Cubic splines to model relationships between continuous variables and outcomes: a guide for clinicians.

SERIES EDITORS' NOTE: We are pleased to add this typescript to the Bone Marrow Transplantation Statistics Series. We realize the term cubic splines may be a bit off-putting to some readers, but stay with us and don't get lost in polynomial equations. What the authors describe is important conceptually and in practice. Have you ever tried to buy a new pair of hiking boots? Getting the correct fit is critical; shoes that are too small or too large will get you in big trouble! Now imagine if hiking shoes came in only 2 sizes, small and large, and your foot size was somewhere in between. You are in trouble. Sailing perhaps?Transplant physicians are often interested in the association between two variables, say pre-transplant measurable residual disease (MRD) test state and an outcome, say cumulative incidence of relapse (CIR). We typically reduce the results of an MRD test to a binary, negative or positive, often defined by an arbitrary cut-point. However, MRD state is a continuous biological variable, and reducing it to a binary discards what may be important, useful data when we try to correlate it with CIR. Put otherwise, we may miss the trees from the forest.Another way to look at splines is a technique to make smooth curves out of irregular data points. Consider, for example, trying to describe the surface of an egg. You could do it with a series of straight lines connecting points on the egg surface but a much better representation would be combining groups of points into curves and then combining the curves. To prove this try drawing an egg using the draw feature in Microsoft Powerpoint; you are making splines.Gauthier and co-workers show us how to use cubic splines to get the maximum information from data points, which may, unkindly, not lend themselves to dichotomization or a best fit line. Please read on. We hope readers will find their typescript interesting and exciting, and that it will give them a new way to think about how to analyse data. And no, a spline is not a bunch of cactus spines. Robert Peter Gale, Imperial College London, and Mei-Jie Zhang, Medical College of Wisconsin and CIBMTR.

A randomized study of melphalan 200 mg/m(2) vs 280 mg/m(2) as a preparative regimen for patients with multiple myeloma undergoing auto-SCT.

We aimed to examine whether doses of melphalan higher than 200 mg/m(2) improve response rates when used as conditioning before autologous transplant (ASCT) in multiple myeloma (MM) patients. Patients with MM, n=131, were randomized to 200 mg/m(2) (mel200) vs 280 mg/m(2) (mel280) using amifostine pretreatment. The primary end point was the proportion of patients achieving near complete response (⩾nCR). No treatment-related deaths occurred in this study. Responses following ASCT were for mel200 vs mel280, respectively, ⩾nCR 22 vs 39%, P=0.03, ⩾PR 57 vs 74%, P=0.04. The hazard of mortality was not statistically significantly different between groups (mel200 vs mel280; hazard ratio (HR)=1.15 (95% confidence interval (CI), 0.62-2.13, P=0.66)) nor was the rate of progression/mortality (HR=0.81 (0.52-1.27, P=0.36)). The estimated PFS at 1 and 3 years were 83 and 46%, respectively, for mel200 and 78 and 54%, respectively, for mel280. Amifostine and mel280 were well tolerated, with no grade 4 regimen-related toxicities and only one grade 3 mucositis (none with mel200) and three grade 3 gastrointestinal (GI) toxicities (two in mel200). Hospitalization rates were more frequent in the mel280 group (59 vs 43%, P=0.08). Mel280 resulted in a higher major response rate (CR+nCR) and should be evaluated in larger studies.

Effects on tumor development and metastatic dissemination by the NKG2D lymphocyte receptor expressed on cancer cells.

The stimulatory NKG2D lymphocyte receptor together with its tumor-associated ligands enable the immune system to recognize and destroy cancer cells. However, with dynamic changes unfolding, cancers exploit NKG2D and its ligands for immune evasion and suppression. Recent findings have added yet another functional dimension, wherein cancer cells themselves co-opt NKG2D for their own benefit to complement the presence of its ligands for self-stimulation of parameters of tumorigenesis. Those findings are here extended to in vivo tumorigenicity testing by employing orthotopic xenotransplant breast cancer models in mice. Using human cancer lines with ectopic NKG2D expression and RNA interference (RNAi)-mediated protein depletion among other controls, we show that NKG2D self-stimulation has tumor-promoting capacity. NKG2D signals had no notable effects on cancer cell proliferation and survival but acted at the level of angiogenesis, thus promoting tumor growth, tumor cell intravasation and dissemination. NKG2D-mediated effects on tumor initiation may represent another factor in the observed overall enhancement of tumor development. Altogether, these results may have an impact on immunotherapy approaches, which currently do not account for such NKG2D effects in cancer patients and thus could be misdirected as underlying assumptions are incomplete.

