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Neuroblastomas harbor ALKaberrations clinically resistant to crizotinib yet sensitive pre-clinically to the third-generation ALK inhibitor lorlatinib. We conducted a first-in-child study evaluating lorlatinib with and without chemotherapy in children and adults with relapsed or refractory ALK-driven neuroblastoma. The trial is ongoing, and we report here on three cohorts that have met pre-specified primary endpoints: lorlatinib as a single agent in children (12 months to <18 years); lorlatinib as a single agent in adults (≥18 years); and lorlatinib in combination with topotecan/cyclophosphamide in children (<18 years). Primary endpoints were safety, pharmacokinetics and recommended phase 2 dose (RP2D). Secondary endpoints were response rate and 123I-metaiodobenzylguanidine (MIBG) response. Lorlatinib was evaluated at 45–115 mg/m2/dose in children and 100–150 mg in adults. Common adverse events (AEs) were hypertriglyceridemia (90%), hypercholesterolemia (79%) and weight gain (87%). Neurobehavioral AEs occurred mainly in adults and resolved with dose hold/reduction. The RP2D of lorlatinib with and without chemotherapy in children was 115 mg/m2. The single-agent adult RP2D was 150 mg. The single-agent response rate (complete/partial/minor) for <18 years was 30%; for ≥18 years, 67%; and for chemotherapy combination in <18 years, 63%; and 13 of 27 (48%) responders achieved MIBG complete responses, supporting lorlatinib’s rapid translation into active phase 3 trials for patients with newly diagnosed high-risk, ALK-driven neuroblastoma. ClinicalTrials.gov registration: NCT03107988.

Consumers served large meals are often confronted with the trade-off between wasting food and overconsumption. Although these outcomes are negatively correlated, researchers have rarely considered how individuals who subscribe to different political ideologies resolve this trade-off. Two experiments show that when consumers are served large portions, making the potential for food waste salient produces opposing reactions among liberals and conservatives. Liberals become concerned with avoiding the negative environmental consequences of food waste and eat more than normal (i.e., they overconsume) to prevent waste. In contrast, conservatives focus more on taking personal responsibility to avoid the negative consequences of overconsumption and eat less than normal, avoiding overconsumption but producing more food waste. Practical implications for marketers and researchers are discussed.

Many would contend that drinking to the point of intoxication is maladaptive. Further, almost all would agree that criminal behavior is maladaptive. Thus, one might assume that someone who commits a crime while intoxicated would be judged more harshly by others, given that such a perpetrator enacted two maladaptive behaviors. Yet, across two studies, including one analysis of archival criminal sentencing data, we demonstrate that this is often not the case. Critically, the type of crime committed influences whether intoxication mitigates versus exacerbates censure from others. For crimes predominately believed to be driven by emotion, intoxication discounts dispositional attributions and reduces perceived culpability. However, for other actions, intoxication has no effect, and can sometimes increase culpability. We conclude by discussing the important theoretical, practical, and policy implications of these findings, in addition to why a link between certain crimes and intoxication may enable abuse to the self and others.

People generally respond to resource scarcity through one of two pathways: scarcity-reduction or control-restoration. We draw from recent work on the solidity (versus liquidity) of consumption opportunities to offer a new lens through which to view how the two pathways that follow from resource scarcity relate to object attachment. In this review, we discuss when each pathway predicts stronger (versus weaker) object attachment. We also offer several open questions for when resource scarcity might prompt consumers to forgo the security afforded by stronger attachments for the flexibility afforded by weaker attachments.

Although many individuals endorse prosocial causes, little is understood about the factors that determine when people’s desire to consume in ways that benefit society as a whole will exceed their desire to accrue direct benefits to the self. We address this question by testing how reminders of resource scarcity affect consumers’ interest in sustainable products, and whether highlighting the prosocial or personal benefits associated with such products moderates this effect. We demonstrate the novel result that consumers exposed to reminders of resource scarcity are more likely to choose a sustainable product when the product’s prosocial (vs. personal) benefits are emphasized and the associated costs to the self are low. We provide evidence that this effect occurs because reminders of resource scarcity promote an abstract level of construal, which can carry over to affect decision making.

