Lack of association of tumor-associated macrophages with clinical outcome in patients with classical Hodgkin's lymphoma.

BACKGROUND: A recent study demonstrated that an increased number of CD68+ macrophages were correlated with primary treatment failure, shortened progression-free survival (PFS) and disease-specific survival (DSS) in patients with classical Hodgkin's lymphoma (cHL). PATIENTS AND METHODS: The aim of the present study was to verify the relationship between the number of CD68+ and CD163+ macrophages with clinical outcomes in a cohort of 265 well-characterized patients with cHL treated uniformly with the standard doxorubicin, bleomycin, vinblastine and dacarbazine chemotherapy regimen. Two pairs of hematopathologists carried out independent pathological evaluations of tissue microarray slides. RESULTS: There were no associations between clinical characteristics and the expression of CD68 or CD163. However, higher levels of CD68 and CD163 expression were correlated with the presence of Epstein-Barr virus-positive Hodgkin tumor cells (P = 0.01 and 0.037, respectively). The expression of CD68 or CD163 was not associated with either the PFS or the DSS. CONCLUSION: CD68 and CD163 expression require further evaluation before their use can be recommended for prognostic stratification of patients with cHL.

High incidence of false-positive PET scans in patients with aggressive non-Hodgkin's lymphoma treated with rituximab-containing regimens.

BACKGROUND: Positron emission tomography (PET) is a powerful predictor of relapse and survival in non-Hodgkin's lymphomas (NHLs) based on studies carried out in the prerituximab era. Little is known about the predictive power of PET in rituximab-treated patients. PATIENTS AND METHODS: Patients with aggressive B-cell NHL with baseline and follow-up PET studies were included. Clinical characteristics, PET and computed tomography scans, biopsy results, and outcomes were reviewed. PET was defined as positive if higher than mediastinal or background activity was observed. RESULTS: In all, 51 patients (diffuse large B cell-38; mantle cell lymphoma-13) treated with rituximab-containing regimens were included. For 13 of 40 patients (32.5%), mid-therapy PET studies were positive and 9 of 48 patients (18.7%) had positive posttherapy PET. The positive predictive value (PPV), negative predictive value (NPV), sensitivity (Se), and specificity (Sp) of the mid-therapy PET for predicting relapse were 33% [95% confidence interval (CI) 19% to 49%], 68% (95% CI 51% to 81%), 33% (95% CI 6% to 76%), and 68% (95% CI 49% to 82%), respectively. For posttherapy PET, the relapse PPV, NPV, Se and Sp were 19% (95% CI 9% to 33%), 81% (95% CI 67% to 91%), 13% (95% CI 0.6% to 53%), and 80%(95% CI 64% to 90%), respectively. CONCLUSIONS: Compared with previous reports in prerituximab era, addition of rituximab resulted in reduced PPV and sensitivity of mid- and posttherapy PET in patients with aggressive B-cell NHL.

Molecular impact of selective NFKB1 and NFKB2 signaling on DLBCL phenotype.

Diffuse large B-cell lymphoma (DLBCL) has been categorized into two molecular subtypes that have prognostic significance, namely germinal center B-cell like (GCB) and activated B-cell like (ABC). Although ABC-DLBCL has been associated with NF-κB activation, the relationships between activation of specific NF-κB signals and DLBCL phenotype remain unclear. Application of novel gene expression classifiers identified two new DLBCL categories characterized by selective p100 (NF-κB2) and p105 (NF-κB1) signaling. Interestingly, our molecular studies showed that p105 signaling is predominantly associated with GCB subtype and histone mutations. Conversely, most tumors with p100 signaling displayed ABC phenotype and harbored ABC-associated mutations in genes such as MYD88 and PIM1. In vitro, MYD88 L265P mutation promoted p100 signaling through TAK1/IKKα and GSK3/Fbxw7a pathways, suggesting a novel role for this protein as an upstream regulator of p100. p100 signaling was engaged during activation of normal B cells, suggesting p100's role in ABC phenotype development. Additionally, silencing p100 in ABC-DLBCL cells resulted in a GCB-like phenotype, with suppression of Blimp, IRF4 and XBP1 and upregulation of BCL6, whereas introduction of p52 or p100 into GC cells resulted in differentiation toward an ABC-like phenotype. Together, these findings identify specific roles for p100 and p105 signaling in defining DLBCL molecular subtypes and posit MYD88/p100 signaling as a regulator for B-cell activation.

