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Background: Histologic transformation (HT) is an important event with adverse prognosis in the natural history of indolent lymphomas. There are minimal data on HT in the Australian setting. Aim(s): To characterise patients with biopsy-proven HT and their outcomes identified at a tertiary Australian Hospital. Method(s): All patients with biopsy-proven HT during a 15-year period (2002-2017) were included. Clinico-pathological data were systematically collected from review of patient records. Survival estimates were assessed using the Kaplan-Meier method and compared using the log-rank test. Associations between variables and clinical outcomes were evaluated using Cox's proportional hazards model. Result(s): A cohort of 45 patients was identified with a median age of 66 years and the majority (59%) having high-risk disease (Revised-International Prognostic Index score >=3). R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone) induction was used in 69%, with an overall response rate of 82% (complete response (CR), 75%). Sixty-one percent of these induction responders received consolidation, with autologous stem cell transplant (ASCT) performed in only 17% and rituximab maintenance given to 31%. With a median follow up of 47 months (range: 4-136), the 5-year overall survival (OS) was 69% (95% CI: 52%, 81%). Chemotherapy-naivety at HT was associated with a superior rate of CR (84% vs 54%, P = 0.057) and 5-year OS (82% vs 46%, P = 0.012). Rituximab maintenance was associated with a durable progression-free survival in induction responders. Conclusion(s): Excellent OS was observed in this modern cohort of patients treated with rituximab-containing induction and low rate of consolidation by ASCT, particularly in those who were chemotherapy-naive at HT.Copyright © 2020 Royal Australasian College of Physicians

The last decade has seen a significant leap in our understanding of the wide range of genetic lesions underpinning acute lymphoblastic leukaemia (ALL). Next generation sequencing has led to the identification of driver mutations with significant implications on prognosis and has defined entities such as BCR-ABL-like ALL, where targeted therapies such as tyrosine kinase inhibitors (TKIs) and JAK inhibitors may play a role in its treatment. In Philadelphia positive ALL, the introduction of TKIs into frontline treatment regimens has already transformed patient outcomes. In B-ALL, agents targeting surface receptors CD19, CD20 and CD22, including monoclonal antibodies, bispecific T cell engagers, antibody drug conjugates and chimeric antigen receptor (CAR) T cells, have shown significant activity but come with unique toxicities and have implications for how treatment is sequenced. Advances in T-ALL have lagged behind those seen in B-ALL. However, agents such as nelarabine, bortezomib and CAR T cell therapy targeting T cell antigens have been examined with promising results seen. As our understanding of disease biology in ALL grows, as does our ability to target pathways such as apoptosis, through BH3 mimetics, chemokines and epigenetic regulators. This review aims to highlight a range of available and emerging targeted therapeutics in ALL, to explore their mechanisms of action and to discuss the current evidence for their use.Copyright © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

The last decade has seen a significant leap in our understanding of the wide range of genetic lesions underpinning acute lymphoblastic leukaemia (ALL). Next generation sequencing has led to the identification of driver mutations with significant implications on prognosis and has defined entities such as BCR-ABL-like ALL, where targeted therapies such as tyrosine kinase inhibitors (TKIs) and JAK inhibitors may play a role in its treatment. In Philadelphia positive ALL, the introduction of TKIs into frontline treatment regimens has already transformed patient outcomes. In B-ALL, agents targeting surface receptors CD19, CD20 and CD22, including monoclonal antibodies, bispecific T cell engagers, antibody drug conjugates and chimeric antigen receptor (CAR) T cells, have shown significant activity but come with unique toxicities and have implications for how treatment is sequenced. Advances in T-ALL have lagged behind those seen in B-ALL. However, agents such as nelarabine, bortezomib and CAR T cell therapy targeting T cell antigens have been examined with promising results seen. As our understanding of disease biology in ALL grows, as does our ability to target pathways such as apoptosis, through BH3 mimetics, chemokines and epigenetic regulators. This review aims to highlight a range of available and emerging targeted therapeutics in ALL, to explore their mechanisms of action and to discuss the current evidence for their use.Copyright © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

BACKGROUND: Histologic transformation (HT) is an important event with adverse prognosis in the natural history of indolent lymphomas. There is minimal data on HT in the Australian setting. AIM: To characterise patients with biopsy-proven HT and their outcomes identified at a tertiary Australian Hospital. METHOD(S): All patients with biopsy-proven HT during a 15-year period (2002-2017) were included. Clinico-pathological data were systematically collected from review of patient records. Survival estimates were assessed by the Kaplan-Meier method and compared using the log-rank test. Associations between variables and clinical outcomes were evaluated using Cox's proportional hazards model. RESULT(S): A cohort of 45 patients was identified with a median age of 66years and the majority (59%) having high-risk disease (Revised-International Prognostic Index score >=3). R-CHOP induction was used in 69% with an overall response rate of 82% (complete response (CR), 75%). 61% of these induction-responders received consolidation, with autologous stem cell transplant (ASCT) performed in only 17% and rituximab maintenance given to 31%. With a median follow-up of 47 months (range: 4-136), the 5-year overall survival (OS) was 69% (95% CI: 52%, 81%). Chemotherapy-naivety at HT was associated with a superior rate of CR (84% vs. 54%, p = 0.057) and 5-year OS (82% vs. 46%, p = 0.012). Rituximab maintenance was associated with a durable progression-free survival in induction-responders. CONCLUSION(S): Excellent OS was observed in this modern cohort of patients treated with rituximab-containing induction and low rate of consolidation by ASCT, particularly in those who were chemotherapy-naive at HT. This article is protected by copyright. All rights reserved.

