Dispenzieri, Angela

AbstractPrimary systemic amyloidosis (AL) is a rare monoclonal plasma cell (PC) disorder characterized by the deposition of misfolded immunoglobulin (Ig) light chains (LC) in vital organs throughout the body. To our knowledge, no cell lines have ever been established from AL patients. Here we describe the establishment of the ALMC-1 and ALMC-2 cell lines from an AL patient. Both cell lines exhibit a PC phenotype and display cytokine-dependent growth. Using a comprehensive genetic approach, we established the genetic relationship between the cell lines and the primary patient cells, and we were also able to identify new genetic changes accompanying tumor progression that may explain the natural history of this patient's disease. Importantly, we demonstrate that free lambda LC secreted by both cell lines contained a beta structure and formed amyloid fibrils. Despite absolute Ig LC variable gene sequence identity, the proteins show differences in amyloid formation kinetics that are abolished by the presence of Na2SO4. The formation of amyloid fibrils from these naturally secreting human LC cell lines is unprecedented. Moreover, these cell lines will provide an invaluable tool to better understand AL, from the combined perspectives of amyloidogenic protein structure and amyloid formation, genetics, and cell biology.

AbstractThe immunoglobulin free light chain (FLC) assay is an invaluable tool for following patients with oligosecretory plasma cell dyscrasia. Baseline values have also been shown to be prognostic in all plasma cell disorders tested. A looming question, however, is the role it should play in following myeloma patients with disease that is measurable using serum and urine electrophoresis. We used the data and stored samples from a mature Eastern Cooperative Oncology Group clinical trial (E9486) to assess serum levels of FLC at baseline and after 2 months of alkylator-based therapy. For serial determinations, the absolute level of involved serum FLC or the difference of the involved and uninvolved FLC is preferred over the ratio of involved to uninvolved FLC. FLC response after 2 months of therapy was superior to early M-protein measurement to predict overall response. The ideal cut-point for FLC change appears to be between 40% and 50% reduction. The correlation between serial measurements of serum FLC and urine M-protein is inadequate to abolish the serial 24-hour urine protein. Although baseline values of FLC are prognostic in newly diagnosed myeloma patients, serial measurements do not appear to have added value in patients who have M-proteins measurable by electrophoresis.

Abstract The Multiple Myeloma Research Consortium has established a tissue bank for the deposition of bone marrow samples from patients with multiple myeloma to be mailed and processed under good laboratory practices. To date, over 1,000 samples have been collected. At this time, limited information is available on shipped bone marrow aspirates in regards to cell viability, yield, purity, and subsequent RNA yield and quality. To test these determinants, we did a pilot study on behalf of the Multiple Myeloma Research Consortium where samples were drawn at Mayo Clinic Rochester (MCR) pooled and split into two equal aliquots. One-half of each sample was processed following good laboratory practices compliant standard operating procedures, immediately after sample procurement, at MCR. The CD138+ cells were stored at -80°C as a Trizol lysate. The other half of the aspirate was sent overnight to Mayo Clinic Scottsdale where they were processed using identical standard operating procedures. The RNA was extracted and analyzed in a single batch at MCR. At both locations, samples were assayed for the following quality determinants: Viability was assessed using a three-color flow cytometric method (CD45, CD38, and 7-AAD). Cell counts were done to determine plasma cell recovery and post-sort purity determined by means of a slide-based immunofluorescent assay. RNA recovery and integrity was assessed using the Agilent Bioanalyzer. Lastly, gene expression profiles were compared to determine the signature emanating from the shipment of samples. Despite minor differences, our results suggest that shipment of samples did not significantly affect these quality determinants in aggregate. (Cancer Epidemiol Biomarkers Prev 2008;17(3):666–73)

Treatments for myeloma have expanded in the last decade, but it is not clear if the introduction of novel therapies and the increased use of high-dose therapy have translated into better outcome for patients with myeloma. We examined the outcome of 2 groups of patients seen at a single institution, one from time of diagnosis and the other from the time of relapse, to examine the survival trends over time. Among 387 patients relapsing after stem-cell transplantation, a clear improvement in overall survival from the time of relapse was seen, with those relapsing after 2000 having a median overall survival of 23.9 versus 11.8 months (P < .001) for those who relapsed prior to this date. This improvement was independent of other prognostic factors. Patients treated with one or more of the newer drugs (thalidomide, lenalidomide, bortezomib) had longer survival from relapse (30.9 vs 14.8 months; P < .001). In a larger group of 2981 patients with newly diagnosed myeloma, those diagnosed in the last decade had a 50% improvement in overall survival (44.8 vs 29.9 months; P < .001). In this study, we demonstrate improved outcome of patients with myeloma in recent years, both in the relapsed setting as well as at diagnosis.