Apoptosis-stimulating protein of p53-2 (ASPP2/53BP2L) is an E2F target gene.

The p53 pathway is a central apoptotic regulator. Deregulation of the Rb/E2F pathway occurs in a majority of tumors, resulting in both unrestrained proliferation and enhanced apoptosis sensitivity via p53-dependent and independent mechanisms. However, the mechanisms coupling the p53 and Rb/E2F pathways remain incompletely understood. We report that ASPP2/53BP2L, a p53/p73-binding protein that promotes p53/p73-dependent apoptosis, is an E2F target gene. The ASPP2/53BP2L promoter was identified and ectopic expression of transcription-competent E2F-1 (E2F-2 and E2F-3) stimulated an ASPP2/53BP2L promoter-luciferase reporter. Mutational analysis of the ASPP2/53BP2L promoter identified E2F-binding sites that cooperate for E2F-1 induction and basal repression of ASPP2/53BP2L. Moreover, endogenous ASPP2/53BP2L levels increased after E2F-1 expression, and E2F-1 bound the endogenous ASPP2/53BP2L promoter after chromatin immunoprecipitation. Typical for an E2F target, ASPP2/53BP2L expression was maximal in early S-phase. Thus, ASPP2/53BP2L is downstream of E2F, suggesting that it functions as a common link between the p53/p73 and Rb/E2F apoptotic pathways.

AID is expressed in germinal center B-cell-like and activated B-cell-like diffuse large-cell lymphomas and is not correlated with intraclonal heterogeneity.

Activation-induced cytidine deaminase (AID), highly expressed in germinal center (GC)-lymphocytes, is involved in somatic hypermutation (SHM). We examined AID expression in diffuse large B-cell lymphomas (DLBCL) of germinal center B-cell (GCB)-like and activated B-cell (ABC)-like subtypes. These two types of DLBCL are characterized by high and low expression of GC-specific genes, respectively. AID expression was detected in both GCB- and ABC-like DLBCL, thus demonstrating a dissociation between AID expression and that of other GC genes. We also tested for the presence of intraclonal heterogeneity in immunoglobulin and BCL6 genes in those same tumors and in follicle center lymphomas (FCL) that transformed to DLBCL. The level of AID expression did not correlate with the presence of intraclonal sequence heterogeneity in either IgV(H) or BCL6. Our findings suggest that lymphomas maintain some but not all of the gene expression signatures of their normal B-cell counterparts. The fact that AID expression can be elevated without intraclonal sequence heterogeneity raises the possibility that other factors are required for SHM in these tumors. We found decreased levels of AID expression in DLBCL that evolved from FCL and which had acquired new mutations in their BCL6 genes. This dissociation suggests that AID expression and SHM may occur at the time prior to the clinical detection of transformed lymphoma.

BCL-6 mRNA expression in higher grade transformation of follicle center lymphoma: correlation with somatic mutations in the 5' regulatory region of the BCL-6 gene.

Follicle center lymphoma (FCL) is an indolent low-grade B cell non-Hodgkin's lymphoma (NHL) that frequently transforms to aggressive diffuse large B cell lymphoma (DLBCL). Histological transformation of FCL is commonly associated with accumulation of secondary genetic alterations. The BCL-6 gene is commonly implicated in the pathogenesis of DLBCL and its expression may be altered by clonal rearrangements and somatic point mutations in its 5' non-translated regulatory region. Recently, somatic mutations of the BCL-6 gene were associated with the transformation process. Here, we examined BCL-6 mRNA expression and BCL-6 mutations in paired biopsies from the same patients obtained at the time of FCL diagnosis and after transformation. BCL-6 mRNA expression markedly increased upon transformation (1.9- to 4.8-fold) in three cases, remained unchanged in one case and decreased compared to the diagnosis FCL specimens in four cases. The three specimens that demonstrated an increase in the BCL-6 mRNA expression upon transformation harbored BCL-6 gene mutations in the 5' region of the first intron that overlapped with the previously reported negative regulatory region of the gene. Accumulation of new mutations in this region was not observed in DLBCL biopsies in which the BCL-6 mRNA expression did not increase. The present study demonstrates that although BCL-6 gene mutations do accumulate during the transformation process and, depending on their location within the first intron, may deregulate BCL-6 mRNA expression, increase in BCL-6 mRNA expression is not uniformly required for transformation from FCL to DLBCL.