Multiple myeloma is an incurable and fatal cancer of immunoglobulin-secreting plasma cells. Most conventional therapies aim to induce apoptosis in myeloma cells but resistance to these drugs often arises and drives relapse. In this study, we sought to identify the best adjunct targets to kill myeloma cells resistant to conventional therapies using deep profiling by mass cytometry (CyTOF). We validated probes to simultaneously detect 26 regulators of cell death, mitosis, cell signaling, and cancer-related pathways at the single-cell level following treatment of myeloma cells with dexamethasone or bortezomib. Time-resolved visualization algorithms and machine learning random forest models (RFMs) delineated putative cell death trajectories and a hierarchy of parameters that specified myeloma cell survival versus apoptosis following treatment. Among these parameters, increased amounts of phosphorylated cAMP response element-binding protein (CREB) and the pro-survival protein, MCL-1, were defining features of cells surviving drug treatment. Importantly, the RFM prediction that the combination of an MCL-1 inhibitor with dexamethasone would elicit potent, synergistic killing of myeloma cells was validated in other cell lines, in vivo preclinical models and primary myeloma samples from patients. Furthermore, CyTOF analysis of patient bone marrow cells clearly identified myeloma cells and their key cell survival features. This study demonstrates the utility of CyTOF profiling at the single-cell level to identify clinically relevant drug combinations and tracking of patient responses for future clinical trials.Copyright © 2020, The Author(s), under exclusive licence to ADMC Associazione Differenziamento e Morte Cellulare.

BACKGROUND: Histologic transformation (HT) is an important event with adverse prognosis in the natural history of indolent lymphomas. There is minimal data on HT in the Australian setting. AIM: To characterise patients with biopsy-proven HT and their outcomes identified at a tertiary Australian Hospital. METHOD(S): All patients with biopsy-proven HT during a 15-year period (2002-2017) were included. Clinico-pathological data were systematically collected from review of patient records. Survival estimates were assessed by the Kaplan-Meier method and compared using the log-rank test. Associations between variables and clinical outcomes were evaluated using Cox's proportional hazards model. RESULT(S): A cohort of 45 patients was identified with a median age of 66years and the majority (59%) having high-risk disease (Revised-International Prognostic Index score >=3). R-CHOP induction was used in 69% with an overall response rate of 82% (complete response (CR), 75%). 61% of these induction-responders received consolidation, with autologous stem cell transplant (ASCT) performed in only 17% and rituximab maintenance given to 31%. With a median follow-up of 47 months (range: 4-136), the 5-year overall survival (OS) was 69% (95% CI: 52%, 81%). Chemotherapy-naivety at HT was associated with a superior rate of CR (84% vs. 54%, p = 0.057) and 5-year OS (82% vs. 46%, p = 0.012). Rituximab maintenance was associated with a durable progression-free survival in induction-responders. CONCLUSION(S): Excellent OS was observed in this modern cohort of patients treated with rituximab-containing induction and low rate of consolidation by ASCT, particularly in those who were chemotherapy-naive at HT. This article is protected by copyright. All rights reserved.

Multiple myeloma is an incurable and fatal cancer of immunoglobulin-secreting plasma cells. Most conventional therapies aim to induce apoptosis in myeloma cells but resistance to these drugs often arises and drives relapse. In this study, we sought to identify the best adjunct targets to kill myeloma cells resistant to conventional therapies using deep profiling by mass cytometry (CyTOF). We validated probes to simultaneously detect 26 regulators of cell death, mitosis, cell signaling, and cancer-related pathways at the single-cell level following treatment of myeloma cells with dexamethasone or bortezomib. Time-resolved visualization algorithms and machine learning random forest models (RFMs) delineated putative cell death trajectories and a hierarchy of parameters that specified myeloma cell survival versus apoptosis following treatment. Among these parameters, increased amounts of phosphorylated cAMP response element-binding protein (CREB) and the pro-survival protein, MCL-1, were defining features of cells surviving drug treatment. Importantly, the RFM prediction that the combination of an MCL-1 inhibitor with dexamethasone would elicit potent, synergistic killing of myeloma cells was validated in other cell lines, in vivo preclinical models and primary myeloma samples from patients. Furthermore, CyTOF analysis of patient bone marrow cells clearly identified myeloma cells and their key cell survival features. This study demonstrates the utility of CyTOF profiling at the single-cell level to identify clinically relevant drug combinations and tracking of patient responses for future clinical trials.Copyright © 2020, The Author(s), under exclusive licence to ADMC Associazione Differenziamento e Morte Cellulare.

Plasma cells can provide lifelong immunity by the continued production of high-affinity, antigen-specific antibody. Low et al. reveal that the transcription factor IRF4 acts within established plasma cells to prevent apoptosis and maintain cellular identity and mitochondrial homeostasis. The transcription factor interferon regulatory factor 4 (IRF4) is critical for the development, maintenance, and function of plasma cells. The mechanism by which IRF4 exerts its action in mature plasma cells has been elusive due to the death of all such cells upon IRF4 loss. While we identify apoptosis as a critical pathway for the death of plasma cells caused by IRF4 loss, we also determine that IRF4 did not regulate the intrinsic apoptotic pathway directly. By using an inducible IRF4 deletion system in the presence of the overexpression of anti-apoptotic BCL2, we identify genes whose expression is coordinated by IRF4 and that in turn specify plasma cell identity and mitochondrial homeostasis.Copyright © 2019 The Authors

