Serum BCMA levels predict outcomes in MGUS and smoldering myeloma patients.

Soluble BCMA (sBCMA) levels are elevated in monoclonal gammopathy of undetermined significance (MGUS) and smoldering multiple myeloma (SMM). However, the association between sBCMA levels and prognosis in MGUS and SMM has not been studied. We retrospectively analyzed sBCMA levels in stored samples from 99 MGUS and 184 SMM patients. Baseline sBCMA levels were significantly higher in MGUS and SMM patients progressing to MM during clinical follow up. When stratified according to the median baseline sBCMA level for each cohort, higher levels were associated with a shorter PFS for MGUS (HR 3.44 comparing sBCMA ≥77 vs <77 ng/mL [95% CI 2.07-5.73, p < 0.001] and SMM (HR 2.0 comparing sBCMA ≥128 vs <128 ng/mL, 95% 1.45-2.76, p < 0.001) patients. The effect of sBCMA on PFS was similar even after adjusting for the baseline MGUS or SMM risk stratification. We evaluated paired serum samples and found that sBCMA increased significantly in MGUS and SMM patients who eventually progressed to MM, whereas among MGUS non-progressors the sBCMA level remained stable. While our results require independent validation, they suggest that sBCMA may be a useful biomarker to identify MGUS and SMM patients at increased risk of progression to MM independent of the established risk models.

Kaposi's sarcoma-associated herpesvirus infection of bone marrow dendritic cells from multiple myeloma patients.

Kaposi's sarcoma-associated herpesvirus (KSHV) was found in the bone marrow dendritic cells of multiple myeloma patients but not in malignant plasma cells or bone marrow dendritic cells from normal individuals or patients with other malignancies. In addition the virus was detected in the bone marrow dendritic cells from two out of eight patients with monoclonal gammopathy of undetermined significance (MGUS), a precursor to myeloma. Viral interleukin-6, the human homolog of which is a growth factor for myeloma, was found to be transcribed in the myeloma bone marrow dendritic cells. KSHV may be required for transformation from MGUS to myeloma and perpetuate the growth of malignant plasma cells.

Interference with nuclear factor kappa B and c-Jun NH2-terminal kinase signaling by TRAF6C small interfering RNA inhibits myeloma cell proliferation and enhances apoptosis.

The tumor necrosis factor receptor (TNFR)-associated factor (TRAF) family of six adaptor proteins (TRAF1-6) links the TNFR superfamily to the nuclear factor kappa B (NF-kappaB) and activator protein-1 (AP-1) transcriptional activators. Unlike other TRAFs, TRAF6 is also involved in Toll-like/interleukin (IL)-1 receptor (TIR) signal transduction. Thus, inhibition of TRAF6 function could interrupt both CD40 (TNFR family) and IL-1 growth signals, pathways critical to myeloma proliferation. To block TRAF6-mediated IL-1 signaling, we constructed small interfering RNA (siRNA) against TRAF6. We found that siRNA targeting the TRAF6 C-terminal (siTRAF6C) receptor interaction domain specifically reduced only TRAF6 protein expression, without affecting TRAF2 or 5 levels, and substantially interfered with IL-1-induced NF-kappaB and c-Jun/AP-1 activation. Inhibition by siTRAF6C was concentration-dependent. SiTRAF6C also significantly reduced myeloma proliferation and enhanced apoptosis in a similar dose-dependent fashion in vitro. More importantly, marked siTRAF6C growth inhibition was detected in vivo when these cells were implanted into the bone marrow of irradiated normal mice. In contrast, introduction of siRNA derived from the TRAF6 Zn-finger domain or an irrelevant siRNA construct failed to alter cell growth or cell death. These studies suggest that TRAF6 may be a new molecular target to block cell signal transduction important for the survival and proliferation of multiple myeloma cells.

Identification of malignant cells in multiple myeloma bone marrow with immunoglobulin VH gene probes by fluorescent in situ hybridization and flow cytometry.

