Drug response prediction in high-risk multiple myeloma.

A Drug Response Prediction (DRP) score was developed based on gene expression profiling (GEP) from cell lines and tumor samples. Twenty percent of high-risk patients by GEP70 treated in Total Therapy 2 and 3A have a progression-free survival (PFS) of more than 10years. We used available GEP data from high-risk patients by GEP70 at diagnosis from Total Therapy 2 and 3A to predict the response by the DRP score of drugs used in the treatment of myeloma patients. The DRP score stratified patients further. High-risk myeloma with a predicted sensitivity to melphalan by the DRP score had a prolonged PFS, HR=2.4 (1.2-4.9, P=0.014) and those with predicted sensitivity to bortezomib had a HR 5.7 (1.2-27, P=0.027). In case of predicted sensitivity to bortezomib, a better response to treatment was found (P=0.022). This method may provide us with a tool for identifying candidates for effective personalized medicine and spare potential non-responders from suffering toxicity.

Gene expression risk signatures maintain prognostic power in multiple myeloma despite microarray probe set translation.

INTRODUCTION: Gene expression profiling (GEP) risk models in multiple myeloma are based on 3'-end microarrays. We hypothesized that GEP risk signatures could retain prognostic power despite being translated and applied to whole-transcript microarray data. METHODS: We studied CD138-positive bone marrow plasma cells in a prospective cohort of 59 samples from newly diagnosed patients eligible for high-dose therapy (HDT) and 67 samples from previous HDT patients with progressive disease. We used Affymetrix Human Gene 1.1 ST microarrays for GEP. Nine GEP risk signatures were translated by probe set match and applied to our data in multivariate Cox regression analysis for progression-free survival and overall survival in combination with clinical, cytogenetic and biochemical risk markers, including the International Staging System (ISS). RESULTS: Median follow-up was 66 months (range 42-87). Various translated GEP risk signatures or combinations hereof were significantly correlated with survival: among newly diagnosed patients mainly in combination with cytogenetic high-risk markers and among relapsed patients mainly in combination with ISS stage III. CONCLUSION: Translated GEP risk signatures maintain significant prognostic power in HDT myeloma patients. We suggest probe set matching for GEP risk signature translation as part of the efforts towards a microarray-independent GEP risk standard. (ClicinalTrials.gov identifier: NCT00639054).

Increased level of both CD4+FOXP3+ regulatory T cells and CD14+HLA-DR⁻/low myeloid-derived suppressor cells and decreased level of dendritic cells in patients with multiple myeloma.

Patients with multiple myeloma (MM) suffer from a general impaired immunity comprising deficiencies in humoral responses, T-cell responses as well as dendritic cell (DC) function. Thus, to achieve control of tumour growth through immune therapy constitutes a challenge. Careful evaluation of the immune status in patients with MM seems crucial prior to active immune therapy. We evaluated the proportion of both, DC, Treg cells and myeloid-derived suppressor cells (MDSC) in peripheral blood from patients with MM at diagnosis and in remission as well as patients with monoclonal gammopathy of undetermined significance (MGUS). We found that the proportion of both myeloid (m) DC and plasmacytoid (p) DC in patients at diagnosis was lowered compared to control donors, while only the proportion of pDC in patients in remission and with MGUS was significantly lower than in controls. The proportion of CD4+FOXP3+ Treg cells was increased in patients at diagnosis and not in patients in remission or with MGUS. Also, Treg cells from patients with MM were functionally intact as they were able to inhibit proliferation of both CD4 and CD8 T cells. Finally, we observed an increase in the proportion of CD14+HLA-DR⁻/low MDSC in patients with MM at diagnosis, illustrating that this cell fraction is also distorted in patients with MM. Taken together, our results illustrate that, both mDC, pDC, Treg cells and MDSC are affected in patients with MM underlining the fact that the immune system is dysregulated as a consequence of the disease.

Early infections in patients undergoing high-dose treatment with stem cell support: a comparison of patients with non-Hodgkin lymphoma and multiple myeloma.

BACKGROUND: Infections are life-threatening complications in patients undergoing high-dose chemotherapy with stem cell support (HDT). Knowledge of the infectious pathogens is essential to make a safe outpatient setting. METHODS: We conducted a retrospective study of 208 patients treated with HDT. The population included non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) patients. No patients received prophylactic antibacterial treatment. RESULTS: Pathogens were isolated from 44% of all patients. MM patients more frequently had multiple pathogens in blood cultures (38% versus 25%). Transplantation related mortality was similar between the groups. CONCLUSION: The frequency of isolated pathogens, positive blood cultures, and the diversity of pathogens were higher in MM patients as compared to NHL patients. However, this did not translate into higher transplantation-related mortality, probably because broad-spectrum antibiotic treatment could be initiated immediately. A safe outpatient setting with prophylactic antibiotic treatment is dependent on continuous collection and registration of microbiological findings.