Characteristics of chronic GVHD after cord blood transplantation.

Most reports of chronic GVHD after cord blood transplantation (CBT) have utilized traditional diagnostic criteria. We used traditional criteria and National Institutes of Health (NIH) criteria prospectively to evaluate chronic GVHD in a cohort of 87 adult and pediatric recipients of single or double unrelated CBT for treatment of hematologic malignancies. Fifty-four patients developed traditionally defined chronic GVHD, for an estimated 2-year probability of 64%. Among 54 patients, 25 (46%) met the NIH criteria for persistent, recurrent or late acute GVHD at onset. Twenty-four (44%) had overlap chronic GVHD, including one who presented initially with late acute GVHD, and only seven (13%) had classic chronic GVHD, including one who also presented initially with late acute GVHD. Among patients who successfully discontinued all systemic immunosuppression (SI), the median time to discontinuation of corticosteroid treatment was 315 days (range 28-977), and the median time to discontinuation of all SI was 353 days (range 67-977). Chronic GVHD diagnosed by traditional criteria after CBT had a predominance of acute GVHD clinical features.

Predicting relapse prior to transplantation in chronic myeloid leukemia by integrating expert knowledge and expression data.

MOTIVATION: Selecting a small number of signature genes for accurate classification of samples is essential for the development of diagnostic tests. However, many genes are highly correlated in gene expression data, and hence, many possible sets of genes are potential classifiers. Because treatment outcomes are poor in advanced chronic myeloid leukemia (CML), we hypothesized that expression of classifiers of advanced phase CML when detected in early CML [chronic phase (CP) CML], correlates with subsequent poorer therapeutic outcome. RESULTS: We developed a method that integrates gene expression data with expert knowledge and predicted functional relationships using iterative Bayesian model averaging. Applying our integrated method to CML, we identified small sets of signature genes that are highly predictive of disease phases and that are more robust and stable than using expression data alone. The accuracy of our algorithm was evaluated using cross-validation on the gene expression data. We then tested the hypothesis that gene sets associated with advanced phase CML would predict relapse after allogeneic transplantation in 176 independent CP CML cases. Our gene signatures of advanced phase CML are predictive of relapse even after adjustment for known risk factors associated with transplant outcomes.

Autologous hematopoietic cell transplantation following high-dose immunosuppressive therapy for advanced multiple sclerosis: long-term results.

The purpose of the study was to determine the long-term safety and effectiveness of high-dose immunosuppressive therapy (HDIT) followed by autologous hematopoietic cell transplantation (AHCT) in advanced multiple sclerosis (MS). TBI, CY and antithymocyte globulin were followed by transplantation of autologous, CD34-selected PBSCs. Neurological examinations, brain magnetic resonance imaging and cerebrospinal fluid (CSF) for oligoclonal bands (OCB) were serially evaluated. Patients (n=26, mean Expanded Disability Status Scale (EDSS)=7.0, 17 secondary progressive, 8 primary progressive, 1 relapsing/remitting) were followed for a median of 48 months after HDIT followed by AHCT. The 72-month probability of worsening ≥1.0 EDSS point was 0.52 (95% confidence interval, 0.30-0.75). Five patients had an EDSS at baseline of ≤6.0; four of them had not failed treatment at last study visit. OCB in CSF persisted with minor changes in the banding pattern. Four new or enhancing lesions were seen on MRI, all within 13 months of treatment. In this population with high baseline EDSS, a significant proportion of patients with advanced MS remained stable for as long as 7 years after transplant. Non-inflammatory events may have contributed to neurological worsening after treatment. HDIT/AHCT may be more effective in patients with less advanced relapsing/remitting MS.

Extracorporeal photopheresis in addition to pentostatin in conditioning for canine hematopoietic cell transplantation: role in engraftment.

Extracorporeal photopheresis (ECP) and the purine analog pentostatin exert potent immunomodulatory effects, but have not been evaluated for their ability to enhance engraftment of hematopoietic stem cells. We evaluated, in a canine model of dog leukocyte antigen (DLA)-identical hematopoietic cell transplantation (HCT), whether ECP in combination with pentostatin could enhance engraftment using a nonmyeloablative regimen consisting of 100 cGy TBI and postgrafting immunosuppression with mycophenolate mofetil and CYA. We have shown previously that with 100 cGy TBI alone as conditioning, all of the six dogs rejected their grafts 2-12 weeks after HCT. With the addition of pentostatin to 100 cGy TBI, 6 of 10 dogs rejected their graft. We now tested the additional use of ECP alone (n=2) or ECP and 3-6 doses of pentostatin (n=7) before 100 cGy TBI and HCT. Eight out of nine dogs rejected their grafts within 6-11 weeks after HCT. Compared with data without ECP, we failed to demonstrate a positive impact of the use of either ECP or pentostatin for prevention of rejection.