Because it has been difficult to identify and separate malignant cells in human lymphoid malignancies, we have developed a flow cytometry-based fluorescent in situ hybridization (FISH) technique using immunoglobulin (Ig) heavy chain variable region (VH) gene probes. After obtaining the specific VH gene sequence expressed by the multiple myeloma IM-9 cell line and the malignant cells in five multiple myeloma patients, sense and antisense biotinylated single-stranded RNA probes were prepared by transcription from the malignant clone's VH DNA sequences. The cells from the IM-9 cell line and from the mononuclear bone marrow cells of multiple myeloma patients were fixed, hybridized with the above biotinylated RNA probes, incubated with streptavidin-phycoerythrin, and analyzed by FACS analysis. The myeloma cells stained positive with their own specific antisense VH biotinylated RNa probes, whereas sense and irrelevant antisense biotinylated probes demonstrated only background staining. Dilutional concentrations of the IM-9 cell line with normal bone marrow cells were also accurately quantitated by this procedure. The application of this technique will allow a more accurate assessment of tumor burden in patients with multiple myeloma and should permit an accurate method of tumor cell purification for clinical as well as biological studies. Furthermore, this technological advance should be equally effective at identifying specific VH gene-expressing cells in other lymphoid malignancies, as well as in nonmalignant B cell disorders.

Resistance of human ovarian cancer cells to tumor necrosis factor and lymphokine-activated killer cells: correlation with expression of HER2/neu oncogenes.

Since overexpression of HER2/neu oncogenes in breast cancer cells is associated with resistance to the cytotoxic effect of tumor necrosis factor (TNF), we investigated whether this correlation also existed for ovarian cancer targets. Nine continuously cultured human ovarian cancer lines were studied and compared to 3 breast cancer lines. Three of the ovarian and 1 breast cancer line demonstrated amplified HER2/neu genes by Southern analysis, increased HER2/neu RNA by Northern analysis, and marked immunoperoxidase staining for HER2/neu protein. The other 8 lines contained unamplified genes and undetectable RNA and protein. All 4 overexpressed lines were relatively resistant to the cytotoxic effects of TNF. Interestingly, they were also resistant to lymphokine-activated killer cells. In contrast, 7 of 8 nonexpressed lines showed sensitivity to TNF and all 8 were sensitive to lymphokine-activated killer cells. There was no difference in sensitivity to lysis by hydrogen peroxide or peptide defensins between over- and nonexpressed lines. These data indicate that expression of HER2/neu oncogenes may impart a proliferative advantage in tumor cells due to induction of resistance to several different cytotoxic mechanisms.

BCL-X expression in multiple myeloma: possible indicator of chemoresistance.

Because murine myeloma plasma cells and normal human lymph node plasma cells express BCL-X, we evaluated BCL-X expression in malignant human plasma cells. BCL-X expression was detected in several human myeloma cell lines, as well as in CD38-sorted bone marrow cells obtained from some patients. Only the antiapoptotic long form of BCL-X (BCL-X-L), was detected. Because BCL-X-L expression can protect tumor cells from apoptotic death induced by chemotherapeutic agents, we tested the clinical relevance of expression in 55 archival bone marrow biopsies. The biopsies were stained by immunohistochemistry, and BCL-X expression was correlated with the subsequent response to treatment. BCL-X expression in malignant plasma cells strongly correlated with decreased response rates in patient groups treated with either melphalan and prednisone or vincristine, Adriamycin, and dexamethasone. Response rates were 83-87% in non-BCL-X-expressing cases and 20-31% in BCL-X-expressing cases. In addition, BCL-X expression was more frequent in specimens taken from patients at relapse (77%), when compared to those at initial diagnosis (29%). Further support for the association of drug resistance with BCL-X-L expression came from studies of the 8226 dox-40 cell line. This line, which expresses p-glycoprotein and serves as a model of multidrug resistance in multiple myeloma cells, demonstrated an up-regulated expression of BCL-X-L, which was relatively specific, in that BCL-2 or BAX expression was not altered. In addition, dox-40 cells demonstrated a generalized resistance to apoptosis that was induced by several different agents. These results indicate that malignant plasma cells can express BCL-X-L and that such expression may be a marker of chemoresistant disease.