The polymorphism IL-1beta T-31C is associated with a longer overall survival in patients with multiple myeloma undergoing auto-SCT.

Proinflammatory cytokines are suspected to play a role in the pathogenesis of multiple myeloma (MM). Therefore, it is possible that inborn genetic variations leading to a modified expression of these cytokines will influence the outcome for these patients. We investigated 348 MM patients undergoing high-dose melphalan treatment followed by Auto-SCT and examined the influence of single nucleotide polymorphisms (SNPs) in genes involved in the inflammatory response. We found that the polymorphism IL-1beta T-31C significantly influenced overall survival (OS; P=0.02) and that carriers of the variant C-allele had a significantly longer survival than homozygous wild-type allele TT-carriers (relative risk 0.6 (95% CI=0.5-0.9); P=0.008). The polymorphisms IL-6 G-174C, IL-10 C592A, PPARgamma2 Pro(12)Ala, COX-2 A-1195G, COX-2 T8473C and NFKB1 ins/del did not influence the OS in this group of patients. Furthermore, homozygous carriers of the variant allele of IL-1beta T-31C were at 1.37-fold (CI=1.05-1.80) increased risk of MM as compared with population-based controls (P=0.02). Our results indicate that IL-1beta is involved in the pathogenesis of MM.

Serological tumor markers for small cell lung cancer and their therapeutic implications.

Serological tumor markers may become widely used as inexpensive and non-invasive methods of cancer detection. Markers of current interest for small cell lung cancer (SCLC) comprise enzymes, peptides, proteins, and carbohydrates. None of the serological markers for SCLC have yet proven to be of diagnostic value and at present their use is limited to monitoring disease and indicating prognosis. However, whilst serological markers related to the metabolic state of SCLC cells, such as neuron-specific enolase, serum thymidine kinase and tissue polypeptide antigen, may only be used for monitoring patients and for estimating prognosis, the other serological markers under current investigation may be used to indicate new treatment forms. Several novel approaches, including interference in the autocrine growth-regulating loop of SCLC by either peptides or antibodies, have been tried, SCLC is a highly heterogeneous tumor with respect to antigen expression, regulation of growth, and differentiation state. It is therefore important that new interventions are directed against both antigen-positive and antigen-negative tumor cells. For instance, radioisotopes or enzymes coupled to antibodies may be effective by exerting toxicity at some distance from the target. Antigens expressed on SCLC cells, such as peptide receptors involved in growth regulation, carbohydrate antigens like Lewis antigens, carcinoembryonic antigen and the ganglioside fucosylGM1, provide potential targets for antibody-conjugated therapy.

Monoclonal antibodies for diagnosis and potential therapy of small cell lung cancer--the ganglioside antigen fucosyl-GM1.

Lung carcinomas represent a highly heterogenous group of tumors, which includes small cell lung cancer (SCLC). The ganglioside fucosyl-GM1 (FucGM1) was originally identified as a highly selective marker for SCLC. A number of monoclonal antibodies against FucGM1 have been produced against the purified ganglioside, and their specificity validated. With monoclonal antibodies, FucGM1 has been detected in tissues as well as in serum samples from SCLC patients. A sensitive, quantitative assay method for FucGM1 was developed based on scintillation proximity immunoassay (SPA): A specific monoclonal antibody against FucGM1 is bound to immunosorbent particles that contain a fluor. When radiolabelled FucGM1 binds to these particles, the fluor is excited, and the photons emitted can be measured directly in a beta-counter. This sensitive assay for FucGM1 has several potential diagnostic applications: differential diagnosis of SCLC, development of serum assays for diagnosis and monitoring of disease progression, and monitoring of patient response to therapy. The expression of FucGM1 in SCLC cells is heterogeneous, and may depend on the differentiated state of tumor cells. However, monoclonal antibodies specific for FucGM1 may have important future applications for radioimmunoimaging as well as in immunotherapy for SCLC.

Gastrin releasing peptide GRP(14-27) in human breast cancer cells and in small cell lung cancer.

