Jeffrey R. Sawyer

Cytogenetic findings in 200 patients with multiple myeloma

Cytogenetic studies were performed in 200 consecutive patients with multiple myeloma and related disorders. Structurally or numerically abnormal clones were found in 63 patients (32%), including 8 of 45 untreated patients (18%), and 55 of 155 treated patients (35%). The abnormal karyotypes generally showed numerous numerical and structural aberrations and in some patients multiple abnormal clones. The most striking feature of patients with hyperdiploid karyotypes was the finding of consistent recurring trisomies for chromosomes 3, 5, 7, 9, 11, 15, 19, and 21, cosegregating together in many cases. Monosomy for chromosome 13 was the most common chromosome loss, occurring in 18 abnormal patients (29%), while interstitial deletions involving band 13q14 occurred in an additional 9 patients, indicating a loss of all or part of chromosome 13 in a high percentage of patients with abnormal karyotypes (43%). Structural aberrations of chromosome 1 were most frequent, occurring in 30 of 63 patients (48%), and involved almost equally the short and long arms. The single most frequent chromosome breakpoint involved band 14q32 and was found in 21 patients (33%), including 11 patients with a 14q+ chromosome, 8 with t(11;14)(q13;q32), and 2 with t(8;14)(q24;132).

Magnetic Resonance Imaging in Multiple Myeloma: Diagnostic and Clinical Implications

PURPOSE Magnetic resonance imaging (MRI) permits the detection of diffuse and focal bone marrow infiltration in the absence of osteopenia or focal osteolysis on standard metastatic bone surveys (MBSs). PATIENTS AND METHODS Both baseline MBS and MRI were available in 611 of 668 myeloma patients who were treated uniformly with a tandem autologous transplantation-based protocol and were evaluated to determine their respective merits for disease staging, response assessment, and outcome prediction. RESULTS MRI detected focal lesions (FLs) in 74% and MBS in 56% of imaged anatomic sites; 52% of 267 patients with normal MBS results and 20% of 160 with normal MRI results had FL on MRI and MBS, respectively. MRI- but not MBS-defined FL independently affected survival. Cytogenetic abnormalities (CAs) and more than seven FLs on MRI (MRI-FLs) distinguished three risk groups: 5-year survival was 76% in the absence of both more than seven MRI-FLs and CA (n = 276), 61% in the presence of one MRI-FL (n = 262), and 37% in the presence of both unfavorable parameters (n = 67). MRI-FL correlated with low albumin and elevated levels of C-reactive protein, lactate dehydrogenase, and creatinine, but did not correlate with age, beta-2-microglobulin, and CA. Resolution of MRI-FL, occurring in 60% of cases and not seen with MBS-defined FL, conferred superior survival. CONCLUSION MRI is a more powerful tool for detection of FLs than is MBS. MRI-FL number had independent prognostic implications; additionally, MRI-FL resolution identified a subgroup with superior survival. We therefore recommend that, in addition to MBS, MRI be used routinely for staging, prognosis, and response assessment in myeloma.

Heparanase Enhances Syndecan-1 Shedding A NOVEL MECHANISM FOR STIMULATION OF TUMOR GROWTH AND METASTASIS

When shed from the cell surface, the heparan sulfate proteoglycan syndecan-1 can facilitate the growth, angiogenesis, and metastasis of tumors. Here we report that tumor cell expression of heparanase, an enzyme known to be a potent promoter of tumor progression and metastasis, regulates both the level and location of syndecan-1 within the tumor microenvironment by enhancing its synthesis and subsequent shedding from the tumor cell surface. Heparanase regulation of syndecan-1 is detected in both human myeloma and breast cancer cell lines. This regulation requires the presence of active enzyme, because mutated forms of heparanase lacking heparan sulfate-degrading activity failed to influence syndecan-1 expression or shedding. Removal of heparan sulfate from the cell surface using bacterial heparitinase dramatically accelerated syndecan-1 shedding, suggesting that the effects of heparanase on syndecan-1 expression by tumor cells may be due, at least in part, to enzymatic removal or reduction in the size of heparan sulfate chains. Animals bearing tumors formed from cells expressing high levels of heparanase or animals transgenic for heparanase expression exhibited elevated levels of serum syndecan-1 as compared with controls, indicating that heparanase regulation of syndecan-1 expression and shedding can occur in vivo and impact cancer progression and perhaps other pathological states. These results reveal a new mechanism by which heparanase promotes an aggressive tumor phenotype and suggests that heparanase and syndecan-1 act synergistically to fine tune the tumor microenvironment and ensure robust tumor growth.

Prognostic impact of cytogenetic and interphase fluorescence in situ hybridization-defined chromosome 13 deletion in multiple myeloma: early results of total therapy II.

