Autologous Stem Cell Transplantation In Multiple Myeloma

Background. Multiple myeloma (MM) is the second most common hematologic malignancy after lymphomas In Finland: the annual incidence of MM is approximately 200. For three decades the median survival remained at 3 to 4 years from diagnosis until high-dose melphalan treatment supported by autologous stem cell transplantation (ASCT) became the standard of care for newly diagnosed MM since the mid 1990’s and the median survival increased to 5 – 6 years. This study focuses on three important aspects of ASCT, namely 1) stem cell mobilization, 2) single vs. double ASCT as initial treatment, and 3) the role of minimal residual disease (MRD) for longterm outcome. Aim. The aim of this series of studies was to evaluate the outcomes of MM patients and the ASCT procedure at the Turku University Central Hospital, Finland. First, we tried to identify which factors predict unsuccessful mobilization of autologous stem cells. Second, we compared the use of short-acting granulocyte-colony stimulating factor (GCSF) with long-acting G-CSF as mobilization agents. Third, one and two successive ASCTs were compared in 100 patients with MM. Fourth, for patients in complete response (CR) after stem cell transplantation (SCT), patient-specific probes for quantitative allele-specific oligonucleotide polymerase-chain reaction (qASO-PCR) measurements were designed to evaluate MRD and its importance for long-term outcome. Results. The quantity of previous chemotherapy and previous interferon use were significant pre-mobilization factors that predicted mobilization failure, together with some factors related to mobilization therapy itself, such as duration and degree of cytopenias and occurrence of sepsis. Short-acting and long-acting G-CSF combined with chemotherapy were comparable as stem cells mobilizers. The progression free (PFS) and overall survival (OS) tended to be longer after double ASCT than after single ASCT with a median follow-up time of 4 years, but this difference disappeared as the follow-up time increased. qASO-PCR was a good and sensitive divider of the CR patients into two prognostic groups: MRD low/negative (≤ 0.01%) and MRD high (>0.01%) groups with a significant difference in PFS and suggestively also in OS. Conclusions. When the factors prediciting a poor outcome of stem cell mobilization prevail, it is possible to identify those patients who need specific efforts to maximize the mobilization efficacy. Long-acting pegfilgrastim is a practical and effective alternative to short-acting filgrastim for mobilization therapy. There is no need to perform double ASCT on all eligible patients. MRD assessment with qASO-PCR is a sensitive method for evaluation of the depth of the CR response and can be used to predict long-term outcome after ACST.

Homogeneous Assays for Simplified Screening of HLA-conferred Genetic Disease Risk

The increasing incidence of type 1 diabetes has led researchers on a quest to find the reason behind this phenomenon. The rate of increase is too great to be caused simply by changes in the genetic component, and many environmental factors are under investigation for their possible contribution. These studies require, however, the participation of those individuals most likely to develop the disease, and the approach chosen by many is to screen vast populations to find persons with increased genetic risk factors. The participating individuals are then followed for signs of disease development, and their exposure to suspected environmental factors is studied. The main purpose of this study was to find a suitable tool for easy and inexpensive screening of certain genetic risk markers for type 1 diabetes. The method should be applicable to using whole blood dried on sample collection cards as sample material, since the shipping and storage of samples in this format is preferred. However, the screening of vast sample libraries of extracted genomic DNA should also be possible, if such a need should arise, for example, when studying the effect of newly discovered genetic risk markers. The method developed in this study is based on homogeneous assay chemistry and an asymmetrical polymerase chain reaction (PCR). The generated singlestranded PCR product is probed by lanthanide-labelled, LNA (locked nucleic acid)-spiked, short oligonucleotides with exact complementary sequences. In the case of a perfect match, the probe is hybridised to the product. However, if even a single nucleotide difference occurs, the probe is bound instead of the PCR product to a complementary quencher-oligonucleotide labelled with a dabcyl-moiety, causing the signal of the lanthanide label to be quenched. The method was applied to the screening of the well-known type 1 diabetes risk alleles of the HLA-DQB1 gene. The method was shown to be suitable as an initial screening step including thousands of samples in the scheme used in the TEDDY (The Environmental Determinants of Diabetes in the Young) study to identify those individuals at increased genetic risk. The method was further developed into dry-reagent form to allow an even simpler approach to screening. The reagents needed in the assay were in dry format in the reaction vessel, and performing the assay required only the addition of the sample and, if necessary, water to rehydrate the reagents. This allows the assay to be successfully executed even by a person with minimal laboratory experience.