Recent studies have demonstrated that the immunomodulatory drugs (IMiDs) lead to the degradation of the transcription factors Ikaros and Aiolos. However, why their loss subsequently leads to multiple myeloma (MM) cell death remains unclear. Using CRISPR-Cas9 genome editing, we have deleted IKZF1/Ikaros and IKZF3/Aiolos in human MM cell lines to gain further insight into their downstream gene regulatory networks. Inactivation of either factor alone recapitulates the cell intrinsic action of the IMiDs, resulting in cell cycle arrest and induction of apoptosis. Furthermore, evaluation of the transcriptional changes resulting from their loss demonstrates striking overlap with lenalidomide treatment. This was not dependent on reduction of the IRF4-MYC "axis," as neither protein was consistently downregulated, despite cell death occurring, and overexpression of either factor failed to rescue for Ikaros loss. Importantly, Ikaros and Aiolos repress the expression of interferon-stimulated genes (ISGs), including CD38, and their loss led to the activation of an interferon-like response, contributing to MM cell death. Ikaros/Aiolos repressed CD38 expression through interaction with the nucleosome remodeling and deacetylase complex in MM. IMiD-induced loss of Ikaros or treatment with interferon resulted in an upregulation of CD38 surface expression on MM cells, priming for daratumumab-induced NK cell-mediated antibody-dependent cellular cytotoxicity. These results give further insight into the mechanism of action of the IMiDs and provide mechanistic rationale for combination with anti-CD38 monoclonal antibodies. (Blood. 2018;132(20):2166-2178)Copyright © 2018 by The American Society of Hematology.

Plasma cells can provide lifelong immunity by the continued production of high-affinity, antigen-specific antibody. Low et al. reveal that the transcription factor IRF4 acts within established plasma cells to prevent apoptosis and maintain cellular identity and mitochondrial homeostasis. The transcription factor interferon regulatory factor 4 (IRF4) is critical for the development, maintenance, and function of plasma cells. The mechanism by which IRF4 exerts its action in mature plasma cells has been elusive due to the death of all such cells upon IRF4 loss. While we identify apoptosis as a critical pathway for the death of plasma cells caused by IRF4 loss, we also determine that IRF4 did not regulate the intrinsic apoptotic pathway directly. By using an inducible IRF4 deletion system in the presence of the overexpression of anti-apoptotic BCL2, we identify genes whose expression is coordinated by IRF4 and that in turn specify plasma cell identity and mitochondrial homeostasis.Copyright © 2019 The Authors

Recent studies have demonstrated that the immunomodulatory drugs (IMiDs) lead to the degradation of the transcription factors Ikaros and Aiolos. However, why their loss subsequently leads to multiple myeloma (MM) cell death remains unclear. Using CRISPR-Cas9 genome editing, we have deleted IKZF1/Ikaros and IKZF3/Aiolos in human MM cell lines to gain further insight into their downstream gene regulatory networks. Inactivation of either factor alone recapitulates the cell intrinsic action of the IMiDs, resulting in cell cycle arrest and induction of apoptosis. Furthermore, evaluation of the transcriptional changes resulting from their loss demonstrates striking overlap with lenalidomide treatment. This was not dependent on reduction of the IRF4-MYC "axis," as neither protein was consistently downregulated, despite cell death occurring, and overexpression of either factor failed to rescue for Ikaros loss. Importantly, Ikaros and Aiolos repress the expression of interferon-stimulated genes (ISGs), including CD38, and their loss led to the activation of an interferon-like response, contributing to MM cell death. Ikaros/Aiolos repressed CD38 expression through interaction with the nucleosome remodeling and deacetylase complex in MM. IMiD-induced loss of Ikaros or treatment with interferon resulted in an upregulation of CD38 surface expression on MM cells, priming for daratumumab-induced NK cell-mediated antibody-dependent cellular cytotoxicity. These results give further insight into the mechanism of action of the IMiDs and provide mechanistic rationale for combination with anti-CD38 monoclonal antibodies. (Blood. 2018;132(20):2166-2178)Copyright © 2018 by The American Society of Hematology.

Antibody Secreting Cells (ASCs) are a fundamental component of humoral immunity, however, deregulated or excessive antibody production contributes to the pathology of autoimmune diseases, while transformation of ASCs results in the malignancy Multiple Myeloma (MM). Despite substantial recent improvements in treating these conditions, there is as yet no widely used ASC-specific therapeutic approach, highlighting a critical need to identify novel methods of targeting normal and malignant ASCs. Surface molecules specifically expressed by the target cell population represent ideal candidates for a monoclonal antibody-based therapy. By interrogating the ASC gene signature that we previously defined we identified three surface proteins, Plpp5, Clptm1l and Itm2c, which represent potential targets for novel MM treatments. Plpp5, Clptm1l and Itm2c are highly and selectively expressed by mouse and human ASCs as well as MM cells. To investigate the function of these proteins within the humoral immune system we have generated three novel mouse strains, each carrying a loss-of-function mutation in either Plpp5, Clptm1l or Itm2c. Through analysis of these novel strains, we have shown that Plpp5, Clptm1l and Itm2c are dispensable for the development, maturation and differentiation of B-lymphocytes, and for the production of antibodies by ASCs. As adult mice lacking either protein showed no apparent disease phenotypes, it is likely that targeting these molecules on ASCs will have minimal on-target adverse effects.Copyright © 2018 by the authors. Licensee MDPI, Basel, Switzerland.