Comparison of matched unrelated and matched related donor myeloablative hematopoietic cell transplantation for adults with acute myeloid leukemia in first remission.

Hematopoietic cell transplantation (HCT) from a matched related donor (MRD) benefits many adults with acute myeloid leukemia (AML) in first complete remission (CR1). The majority of patients does not have such a donor and will require an alternative donor if HCT is to be undertaken. We retrospectively analyzed 226 adult AML CR1 patients undergoing myeloablative unrelated donor (URD) (10/10 match, n=62; 9/10, n=29) or MRD (n=135) HCT from 1996 to 2007. The 5-year estimates of overall survival, relapse and nonrelapse mortality (NRM) were 57.9, 29.7 and 16.0%, respectively. Failure for each of these outcomes was slightly higher for 10/10 URD than MRD HCT, although statistical significance was not reached for any end point. The adjusted hazard ratios (HRs) were 1.43 (0.89-2.30, P=0.14) for overall mortality, 1.17 (0.66-2.08, P=0.60) for relapse and 1.79 (0.86-3.74, P=0.12) for NRM, respectively, and the adjusted odds ratio for grades 2-4 acute graft-versus-host disease was 1.50 (0.70-3.24, P=0.30). Overall mortality among 9/10 and 10/10 URD recipients was similar (adjusted HR 1.16 (0.52-2.61), P=0.71). These data indicate that URD HCT can provide long-term survival for CR1 AML; outcomes for 10/10 URD HCT, and possibly 9/10 URD HCT, suggest that this modality should be considered in the absence of a suitable MRD.

Influence of oral beclomethasone dipropionate on early non-infectious pulmonary outcomes after allogeneic hematopoietic cell transplantation: results from two randomized trials.

Early non-infectious pulmonary complications represent a significant cause of mortality after hematopoietic cell transplantation (HCT). We tested the hypothesis that oral beclomethasone dipropionate (BDP) is effective for preventing early non-infectious pulmonary complications after allogeneic HCT. We retrospectively reviewed the medical records of 120 patients, 60 in each treatment arm, to identify non-infectious and infectious pulmonary events and pulmonary function test results from all patients who participated in two randomized trials of oral BDP for treatment of acute gastrointestinal GVHD. 17-Beclomethasone monopropionate (17-BMP), the active metabolite of BDP, was evaluated in blood from the right atrium in four patients. Thirty-three of 42 (79%) placebo-treated patients experienced a decrease of the DL(CO) from pretransplant to day 80 after transplant, compared with 27 of 49 (55%) BDP-treated patients (P=0.02). In the first 200 days after randomization, there were no cases of non-infectious pulmonary complications in BDP-treated patients, vs four cases among placebo-treated patients (P=0.04). Levels of 17-BMP were detected in atrial blood at steady state. Delivery of a potent glucocorticoid such as 17-BMP to the pulmonary artery after oral dosing of BDP may be useful in modulating pulmonary inflammation and preventing the development of non-infectious pulmonary complications after allogeneic HCT.

The impact of histologic grade on the outcome of high-dose therapy and autologous stem cell transplantation for follicular lymphoma.

The impact of the follicular lymphoma (FL) histologic grade on outcomes after high-dose therapy (HDT) and autologous stem cell transplantation (ASCT) is unknown. We evaluated 219 consecutive patients with grades 1-3 FL who underwent HDT and ASCT at our center. Overall survival (OS), progression-free survival (PFS), relapse and non-relapse mortality (NRM) was estimated for each grade after controlling for other predictive factors. The number of patients with grades 1, 2 and 3 FL was 106 (48%), 75 (34%) and 38 (17%), respectively. Five-year outcome estimates for the entire cohort included 60% OS, 39% PFS and 46% relapse (median follow-up=7.8 years). PFS and relapse were nearly identical among patients with grade 3 FL versus grades 1-2 FL after adjusting for other contributing factors (hazard ratio (HR)=0.90, P=0.68; HR=1.07, P=0.80, respectively). The hazard for mortality (HR=0.70, P=0.23) and NRM (HR=0.33, P=0.07) was non-significantly lower among patients with grade 3 FL compared to patients with grades 1-2 disease. Factors associated with inferior PFS included elevated lactate dehydrogenase (HR=1.52, P=0.03), chemoresistance (HR=1.82, P=0.02), > or =2 prior therapies (HR=1.8, P=0.03) and prior radiation (HR=1.99, P=0.003). These data suggest that the histologic grade of FL does not impact PFS or relapse following HDT and ASCT.