Frequent amplification of the bcl-1 locus in head and neck squamous cell carcinomas.

The bcl-1 locus on chromosome 11 at band q13 has been shown to be rearranged in some chronic B lymphoid malignancies with the t(11;14)(q13;q32). Chromosome 11 abnormalities also occur in squamous cell carcinomas including tumors from the head and neck region. In this investigation, we analysed the arrangement, copy number and expression of this locus in 23 fresh head and neck cancers and six established head and neck squamous cell carcinoma cell lines. Bcl-1 was found to be amplified two- to 10-fold in eight fresh tumors and two cell lines, UM-SCC-8 and UM-SCC-22A. Moreover, amplification was more frequently observed in poorly differentiated fresh tumors. None of the fresh tumors or cell lines showed rearrangement of bcl-1. In addition, these tumors and cell lines did not contain amplification of the c-myc or Ha-ras oncogenes.

Oncogenes in multiple myeloma: point mutation of N-ras.

Alterations of ras, c-myc and bcl-1 have been described in hematologic malignancies of lymphoid origin. We investigated the structure of these genes and evaluated the frequency of point mutations involving H-, K- or N-ras in bone marrow samples from patients with multiple myeloma. No abnormalities were detected in the c-myc and bcl-1 genes, but two of 17 patients were found to have N-ras mutations by differential oligonucleotide hybridization and dideoxynucleotide sequencing following amplification by polymerase chain reaction. Bone marrow DNA from both patients had identical missense mutations of N-ras codon 61 changing CAA to AAA, resulting in a substitution of lysine for glutamine in the encoded protein. Multiple myeloma is the first mature B cell neoplasm found to harbor ras mutations.

Frequent amplification of the bcl-1 locus in poorly differentiated squamous cell carcinoma of the lung. The Lung Cancer Study Group.

The bcl-1 locus on chromosome 11 at band q13 has been shown to be rearranged in some chronic B lymphoid malignancies with the t(11;14)(q13;q32). Chromosome 11 abnormalities occur in solid tumors including squamous cell cancers and adenocarcinomas. We have recently reported the frequency amplification of the bcl-1 locus in squamous cell carcinomas of head and neck origin, and increased copy number also has been found in some breast adenocarcinomas. In this investigation, we analyzed the arrangement, copy number and expression of this locus in 111 primary non-small cell lung cancers. Bcl-1 was found to be amplified three- to 10-fold in seven fresh tumors. Whereas none of 51 adenocarcinomas showed bcl-1 amplification, six of 46 squamous cell carcinomas revealed an increase in bcl-1 copy number. This amplification was more frequently associated with larger and more poorly differentiated tumors. None of the cancers showed rearrangement of this locus.

American Society of Clinical Oncology guideline on the role of bisphosphonates in breast cancer. American Society of Clinical Oncology Bisphosphonates Expert Panel.