Introduction Current DLBCL prognostic scores are poor predictors of individual risk with 26% of "very good" and "good" risk patients having unfavourable outcomes. Hypercalcaemia has been associated with poor outcomes in several cancers including multiple myeloma, but is not included in current DLBCL prognostic scoring models. Aim To determine the prognostic significance of hypercalcaemia in newly diagnosed DLBCL and identify relationships between hypercalcaemia and other established prognostic variables including cell of origin, and the components of the revised international prognostic index (R-IPI including impaired performance status, older age, advanced stage, elevated lactate dehydrogenase, and extranodal involvement. Methods Retrospective cohort study at two academic healthcare networks in Melbourne, Australia. All patients with a newly diagnosed DLBCL by WHO 2008 criteria were eligible for inclusion. Cases were identified from hospital lymphoma databases. Only cases with adequate clinical information including baseline characteristics, therapy received and outcomes were included. Cell of origin was determined by the modified Hans criteria. At Monash Health, patients were included from September 2002 until December 2015, and from Western Health, patients were included from January 2012 until December 2015. Hypercalcaemia was defined as a serum calcium >2.6mmol/L within 30 days prior to, or 15 days following the diagnosis of DLBCL. Serum calcium levels were corrected for hypoalbuminaemia if present. Immunohistochemical cell of origin was recorded using modified Hans criteria (Meyer et al. 2011). Survival correlates were estimated by Kaplan-Meier log rank method. Multivariable analysis was performed by Cox regression. Relationships between serum calcium and other covariates were examined by t test for continuous variables and chi for categorical variables. This project was approved by the human research review boards at each institution. Results A total of 514 p

The transcription factors Ikaros (IKZF1) and Aiolos (IKZF3) have recently been found to be targeted for destruction by the immunomodulatory drugs (IMiDs). However, the role of Ikaros and Aiolos and why their loss leads to multiple myeloma (MM) cell death remains unclear. We have used CRISPRCas9 genome editing to delete IKZF1 and IKZF3 in human MM cell lines to gain further insight into their downstream gene regulatory networks. Consistent with the action of the IMiDs, loss of either of these genes resulted in both a G1/S cell cycle arrest and induction of apoptosis. This was not dependent on the subsequent reduction of the IRF4-MYC "axis", as neither IRF4 or MYC were consistently downregulated across all cell lines examined (despite cell death occurring) and overexpression of either factor failed to rescue for Ikaros loss. This led us to further investigate the transcriptional changes resulting from deletion of Ikaros, Aiolos or treatment with the IMiD lenalidomide using RNA-sequencing. Supporting their central role in the action of the IMiDs, significant overlap was seen on loss of Ikaros, Aiolos, or lenalidomide treatment; with loss of Ikaros/Aiolos accounting for approximately 75% of differential gene expression following lenalidomide. Importantly both Ikaros and Aiolos were found to repress the expression of interferon stimulated genes (ISGs) and their loss led to the activation of an interferon-like response, possibly contributing to cell death. In keeping with this, treatment with lenalidomide and low dose IFNb resulted in synergistic cell death. Furthermore, CD38 appeared to be both an ISG and repressed target of Ikaros/Aiolos through their interaction with the nucleosome remodelling and deacetylase complex, and its expression is increased at both mRNA and protein level on their loss. Given recent clinical studies have shown improved outcomes with combination treatment with lenalidomide and the anti-CD38 monoclonal antibody daratumumab, we wondered whether th

Antibody Secreting Cells (ASCs) are a fundamental component of humoral immunity, however, deregulated or excessive antibody production contributes to the pathology of autoimmune diseases, while transformation of ASCs results in the malignancy Multiple Myeloma (MM). Despite substantial recent improvements in treating these conditions, there is as yet no widely used ASC-specific therapeutic approach, highlighting a critical need to identify novel methods of targeting normal and malignant ASCs. Surface molecules specifically expressed by the target cell population represent ideal candidates for a monoclonal antibody-based therapy. By interrogating the ASC gene signature that we previously defined we identified three surface proteins, Plpp5, Clptm1l and Itm2c, which represent potential targets for novel MM treatments. Plpp5, Clptm1l and Itm2c are highly and selectively expressed by mouse and human ASCs as well as MM cells. To investigate the function of these proteins within the humoral immune system we have generated three novel mouse strains, each carrying a loss-of-function mutation in either Plpp5, Clptm1l or Itm2c. Through analysis of these novel strains, we have shown that Plpp5, Clptm1l and Itm2c are dispensable for the development, maturation and differentiation of B-lymphocytes, and for the production of antibodies by ASCs. As adult mice lacking either protein showed no apparent disease phenotypes, it is likely that targeting these molecules on ASCs will have minimal on-target adverse effects.Copyright © 2018 by the authors. Licensee MDPI, Basel, Switzerland.

