungi. The study aim was to evaluate whether measurement of the responses of antigen-specific T-cells, recognizing infectious pathogens would correlate to protective functions in the stem cell recipient post-transplant. MATERIALS AND METHODS: Twenty-one grafts were analysed by flow cytometry and cells were stimulated in vitro with relevant infectious antigens, followed by evaluation of T-cell proliferation and cytokine production. Results were compared to the recipients' clinical records 1-year post-transplantation. RESULTS: We show that an extensive repertoire of transferred antigen-specific T-cells from allogeneic donor grafts against infectious agents, involved in post-transplant infections, are linked to an absence of infectious complications for the recipient up-to 1-year post-transplant. The protective effect was associated with antigen-specific T-cell proliferation and IL-1ß secretion. CONCLUSION: Our results suggest that assaying T-cell function before HSCT could determine individual risks for infectious complications and thus aid in clinical decision-making regarding prophylactic and pre-emptive anti-infective therapy.

No breast cancer subgroup can be spared postoperative radiotherapy after breast-conserving surgery. Fifteen-year results from the Swedish Breast Cancer Group randomised trial, SweBCG 91 RT.

BACKGROUND: Breast-conserving surgery (BCS) followed by radiotherapy (RT) is an established treatment for women with T1-2N0 breast cancers. Since subgroups of patients have low ipsilateral breast tumour recurrence (IBTR) rates, it is important to study whether RT is necessary for all patients. PATIENTS AND METHODS: A total of 1187 women with primary T1-2N0M0 breast cancer were randomised, after standardised sector resection, to postoperative whole breast RT or no local treatment. Adjuvant systemic therapy was offered to patients with stage II cancers. Patients were followed with clinical examinations and annual mammography for 10 years and thereafter referred to the Swedish mammography screening program. RESULTS: After 15 years of follow-up, a higher cumulative incidence of IBTR was observed in control patients, 23.9%, versus irradiated patients, 11.5%, P<0.001. Recurrence-free survival was inferior, 51.7% versus 60.4%, P=0.0013. The main effect of RT was seen during the first 5 years. However, overall survival was not significantly lower 68.4% versus 71.1%, P=0.68, nor was breast cancer-specific mortality significantly higher. CONCLUSIONS: RT after BCS significantly reduced the incidence of IBTR at 15 years of follow-up. We were unable to identify subgroups which could be spared RT. Breast cancer mortality was not significantly reduced after RT. Good predictive markers for radiation sensitivity and improved adjuvant systemic therapy are needed to omit RT after BCS.

Prolonged outbreak of adenovirus A31 in allogeneic stem cell transplant recipients.

BACKGROUND: An outbreak of human adenovirus (HAdV) A31 occurred from December 2011 to March 2012 at the Center for Allogeneic Stem Cell Transplantation (CAST), Karolinska University Hospital in Sweden. We analyzed the outbreak, the routes of transmission, and report the medical consequences. METHODS: The medical records of all patients admitted to CAST during the outbreak period were studied. Phylogenetic analysis of the patient HAdV strains was performed by sequencing the hexon gene and the more variable E3 gene. RESULTS: We identified 9 cases of HAdV A31. Hygiene measures were implemented, but transmission continued for 2 months. All 9 patients had been admitted to the ward, but 2 had no connection in time to other known HAdV A31 cases. DNA sequencing of the patient strains strongly suggested nosocomial transmission. Transplantation was postponed and then cancelled in 1 patient, and 5 patients were treated with cidofovir because of high levels of viremia. In 7 patients, concomitant graft-versus-host disease (GVHD) grade II-V complicated the clinical picture, as it was difficult to distinguish symptoms of GVHD from those of HAdV infection. CONCLUSION: An outbreak of HAdV in HSCT recipients can be difficult to control. Although none of the patients had severe disease, the medical consequences were significant. It is possible that unidentified cases with mild symptoms may have caused continuous transmission at the unit. Regular testing of all patients several weeks beyond the last case identified may be an important measure to control transmission.

Cholera toxin adjuvant promotes a balanced Th1/Th2/Th17 response independently of IL-12 and IL-17 by acting on Gsα in CD11b⁺ DCs.

