Variability in Cytogenetic Testing for Multiple Myeloma: A Comprehensive Analysis From Across the United States.

PURPOSE: Multiple myeloma (MM) treatment has changed tremendously, with significant improvement in patient out-comes. One group with a suboptimal benefit is patients with high-risk cytogenetics, as tested by conventional karyotyping or fluorescence in situ hybridization (FISH). Methodology for these tests has been published, but not necessarily standardized. METHODS: We address variability in the testing and reporting methodology for MM cytogenetics in the United States using the ongoing African American Multiple Myeloma Study (AAMMS). We evaluated clinical and cytogenetic data from 1,221 patients (1,161 with conventional karyotyping and 976 with FISH) tested between 1998 and 2016 across 58 laboratories nationwide. RESULTS: Interlab and intralab variability was noted for the number of cells analyzed for karyotyping, with a significantly higher number of cells analyzed in patients in whom cytogenetics were normal (P 5.0025). For FISH testing, CD138-positive cell enrichment was used in 29.7% of patients and no enrichment in 50% of patients, whereas the remainder had unknown status. A significantly smaller number of cells was analyzed for patients in which CD138 cell enrichment was used compared with those without such enrichment (median, 50 v 200; P, .0001). A median of 7 loci probes (range, 1-16) were used for FISH testing across all laboratories, with variability in the loci probed even within a given laboratory. Chromosome 13-related abnormalities were the most frequently tested abnormality (n5956; 97.9%), and t(14;16) was the least frequently tested abnormality (n 5 119; 12.2%). CONCLUSIONS: We report significant variability in cytogenetic testing across the United States for MM, potentially leading to variability in risk stratification, with possible clinical implications and personalized treatment approaches.

Serum cholesterol trajectories in the 10 years prior to lymphoma diagnosis.

PURPOSE: Many studies suggest a role for cholesterol in cancer development. Serum cholesterol levels have been observed to be low in newly diagnosed lymphoma cases. The objective of these analyses was to examine the time-varying relationship of cholesterol with lymphomagenesis in the 10 years prior to diagnosis by lymphoma subtype. METHODS: Participants were selected from the combined membership of six National Cancer Institute-funded Cancer Research Network health plans from 1998 to 2008, excluding members with human immunodeficiency virus, cancer (except lymphoma), or organ transplants. Incident lymphoma cases within this population were ascertained and matched with up to five controls. Total serum cholesterol, high-density lipoprotein, and low-density lipoprotein were collected from plan databases. Multilevel, multivariable longitudinal models were fit after choosing the best polynomial order by deviance statistics for selected lymphoma histotypes to examine pre-diagnosis cholesterol trajectories: Hodgkin lymphoma (n = 519) and all non-Hodgkin lymphomas combined (n = 12,635) as well as six subtypes of the latter. RESULTS: For all categories, lymphoma cases had statistically significantly lower estimated total serum cholesterol, high-density lipoprotein, and low-density lipoprotein levels than controls in the years prior to diagnosis/index date. Between-group differences were most pronounced 3-4 years prior to diagnosis, when cases' cholesterol levels declined steeply. CONCLUSIONS: This analysis is the first to examine changes in serum cholesterol for a decade prior to lymphoma diagnosis. A drop in cholesterol levels was evident several years before diagnosis. Our results suggest that cholesterol-related pathways have an important relationship with lymphomagenesis and low cholesterol could be a preclinical lymphoma marker.

Chimerism in Myeloid Malignancies following Stem Cell Transplantation Using FluBu4 with and without Busulfan Pharmacokinetics versus BuCy.

In the era of precision medicine, the impact of personalized dosing of busulfan is not clear. We undertook a retrospective analysis of 78 patients with myeloid malignancies who received fludarabine and busulfan (FluBu4) with or without measuring Bu pharmacokinetics (Bu PK) and those who received busulfan with cyclophosphamide (BuCy). Fifty-five patients received FluBu4, of whom 21 had Bu PK measured, and 23 patients received BuCy. Total donor cell chimerism showed that the percentage of patients maintaining 100% donor chimerism on day 100 was 66.7%, 38.2%, and 73.9% in the FluBu4 with PK, FluBu4 with no PK, and BuCy, respectively (P = .001). Patients who had decreasing donor chimerism by day 100 were 23.8%, 52.9%, and 26.1% in the FluBu4 with PK, FluBu4 with no PK, and BuCy, respectively (P = .04). Bu PK group had fewer patients with less than 95% donor chimerism on day 30, which was not statistically significant, 5% (FluBu4 PK), 31% (FluBu4 with no PK), and 21% (BuCy) (P = .18). Survival distributions were not statistically significant (P = .11). Thus, personalized drug dosing can impact donor chimerism in myeloid malignancies. This will need to be examined in larger retrospective multicenter studies and prospective clinical trials.