Introduction Current DLBCL prognostic scores are poor predictors of individual risk with 26% of "very good" and "good" risk patients having unfavourable outcomes. Hypercalcaemia has been associated with poor outcomes in several cancers including multiple myeloma, but is not included in current DLBCL prognostic scoring models. Aim To determine the prognostic significance of hypercalcaemia in newly diagnosed DLBCL and identify relationships between hypercalcaemia and other established prognostic variables including cell of origin, and the components of the revised international prognostic index (R-IPI including impaired performance status, older age, advanced stage, elevated lactate dehydrogenase, and extranodal involvement. Methods Retrospective cohort study at two academic healthcare networks in Melbourne, Australia. All patients with a newly diagnosed DLBCL by WHO 2008 criteria were eligible for inclusion. Cases were identified from hospital lymphoma databases. Only cases with adequate clinical information including baseline characteristics, therapy received and outcomes were included. Cell of origin was determined by the modified Hans criteria. At Monash Health, patients were included from September 2002 until December 2015, and from Western Health, patients were included from January 2012 until December 2015. Hypercalcaemia was defined as a serum calcium >2.6mmol/L within 30 days prior to, or 15 days following the diagnosis of DLBCL. Serum calcium levels were corrected for hypoalbuminaemia if present. Immunohistochemical cell of origin was recorded using modified Hans criteria (Meyer et al. 2011). Survival correlates were estimated by Kaplan-Meier log rank method. Multivariable analysis was performed by Cox regression. Relationships between serum calcium and other covariates were examined by t test for continuous variables and chi for categorical variables. This project was approved by the human research review boards at each institution. Results A total of 514 p

The transcription factors Ikaros (IKZF1) and Aiolos (IKZF3) have recently been found to be targeted for destruction by the immunomodulatory drugs (IMiDs). However, the role of Ikaros and Aiolos and why their loss leads to multiple myeloma (MM) cell death remains unclear. We have used CRISPRCas9 genome editing to delete IKZF1 and IKZF3 in human MM cell lines to gain further insight into their downstream gene regulatory networks. Consistent with the action of the IMiDs, loss of either of these genes resulted in both a G1/S cell cycle arrest and induction of apoptosis. This was not dependent on the subsequent reduction of the IRF4-MYC "axis", as neither IRF4 or MYC were consistently downregulated across all cell lines examined (despite cell death occurring) and overexpression of either factor failed to rescue for Ikaros loss. This led us to further investigate the transcriptional changes resulting from deletion of Ikaros, Aiolos or treatment with the IMiD lenalidomide using RNA-sequencing. Supporting their central role in the action of the IMiDs, significant overlap was seen on loss of Ikaros, Aiolos, or lenalidomide treatment; with loss of Ikaros/Aiolos accounting for approximately 75% of differential gene expression following lenalidomide. Importantly both Ikaros and Aiolos were found to repress the expression of interferon stimulated genes (ISGs) and their loss led to the activation of an interferon-like response, possibly contributing to cell death. In keeping with this, treatment with lenalidomide and low dose IFNb resulted in synergistic cell death. Furthermore, CD38 appeared to be both an ISG and repressed target of Ikaros/Aiolos through their interaction with the nucleosome remodelling and deacetylase complex, and its expression is increased at both mRNA and protein level on their loss. Given recent clinical studies have shown improved outcomes with combination treatment with lenalidomide and the anti-CD38 monoclonal antibody daratumumab, we wondered whether th

Current screening guidelines for malaria in new refugees include a combination of thick and thin film examination and immunochromatographic antigen test (ICT). However, as the prevalence of malaria in our population has decreased due to changing refugee demographics, we sought to determine if an ICT alone can reliably exclude malaria in our asymptomatic refugee population.A retrospective analysis was conducted of all investigations for malaria performed from 1 August 2011 to 31 July 2013, including thick and thin blood film examination, BinaxNOW ICT, and external morphological and polymerase chain reaction (PCR) validation where applicable.Malaria was diagnosed in 45 of 1248 (3.6%) patients investigated, all of whom were symptomatic and the majority (71.1%) returned travellers. All 599 asymptomatic refugees screened were negative. Overall, 42 of 45 malaria cases were detected by the ICT; sensitivity 93.3% (95% CI 80.7-98.3%) and negative predictive value (NPV) 99.8% (99.2-99.9%). All 21 cases of Plasmodium falciparum and 20 of 22 cases of Plasmodium vivax were detected, giving a sensitivity of 100% (80.8-100%) and 90.9% (69.4-98.4%) respectively. Too few cases of Plasmodium malariae and no cases of Plasmodium ovale or Plasmodium knowlesi were diagnosed for adequate assessment to be carried out.These data suggest that full malaria screening in all asymptomatic refugees with the combination of thick and thin blood films and rapid antigen test may not be warranted. Alternative screening approaches should be considered, including the use of ICT alone, or limiting screening of asymptomatic refugees to only those originating from countries with high incidence of malaria.

Current screening guidelines for malaria in new refugees include a combination of thick and thin film examination and immunochromatographic antigen test (ICT). However, as the prevalence of malaria in our population has decreased due to changing refugee demographics, we sought to determine if an ICT alone can reliably exclude malaria in our asymptomatic refugee population.A retrospective analysis was conducted of all investigations for malaria performed from 1 August 2011 to 31 July 2013, including thick and thin blood film examination, BinaxNOW ICT, and external morphological and polymerase chain reaction (PCR) validation where applicable.Malaria was diagnosed in 45 of 1248 (3.6%) patients investigated, all of whom were symptomatic and the majority (71.1%) returned travellers. All 599 asymptomatic refugees screened were negative. Overall, 42 of 45 malaria cases were detected by the ICT; sensitivity 93.3% (95% CI 80.7-98.3%) and negative predictive value (NPV) 99.8% (99.2-99.9%). All 21 cases of Plasmodium falciparum and 20 of 22 cases of Plasmodium vivax were detected, giving a sensitivity of 100% (80.8-100%) and 90.9% (69.4-98.4%) respectively. Too few cases of Plasmodium malariae and no cases of Plasmodium ovale or Plasmodium knowlesi were diagnosed for adequate assessment to be carried out.These data suggest that full malaria screening in all asymptomatic refugees with the combination of thick and thin blood films and rapid antigen test may not be warranted. Alternative screening approaches should be considered, including the use of ICT alone, or limiting screening of asymptomatic refugees to only those originating from countries with high incidence of malaria.