Extensive use of incentives in practice suggests that they play a key role in motivating behavior. However, conflicting findings have emerged about the effectiveness of various types of incentives (e.g., a cash incentive vs. a donation to charity). We propose a theoretical framework to explain when and why different types of incentives may be more effective. We posit that motivational efficacy of an incentive is jointly determined by its cognitive and affective value. Thus, an incentive offering a donation to charity can be more motivating than an equivalent cash incentive for the self when the incentive amount is low because, although both lack a motivating cognitive valuation, superior affective valuation of the prosocial incentive can increase its relative effectiveness. Consistent with this account, we demonstrate that low-amount self-benefiting incentives become more motivating when their affective value is enhanced, and low-amount prosocial incentives become less motivating as their affective value is attenuated.

ABSTRACTPediatric patients with high-risk neuroblastoma often relapse with chemotherapy-resistant, incurable disease. Relapsed neuroblastomas harbor chemo-resistant mesenchymal tumor cells and increased expression/activity of the transcriptional co-regulator, the Yes-Associated Protein (YAP). Patients with relapsed neuroblastoma are often treated with immunotherapy such as the anti-GD2 antibody, dinutuximab, in combination with chemotherapy. We have previously shown that YAP mediates both chemotherapy and MEK inhibitor resistance in relapsed RASmutated neuroblastoma and so posited that YAP might also be involved in anti-GD2 antibody resistance. We now show that YAP genetic inhibition significantly enhances sensitivity of mesenchymal neuroblastomas to dinutuximab and gamma delta (γδ) T cells both in vitroand in vivo. Mechanistically, YAP inhibition induces increased GD2 cell surface expression through upregulation of ST8SIA1, the gene encoding GD3 synthase and the rate-limiting enzyme in GD2 biosynthesis. The mechanism of ST8SIA1suppression by YAP is independent of PRRX1expression, a mesenchymal master transcription factor, suggesting YAP may be the downstream effector of mesenchymal GD2 resistance. These results therefore identify YAP as a therapeutic target to augment GD2 immunotherapy responses in patients with neuroblastoma.

Consumers often have multiple goals that are active simultaneously and make choices to satisfy those goals. However, no work to date has studied how people choose when all available options serve a goal (e.g., a choice-set goal) that conflicts with another goal they hold (e.g., an incidental goal). We demonstrate that in such contexts, consumers are more likely to choose the option that is most instrumental for attaining the choice-set goal, even when that option poses the greatest violation of the incidental goal. This occurs because the experience of goal conflict increases consumers’ need to justify their choices. Since the consumer will violate their incidental goal by choosing any of the available alternatives, we propose that the most justifiable reason for violating the incidental goal would be to maximize on the choice-set goal. Six experiments provide evidence for these effects and the underlying theoretical mechanism.

Academic research in several disciplines has demonstrated that consumers generally show a preference for certainty in the domain of gains. The current research provides evidence for an important psychological antecedent to this effect. Specifically, the authors find that the likelihood of choosing a certain reward over a risky or uncertain reward with a greater expected value is affected by whether consumers attend to the gist of the choice options or their associated details. Five experiments reveal that shifting attention to the gist of the choice options accentuates the preference for certainty and, conversely, shifting attention to the details of the choice options attenuates it. The authors provide convergent evidence for this using a variety of different means to manipulate consumers’ attention, and offer novel insights into when consumers are more or less likely to show a preference for certainty and an aversion to risk and uncertainty in common, retail settings.

GD2-targeting immunotherapies have improved survival in children with neuroblastoma, yet on-target, off-tumor toxicities can occur and a subset of patients cease to respond. The majority of neuroblastoma patients who receive immunotherapy have been previously treated with cytotoxic chemotherapy, making it paramount to identify neuroblastoma-specific antigens that remain stable throughout standard treatment. Cell surface glycoproteomics performed on human-derived neuroblastoma tumors in mice following chemotherapy treatment identified protein tyrosine kinase 7 (PTK7) to be abundantly expressed. Furthermore, PTK7 shows minimal expression on pediatric-specific normal tissues. We developed an anti-PTK7 chimeric antigen receptor (CAR) and find PTK7 CAR T cells specifically target and kill PTK7-expressing neuroblastoma in vitro. In vivo, human/murine binding PTK7 CAR T cells regress aggressive neuroblastoma metastatic mouse models and prolong survival with no toxicity. Together, these data demonstrate preclinical efficacy and tolerability for targeting PTK7 and support ongoing investigations to optimize PTK7-targeting CAR T cells for neuroblastoma.