PURPOSE: To determine clinical practice guidelines for the use of bisphosphonates in the prevention and treatment of bone metastases in breast cancer and their role relative to other therapies for this condition. METHODS: An expert multidisciplinary panel reviewed pertinent information from the published literature and meeting abstracts through May 1999. Additional data collected as part of randomized trials and submitted to the United States Food and Drug Administration were also reviewed, and investigators were contacted for more recent information. Values for levels of evidence and grade of recommendation were assigned by expert reviewers and approved by the panel. Expert consensus was used if there were insufficient published data. The panel addressed which patients to treat and when in their course of disease, specific drug delivery issues, duration of therapy, management of bony metastases with other therapies, and the public policy implications. The guideline underwent external review by selected physicians, members of the American Society of Clinical Oncology (ASCO) Health Services Research Committee, and the ASCO Board of Directors. RESULTS: Bisphosphonates have not had an impact on the most reliable cancer end point: overall survival. The benefits have been reductions in skeletal complications, ie, pathologic fractures, surgery for fracture or impending fracture, radiation, spinal cord compression, and hypercalcemia. Intravenous (IV) pamidronate 90 mg delivered over 1 to 2 hours every 3 to 4 weeks is recommended in patients with metastatic breast cancer who have imaging evidence of lytic destruction of bone and who are concurrently receiving systemic therapy with hormonal therapy or chemotherapy. For women with only an abnormal bone scan but without bony destruction by imaging studies or localized pain, there is insufficient evidence to suggest starting bisphosphonates. Starting bisphosphonates in patients without evidence of bony metastasis, even in the presence of other extraskeletal metastases, is not recommended. Studies of bisphosphonates in the adjuvant setting have yielded inconsistent results. Starting bisphosphonates in patients at any stage of their nonosseous disease, outside of clinical trials, despite a high risk for future bone metastasis, is currently not recommended. Oral bisphosphonates are one of several options which can be used for preservation of bone density in premenopausal patients with treatment-induced menopause. The panel suggests that, once initiated, IV bisphosphonates be continued until evidence of substantial decline in a patient's general performance status. The panel stresses that clinical judgment must guide what is a substantial decline. There is no evidence addressing the consequences of stopping bisphosphonates after one or more adverse skeletal events. Symptoms in the spine, pelvis, or femur require careful evaluation for spinal cord compression and pathologic fracture before bisphosphonate use and if symptoms recur, persist, or worsen during therapy. The panel recommends that current standards of care for cancer pain, analgesics and local radiation therapy, not be displaced by bisphosphonates. IV pamidronate is recommended in women with pain caused by osteolytic metastasis to relieve pain when used concurrently with systemic chemotherapy and/or hormonal therapy, since it was associated with a modest pain control benefit in controlled trials. CONCLUSION: Bisphosphonates provide a meaningful supportive but not life-prolonging benefit to many patients with bone metastases from cancer. Further research is warranted to identify clinical predictors of when to start and stop therapy, to integrate their use with other treatments for bone metastases, to identify their role in the adjuvant setting in preventing bone metastases, and to better determine their cost-benefit consequences.

Purging of autologous peripheral-blood stem cells using CD34 selection does not improve overall or progression-free survival after high-dose chemotherapy for multiple myeloma: results of a multicenter randomized controlled trial.

PURPOSE: Although high-dose chemotherapy supported by autologous peripheral-blood progenitor-cell (PBPC) transplantation improves response rates and survival for patients with multiple myeloma, all patients eventually develop progressive disease after transplantation. It has been hypothesized that depletion of malignant plasma cells from autografts may improve outcome by reducing infused cells contributing to relapse. PATIENTS AND METHODS: A randomized phase III study using the CEPRATE SC System (Cellpro, Bothell, WA) to enrich CD34(+) autograft cells and passively purge malignant plasma cells was completed in 190 myeloma patients randomized to receive an autograft of CD34-selected or unselected PBPCs. RESULTS: After CD34 selection, tumor burden was reduced by 1.6 to 6.0 logs (median, 3.1), with 54% of CD34-enriched products having no detectable tumor. Median time to count recovery, number of transfusions, transplantation-related mortality, and days in hospital were equivalent between the two transplantation arms. With a median follow-up of 37 months, 33 patients (36%) in the selected and 34 patients (35%) in the unselected arm had died (P =.784). Median overall survival in the selected arm was reached at 50 months and is not reached at this time in the unselected arm (P =.78). Median disease-free survival was 100 versus 104 weeks (P =.82), with 67% of patients in the selected arm and 66% of patients in the unselected arm relapsing. CONCLUSION: This phase III trial demonstrates that although CD34 selection significantly reduces myeloma cell contamination in PBPC collections, no improvement in disease-free or overall survival was achieved.