BACKGROUND: Demand for platelet (PLT) and plasma transfusions is increasing. Improved clinical supply and contingency planning requires greater understanding of usage profiles and urgency of clinical requirement. STUDY DESIGN AND METHODS: This study was a random-sample survey of PLT and plasma units produced in Victoria, Australia, to determine product disposition, recipient demographics, clinical indications for transfusion, and urgency (or "deferability") of need. PLTs and fresh-frozen plasma (FFP) were tagged with a case report form before distribution. RESULT(S): A total of 1252 PLT and 1837 FFP units were tagged, comprising 8.3 and 13.3% of all products issued during the study period. The fate of 1243 PLT and 1808 FFP units was determined. Of products issued, 72.2% of PLTs and 87.8% of FFP were transfused. Hematologic and oncologic disorders accounted for 63.9% of PLT transfusions, with acute myeloid leukemia alone accounting for 26%. Conversely, surgical patients received the largest proportion of FFP (40.4%), predominantly for cardiothoracic, solid organ transplant, and vascular surgery. Approximately 15% of PLT transfusions and 35% of plasma transfusions were required within 1 hour, and 80% of PLT transfusions and 90% of FFP transfusions were required within 24 hours. Wastage rates were higher in regional blood banks. CONCLUSION(S): The PUPPY study is a comprehensive and detailed population-based assessment of PLT and plasma usage, including urgency of use. It identifies specific clinical areas with high demand for PLT and FFP transfusion and demonstrates the high urgency of need for both products. These data inform clinical supply and contingency planning activities.Copyright © 2016 AABB

BACKGROUND: Demand for platelet (PLT) and plasma transfusions is increasing. Improved clinical supply and contingency planning requires greater understanding of usage profiles and urgency of clinical requirement. STUDY DESIGN AND METHODS: This study was a random-sample survey of PLT and plasma units produced in Victoria, Australia, to determine product disposition, recipient demographics, clinical indications for transfusion, and urgency (or "deferability") of need. PLTs and fresh-frozen plasma (FFP) were tagged with a case report form before distribution. RESULT(S): A total of 1252 PLT and 1837 FFP units were tagged, comprising 8.3 and 13.3% of all products issued during the study period. The fate of 1243 PLT and 1808 FFP units was determined. Of products issued, 72.2% of PLTs and 87.8% of FFP were transfused. Hematologic and oncologic disorders accounted for 63.9% of PLT transfusions, with acute myeloid leukemia alone accounting for 26%. Conversely, surgical patients received the largest proportion of FFP (40.4%), predominantly for cardiothoracic, solid organ transplant, and vascular surgery. Approximately 15% of PLT transfusions and 35% of plasma transfusions were required within 1 hour, and 80% of PLT transfusions and 90% of FFP transfusions were required within 24 hours. Wastage rates were higher in regional blood banks. CONCLUSION(S): The PUPPY study is a comprehensive and detailed population-based assessment of PLT and plasma usage, including urgency of use. It identifies specific clinical areas with high demand for PLT and FFP transfusion and demonstrates the high urgency of need for both products. These data inform clinical supply and contingency planning activities.Copyright © 2016 AABB

Background: Multiple myeloma prognostication is largely defined by the International Staging System (ISS)1 using serum beta-2 microglobulin (beta2M) and albumin results. Local external quality assurance data showed substantial, method-dependant variations in beta2M results. Aim(s): We determined the variability of different beta2M methods using patient samples, and whether this influences ISS prognostic scores. Method(s): beta2M stability after freeze-thawing was established first. Specimens (n=21) were then sent frozen to four laboratories using different methods/instruments for beta2M: Lab1, nephelometric (Beckman Coulter Immage); Lab2, chemiluminescent (Immulite 2000); Lab3, turbidimetric (Roche Cobas c502) and Lab 4 turbidimetric (Abbott Architect c16200). All laboratories use the same albumin method (bromocresol purple). Result(s): Lab1 produced beta2M results consistently higher than other laboratories (Figure 1): mean difference +1.55 mg/L (28.5%) (95%CI: 1.02 - 2.09, p<0.0001) compared to Lab2; +1.09 mg/L (20.1%) (95% CI: 0.78 - 1.41, p<0.0001) to Lab3; and +0.51 mg/L (9.3%) (95% CI: 0.29 - 0.73, p<0.0001) to Lab4. Compared to Lab1, ISS scores were classified one stage lower in 7, 5 and 3 patients by Lab2, Lab3 and Lab4, respectively. One patient could have been classified either stage I, II or III depending on which laboratory performed the tests. Albumin methods aligned well and had minimal impact on ISS score. Summary and Conclusion(s): Results from different beta2M methods are highly variable and harmonisation is required. This study has important implications on the validity of ISS staging for myeloma.