Although campaigns designed to promote pro-environmental behaviours increasingly highlight self-interest, recent research suggests that such appeals may not always be effective. For example, individuals are more likely to check their tyre pressure when prompted with self-transcendent (that is, benefits to the environment) versus economic motives; and, self-transcendent appeals are more likely to promote recycling behaviours than self-interested appeals. The present experiments identify an important psychological factor that helps to explain when highlighting economic benefits will be more or less effective in encouraging pro-environmental behaviours. Specifically, we demonstrate that highlighting economic benefits (for example, the money a consumer can save) reduces consumer interest in sustainable products when individuals are in more abstract mindsets compared with when the evaluation is more immediate (that is, their mindset is more concrete). Further, we provide evidence that this shift in interest is driven by the lack of ‘fit’ between abstract thinking and economic motivations, in the context of pro-environmental behaviour.

ABSTRACTγδ T lymphocytes represent an emerging class of cellular immunotherapy with preclinical promise to treat cancer, notably neuroblastoma. The innate-like immune cell subset demonstrates inherent cytoxicity toward tumor cells independent of MHC recognition, enabling allogeneic administration of healthy donor-derived γδ T cell therapies. A current limitation is the substantial interindividual γδ T cell expansion variation among leukocyte collections. Overcoming this limitation will enable realization of the full potential of allogeneic γδ T-based cellular therapy. Here, we characterize γδ T cell expansions from healthy adult donors and observe that highly potent natural killer (NK) lymphocytes expand with γδ T cells under zoledronate and IL-2 stimulation. The presence of NK cells correlates with both the expansion potential of γδ T cells and the overall potency of the γδ T cell therapy. However, the potency of the cell therapy in combination with an antibody-based immunotherapeutic, dinutuximab, appears to be independent of γδ T/NK cell content both in vitroand in vivo, which minimizes the implication of interindividual expansion differences toward efficacy. Collectively, these studies highlight the utility of maintaining the NK cell population within expanded γδ T cell therapies and suggest a synergistic action of combined innate cell immunotherapy toward neuroblastoma.

The pediatric solid tumor neuroblastoma (NB) often depends on the anti-apoptotic protein, Mcl-1, for survival through Mcl-1 sequestration of pro-apoptotic Bim. High affinity Mcl-1 inhibitors currently do not exist such that novel methods to inhibit Mcl-1 clinically are in high demand. Receptor tyrosine kinases (RTK) regulate Mcl-1 in many cancers and play a role in NB survival, yet how they regulate Bcl-2 family interactions in NB is unknown. We found that NB cell lines derived to resist the Bcl-2/-xl/-w antagonist, ABT-737, acquire a dependence on Mcl-1 and show increased expression and activation of the RTK, EGFR. Mcl-1 dependent NB cell lines derived at diagnosis and from the same tumor following relapse also have increased EGFR expression compared to those dependent on Bcl-2. Inhibition of EGFR by shRNA or erlotinib in Mcl-1 dependent NBs disrupts Bim binding to Mcl-1 and enhances its affinity for Bcl-2, restoring sensitivity to ABT-737 as well as cytotoxics in vitro. Mechanistically treatment of NBs with small molecule inhibitors of EGFR (erlotinib, cetuximab) and ERK (U0126) increases Noxa expression and dephosphorylates Bim to promote Bim binding to Bcl-2. Thus, EGFR regulates Mcl-1 dependence in high-risk NB via ERK-mediated phosphorylation of Bim such that EGFR/ERK inhibition renders Mcl-1 dependent tumors now reliant on Bcl-2. Clinically, EGFR inhibitors are ineffective as single agent compounds in patients with recurrent NB, likely due to this transferred survival dependence to Bcl-2. Likewise, EGFR or ERK inhibitors warrant further testing in combination with Bcl-2 antagonists in vivoas a novel future combination to overcome therapy resistance in the clinic.