Optimization of quantitative real-time RT-PCR parameters for the study of lymphoid malignancies.

Real-time quantitative reverse transcription polymerase chain reaction (RT-PCR) is a powerful method for measurement of gene expression for diagnostic and prognostic studies of non-Hodgkin's lymphomas (NHL). In order for this technique to gain wide applicability, it is critically important to establish a uniform method for normalization of RNA input. In this study, we have determined the best method to quantify the RNA/cDNA input per reaction and searched for the most useful endogenous control genes for normalization of the measurements, based on their abundance and lowest variability between different types of lymphoid cells. To accomplish these aims, we have analyzed the RNA expression of 11 potential endogenous control genes (glyceraldehyde-3-phosphate dehydrogenase, beta-actin, peptidylprolyl isomerase A, beta 2 microglobulin, protein kinase cGMP-dependent, type I, hypoxanthine phosphoribosyltransferase 1, TATA box binding protein, transferrin receptor, large ribosomal protein, beta-glucoronidase and 18S ribosomal RNA). In all, 12 different B- and T-cell lymphoma/leukemia cell lines, 80 B- and T-cell NHL specimens, and resting and activated normal B and T lymphocytes were screened. Normalization of the nucleic acid input by spectrophotometric OD(260) measurement of RNA proved more reliable than spectrophotometric or fluorometric measurements of cDNA or than electrophoretic estimation of the ribosomal and mRNA fractions. The protein kinase cGMP-dependent, type I (PRKG1) and the TBP genes were expressed at common abundance and exhibited the lowest variability among the cell specimens. We suggest that for further lymphoma studies based on the real-time RT-PCR quantification of gene expression, that RNA input in each reaction be equalized between the specimens by spectrophotometric OD(260) measurements. The expression of the gene of interest in different samples should be normalized by concomitant measurement of the PRKG1 and/or the TBP gene products.

The BCL6 gene in B-cell lymphomas with 3q27 translocations is expressed mainly from the rearranged allele irrespective of the partner gene.

The BCL6 gene, which functions as a transcription repressor, is the target of multiple chromosomal translocations in non-Hodgkin's lymphomas (NHL). These translocations occur in the nontranslated region of the BCL6 gene, juxtaposing regulatory sequences of the diverse partner genes to the open reading frame of the BCL6 gene and thus are thought to deregulate BCL6 gene expression. The levels of expression of the BCL6 gene and protein have been demonstrated to predict the clinical outcome of diffuse large B-cell lymphomas. By contrast, the prognostic significance of BCL6 gene translocations is unclear. In this study we have sought an explanation for this apparent discrepancy. We examined tumors with a variety of different BCL6 translocations and therefore with a variety of potentially substituted promoters. We found no increase in total BCL6 mRNA levels in the NHL specimens harboring BCL6 gene translocation. Indeed, some of these tumors expressed relatively low quantities of the BCL6 mRNA. We also sought to determine whether BCL6 transcription occurs from the rearranged or from the normal untranslocated allele in these tumors. We demonstrate that lymphoma cell lines and majority of NHL tumor specimens expressed BCL6 mRNA predominantly from the rearranged allele that may come under the control of various partner gene promoters. However, few NHL tumors with BCL6 gene translocations expressed BCL6 mRNA equally from the rearranged and the nonrearranged alleles. Neither the nature of the substituted promoters nor the presence of activating mutations in the BCL6 regulatory sequences correlated with the allelic expression of the BCL6 gene in these tumors.

T cell activation following infection of primary follicle center lymphoma B cells with adenovirus encoding CD154.