Despite an extensive literature on the mechanism of action of cholera toxin (CT), we still lack critical information about how the toxin acts as an adjuvant and, especially, which dendritic cells (DCs) are the target cells. Although a T helper type 2 (Th2)-skewing effect of CT is most commonly reported, effective priming of Th17 cells as well as suppression of Th1 responses are well documented. However, the ability of CT to block interferon regulatory factor 8 (IRF8) function and interleukin (IL)-12 production in DCs, which blocks CD8α DC and Th1 cell development, is inconsistent with priming of Th1 and CD8 T cells in many other reports. This prompted us to investigate the adjuvant effect of CT in wild-type, IL-12p40-/-, Batf3-/-, and IL-17A-/- mice and in mice that selectively lack the Gsα target protein for CT adenosine diphosphate (ADP)-ribosylation in DCs. We found that CT promoted Th1 priming independently of IL-12, and whereas Th2 and also Th17 responses were augmented, the gut IgA responses did not require IL-17A. Adjuvanticity was intact in Batf3-/- mice, lacking CD8α(+) DCs, but completely lost in mice with Gsα-deficient CD11c cells. Thus, our data demonstrate that the adjuvant effect requires Gsα expression in CD11b(+) DCs, and that priming of mucosal IgA and CD4 T cells appears unbiased and is independent of IL-12 and IL-17A.

Alpha/beta T-cell depleted grafts as an immunological booster to treat graft failure after hematopoietic stem cell transplantation with HLA-matched related and unrelated donors.

Allogeneic hematopoietic stem cell transplantation is associated with several complications and risk factors, for example, graft versus host disease (GVHD), viral infections, relapse, and graft rejection. While high levels of CD3+ cells in grafts can contribute to GVHD, they also promote the graft versus leukemia (GVL) effect. Infusions of extra lymphocytes from the original stem cell donor can be used as a treatment after transplantation for relapse or poor immune reconstitution but also they increase the risk for GVHD. In peripheral blood, 95% of T-cells express the αß T-cell receptor and the remaining T-cells express the γδ T-cell receptor. As αß T-cells are the primary mediators of GVHD, depleting them from the graft should reduce this risk. In this pilot study, five patients transplanted with HLA-matched related and unrelated donors were treated with αß T-cell depleted stem cell boosts. The majority of γδ T-cells in the grafts expressed Vδ2 and/or Vγ9. Most patients receiving αß-depleted stem cell boosts increased their levels of white blood cells, platelets, and/or granulocytes 30 days after infusion. No signs of GVHD or other side effects were detected. A larger pool of patients with longer follow-up time is needed to confirm the data in this study.

Liquid 1-propanol studied by neutron scattering, near-infrared, and dielectric spectroscopy.

Liquid monohydroxy alcohols exhibit unusual dynamics related to their hydrogen bonding induced structures. The connection between structure and dynamics is studied for liquid 1-propanol using quasi-elastic neutron scattering, combining time-of-flight and neutron spin-echo techniques, with a focus on the dynamics at length scales corresponding to the main peak and the pre-peak of the structure factor. At the main peak, the structural relaxation times are probed. These correspond well to mechanical relaxation times calculated from literature data. At the pre-peak, corresponding to length scales related to H-bonded structures, the relaxation times are almost an order of magnitude longer. According to previous work [C. Gainaru, R. Meier, S. Schildmann, C. Lederle, W. Hiller, E. Rössler, and R. Böhmer, Phys. Rev. Lett. 105, 258303 (2010)] this time scale difference is connected to the average size of H-bonded clusters. The relation between the relaxation times from neutron scattering and those determined from dielectric spectroscopy is discussed on the basis of broad-band permittivity data of 1-propanol. Moreover, in 1-propanol the dielectric relaxation strength as well as the near-infrared absorbance reveal anomalous behavior below ambient temperature. A corresponding feature could not be found in the polyalcohols propylene glycol and glycerol.

Cord blood graft composition impacts the clinical outcome of allogeneic stem cell transplantation.