In this article, we review research on the discrepancy between consumers' high self-reported interest in sustainable products and these products’ poor performance in the marketplace. We offer theoretically derived reasons for why framing sustainable products as “win-wins” (i.e., offering benefits to the self and to the greater good) might present a solution to this intention-behavior gap. Although prior research has typically found negative effects of win–win framing on sustainable consumption, we propose framing sustainable consumption as a win–win provides greater environmental, behavioral, and psychological benefits than framing it only in terms of benefits to the self or to the greater good. We discuss how these outcomes may operate due to differences in moral hypocrisy, licensing, and goal representation, and offer avenues for future research to test these moderators in an effort to improve the efficacy of win–win framing.

Neuroblastoma (NB) is a common, highly lethal pediatric cancer, with treatment failures largely attributable to the emergence of chemoresistance. The pro-survival Bcl2 homology (BH) proteins critically regulate apoptosis, and may represent important therapeutic targets for restoring drug sensitivity in NB. We used a human NB tumor tissue microarray to survey the expression of pro-survival BH proteins Mcl1 and Bcl2, and correlated expression to clinical prognostic factors and survival. Primary NB tumors heterogeneously expressed Mcl1 or Bcl2, with high expression correlating to high-risk phenotype. Co-expression is infrequent (11%), but correlates to reduced survival. Using RNA interference, we investigated the functional relevance of Mcl1 and Bcl2 in high-risk NB cell lines (SK-N-AS, IMR-5, NLF). Mcl1 knockdown induced apoptosis in all NB cell lines, while Bcl2 knockdown inhibited only NLF, suggesting functional heterogeneity. Finally, we determined the relevance of Mcl1 in resistance to conventional chemotherapy (etoposide, doxorubicin) and small molecule Bcl2-family antagonists (ABT-737 and AT-101). Mcl1 silencing augmented sensitivity to chemotherapeutics 2- to 300-fold, while Bcl2 silencing did not, even in Bcl2-sensitive NLF cells. Resistance to ABT-737, which targets Bcl2/-w/-x, was overcome by Mcl1 knockdown. AT-101, which also neutralizes Mcl1, had single-agent cytotoxicity, further augmented by Mcl1 knockdown. In conclusion, Mcl1 appears a predominant pro-survival protein contributing to chemoresistance in NB, and Mcl1 inactivation may represent a novel therapeutic strategy. Optimization of compounds with higher Mcl1 affinity, or combination with additional Mcl1 antagonists, may enhance the clinical utility of this approach.

Autoimmune lymphoproliferative syndrome (ALPS) is a disorder of abnormal lymphocyte survival caused by defective Fas-mediated apoptosis, leading to lymphadenopathy, hepatosplenomegaly, and an increased number of double-negative T cells (DNTs). Treatment options for patients with ALPS are limited. Rapamycin has been shown to induce apoptosis in normal and malignant lymphocytes. Since ALPS is caused by defective lymphocyte apoptosis, we hypothesized that rapamycin would be effective in treating ALPS. We tested this hypothesis using rapamycin in murine models of ALPS. We followed treatment response with serial assessment of DNTs by flow cytometry in blood and lymphoid tissue, by serial monitoring of lymph node and spleen size with ultrasonography, and by enzyme-linked immunosorbent assay (ELISA) for anti–double-stranded DNA (dsDNA) antibodies. Three-dimensional ultrasound measurements in the mice correlated to actual tissue measurements at death (r= .9648). We found a dramatic and statistically significant decrease in DNTs, lymphadenopathy, splenomegaly, and autoantibodies after only 4 weeks when comparing rapamycin-treated mice with controls. Rapamycin induced apoptosis through the intrinsic mitochondrial pathway. We compared rapamycin to mycophenolate mofetil, a second-line agent used to treat ALPS, and found rapamycin's control of lymphoproliferation was superior. We conclude that rapamycin is an effective treatment for murine ALPS and should be explored as treatment for affected humans.