Choice of Unmanipulated T Cell Replete Graft for Haploidentical Stem Cell Transplant and Posttransplant Cyclophosphamide in Hematologic Malignancies in Adults: Peripheral Blood or Bone Marrow-Review of Published Literature.

Allogeneic hematopoietic stem cell transplantation (SCT) is often the only curative option for many patients with malignant and benign hematological stem cell disorders. However, some issues are still of concern regarding finding a donor like shrinking family sizes in many societies, underrepresentation of the ethnic minorities in the registries, genetic variability for some races, and significant delays in obtaining stem cells after starting the search. So there is a considerable need to develop alternate donor stem cell sources. The rapid and near universal availability of the haploidentical donor is an advantage of the haploidentical SCT and an opportunity that is being explored currently in many centers especially using T cell replete graft and posttransplant cyclophosphamide. This is probably because it does not require expertise in graft manipulation and because of the lower costs. However, there are still lots of unanswered questions, like the effect of use of bone marrow versus peripheral blood as the source of stem cells on graft-versus-host disease, graft versus tumor, overall survival, immune reconstitution, and quality of life. Here we review the available publications on bone marrow and peripheral blood experience in the haploidentical SCT setting.

Early failure of frontline rituximab-containing chemo-immunotherapy in diffuse large B cell lymphoma does not predict futility of autologous hematopoietic cell transplantation.

The poor prognosis for patients with diffuse large B cell lymphoma (DLBCL) who relapse within 1 year of initial diagnosis after first-line rituximab-based chemo-immunotherapy has created controversy about the role of autologous transplantation (HCT) in this setting. We compared autologous HCT outcomes for chemosensitive DLBCL patients between 2000 and 2011 in 2 cohorts based on time to relapse from diagnosis. The early rituximab failure (ERF) cohort consisted of patients with primary refractory disease or those with first relapse within 1 year of initial diagnosis. The ERF cohort was compared with those relapsing >1 year after initial diagnosis (late rituximab failure [LRF] cohort). ERF and LRF cohorts included 300 and 216 patients, respectively. Nonrelapse mortality (NRM), progression/relapse, progression-free survival (PFS), and overall survival (OS) of ERF versus LRF cohorts at 3 years were 9% (95% confidence interval [CI], 6% to 13%) versus 9% (95% CI, 5% to 13%), 47% (95% CI, 41% to 52%) versus 39% (95% CI, 33% to 46%), 44% (95% CI, 38% to 50%) versus 52% (95% CI, 45% to 59%), and 50% (95% CI, 44% to 56%) versus 67% (95% CI, 60% to 74%), respectively. On multivariate analysis, ERF was not associated with higher NRM (relative risk [RR], 1.31; P = .34). The ERF cohort had a higher risk of treatment failure (progression/relapse or death) (RR, 2.08; P < .001) and overall mortality (RR, 3.75; P <.001) within the first 9 months after autologous HCT. Beyond this period, PFS and OS were not significantly different between the ERF and LRF cohorts. Autologous HCT provides durable disease control to a sizeable subset of DLBCL despite ERF (3-year PFS, 44%) and remains the standard-of-care in chemosensitive DLBCL regardless of the timing of disease relapse.

Metastatic squamous cell carcinoma in a bone marrow transplant recipient.

We report a case of 53year old female with history of acute myelogenous leukemia, for which she underwent allogeneic peripheral blood stem cell transplant; her course was complicated by chronic graft versus host disease. Seven years later she presents with squamous cell carcinoma of the skin which metastasize to her heart. Here, in this case we tend to highlight the aggressive nature of the squamous cell carcinoma in an immunocompromised individual.

Iron overload in allogeneic hematopoietic stem cell transplant recipients.