Imatinib therapy prior to myeloablative allogeneic stem cell transplantation.

It is unknown whether imatinib prior to myeloablative haematopoietic stem cell transplantation (HSCT) increases transplant-related toxicity. Among the side effects induced by imatinib, myelosuppression and liver injury might worsen HSCT outcomes. We retrospectively analysed engraftment, liver toxicity, acute graft-versus-host disease (aGVHD) incidence and 100-day mortality in 30 patients with BCR/ABL-positive leukaemias who received imatinib before HSCT and compared results of 48 age-matched controls who did not receive preceding imatinib. Both neutrophil and platelet engraftment occurred more rapidly among imatinib patients but the differences adjusted for Gratwohl scale were not statistically significant (P = 0.18 and 0.22, respectively). The adjusted hazards of having liver function tests (LFTs) >1.5 normal increased and the adjusted durations of elevated LFTs were not significantly different. The estimated adjusted difference in mean peak bilirubin values was also not significantly different (P = 0.48). However, the adjusted hazard of increased creatinine >1.5 normal was significantly higher in the imatinib group (HR = 4.09, P = 0.02). The adjusted odds of grades II-IV aGVHD were similar in both groups (OR = 0.86, P = 0.78), and while the adjusted odds of 100-day mortality were lower among imatinib patients, the difference was not significant (OR = 0.65, P = 0.60). These data do not provide any evidence that imatinib preceding HSCT increases acute transplant-related toxicities.

Outcome of allogeneic bone marrow transplantation for children with advanced acute myeloid leukemia.

Allogeneic bone marrow transplantation (BMT) may offer the only chance of cure for children with acute myeloid leukemia (AML) in second complete remission (CR2) or with relapsed disease, but the outcome of these patients has not been clearly defined. We conducted a retrospective study of 58 children, median age 7.4 years (range 0.8-17.3), who received matched related or unrelated BMT at our institution for AML in CR2 (n = 12), in untreated first relapse (n = 11) or with refractory disease (n = 35), to identify risk factors associated with disease-free survival (DFS). Life threatening to fatal regimen-related toxicity was observed in 22% of patients. Estimates of DFS at 5 years (95% confidence interval) for patients in CR2, with untreated first relapse and refractory disease were 58% (27-80%), 36% (11-63%) and 9% (2-21%), respectively. Non-relapse mortality estimates were 0%, 27% (0-54%) and 17% (5-30%), and relapse estimates were 42% (14-70%), 36% (8-65%) and 74% (60-89%), respectively. Advanced disease phase and cytogenetic abnormalities at the time of transplantation were each associated with decreased DFS and increased relapse in multivariable regression models. Survival for children transplanted in CR2 or untreated first relapse is higher than that previously reported, but relapse remains the major cause of treatment failure regardless of disease stage.

Allogeneic bone marrow transplantation in children with myelodysplastic syndrome or juvenile myelomonocytic leukemia: the Seattle experience.

The purpose of this study was to evaluate the role of allogeneic bone marrow transplantation (BMT) in children with myelodysplastic syndrome (MDS). In total, 94 consecutive pediatric patients with MDS received an allogeneic BMT from 1976 to 2001 for refractory anemia (RA) (n=25), RA with ringed sideroblasts (RARS) (n=2), RA with excess blasts (RAEB) (n=20), RAEB in transformation (RAEB-T) (n=14), juvenile myelomonocytic leukemia (JMML) (n=32) or chronic myelomonocytic leukemia (CMML) (n=1). The estimated 3-year probabilities of survival, event-free survival (EFS), nonrelapse mortality and relapse were 50, 41, 28 and 29%, respectively. Patients with RA/RARS had an estimated 3-year survival of 74% compared to 68% in those with RAEB and 33% in patients with JMML/CMML. In multivariable analysis, patients with RAEB-T or JMML were 3.9 and 3.7 times more likely to die compared to those with RA/RARS and RAEB (P=0.005 and 0.004, respectively). Patients with RAEB-T were 5.5 times more likely to relapse (P=0.01). The median follow-up among the 43 surviving patients is 10 years (range 1-25). We conclude that allogeneic BMT for children with MDS is well tolerated and can be curative.

Human herpesvirus 6 reactivation and encephalitis in allogeneic bone marrow transplant recipients.