Autoimmune lymphoproliferative syndrome (ALPS) is a disorder of abnormal lymphocyte survival caused by defective Fas-mediated apoptosis, leading to lymphadenopathy, hepatosplenomegaly, and an increased number of double-negative T cells (DNTs). Treatment options for patients with ALPS are limited. Rapamycin has been shown to induce apoptosis in normal and malignant lymphocytes. Since ALPS is caused by defective lymphocyte apoptosis, we hypothesized that rapamycin would be effective in treating ALPS. We tested this hypothesis using rapamycin in murine models of ALPS. We followed treatment response with serial assessment of DNTs by flow cytometry in blood and lymphoid tissue, by serial monitoring of lymph node and spleen size with ultrasonography, and by enzyme-linked immunosorbent assay (ELISA) for anti–double-stranded DNA (dsDNA) antibodies. Three-dimensional ultrasound measurements in the mice correlated to actual tissue measurements at death (r = .9648). We found a dramatic and statistically significant decrease in DNTs, lymphadenopathy, splenomegaly, and autoantibodies after only 4 weeks when comparing rapamycin-treated mice with controls. Rapamycin induced apoptosis through the intrinsic mitochondrial pathway. We compared rapamycin to mycophenolate mofetil, a second-line agent used to treat ALPS, and found rapamycin's control of lymphoproliferation was superior. We conclude that rapamycin is an effective treatment for murine ALPS and should be explored as treatment for affected humans.

Strategies that enable E1-defective recombinant adenoviruses to selectively undergo replication in neoplastic tissue may be useful for future investigations or therapies of malignancies. A growing body of evidence suggests that some molecular alterations commonly associated with malignancies, such as p53 mutations, can modify the specific E1 requirements for replication of human serotype adenoviruses. In the studies reported here, a panel of human non-small cell lung cancer cell lines with previously defined p53 status were characterized for basal interleukin-6 (IL-6) andbcl-2content because previous studies have indicated both proteins can functionally substitute for the replication requirements provided by native E1 viral proteins. Cell lines were infected with E1-defective adenovirus 5 and simultaneously transfected with different combinations of E1 plasmids, or abcl-2expression plasmid, and adenovirus present in the cells was quantified 6 days later. These assays demonstrated that E1A with both 19- and 55-kDa E1B-encoding plasmids were required for maximal adenoviral replication, independent of the varying p53/IL-6/basalbcl-2phenotypes of the host cell lines. E1A was required for maximal replication enablement, independent of the basal IL-6 content of these cell lines, and exogenous IL-6 also did not obviate the E1A requirement. Interestingly, thebcl-2expression plasmid did not consistently substitute for the 19-kDa expression plasmid in the context of this replication complementation assay. These results suggest that (1) basal levels of IL-6 greater than that present in these cell lines are necessary for functional replacement of the E1A replication function and (2)bcl-2does not predictably substitute for the 19-kDa E1B replication function in the context oftranscomplementation.

n-Butyrate (butyrate) has been shown to amplify transgene expression in cells infected with E1-defective adenoviruses. The present studies were undertaken in order to better define the actions of butyrate in the context of adenovirus gene expression, and to attempt to elucidate the mechanism by which butyrate mediates the transgene amplification. It was found that butyrate amplified viral transgene expression over a concentration range of 0.5–5 mM,and that the amplification required an exposure of 12–24 hr for maximal effect. Western blot analysis of representative viral proteins showed that butyrate treatment amplified DNA-binding protein, but not fiber protein. A transient adenoviral replication system suggested that butyrate had a modest inhibitory effect on replication of the E1-defective adenovirus. Use of a specific inhibitor of histone deacetylase, trichostatin A (TSA), reproduced the amplification of the viral transgene product achieved with the butyrate. In contrast, adenoviral transgene expression could not be amplified by TSA treatment in a cell line known to have a TSA-resistant histone deacetylase. Butyrate amplified steady-state gene expression of the viral transgene, but had no detectable effects on either DNA-binding protein or fiber steady-state gene expression. Nuclear run-off experiments showed that both butyrate and TSA caused an increase in the viral transgene transcription. It was concluded that inhibitors of histone deacetylase amplify adenoviral transgene expression at the transcriptional level.

ABSTRACTGene therapy may serve as a valuable therapeutic modality for malignancies, such as lung cancer, that are poorly responsive to conventional therapies. Although many methods for transducing new genes into cells have been described, little is known about gene transduction into lung cancer, especially under conditions that might be encountered in clinical use. As a first step in addressing this important issue, the study presented here examined the ability of a recombinant retrovirus to add a selectable marker gene to the A549 non-small cell lung cancer (NSCLC) cell line under a variety of conditions. Examination of viral exposure times ranging from 30 sec to 4 hr revealed that the number of infected cells increased with every increment in time. By increasing the multiplicity of infection to 1.0 and including a polycation, Polybrene, as an infection facilitator, 0.8% of the NSCLC cells were infected with only a 30-sec viral exposure. Nebulization, a potentially attractive route of administration for pulmonary malignancies, had no significant effect on viral titer, proviral structure, or proviral transcripts. A single lyophilization did reduce viral titer by 58 ± 6%, but did not affect the proviral structure or transcripts produced by the surviving viruses. These results suggest that recombinant retroviruses have the potential to add new genes to malignancies accessible by the airways under conditions likely required for clinical use.