Purpose High-dose therapy with autologous stem-cell transplantation (HDT-ASCT) is now almost standard therapy for many patients with multiple myeloma, partly because of higher complete response (CR) rates. Some studies suggest that tandem transplantation gives superior results. The aim of this study was to determine whether the depth of the response to HDT-ASCT leads to an improvement in time to progression (TTP) and overall survival (OS). We hypothesized that patients with CR before HDT-ASCT (BCR) will have their disease burden reduced further and experience a longer TTP and perhaps OS. Patients and Methods All patients who achieved BCR or CR after HDT-ASCT (ACR) were identified. The characteristics and long-term outcome of these patients were evaluated. Results We identified 14 patients with BCR and 103 patients with ACR who were treated in similar fashion. The patients have been followed for more than 6 years, and the median for OS has not been reached (60-month survival, 55% for BCR and 63% for ACR; P = .83). The median TTP was 43 months for BCR and 34 months for ACR (P = .39). Conclusion The depth of the response in myeloma does not necessarily lead to an improvement in TTP and OS. Tumor dynamics considerations show that the yield from sequential cycles of chemotherapy decreases. Patients who achieve CR with the first transplant can be safely observed without jeopardizing OS.

Abstract Purpose: 2-Methoxyestradiol (2ME2) is an endogenous product of estradiol metabolism with antiangiogenic and antineoplastic properties. We report on the first phase II trial of 2ME2 in multiple myeloma. Experimental Design: 2ME2 was administered orally at a dose of 1,000 mg daily. Sixty patients (31 men and 29 women) were treated. After 39 patients were accrued, the dose was increased to 800 mg twice daily for the remaining patients. Results: Thirty-one patients had relapsed or refractory multiple myeloma, and 29 had plateau phase multiple myeloma. Median age was 60 years (range, 27-84 years). Therapy was well tolerated. Common adverse events included anemia (35%), fatigue (35%), nausea (25%), diarrhea (20%), hot flushes (20%), headache (17%), muscle cramps (15%), and upper respiratory tract infection (15%). Most adverse events were mild (grade 1-2); 12% experienced grade 3-4 adverse events. Median time to progression was 3.8 months, with 5.6 months for plateau phase disease and 2.3 months for relapsed multiple myeloma. Estimated progression-free survival rates for all patients at 1, 2, and 3 years were 24%, 17%, and 11%, respectively. Three patients, all with plateau phase disease, have been on study for over 4 years without progression at 50, 60, and 63 months, respectively. Minor response was noted in 2 patients. Conclusions: Although no partial responses have been seen thus far, the minor responses and prolonged stable disease seen with 2ME2 therapy are promising. Plasma levels indicate that the dose of 2ME2 was inadequate. A new formulation with better bioavailability will be tested soon in multiple myeloma.

We hypothesized that increased monoclonal free kappa or lambda immunoglobulin light chains in smoldering multiple myeloma (SMM), as detected by the serum free light chain (FLC) assay, indicates an increased risk of progression to active myeloma. Baseline serum samples obtained within 30 days of diagnosis were available in 273 patients with SMM seen from 1970 to 1995. At a median follow-up of surviving patients of 12.4 years, transformation to active disease has occurred in 59%. The best breakpoint for predicting risk of progression was an FLC ratio of 0.125 or less, or 8 or more (hazard ratio, 2.3; 95% CI, 1.6-3.2). The extent of abnormality of FLC ratio was independent of SMM risk categories defined by number of bone marrow plasma cells (BMPCs) and size of serum M proteins (BMPC ≥ 10% and serum M protein ≥ 3 g/dL; BMPC ≥ 10% but serum M protein < 3 g/dL; and serum M protein≥ 3 g/dL but BMPC < 10%). Incorporating the FLC ratio into the risk model, the 5-year progression rates in high-, intermediate-, and low-risk groups were 76%, 51%, and 25%, respectively. The serum immunoglobulin FLC ratio is an important additional determinant of clinical outcome in patients with SMM.