Long-term pamidronate treatment of advanced multiple myeloma patients reduces skeletal events. Myeloma Aredia Study Group.

PURPOSE: To determine the efficacy and safety of 21 monthly cycles of pamidronate therapy in patients with advanced multiple myeloma. PATIENTS AND METHODS: Patients with stage III myeloma and at least one lytic lesion received either placebo or pamidronate 90 mg intravenously administered as a 4-hour infusion monthly for 21 cycles. At study entry, the patients were stratified according to whether they were to receive first-line (stratum 1) or second-line (stratum 2) antimyeloma chemotherapy. Skeletal events (pathologic fracture, radiation or surgery to bone, and spinal cord compression) and hypercalcemia were assessed monthly. RESULTS: The results of the first nine previously reported cycles are extended to 21 cycles. Of the 392 randomized patients, efficacy could be evaluated in 198 who received pamidronate and 179 who received placebo. After 21 cycles, the proportion of patients who developed any skeletal event was lower in the pamidronate-group (P = .015). The mean number of skeletal events per year was less in the pamidronate-group (1.3) than in placebo-treated patients (2.2; P = .008). Although survival was not different between the pamidronate-treated group and placebo patients overall, stratum 2 patients who received pamidronate lived longer than those who received placebo (14 v 21 months, P = .041). Pamidronate was safe and well tolerated during the 21 cycles of therapy. CONCLUSION: Long-term monthly infusions of pamidronate as an adjunct to chemotherapy are superior to chemotherapy alone in reducing skeletal events in stage III multiple myeloma patients, and may improve the survival of patients on salvage therapy.

Multiple myeloma clones are derived from a cell late in B lymphoid development.

We have previously demonstrated that the immunoglobulin (Ig) heavy chain variable region (VH) sequences expressed by the malignant clone in multiple myeloma (MM) contain a high degree of somatic mutation without clonal diversity. This sequence can be used to identify all members of the malignant clone in this B cell malignancy. We sequenced the variable regions expressed by patients with MM and generated primers from the complementarity determining region (CDR) sequences specific for each patient's tumor. Using these primers, we performed PCR amplification on highly purified subpopulations of cells separated by expression of CD10, CD34 and CD38. The results of these experiments demonstrate: 1) there is a small fraction of CD10-expressing tumor cells in MM patients, 2) CD34-bearing malignant cells do not exist in MM, and 3) although the vast amount of tumor is in the CD38-expressing cells, a small amount of tumor is in the CD38-negative population. We also used these primers to determine whether pre-class switch (i.e., Cmu-expressing lymphocytes) clonal cells exist in these patients. After PCR amplification with CDR1 and Cmu primers, colony hybridization was performed using both framework 3 (FR3) and CDR3 probes. Out of > 200 FR3-hybridizing colonies, < or = 5 colonies also hybridized with the CDR3 probe. Colonies which hybridized with both these probes were sequenced, and none of these sequences matched even closely the CDR3 expressed by the malignant clone. These results make the existence of a pre-class switch malignant cell unlikely in MM. Overall, these results suggest that the malignant clone in MM derives from a cell late in B lymphocyte development.

Clonal rearrangement of the beta-T cell receptor gene in multiple myeloma.

Because clonal rearrangements of the beta-T cell receptor (beta-TCR) gene occur in some patients with B cell chronic lymphocytic leukemia, we studied the arrangement of this gene in fourteen patients with multiple myeloma, a malignancy of the most terminally differentiated B cells. The gene was in germline configuration in peripheral blood lymphocytes (PBLs) and bone marrow samples of thirteen patients. By contrast, it was clonally rearranged in the marrow but not in the PBLs of one patient with stage IIA IgA-lambda myeloma. This patient's bone marrow consisted of 95% morphologically identifiable plasma cells which were CALLA-, OKT10+ (93%), and PCA-1+ (78%). Only 5% of marrow cells were small lymphocytes which contained T cell markers (CD3+ or CD2+). To eliminate the possibility that the small percentage of contaminating T cells contained the gene rearrangement, they were depleted by avidin-biotin immunoadsorption using the Leu4 determinant. Positively selected marrow T cells did not contain beta-TCR gene rearrangements. By contrast, the T cell depleted marrow contained the rearranged gene. This is the first demonstration that rearranged beta-TCR genes can occur in multiple myeloma.