ABSTRACTHuman surfactant protein A (SPA) expression is considered a marker of respiratory epithelial differentiation. Non-small cell lung cancers (NSCLC) are respiratory epithelial derivatives, and it was previously shown that a minority of these cancers expressed SPA, presumably a consequence of their respiratory epithelial origin. In the studies reported here, SPA-I gene transcriptional regulatory sequences were localized to a 2.75-kb genomic 5-flanking region fragment obtained by screening a human genomic library. The 2.75-kb fragment was used to direct a luciferase coding sequence transcriptionally within a plasmid construct. In plasmid transduction experiments, the SPA-directed luciferase plasmid produced significant luciferase activity in the SPA-expressing NSCLC cell line, H441, but only background levels in the non-SPA-expressing A549 cells. Because Northern blot analysis of resected NSCLC showed that the majority expressed SPA, an SPA-transcriptional targeting strategy was investigated using chimeric toxin genes comprising the coding sequence for herpes simplex virus thymidine kinase (HSV-TK) under transcriptional control of SPA or SV40 regulatory sequences. As expected, transduction of the constitutive, SV40-directed plasmid followed by ganciclovir treatment reduced numbers of both the A549 and H441 cells. In contrast, the SPA-directed plasmid reduced only the SPA-expressing H441 cells and had no significant effect on the A549 cells. The results of these in vivoexperiments suggest the concept of transcriptionally directing toxin genes with SPA can produce targeted toxicity in NSCLC.

ABSTRACTReplication-incompetent adenovirus is conventionally produced by cells that supply replication-enabling proteins from viral sequences present in trans. As an alternative means of recombinant adenovirus production, replication-enabling E1A sequences were cotransduced into human prostate carcinoma cells infected with an E1A-deleted adenovirus containing a luciferase expression cassette. The replication-enabling plasmid was cotransduced by ionic linkage to the recombinant adenovirus exterior. Cells cotransduced with the replication-enabling plasmid made new adenovirus with titers up to 8 106in the supernatants 72120 hr after transduction. Like the parent virus, the virus present in the cotransduced supernatants and lysates was capable of transferring luciferase activity to new cells. The virus present in the cotransduced cell supernatants was amplified and shown to be identical to the parent virus by genomic analysis. It was concluded that simultaneous addition of a replication-defective adenovirus and a replication-enabling gene sequence in a transconfiguration converts some of the cotransduced cells into recombinant adenovirus-producing cells.

ABSTRACTAn effective therapy regimen for relapsed/refractory high-risk neuroblastoma (NB) includes the anti-GD2 monoclonal antibody, dinutuximab, in combination with temozolomide and irinotecan, supporting a role for chemo-immunotherapy in NB. γδ T cells are an attractive anti-tumor immunotherapy because of their direct cytotoxic activity mediated through cell surface receptors NKG2D and CD16. NKG2D facilitates the innate recognition of stress-induced ligands whereas CD16 recognizes antibody bound to tumors and activates mechanisms of antibody-dependent cellular cytotoxicity (ADCC). This study demonstrates an efficient method for expanding and storing γδ T cells from NB patient-derived apheresis products at clinically relevant amounts. The expanded patient-derived γδ T cells were cytotoxic against the K562 cell line and multiple NB cell lines. Combining γδ T cells with dinutuximab led to a 30% increase in tumor cell lysis compared to γδ T cells alone. Furthermore, low-dose temozolomide in combination with expanded γδ T cells and dinutuximab resulted in increased IFNγ secretion and increased γδ T-cell surface expression of FasL and CD107a. IMR5 NB cell line xenografts established subcutaneously in NSG mice were treated with a regimen of dinutuximab, temozolomide, and γδ T cells. This combination caused targeted killing of NB xenografts in vivo, reducing tumor burden and prolonging survival. These data support the continued preclinical testing of dinutuximab and temozolomide in conjunction with γδ T-cell immunotherapy for patients with recurrent/refractory NB.