Central nervous system (CNS) relapse in diffuse large B-cell lymphoma (DLBCL) is a devastating complication; the optimal prophylactic strategy remains unclear. We performed a multicentre, retrospective analysis of patients with DLBCL with high risk for CNS relapse as defined by two or more of: multiple extranodal sites, elevated serum LDH and B symptoms or involvement of specific high-risk anatomical sites. We compared three different strategies of CNS-directed therapy: intrathecal (IT) methotrexate (MTX) with (R)-CHOP 'group 1'; R-CHOP with IT MTX and two cycles of high-dose intravenous (IV) MTX 'group 2'; dose-intensive systemic antimetabolite-containing chemotherapy (Hyper-CVAD or CODOXM/IVAC) with IT/IV MTX 'group 3'. Overall, 217 patients were identified (49, 125 and 43 in groups 1-3, respectively). With median follow-up of 3.4 (range 0.2-18.6) years, 23 CNS relapses occurred (12, 10 and 1 in groups 1-3 respectively). The 3-year actuarial rates (95% CI) of CNS relapse were 18.4% (9.5-33.1%), 6.9% (3.5-13.4%) and 2.3% (0.4-15.4%) in groups 1-3, respectively (P=0.009). The addition of high-dose IV MTX and/or cytarabine was associated with lower incidence of CNS relapse compared with IT chemotherapy alone. However, these data are limited by their retrospective nature and warrant confirmation in prospective randomised studies.

Background: Multiple myeloma prognostication is largely defined by the International Staging System (ISS)1 using serum beta-2 microglobulin (beta2M) and albumin results. Local external quality assurance data showed substantial, method-dependant variations in beta2M results. Aim(s): We determined the variability of different beta2M methods using patient samples, and whether this influences ISS prognostic scores. Method(s): beta2M stability after freeze-thawing was established first. Specimens (n=21) were then sent frozen to four laboratories using different methods/instruments for beta2M: Lab1, nephelometric (Beckman Coulter Immage); Lab2, chemiluminescent (Immulite 2000); Lab3, turbidimetric (Roche Cobas c502) and Lab 4 turbidimetric (Abbott Architect c16200). All laboratories use the same albumin method (bromocresol purple). Result(s): Lab1 produced beta2M results consistently higher than other laboratories (Figure 1): mean difference +1.55 mg/L (28.5%) (95%CI: 1.02 - 2.09, p<0.0001) compared to Lab2; +1.09 mg/L (20.1%) (95% CI: 0.78 - 1.41, p<0.0001) to Lab3; and +0.51 mg/L (9.3%) (95% CI: 0.29 - 0.73, p<0.0001) to Lab4. Compared to Lab1, ISS scores were classified one stage lower in 7, 5 and 3 patients by Lab2, Lab3 and Lab4, respectively. One patient could have been classified either stage I, II or III depending on which laboratory performed the tests. Albumin methods aligned well and had minimal impact on ISS score. Summary and Conclusion(s): Results from different beta2M methods are highly variable and harmonisation is required. This study has important implications on the validity of ISS staging for myeloma.

Central nervous system (CNS) relapse in diffuse large B-cell lymphoma (DLBCL) is a devastating complication; the optimal prophylactic strategy remains unclear. We performed a multicentre, retrospective analysis of patients with DLBCL with high risk for CNS relapse as defined by two or more of: multiple extranodal sites, elevated serum LDH and B symptoms or involvement of specific high-risk anatomical sites. We compared three different strategies of CNS-directed therapy: intrathecal (IT) methotrexate (MTX) with (R)-CHOP 'group 1'; R-CHOP with IT MTX and two cycles of high-dose intravenous (IV) MTX 'group 2'; dose-intensive systemic antimetabolite-containing chemotherapy (Hyper-CVAD or CODOXM/IVAC) with IT/IV MTX 'group 3'. Overall, 217 patients were identified (49, 125 and 43 in groups 1-3, respectively). With median follow-up of 3.4 (range 0.2-18.6) years, 23 CNS relapses occurred (12, 10 and 1 in groups 1-3 respectively). The 3-year actuarial rates (95% CI) of CNS relapse were 18.4% (9.5-33.1%), 6.9% (3.5-13.4%) and 2.3% (0.4-15.4%) in groups 1-3, respectively (P=0.009). The addition of high-dose IV MTX and/or cytarabine was associated with lower incidence of CNS relapse compared with IT chemotherapy alone. However, these data are limited by their retrospective nature and warrant confirmation in prospective randomised studies.

Introduction Plasmablastic lymphoma (PBL) is a rare, aggressive large cell lymphoma, first described in 1997. PBL is strongly associated with immunodeficient states, such as HIV infection and solid organ transplantation, but up to one third of cases are reported to occur in immunocompetent patients. The pathogenesis of PBL is incompletely understood, though the oncogenic impact of EBV, in particular in the context of dysregulated immune surveillance, together with acquired abnormalities in the MYC pathway appear to play key roles in many cases. Plasma cell markers such as CD138 and CD38 are typically positive, as well as CD30 in a significant subset. Classical B cell markers such as CD20, CD19 and PAX5 are typically absent. The literature on clinical outcomes in PBL is generally limited to small, single-centre case series. Reports describe an aggressive disease of poor prognosis, with median survival of 8 to 15 months, with one series reporting a longer median survival of 32 months. Methods We retrospectively identified patients diagnosed with PBL between 1999 and 2019 from 16 sites across Australia, the United Kingdom and Canada. Patients aged >=18 years with confirmed tissue diagnosis of PBL at their local treating centre were included. Factors associated with overall survival (OS) were analysed using Cox regression, stratified by site to account for heterogeneity across sites. Risk time for mortality began on the date of diagnosis and ended on the date of death. Patients who were alive, lost to follow-up or transferred to another centre for care, were censored on the date of last follow-up. Risk factors analysed included age, year of diagnosis, HIV status, MYC rearrangement status, CD30 status, lactate dehydrogenase level, disease stage by Lugano consensus criteria, and bone marrow involvement. Results We identified 197 patients with PBL (Table 1). The median age at diagnosis was 55 years (range 18-95) and there was a male predominance (69%). 37% of patients were