Purified, high-titer adenovirus encoding murine CD154 (Ad-CD154) or human CD154 (Ad-hCD154) was used to infect lymph node cells isolated from patients with follicle center lymphoma. Infection of lymphoma B cells with Ad-CD154 at a multiplicity of infection (MOI) ratio of 100 or higher resulted in high-level transgene expression. Additionally, upon infection of lymphoma B cells, only Ad-CD154 resulted in surface expression of CD154, despite similar, high-level expression of either human or mouse CD154 by HeLa cells infected with Ad-hCD154 or Ad-CD154, respectively. Moreover, infection of lymphoma B cells with Ad-CD154, but not Ad-hCD154 or adenovirus encoding Eschericheria coli beta-galactosidase (Ad-LacZ), induced the neoplastic B cells to express higher levels of immune co-stimulatory molecules that are required for proficient presentation of antigen to T cells. Consistent with this, we found that Ad-CD154 infected lymphoma B cells could stimulate T cells to proliferate or produce interferon-gamma in allogeneic or autologous mixed lymphocyte interactions. We conclude that lymphoma B cells can be infected with Ad-CD154 and that this significantly enhances their recognition by allogeneic or autologous T cells. As such, Ad-CD154-transduced lymphoma B cells may have potential for the active immune therapy of patients with follicle center lymphoma.

Food-borne outbreak of group A beta-hemolytic streptococcal pharyngitis.

An outbreak of pharyngitis due to group A beta-hemolytic streptococci type T 12 occurred at a military base. An epidemiologic investigation indicated that the outbreak was food borne. Consumption of boiled egg salad at lunch was significantly associated with the illness. Immediate institution of antibiotic therapy and isolation of the patients prevented secondary respiratory spread of the infection. No cases of poststreptococcal suppurative and nonsuppurative complications were found during a 6-week period after the outbreak. Medical personnel should be aware of the possibility of food-borne streptococcal pharyngitis. Regular health surveillance of food handlers and food preparation processes are important for prevention of such outbreaks.

Chlamydophila psittaci-negative ocular adnexal marginal zone lymphomas express self polyreactive B-cell receptors.

The pathogenesis of Chlamydophila psittaci-negative ocular adnexal extranodal marginal zone lymphomas (OAEMZLs) is poorly understood. OAEMZLs are monoclonal tumors expressing a biased repertoire of mutated surface immunoglobulins. Antigenic activation of the B-cell receptor (BCR) may have a role in the pathogenesis of these lymphomas. We have analyzed the reactivity of recombinant OAEMZL immunoglobulins. OAEMZL antibodies reacted with self-human antigens, as demonstrated by enzyme-linked immunosorbent assays, HEp-2 immunofluorescence and human protein microarrays. All the analyzed recombinant antibodies (rAbs) exhibited polyreactivity by comprehensive protein array antibody reactivity and some rAbs also demonstrated rheumatoid factor activity. The identity of several reactive antigens was confirmed by microcapillary reverse-phase high-performance liquid chromatography nano-electrospray tandem mass spectrometry. The tested rAbs frequently reacted with shared intracellular and extracellular self-antigens (for example, galectin-3). Furthermore, these self-antigens induced BCR signaling in B cells expressing cognate surface immunoglobulins derived from OAEMZLs. These findings indicate that interactions between self-antigens and cognate OAEMZL tumor-derived BCRs are functional, inducing intracellular signaling. Overall, our findings suggest that self-antigen-induced BCR stimulation may be implicated in the pathogenesis of C. psittaci-negative OAEMZLs.

Septic arthritis of the glenohumeral joint. A report of 11 cases and review of the literature.

Eleven cases (6 adults and 5 pediatrics) of shoulder septic arthritis are described, and the English literature from 1960 to 1997 reviewed, for a total of 168 cases. Shoulder septic arthritis is an uncommon and difficult diagnosis requiring a high index of suspicion and early evaluation of the affected shoulder by the clinician. The disease usually involves very young infants or elderly patients (65-75 years old). Associated medical conditions were identified in 60% of the patients and include systemic disorders such as liver diseases, alcoholism, and malignancies in 46%; preceding chronic arthritic disorders in 24%; and associated infectious focus in 13% of the patients. Associated infections were more prevalent in the pediatric population. Intravenous drug abuse appears not to constitute a major risk factor; it was identified in less than 5% of patients. All patients presented with acute shoulder ache or with exacerbation of existing chronic pain in joints previously damaged. Elevated body temperature (over 38 degrees C) appeared in 67% of the adult patients and in over 90% of the pediatric patients. Shoulder arthritis was frequently accompanied by an accelerated erythrocyte sedimentation rate that may rise above 100 mm/hr. Increased white blood cell count was found in approximately 40% of patients. The initial X-rays were frequently normal, while ultrasonography supported the diagnosis in some cases by demonstrating accumulation of fluid inside the joint space. Aspiration of synovial fluid from the affected glenohumeral joint was necessary to evaluate the offending pathogen. False-negative Gram stain appeared in approximately 90% of the patients, whereas synovial fluid cultures demonstrated the pathogen in 88% of patients. Blood cultures were positive in 50% of adult patients and 90% of pediatric patients. The most common isolated pathogen was Staphylococcus aureus, which accounted for 41% of infections. Gram-negative bacilli, which accounted for about 20% of infections, are more prevalent in the pediatric population, especially the neonates. Pyogenic shoulder arthritis should first be treated with intravenous antibiotics, effective at least against staphylococcal infections, until the organisms and sensitivities are identified. Duration of antibiotic therapy should be 3-6 weeks. Unfortunately, our experience in addition to the literature summary does not allow statistical analysis and firm conclusions concerning the best therapeutic approach. However, it appears that in the adult population an operative draining procedure is preferred, whereas in the pediatric population, a closed needle aspiration, if needed at all, is the optimal treatment. With prompt antibiotic therapy and drainage of the shoulder, the patient can be expected to improve clinically, with no serious long-term debilitating effects from the disease.