Summary. Cytogenetic abnormalities of chromosome 13 (CA 13) and those detected by fluorescence in situ hybridization (FISH 13) have both been associated with poor prognosis in multiple myeloma (MM) patients. The prognostic implications of CA, FISH 13 and other standard laboratory parameters were examined in the first 231 patients enrolled in Total Therapy II, an intensive cytotoxic chemotherapy programme with tandem autotransplants. Three‐year projections of event‐free survival (EFS) and overall survival (OS) were 71% and 77% respectively. CA 13 was detected in 14% and significantly correlated with FISH 13 (present in 51%), tumour burden, proliferative activity and lactic dehydrogenase (LDH). Both EFS and OS were significantly shorter in patients with CA 13, FISH 13, LDH ≥ 190 U/l, β2 microglobulin ≥ 4 mg/l and C reactive protein ≥ 4·0 mg/l; other CA was an additional risk factor for OS. Two‐thirds of CA 13 patients were identified by FISH 13 and plasma‐cell‐labelling index (PCLI) ≥ 0·4%; however, PCLI failed to identify additional risk groups in FISH subsets. Although present in considerably fewer patients, CA 13 imparted more rapid relapse (61% at 3 years) and death (43% at 3 years) than FISH 13 (38% and 35%; P = 0·02 and 0·1 respectively) and should be part of the initial work‐up of patients with MM.

Both hypodiploidy and deletion of chromosome 13 independently confer poor prognosis in multiple myeloma

Summary. Complete or partial deletion of chromosome 13 or translocations involving 13q (Δ13) by conventional cytogenetic analysis confers a poor prognosis in multiple myeloma (MM) patients, even with timely application of tandem autologous transplants. It was recently suggested that the prognostic significance of Δ13 is related to its frequent association with hypodiploidy but by itself does not have a poor prognostic significance. We therefore analysed our experience in 1475 consecutive MM patients in whom we intended treatment with tandem transplants after a melphalan‐based conditioning regimen. Patients with abnormal cytogenetic analysis were grouped into hypodiploid/hypotetraploid, pseudodiploid and hyperdiploid groups, according to their modal chromosome number. Their event‐free and overall survival were compared with those of patients with a normalkaryotype. Both hypodiploidy and Δ13 were found to independently confer poor prognosis in MM patients. Furthermore, these parameters in combination with easily obtained pretransplant levels of β‐2 microglobulin and albumin define three groups of MM patients with clearly distinct outcomes.

Preceding standard therapy is the likely cause of MDS after autotransplants for multiple myeloma.

Myelodysplastic syndromes (MDS) and acute myeloid leukaemia (AML) have been reported after autologous transplantation (AT) for lymphoma. It is not clear whether myeloablative therapy used in conjunction with autologous transplantation contributes to the development of MDS/AML or whether the conventional chemotherapy preceding the transplant, and administered over a prolonged period, causes these secondary malignancies. To address this issue, we examined 188 patients with multiple myeloma (MM) who had received AT. 71 patients with no more than one cycle of standard chemotherapy were enrolled in our Total Therapy program, designed to avoid exposure to alkylating agents prior to peripheral blood stem cell mobilization (group 1). The median duration of pre‐transplant therapy in group 1 was 7.6 months and significantly shorter than the 24 months of 117 patients (group 2) with more prolonged conventional therapy (P = 0.0001). All seven patients developing MDS post‐transplantation belonged to group 2 (P = 0.02); the median durations from initial therapy and first transplant were 66 months (range 38–86) and 24 months (range 9–39), respectively. Our findings provide evidence that prolonged standard‐dose alkylating agent therapy prior to transplantation, rather than autotransplant‐supported myeloablative treatment, is associated with development of MDS/AML. Stem cell damaging alkylator treatment should be avoided, not to compromise PBSC collection, but also to reduce the risk of treatment‐related MDS/AML.

The prognostic significance of cytogenetics and molecular profiling in multiple myeloma

Multiple myeloma (MM) is a plasma cell malignancy characterized by very complex cytogenetic and molecular genetic aberrations. In newly diagnosed symptomatic patients, the modal chromosome number is usually either hyperdiploid with multiple trisomies or hypodiploid with one of several types of immunoglobulin heavy chain (Ig) translocations. The chromosome ploidy status and Ig rearrangements are two genetic criteria that are used to help stratify patients into prognostic groups based on the findings of conventional cytogenetics and fluorescence in situ hybridization (FISH). In general, the hypodiploid group with t(4;14)(p16;q32) or t(14;16)(q32;q23) is considered a high-risk group, while the hyperdiploid patients with t(11;14)(q13;q32) are considered a better prognostic group. As the disease progresses, it becomes more proliferative and develops a number of secondary chromosome aberrations. These secondary aberrations commonly involve MYC rearrangements, del(13q), del(17p), and the deletion of 1p and/or amplification of 1q. Of the secondary aberrations, del(17p) is consistently associated with poor prognosis. All of these cytogenetic aberrations and many additional ones are now identified by means of high resolution molecular profiling. Gene expression profiling (GEP), array comparative genomic hybridization (aCGH), and single-nucleotide polymorphism (SNP) arrays have been able to identify novel genetic aberration patterns that have previously gone unrecognized. With the integration of data from these profiling techniques, new subclassifications of MM have been proposed which define distinct molecular genetic subgroups. In this review, the findings from conventional cytogenetics, interphase FISH, GEP, aCGH, and SNP profiles are described to provide the conceptual framework for defining the emerging molecular genetic subgroups with prognostic significance.