Immunoreactivity related to the gastrin-releasing peptide (GRP) precursor was detected in four different human breast cancer cell lines. The amounts and the characteristics in extracts from different breast carcinoma cells were compared with cell extracts from small cell lung cancer (SCLC) cells. Two different radioimmunoassays were employed, directed against the amino acid sequence 14-27 of GRP (IR-GRP) or the 42-53 amino acid sequence at the C-terminal end of the GRP precursor (GRP precursor fragment). In extracts from T47D cells cultured under serum free conditions, IR-GRP coeluted with GRP(14-27) or GRP(18-27) in Sephadex G-50 chromatography. No immunoreactivity was detected in the fractions containing high molecular weight components. In a total of 41 human breast carcinoma biopsies from different postmenopausal patients, IR-GRP was detected by immunohistological staining in 39% of the samples. When the GRP(14-27) peptide was added exogenously to breast cancer and SCLC cell lines under serum-free culture conditions, (3H)-thymidine incorporation was stimulated by GRP(14-27) in the SCLC cell lines. Of the breast cancer cell lines only the T47D cell line responded with an increase in (3H)-thymidine incorporation comparable to the increase observed with SCLC cells. Recently, it has been reported that GRP-like receptors are present in some human breast cancer cell lines, including the T47D cell line studied here. The breast cancer cell line T47D therefore expresses the GRP peptide and the receptor for GRP. The identification of GRP-like receptors on T47D cells is in accordance with our present observation of a growth response to GRP(14-27) as evaluated by increased (3H)-thymidine incorporation.

Immunochemical detection of a small cell lung cancer-associated ganglioside (FucGM1) antigen in serum.

Recently, the ganglioside FucGM1 (Fuc alpha 1-2Gal beta 1-3GalNAc beta 1-4[NeuAc alpha 2-3]-Gal beta 1-4Glc beta 1-1 Cer) was identified as a small cell lung cancer (SCLC) marker both in chemical and histochemical studies. In order to further determine whether the FucGM1 ganglioside is shed from the tumor site and consequently is present in the serum of SCLC patients, we produced a series of new monoclonal antibodies raised against FucGM1 and related glycolipids. Shedding of the FucGM1 ganglioside was studied both in vitro and in vivo using SCLC cell lines and nude mice xenografts of SCLC cells as model systems, and finally immunochemical analyses were performed on serum samples from patients with SCLC. High-performance thin-layer chromatography immunostaining demonstrated the presence of FucGM1 in conditioned culture media obtained from FucGM1-positive SCLC cell lines. Furthermore, tumor extracts of SCLC cell line xenografts in nude mice were positive for the FucGM1 marker, and more importantly the marker was also present in serum samples from these mice. Twenty serum samples were obtained from patients with histologically verified SCLC. Eight patients had localized disease, and the remaining patients had disseminated cancer involving metastases to other organ sites. Sera from 4 of these patients were clearly positive, and 2 additional cases were found to be weakly positive. The positive patient sera were all from patients with extensive disease. Sera from 12 patients with non-SCLC and 20 healthy individuals were all found to be negative. These results clearly establish the FucGM1 glycolipid as a potential serum marker of SCLC for which a sensitive immunoassay should be developed and tested using a larger series of serum samples.

Production of gastrin-releasing peptide-(18-27) and a stable fragment of its precursor in small cell lung carcinoma cells.

The production and postsecretory stability of gastrin-releasing peptide (GRP) and the C-terminal part of the GRP precursor were studied in small cell lung cancer cell lines using RIAs developed against these two parts of the precursor. In three otherwise different cell lines (NIC-H345, NIC-H69, and NIC-H510), similar chromatographic patterns of mainly GRP-(18-27) and some GRP-(14-27) along with large fragments of the C-terminal counterpart of the precursor were found to be stored in the cells. In tissue culture medium, gel filtration chromatography indicated that postsecretory limited proteolysis of the GRP precursor fragments occurred. The amount of accumulated immunoreactivity varied among the three cell lines and between the two parts of the precursor. In medium in which only low amounts of GRP immunoreactivity accumulated, the radiolabeled GRP was degraded rapidly. When incubated with plasma, GRP-(14-27) disappeared within a few hours, whereas the C-terminal precursor fragments were stable. It is concluded that the postsecretory stability of peptides excised from the GRP precursor in small cell lung cancer cells varies under tissue culture conditions, but epitopes in the C-terminal part of the precursor are more stable in plasma than the small GRP peptides and, thus, may serve as a better indicator than GRP itself for expression of the GRP precursor in cancer cells.