Multiple myeloma: the cells of origin--a two-way street.

Multiple myeloma results from an interplay between the monoclonal malignant plasma cells and supporting nonmalignant cells in the bone marrow. Recent studies suggest that the final transforming event in this B cell disorder occurs at a late stage of B cell differentiation based on the characteristics of the immunoglobulin genes expressed by the malignant clone as well as surface markers present on the tumor cells. Recently, an increasing pathogenic role in this malignancy by the nonmalignant cells in the bone marrow has been suggested by several studies. Specific infection of these supporting cells by the recently identified Kaposi's sarcoma-associated herpes virus (KSHV) suggests a novel mechanism by which this nonmalignant population may lead to the development of this B cell malignancy and support its growth.

In vitro cytoreductive effects on multiple myeloma cells induced by bisphosphonates.

In a double blind randomized study, the bisphosphonate drug Pamidronate (Aredia) significantly protected Durie-Salmon stage III multiple myeloma patients from osteolytic bone disease. In the patient sub-group on salvage chemotherapy. Pamidronate treatment was also significantly associated with prolonged survival. To test if this drug could induce direct antitumor effects, we exposed myeloma cells to increasing concentrations of Pamidronate or a more potent bisphosphonate, Zoledronate. A concentration- and time-dependent cytotoxic effect was detected on four of five myeloma cell lines as well as three specimens obtained directly from myeloma patients. Zoledronate-induced cytotoxicity was significantly greater than that of Pamidronate. Cytotoxicity could not be explained by bisphosphonate-induced chelation of extracellular calcium or secondary decrease in production of the myeloma growth factor interleukin-6. Morphological examination, DNA electrophoresis and cell cycle analysis indicated that the bisphosphonate-induced cytotoxic effect consisted of a combination of cytostasis and apoptotic myeloma cell death. Enforced expression of BCL-2 protected against the apoptotic death but not against cytostasis. Most cytotoxic effects were seen between 10 and 100 microM of drug. The results suggest a possible direct anti-tumor effect in myeloma patients treated with bisphosphonates which may participate in their significantly increased survival. This hypothesis should now be further tested in clinical trials.

A CD10-positive subset of malignant cells is identified in multiple myeloma using PCR with patient-specific immunoglobulin gene primers.

Immunophenotypic studies show the presence of CD10-bearing malignant cells in a small subset of multiple myeloma (MM) patients. We used a sensitive PCR-based technique in order to determine the frequency that MM patients contain a malignant subpopulation which expresses this antigen. The immunoglobulin (Ig) heavy chain variable region (VH) gene sequence expressed by the malignant clone in MM can be used as a tumor specific marker. After determining this sequence in six MM patients, patient specific VH oligonucleotide primers from complementarity determining region (CDR) sequences were generated. Bone marrow mononuclear cells from these patients were incubated with two different anti-CD10 antibodies or isotype identical murine IgG controls. Cells were then sorted by flow cytometry into the 1% brightest cells containing > 99.99% CD10-positive cells and two fractions including the 90 and 10% dimmest staining cells. PCR amplification was performed on DNA from approximately 10(4) cells (0.1 microgram) using patient specific CDR1 and CDR3 primers. Detectable PCR product was obtained in each sorted sample although the intensity of the band was much higher in cells lacking CD10 expression (the 90 and 10% dimmest fractions) than in the CD10-bearing (1% brightest) population. These results imply that there is a small population of CD10-bearing clonal cells in most, if not all patients with MM.

Frequent demonstration of human herpesvirus 8 (HHV-8) in bone marrow biopsy samples from Turkish patients with multiple myeloma (MM).