The transcription factor IRF4 is essential for the survival of both normal plasma cells and the plasma cell malignancy multiple myeloma (MM). However the basis for this dependency remains uncertain, with the prevailing view that IRF4 loss in MM induces "death by a thousand cuts." Here we have explored the genetic basis for IRF4 addiction through CRISPR-Cas9 mediated deletion of IRF4 in three lenalidomide sensitive MM cells lines (MM1S, OPM2 and H929) and one resistant cell line (RPMI-8226). Loss of IRF4 resulted in marked loss of viability, irrespective of lenalidomide sensitivity, which was driven by two distinct mechanisms: 1) induction of apoptosis, which was rescued through prior deletion of apoptotic mediators BAX and BAK, and 2) proliferative arrest, which persisted despite rescue from apoptosis. RNA-sequencing following IRF4 inactivation identified a core group of 86 genes whose dysregulation was shared across all 4 MM cell lines. We identified 31 common down-regulated genes (IRF4 activated), including several with previously described roles in survival (such as KLF2, TNFRSF17/BCMA and MYB). Furthermore, MYC, a previously identified target of IRF4 known to play an important role in MM pathogenesis and survival, was down-regulated in 3 of the 4 cell lines. Most surprisingly, IRF4 inactivation resulted in 55 common up-regulated genes that included two BH3-only pro-apoptotic proteins, BMF and BCL2L11 (BIM). Up-regulation of BMF appeared more marked than BIM with an average fold change of 6.5x (range 2.7 - 10.3) vs 1.9x (range: 1.8 - 2.2) respectively. Remarkably, genetic ablation of BMF in the OPM2 cell line and BMF/BIM in the MM1S largely protected from the cell death observed following IRF4 inactivation, suggesting that IRF4 maintains MM survival through the transcriptional repression of BMF and BIM. Our results confirm the vital role of IRF4 for MM proliferation and survival, and shed further light into its critical downstream transcriptional targets. Most in

Introduction Plasmablastic lymphoma (PBL) is a rare, aggressive large cell lymphoma, first described in 1997. PBL is strongly associated with immunodeficient states, such as HIV infection and solid organ transplantation, but up to one third of cases are reported to occur in immunocompetent patients. The pathogenesis of PBL is incompletely understood, though the oncogenic impact of EBV, in particular in the context of dysregulated immune surveillance, together with acquired abnormalities in the MYC pathway appear to play key roles in many cases. Plasma cell markers such as CD138 and CD38 are typically positive, as well as CD30 in a significant subset. Classical B cell markers such as CD20, CD19 and PAX5 are typically absent. The literature on clinical outcomes in PBL is generally limited to small, single-centre case series. Reports describe an aggressive disease of poor prognosis, with median survival of 8 to 15 months, with one series reporting a longer median survival of 32 months. Methods We retrospectively identified patients diagnosed with PBL between 1999 and 2019 from 16 sites across Australia, the United Kingdom and Canada. Patients aged >=18 years with confirmed tissue diagnosis of PBL at their local treating centre were included. Factors associated with overall survival (OS) were analysed using Cox regression, stratified by site to account for heterogeneity across sites. Risk time for mortality began on the date of diagnosis and ended on the date of death. Patients who were alive, lost to follow-up or transferred to another centre for care, were censored on the date of last follow-up. Risk factors analysed included age, year of diagnosis, HIV status, MYC rearrangement status, CD30 status, lactate dehydrogenase level, disease stage by Lugano consensus criteria, and bone marrow involvement. Results We identified 197 patients with PBL (Table 1). The median age at diagnosis was 55 years (range 18-95) and there was a male predominance (69%). 37% of patients were

The transcription factor IRF4 is essential for the survival of both normal plasma cells and the plasma cell malignancy multiple myeloma (MM). However the basis for this dependency remains uncertain, with the prevailing view that IRF4 loss in MM induces "death by a thousand cuts." Here we have explored the genetic basis for IRF4 addiction through CRISPR-Cas9 mediated deletion of IRF4 in three lenalidomide sensitive MM cells lines (MM1S, OPM2 and H929) and one resistant cell line (RPMI-8226). Loss of IRF4 resulted in marked loss of viability, irrespective of lenalidomide sensitivity, which was driven by two distinct mechanisms: 1) induction of apoptosis, which was rescued through prior deletion of apoptotic mediators BAX and BAK, and 2) proliferative arrest, which persisted despite rescue from apoptosis. RNA-sequencing following IRF4 inactivation identified a core group of 86 genes whose dysregulation was shared across all 4 MM cell lines. We identified 31 common down-regulated genes (IRF4 activated), including several with previously described roles in survival (such as KLF2, TNFRSF17/BCMA and MYB). Furthermore, MYC, a previously identified target of IRF4 known to play an important role in MM pathogenesis and survival, was down-regulated in 3 of the 4 cell lines. Most surprisingly, IRF4 inactivation resulted in 55 common up-regulated genes that included two BH3-only pro-apoptotic proteins, BMF and BCL2L11 (BIM). Up-regulation of BMF appeared more marked than BIM with an average fold change of 6.5x (range 2.7 - 10.3) vs 1.9x (range: 1.8 - 2.2) respectively. Remarkably, genetic ablation of BMF in the OPM2 cell line and BMF/BIM in the MM1S largely protected from the cell death observed following IRF4 inactivation, suggesting that IRF4 maintains MM survival through the transcriptional repression of BMF and BIM. Our results confirm the vital role of IRF4 for MM proliferation and survival, and shed further light into its critical downstream transcriptional targets. Most in