Pacemaker endocarditis. Report of 44 cases and review of the literature.

We conducted a retrospective study to characterize the clinical course, microbiologic spectrum, and risk factors for endocarditis and for associated mortality in a large series of patients with documented pacemaker endocarditis. Using a computerized search through the medical records of 10 major hospitals in Israel from 1982 to 1992, and carefully reviewing the charts, we identified 44 patients with pacemaker endocarditis. The cases were categorized as definite (n = 25), probable (n = 12), or possible (n = 7) infective endocarditis based on strict case definition. Fever and chills were the most common symptoms. Increased ESR, leukocytosis, microscopic hematuria, and anemia were the most common laboratory findings. A relatively high proportion of the patients were diabetic. The most common source of endocarditis was infection acquired by the placement procedure or infection of the pacemaker pouch. Demographic, clinical, and laboratory features were similar to those of endocarditis patients of a similar age range without pacemakers, although the frequency of fever and chills was higher in our patients than in those patients and splenomegaly, vascular embolic phenomena, and new or changing murmurs were rare in our patients. The major pathogens were Staphylococcus aureus and Staphylococcus epidermidis, similar to other series of pacemaker-associated bacteremia and similar to the microbiologic findings of early prosthetic-valve endocarditis. However, this microbiologic profile is different from that of native-valve endocarditis. Although the present series did not show a statistically significant advantage to electrode removal over conservative treatment, when analyzed together with pooled data from other studies, it suggests that the surgical approach is preferable.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacterial pneumonia in recipients of bone marrow transplantation. A five-year prospective study.

Bacterial pneumonia as an important complication of bone marrow transplantation (BMT) has not been subjected to comprehensive analysis. Two hundred fifty-five consecutive allogeneic and autologous BMT recipients, ranging in age from 1 month to 53 years, were prospectively followed for 3 days to 3 years (median, 108 days) for development of bacterial pneumonia. Etiology, place acquired, chest radiography, and outcome were recorded and the association between bacterial pneumonia and demographic and clinical variables was analyzed. Thirty-seven (15%) patients experienced 52 episodes of bacterial pneumonia: onset of 13 episodes occurred within 30 days after transplantation, 10 episodes occurred on days +31 to +100, and 29 episodes occurred thereafter. Bacterial pneumonia was the terminal event or contributed to fatal outcome in 8 patients (22% of bacterial pneumonia cases, 3% total study population). Mortality due to hospital-acquired pneumonia (6/21) was significantly higher than (P = 0.03). Bacterial pathogens were identified in 27 (52%) episodes. During the first 100 days after BMT, hospital-acquired Gram-negative bacteria predominated, caused mainly by Pseudomonas aeruginosa, Klebsiella pneumoniae, Acinetobacter lwoffi, and Enterobacter cloacae. After day +100, community-acquired, Gram-positive bacteria predominated, particularly Streptococcus pneumoniae. Haemophilus influenzae occurred periodically. Considering all episodes, significant association was found between bacterial pneumonia and veno-occlusive disease (VOD) (P < 0.01) and chronic graft-versus-host disease (GVHD) (P < 0.02). For culture-positive episodes, the association between bacterial pneumonia and VOD was significant (P < 0.001) and borderline for acute GVHD (P = 0.07). It is concluded that VOD and GVHD are positively associated with post-BMT bacterial pneumonia. Its incidence, etiology, risk factors, and outcome are important considerations in its prevention and treatment.