INTRODUCTION: Despite routine use of umbilical cord blood (CB) grafts as stem cell source for allogeneic stem cell transplantations, much remains unknown regarding their cell composition and correlation with clinical outcome. METHODS: We analyzed material from 30 CB units used for allogeneic hematopoietic stem cell transplantation by multicolor flow cytometry. Phenotypic data were correlated with various clinical outcomes such as survival, graft-versus-host disease (GVHD), relapse, rejection, viral reactivation, and bacteremia. RESULTS: We found that above-median frequencies of CD69+ T cells, naïve CD8+ T cells, and CD127+ B cells in the CB graft were each associated with significantly improved patient survival. Moreover, a statistically significant correlation was seen between higher levels of CD94+ T cells and herpes simplex virus and varicella zoster virus reactivation post transplantation. A similar correlation was seen for the frequency of CD95+ cells in total CD3+, as well as CD4+ and CD8+ T-cell subsets, and viral reactivation. Finally, a higher frequency of naïve CD8+ T cells was associated with the incidence of acute GVHD. CONCLUSION: Our study highlights the importance of further exploration of graft composition before CB transplantation as a tool for risk prediction.

Incidence, risk factors, and outcome of bloodstream infections during the pre-engraftment phase in 521 allogeneic hematopoietic stem cell transplantations.

BACKGROUND: Bloodstream infection (BSI) after allogeneic hematopoietic stem cell transplantation (HSCT) is a well-known complication during the pre-engraftment phase. Knowledge of trends in etiology and antibiotic susceptibility of BSI is important as the time to effective antibiotic treatment is closely associated with survival in bacteremic patients with septic shock. METHODS: BSI during the pre-engraftment phase was studied retrospectively in 521 patients undergoing HSCT at our center in 2001-2008. Incidence, risk factors, outcome, and microbiology findings were investigated and compared with BSI in a cohort transplanted during 1975-1996. RESULTS: The incidence of at least 1 episode of BSI was 21%, the total attributable mortality of BSI was 3.3%, and crude mortality at day 120 after transplantation was 21%. The rate of gram-positive and gram-negative BSI was 80% and 13%, respectively. Gram-negative BSI was more frequent both in 2001-2004 and in 2005-2008 compared with 1986-1996 (P = 0.023 for 2001-2004, P = 0.001 for 2005-2008), with fluoroquinolone-resistant Escherichia coli as the predominant finding. BSI with viridans streptococci and E. coli occurred significantly earlier after HSCT than BSI with Enterococcus species, with median time of 4, 8, and 11 days, respectively (P < 0.01 both for viridians streptococci vs. Enterococcus species, and E. coli vs. Enterococcus species). Risk factors for BSI in multivariate analysis were transplantation from unrelated donor and cord blood as stem cell source, whereas peripheral blood as stem cell source was protective. CONCLUSIONS: Despite low attributable mortality of BSI, crude mortality at day 120 after transplantation was 21%, indicating an association between BSI and other risk factors for death. The risk of gram-negative BSI increased over time in parallel with an increased rate of quinolone resistance. However, the incidence and attributable mortality of gram-negative BSI remained low. Thus, prophylaxis with ciprofloxacin is still deemed appropriate, but continued assessments of the risk and benefits of fluoroquinolone prophylaxis must be performed.

Isolated hepatic perfusion for ocular melanoma metastasis: registry data suggests a survival benefit.

BACKGROUND: Uveal melanoma is the most common primary intraocular malignancy in adults. Despite successful control of the primary tumor, metastatic disease will ultimately develop in approximately 35% of the patients, with the liver being the most common site for metastases. These metastases are generally refractory to systemic chemotherapy, and the median survival for patients with liver metastases is about 6 months. This phase II trial reports the experience of isolated hepatic perfusion (IHP) as a treatment option. METHOD: A total of 34 patients with isolated liver metastasis from ocular melanoma underwent IHP. An overall survival comparison was made using data retrieved from the National Patient Register managed by the Swedish National Board of Health and Welfare. RESULTS: An overall radiological response was seen in 68% of the patients, with 12% having a complete response. Time to local progression was 7 months; 68% of the patients developed extrahepatic metastases after a median of 13 months, and the median overall survival was 24 months. There was a significant survival advantage of 14 months (p = 0.029) when comparing these patients with a control group consisting of the longest surviving patients in Sweden with uveal melanoma liver metastases not treated with IHP. CONCLUSIONS: IHP is a treatment option with a high response rate and a potential survival benefit of more than 1 year. IHP should be considered an option in the treatment of uveal melanoma metastases. A randomized trial comparing IHP and best alternative care will start during 2013 (the SCANDIUM trial, ClinicalTrials.gov identifier NCT01785316).