To determine whether receipt of an investigational anti-CD3 monoclonal antibody (BC3) increased the risk of human herpesvirus 6 (HHV-6) reactivation and development of encephalitis in bone marrow transplant (BMT) recipients, persons who had and had not received BC3 were compared. Odds of HHV-6 reactivation were higher among BC3 recipients than among control patients (odds ratio, 2.5; 95% confidence interval [CI], 1.3-4.7). In addition, BC3 recipients were more likely than control patients to develop encephalitis (risk ratio [RR], 3.5; 95% CI, 1.3-9.5), and this association followed a BC3 dose-dependent relationship (P=.03, by Mantel-Haenszel chi(2) test). In a multivariable model, HHV-6 reactivation and receipt of BC3 were associated with increased risk of encephalitis (RR, 5.4; 95% CI, 1.9-15.3, and RR, 3.3; 95% CI, 1.2-9.1, respectively). In conclusion, both HHV-6 reactivation and receipt of BC3 for prophylaxis of acute graft-versus-host disease independently increased the risk of encephalitis in allogeneic BMT recipients. Prospective studies to better define the relationship between HHV-6 reactivation and encephalitis in allogeneic BMT recipients are warranted.

Multidrug-resistance phenotype and clinical responses to gemtuzumab ozogamicin.

Expression of multidrug resistance (MDR) features by acute myeloid leukemia (AML) cells predicts a poor response to many treatments. The MDR phenotype often correlates with expression of P-glycoprotein (Pgp), and Pgp antagonists such as cyclosporine (CSA) have been used as chemosensitizing agents in AML. Gemtuzumab ozogamicin, an immunoconjugate of an anti-CD33 antibody linked to calicheamicin, is effective monotherapy for CD33(+) relapsed AML. However, the contribution of Pgp to gemtuzumab ozogamicin resistance is poorly defined. In this study, blast cell samples from relapsed AML patients eligible for gemtuzumab ozogamicin clinical trials were assayed for Pgp surface expression and Pgp function using a dye efflux assay. In most cases, surface expression of Pgp correlated with Pgp function, as indicated by elevated dye efflux that was inhibited by CSA. Among samples from patients who either failed to clear marrow blasts or failed to achieve remission, 72% or 52%, respectively, exhibited CSA-sensitive dye efflux compared with 29% (P =.003) or 24% (P <.001) among samples from responders. In vitro gemtuzumab ozogamicin--induced apoptosis was also evaluated using an annexin V--based assay. Low levels of drug-induced apoptosis were associated with CSA-sensitive dye efflux, whereas higher levels correlated strongly with achievement of remission and marrow blast clearance. In vitro drug-induced apoptosis could be increased by CSA in 14 (29%) of 49 samples exhibiting low apoptosis in the absence of CSA. Together, these findings indicate that Pgp plays a role in clinical resistance to gemtuzumab ozogamicin and suggest that treatment trials combining gemtuzumab ozogamicin with MDR reversal agents are warranted. (Blood. 2001;98:988-994)

Hematopoietic cell transplantation in older patients with hematologic malignancies: replacing high-dose cytotoxic therapy with graft-versus-tumor effects.

Toxicities have limited the use of allogeneic hematopoietic cell transplantation (HCT) to younger, medically fit patients. In a canine HCT model, a combination of postgrafting mycophenolate mofetil (MMF) and cyclosporine (CSP) allowed stable allogeneic engraftment after minimally toxic conditioning with low-dose (200 cGy) total-body irradiation (TBI). These findings, together with the known antitumor effects of donor leukocyte infusions (DLIs), led to the design of this trial. Forty-five patients (median age 56 years) with hematologic malignancies, HLA-identical sibling donors, and relative contraindications to conventional HCT were treated. Immunosuppression involved TBI of 200 cGy before and CSP/MMF after HCT. DLIs were given after HCT for persistent malignancy, mixed chimerism, or both. Regimen toxicities and myelosuppression were mild, allowing 53% of eligible patients to have entirely outpatient transplantations. Nonfatal graft rejection occurred in 20% of patients. Grades II to III acute graft-versus-host disease (GVHD) occurred in 47% of patients with sustained engraftment. With median follow-up of 417 days, survival was 66.7%, nonrelapse mortality 6.7%, and relapse mortality 26.7%. Fifty-three percent of patients with sustained engraftment were in complete remission, including 8 with molecular remissions. This novel allografting approach, based on the use of postgrafting immunosuppression to control graft rejection and GVHD, has dramatically reduced the acute toxicities of allografting. HCT with the induction of potent graft-versus-tumor effects can be performed in previously ineligible patients, largely in an outpatient setting. Future protocol modifications should reduce rejection and GVHD, thereby facilitating studies of allogeneic immunotherapy for a variety of malignancies. (Blood. 2001;97:3390-3400)