CONTEXT: Patients who undergo hematopoietic stem cell transplant are at an increased risk of developing iron overload. OBJECTIVES: To describe the effect of hepatic iron overload on hematopoietic stem cell transplant recipients and to validate the utility of histologic scoring system of iron granules in the liver. DESIGN: Records of 154 post allogeneic hematopoietic stem cell transplant patients were reviewed. Forty-nine patients underwent liver biopsy. Histologic hepatic iron overload was defined as a score of 2 or greater (scale, 0-4). RESULTS: Twenty-eight of 49 patients (57%) evaluated by liver biopsy had hepatic iron overload; 17 had moderate to severe hepatic iron overload (score, 3 or 4). In multivariate analysis, a significant correlation was discovered between hepatic iron overload and the number of transfusions (P < .001), posttransplant serum ferritin levels (P=.004), lactate dehydrogenase levels (P=.03), and the development of blood stream infections (P= .02). There was no correlation between hepatic iron overload and abnormal liver function test results. While 37 patients (76%) died after receiving a transplant, mortality was not influenced by hepatic iron overload but was significantly higher in older patients, in patients with lower serum albumin levels, higher serum bilirubin levels, and higher clinical grade of acute graft-versus-host disease (P=.04, P=.001, P=<.001, and P .004, respectively). CONCLUSIONS: Hepatic iron overload is commonly identified in hematopoietic stem cell transplant patients and can be accurately diagnosed by liver biopsy. In addition, hepatic iron overload has been identified in patients receiving as few as 25 units of packed red blood cells, with elevated posttransplant serum ferritin levels, and with blood stream infections.

Frequent expression of C4d in hepatic graft-versus-host disease: potential clue for diagnosis and distinguishing acute and chronic form.

BACKGROUND: Graft-versus-host disease (GVHD), a common complication of hematopoietic stem cell transplant, is generally regarded to develop through cell-mediated immune response following activation of helper T cells. Since production of antibodies is also mediated by helper T cells, the role of humoral immunity in GVHD is questioned and has not yet been explored in clinical practice. We conducted a pilot study to evaluate the role of antibody production in hepatic H-GVHD and whether it can distinguish acute and chronic forms. RESULTS: C4d expression was increased in portal vessels and hepatic sinusoids of patients with histological proven evidence of GVHD 11/16 (P=0.007). Patients classified as chronic GVHD were statistically more likely to have C4d expression in the portal vasculature and liver sinusoids (P=0.011). CONCLUSION: Humoral activation seems to play a role in pathophysiology of hepatic, especially chronic GVHD.

Single fraction spine radiosurgery for myeloma epidural spinal cord compression.

Radiosurgery delivers highly focused radiation beams to the defined target with high precision and accuracy. It has been demonstrated that spine radiosurgery can be safely used for treatment of spine metastasis with rapid and durable pain control, but without detrimental effects to the spinal cord. This study was carried out to determine the role of single fraction radiosurgery for epidural spinal cord compression due to multiple myeloma. A total of 31 lesions in 24 patients with multiple myeloma, who presented with epidural spinal cord compression, were treated with spine radiosurgery. Single fraction radiation dose of 10-18 Gy (median of 16 Gy) was administered to the involved spine including the epidural or paraspinal tumor. Patients were followed up with clinical exams and imaging studies. Median follow-up was 11.2 months (range 1-55). Primary endpoints of this study were pain control, neurological improvement, and radiographic tumor control. Overall pain control rate was 86%; complete relief in 54%, and partial relief in 32% of the patients. Seven patients presented with neurological deficits. Five patients neurologically improved or became normal after radiosurgery. Complete radiographic response of the epidural tumor was noted in 81% at 3 months after radiosurgery. During the follow-up time, there was no radiographic or neurological progression at the treated spine. The treatment was non-invasive and well tolerated. Single fraction radiosurgery achieved an excellent clinical and radiographic response of myeloma epidural spinal cord compression. Radiosurgery can be a viable treatment option for myeloma epidural compression.

Phase II study of bryostatin 1 and vincristine for aggressive non-Hodgkin lymphoma relapsing after an autologous stem cell transplant.

Bryostatin 1, isolated from a marine bryozoan, enhances the efficacy of cytotoxic agents through modulation of the protein kinase C pathway and is active in combination with vincristine for diffuse large B-cell lymphoma. Further, the apoptotic frequency of peripheral blood T lymphocytes as determined by flow cytometry may predict which patients will respond to this combination. We tested the efficacy and safety of bryostatin 1 50 microg/m(2) given over 24 hr and vincristine 1.4 mg/m(2) on days 1 and 15 every 28 days in aggressive B-cell non-Hodgkin lymphoma (NHL) relapsing after autologous stem cell transplantation. End points included tumor response, toxicity, and survival. Responses were correlated with an increase in apoptotic frequency of CD5+ cells by flow cytometry using annexin V staining. Fourteen patients were enrolled with 13 being evaluable for a response. The overall response rate was 31% with two patients achieving a complete response. The most common toxicities were Grade 3 lymphopenia (seven patients), Grade 3 to 4 neutropenia (two patients), and Grade 3 hypophosphatemia (two patients). Median progression-free and overall survivals for all patients were 5.7 and 21.4 months, respectively. One patient demonstrated an increase in T-cell apoptotic frequency, also achieving a complete response. Bryostatin 1 and vincristine have efficacy in select patients with aggressive NHL. Future investigations of agents targeting the protein kinase C pathway may benefit from early response assessment using flow cytometry to evaluate T-cell apoptosis.