Multicolour spectral karyotyping identifies new translocations and a recurring pathway for chromosome loss in multiple myeloma

Multicolour spectral karyotyping (SKY) was performed on primary tumour specimens from 100 patients with multiple myeloma (MM) that showed complex clonal chromosome aberrations not fully characterized by G‐banding. In this study, SKY was able to identify or revise translocations with breakpoints involving 14q32, 11q13 or 8q24 in 32 patients (32%). Five new recurring translocations were identified, two of which involved chromosome 22. A subtle reciprocal translocation t(14;22) (q32;q11∼12) was identified using SKY in two patients and a second, much larger, translocation t(11;22)(q13;q13) was identified using G‐banding in three patients. A third new translocation was identified in two patients using SKY and G‐banding as der(7)t(7;7)(p15∼22;q22∼32). Twenty‐three patients (23%) showed the loss of 8p by whole‐arm translocations with different whole‐arm donor chromosomes. Among this group, two new recurring whole‐arm translocations involving the centromeric breakpoint 8q10 were identified as der(8;20)(q10;q10) and der(8;18) (q10;q10) in three patients each. In addition, a novel pattern of three‐way translocations involving the clonal evolution of the t(8;22)(q24;q11) by the subsequent loss of 8p by whole‐arm translocations was found in three patients. The chromosome instability identified here demonstrates that the loss of 8p can occur by multiple whole‐arm translocations, indicating a new pathway for the loss of a specific chromosome region in MM.

Predicting long-term (≥ 5 years) event-free survival in multiple myeloma patients following planned tandem autotransplants

Summary. Although outcome in multiple myeloma (MM) patients has improved significantly with the introduction of autotransplants (AT), the curability of this approach remained to be demonstrated. Therefore, we analysed outcome and prognostic factors using a logistic regression model in 515 consecutive newly diagnosed and previously treated patients intended to receive melphalan‐based tandem transplants with follow up of ≥ 5 years. One quarter ofpatients had event‐free survivals (EFS) ≥ 5 years with no further relapses seen after 7 years (46 patients on plateau). On multivariate analysis, factors associated with EFS ≥ 5 years were absence of chromosome 11 and 13 abnormalities (odds ratio: 6·1), ≤ 12 months of preceding standard‐dose therapy (SDT) (OR: 2·6) and β‐2 microglobulin (B2M) level ≤ 2·5 mg/l at time of first AT (OR: 1·7). Patients with only favourable variables (25%) had a 7‐year EFS in excess of 35%, compared with 15% and 10%, respectively, with one (43%) or two unfavourable variables (27%), and 0% for 5% of patients with three unfavourable variables (P < 0·0001). Using a 1‐year landmark analysis to allow for guaranteed time and thereby excluding early treatment failures, attaining a complete remission (CR) had no significant effect on long‐term survival. Our data are consistent with cure in MM patients with a CR duration ≥ 7 years and re‐establishment of a monoclonal gammopathy of undetermined significance (MGUS) phase in those with persistent evidence of disease post transplantation, but without disease progression ≥ 7 years.

Genomic instability in multiple myeloma: Evidence for jumping segmental duplications of chromosome arm 1q

Multiple myeloma (MM) is a malignant plasma cell disorder characterized by complex karyotypes and chromosome 1 instability at the cytogenetic level. Chromosome 1 instability generally involves partial duplications, whole‐arm translocations, or jumping translocations of 1q, identified by G‐banding. To characterize this instability further, we performed spectral karyotyping and fluorescence in situ hybridization with probes for satII/III (1q12), BCL9 (1q21), and IL6R (1q21) on the karyotypes of 44 patients with known 1q aberrations. In eight patients, segmental duplication of 1q12–21 and adjacent bands occurred on nonhomologous chromosomes. In five cases, the 1q first jumped to a nonhomologous chromosome, after which the 1q12–21 segment again duplicated itself 1–3 times. In three other cases, segmental duplications occurred after the 1q first jumped to a nonhomologous chromosome, where the proximal adjacent nonhomologous chromosome segment was duplicated prior to the 1q jumping or inserting itself into a new location. These cases demonstrate that satII/III DNA sequences are not only associated not only with the duplication of adjacent distal chromosome segments after translocation, but are also associated with the duplication and jumping/insertion of proximal nonhomologous chromosome segments. We have designated this type of instability as a jumping segmental duplication.