In order to investigate the frequency of HHV-8 in MM patients from another geographic location, we obtained fresh bone marrow (BM) biopsies from Turkish patients with MM (n = 21), monoclonal gammopathy of undetermined significance (MGUS) (n = 2), plasmacytoma (n = 1) with BM plasma cell infiltration, various hematological disorders (n = 6), and five healthy Turkish controls. The frequency of HHV-8 was analyzed by polymerase chain reaction (PCR) in two independent laboratories in the USA and in Turkey. Using fresh BM biopsies, 17/21 MM patients were positive for HHV-8 whereas all five healthy controls, and six patients with other hematological disorders were negative. Two patients with MGUS, and one patient with a solitary plasmacytoma were also negative. The data from the two laboratories were completely concordant. Also using primer pairs for v IRF and v IL-8R confirmed the results observed with the KS330233 primers. Furthermore, sequence analysis demonstrated a C3 strain pattern in the ORF26 region which was also found in MM patients from the US. Thus, HHV-8 is present in the majority of Turkish MM patients, and the absence of the virus in healthy controls further supports its role in the pathogenesis of MM.

A phase I dose-ranging trial of monthly infusions of zoledronic acid for the treatment of osteolytic bone metastases.

Bisphosphonates are potent inhibitors of bone resorption and provide a therapeutic benefit for patients with bone metastases. Zoledronic acid is a highly potent, nitrogen-containing bisphosphonate. In the present trial, we assessed the safety and tolerability of increasing doses of zoledronic acid and its effects on urinary markers of bone resorption in cancer patients with bone metastases. Fifty-nine cancer patients with bone metastases were enrolled sequentially into one of 8 treatment groups in the core protocol. Each patient received a 5-min i.v. infusion of 0.1, 0.2, 0.4, 0.8, 1.5, 2, 4, or 8 mg zoledronic acid monthly for 3 months. Patients were monitored for clinical findings, adverse events, electrocardiograms, markers of bone resorption, as well as routine hematology, blood chemistries, and urinalysis. Thirty patients who demonstrated a radiographic response to treatment or stable disease in the core protocol were enrolled in a humanitarian extension protocol and continued to receive monthly infusions. Zoledronic acid was well tolerated at all dose levels. Adverse events reported by >10% of patients included skeletal pain, nausea, fatigue, upper respiratory tract infection, constipation, headache, diarrhea, and fever. Three patients in the core protocol and one patient in the extension protocol experienced grade 3 skeletal pain, "flu-like" symptoms, or hypophosphatemia, which were possibly related to treatment; all recovered completely. Adverse events were reported with similar frequency across all of the dosage groups. Zoledronic acid resulted in sustained, dose-dependent decreases in urinary markers of bone resorption. Zoledronic acid was safe and well tolerated and demonstrated potent inhibition of bone resorption.

Human herpesvirus 8 open reading frame 26 and open reading frame 65 sequences from multiple myeloma patients: a shared pattern not found in Kaposi's sarcoma or primary effusion lymphoma.

Human herpesvirus 8 (HHV-8), also known as Kaposi's sarcoma-associated herpesvirus, has been implicated in the pathogenesis of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), multicentric Castleman's disease, and recently multiple myeloma (MM). DNA sequence analyses of HHV-8 suggest that multiple HHV-8 strains exist. We extracted DNA from 24 patients with MM and 3 patients with monoclonal gammopathy of undetermined significance and compared HHV-8 open reading frames (ORFs) 26 and 65 sequences with those derived from patients with KS, PEL, and two HHV-8-positive PEL cell lines KS-1 and BC-1. ORF26 sequence data suggest that MM patients are consistently carriers of HHV-8 strain subtype C3. All MM patients also consistently revealed either a single bp deletion or substitution at position 112197 in ORF65. This unique alteration is not present in patients with KS or PEL or in PEL cell lines. It occurs in the portion of ORF65 that is known to be responsible for a serological response to HHV-8.