Phase II trial of idiotype vaccination in previously treated patients with indolent non-Hodgkin's lymphoma resulting in durable clinical responses.

PURPOSE: To evaluate idiotype (Id) vaccination as a single agent in previously treated patients with indolent non-Hodgkin's lymphoma. PATIENTS AND METHODS: Patients underwent biopsy for determination of their lymphoma-specific Id sequence. Recombinant Id protein was manufactured and covalently linked with keyhole limpet hemocyanin (KLH) to generate Id/KLH. Patients received Id/KLH 1 mg on day 1 subcutaneously, with granulocyte-macrophage colony-stimulating factor 250 mug on days 1 to 4, monthly for 6 months. Booster injections were administered until progression. Both clinical and immune responses were evaluated. RESULTS: Thirty-two previously treated patients received at least one injection of Id/KLH, and 31 were assessed for efficacy. Responses were observed in four patients (one complete response and three partial responses). Median time to onset of response was 5.9 months (range, 2.3 to 14.1 months). Median duration of response has not been reached but should be at least 19.4 months (range, 10.4 to 27.2+ months). Median time to progression is 13.5 months. The most common adverse events were mild to moderate injection site reactions. Six (67%) of nine patients tested demonstrated a cellular immune response, and four (20%) of 20 patients demonstrated an antibody response against their Id. CONCLUSION: This trial demonstrates that Id/KLH alone can induce tumor regression and durable objective responses. Further study of Id/KLH is recommended in other settings where efficacy may be further enhanced as in first-line therapy or after cytoreductive therapy.

Hematologic malignancies with primary retroperitoneal presentation: clinicopathologic study of 32 cases.

CONTEXT: The retroperitoneum is an uncommon location for primary lymphomatous involvement. Initial presentation of disease in this site (primary retroperitoneal lymphoma) is considered to be rare. Because of the uncommon anatomic location, the diagnosis and subsequent management of these patients tend to be difficult for both pathologists and clinicians. OBJECTIVE: This report describes our experience during a period of 6 years 4 months with patients with hematologic malignancies primarily presenting in the retroperitoneum. DESIGN: A retrospective search of our medical records generated 32 patients who presented initially with abdominal pain or discomfort as their predominant symptom and who were found by imaging studies to have retroperitoneal mass or masses. All the histopathology slides were reviewed and classified based on the World Health Organization classification. RESULTS: There were 13 male and 19 female patients. Tumor types included diffuse large B-cell lymphoma (n = 12); grade 1 follicular lymphoma (n = 4); grade 3 follicular lymphoma (n = 1); B chronic lymphocytic leukemia/small lymphocytic lymphoma (n = 2); multiple myeloma (n = 1); mixed-cellularity Hodgkin lymphoma (n = 1); nodular sclerosis Hodgkin lymphoma (n = 1); aggressive B-cell lymphoma (n = 4); low-grade B-cell lymphoma (n = 4); lymphoblastic lymphoma, null cell type (n = 1); and precursor B-lymphoblastic lymphoma/leukemia (n = 1). More than half of the cases (17/32) were diagnosed on needle biopsy with immunophenotyping. CONCLUSIONS: Although open lymph node biopsy is a preferred method for diagnosis, needle biopsy plays a significant role in this setting, and, coupled with other information such as flow cytometry and immunohistochemistry, it is considered a practical and reliable method.

In vitro analysis of the antileukemic effect of tumor necrosis factor-alpha gene therapy with myeloid progenitor cells: the role of dendritic cells.

We have previously demonstrated that tumor necrosis factor-alpha (TNF-alpha) gene therapy with transgene-expressing myeloid progenitor cells (32DTNF-alpha) is effective in inhibiting the progression of leukemia with a lethal dose of murine 32Dp210 myeloid leukemia cells. Because TNF-alpha has been shown to induce the activation and maturation of dendritic cells (DCs), we investigated the effect of TNF-alpha secreted by transduced cells (32DTNF-alpha cells) on the activation of DCs and their role in the production of antileukemic cytotoxic T lymphocytes (CTLs). We demonstrate that administration of 32DTNF-alpha cells to the mice enhances the allo-stimulatory capacity of the splenic (CD11c+) and bone marrow-derived DCs in both mixed leukocyte response and CTL development. The enhanced allo-stimulatory capacity of splenic DCs from mice injected with 32DTNF-alpha cells correlated with increase in the cell-surface expression of the costimulatory molecules CD40, CD80, CD86, and major histocompatibility complex (MHC) class II molecules (I-Ak), and production of interleukin-12 (IL-12). Furthermore, administration of 32DTNF-alpha cells during immunization with irradiated 32Dp210 leukemia cells augmented the capacity of splenic DCs to stimulate antileukemic CTL response in spleen cells. Collectively, these data suggest that in vivo production of TNF-alpha by transduced cells enhances the phenotypic and functional activation of DCs, resulting in induction of a stronger antileukemic cytotoxic T-cell immune response.

Curcumin (diferuloyl-methane) enhances tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in LNCaP prostate cancer cells.

The role of natural food products in prevention of prostate cancer has been confirmed in recent epidemiological studies; however, the mechanism of chemoprevention by the dietary constituents largely remains unknown. Curcumin, the yellow pigment and active component of turmeric (Curcuma longa), exhibits chemopreventive and growth inhibitory activity against several tumor cell lines. The androgen-sensitive human prostate cancer cell line LNCaP is only slightly susceptible to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a member of the tumor necrosis factor family of cell death-inducing ligands. In this study, we investigated whether curcumin and TRAIL cooperatively interact to promote death of LNCaP cells. At low concentrations (10 micro M curcumin and 20 ng/ml TRAIL), neither of the two agents alone produced significant cytotoxicity (curcumin, <10%; TRAIL, approximately 15%) in LNCaP cells, as measured by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfonyl)-2H-tetrazolium dye reduction assay. On the other hand, cell death was markedly enhanced (2-3-fold) if tumor cells were treated with curcumin and TRAIL together. The combined curcumin and TRAIL treatment increased the number of hypodiploid cells and induced DNA fragmentation in LNCaP cells. The combined treatment induced cleavage of procaspase-3, procaspase-8, and procaspase-9, truncation of Bid, and release of cytochrome c from the mitochondria, indicating that both the extrinsic (receptor-mediated) and intrinsic (chemical-induced) pathways of apoptosis are triggered in prostate cancer cells treated with a combination of curcumin and TRAIL. These results define a potential use of curcumin to sensitize prostate cancer cells for TRAIL-mediated immunotherapy.

Disparate in vitro and in vivo antileukemic effects of resveratrol, a natural polyphenolic compound found in grapes.

Resveratrol (trans-3,4',5-trihydroxystilbene), a polyphenol found in grapes and grape wine, has been reported to exhibit cardioprotective and chemopreventive activity against chemical carcinogenesis. It has also been shown to have growth inhibitory activity toward solid tumors in vivo. However, the antitumor activity of resveratrol against hematologic tumors in vivo has not been examined. In this study, the antileukemic activity of resveratrol in vitro and in vivo was examined using a mouse myeloid leukemia cell line (32Dp210). Treatment of 32Dp210 leukemia cells with resveratrol at micromolar concentrations (25-50 micromol/L) significantly and irreversibly inhibited their clonal growth in vitro. The clonal growth inhibition by resveratrol was associated with extensive cell death and an increase in hypodiploid (sub-G1) cells. Resveratol caused internucleosomal DNA fragmentation, suggesting apoptosis as the mode of cell death in 32Dp210 cells. DNA fragmentation was associated with activation of caspase-3, because cleavage of procaspase-3 was detected in resveratrol-treated cells. Although 32Dp210 cells treated with resveratrol in vitro did not produce leukemia in vivo, only a weak antileukemic effect of resveratrol was observed when administered orally. At doses of 8 mg or 40 mg/kg body daily, five times/wk, resveratrol did not affect the survival of mice injected with leukemia cells. Weak potential antileukemic activity of resveratrol was suggested only at a dose of 80 mg/kg body (2 survivors of 14 mice treated). Thus, despite strong antiproliferative and proapoptotic activities of resveratrol against 32Dp210 cells in vitro, a potential antileukemia effect in vivo, if present, occurs only in a small fraction of mice.