On the inheritance of primary spontaneous pneumothorax.

We report on 15 families with familial primary spontaneous pneumothorax detected through a retrospective study of a large group of male patients. Genetic analysis of these families together with 14 families from the literature in which enough details were available suggested 2 possible models of inheritance. First, the familial cases are due to an autosomal dominant gene with incomplete penetrance, the penetrance being lower in females (21%) than in males (50%). Second, the familial cases represent a heterogeneous group of patients: some of the familial cases are due to an X-linked recessive gene and others to an autosomal dominant gene with the incomplete penetrance (35% in females and 50% in males). The second model may explain the excess of female carriers found in the families of the patients and is supported by the existence of clinical differences between the patients in each of the sub-groups. In particular the number of episodes of pneumothorax per patient was significantly higher in the sub-group with the autosomal dominant form than in the sub-group with the X-linked recessive form of the disease.

Spontaneous pneumomediastinum. A report of 25 cases.

Spontaneous pneumomediastinum (SPM) is a relatively uncommon, infrequently reported entity. To determine the clinical presentation and sequelae of SPM, data were obtained from 25 patients: 14 from Hadassah University Hospital, and 11 from other medical institutions. The mean age was 18.8 +/- 5.2 years (+/- SD), with a range of 8 to 31 years. The most common presenting complaint was retrosternal pain in 22 patients (88 percent), dyspnea in 15 (60 percent), dysphagia in 10 (40 percent), and weakness in 10 (40 percent). Predisposing factors for the development of SPM could be identified in 18 patients (72 percent). Information on the sequelae of SPM during a mean period of 87.4 +/- 38.0 months following the initial SPM episode was obtained from 23 patients. Recurrent SPM occurred in 1 patient at 18 months, and another patient experienced 4 episodes of recurrent spontaneous pneumothorax. No other long-term sequelae were reported. We conclude that SPM is a benign self-limited disease with diverse clinical manifestations. Although uncommon, recurrences of SPM may be observed.

Differential diagnosis of pleural effusion by lactate dehydrogenase isoenzyme analysis.

STUDY OBJECTIVE: To determine the diagnostic value of pleural fluid lactate dehydrogenase (LDH) isoenzyme analysis in the differential diagnosis of pleural fluid. PATIENTS AND METHODS: Eighty-seven consecutive patients with pleural effusion caused by congestive heart failure (33), infection (33), and malignancy (21) comprised a derivation set of patients. Pleural fluid LDH activity and isoenzyme pattern were established in all patients and analyzed by the classification and regression trees (CART) method. An additional group of 20 consecutive patients comprised a validation set that was used for cross-validation of CART-derived decision tree. RESULTS: A decision tree, with a positive predictive value of 83%, was constructed and validated by data from a validation set of patients. CONCLUSIONS: Pleural fluid LDH isoenzyme pattern may be helpful for the differential diagnosis of the most common causes of pleural effusions: congestive heart failure, infections, and malignancy.

A novel translocation (1;2)(p34;p21-22) in acute myelomonoblastic leukemia.

A novel and as yet unrecorded translocation, (1;2)(p34;p21-22), detected in a patient with acute myeloid leukemia (AML) is reported. The leukemia--in this case, AML-M4--showed a rapidly progressive fatal course despite an early transient response to aggressive chemotherapy. In this patient, the leukemic cells showed a novel balanced translocation, (1;2)(p34;p21-22), in most of the metaphases at the time of diagnosis and during subsequent relapse. Interferon-inducible double-stranded RNA-dependent protein kinase (ds RNA-PK) is located in the chromosome region, 2p21-22, that was involved in the translocation in this case. The possible role of ds RNA-PK in leukemogenesis is briefly mentioned.

Salvage chemotherapy using a combination of fludarabine and cyclophosphamide for refractory or relapsing indolent and aggressive non-Hodgkin's lymphomas.

The prognosis of patients with refractory or relapsing non-Hodgkin's lymphoma (NHL) after primary therapy is poor and multi-drug salvage treatments are associated with less than 60% response rates, usually of short duration. Here we report the results of a phase II study using a fludarabine-cyclophosphamide (FAMP-Cy) combination as a salvage failure regimen in refractory and relapsing low-grade (6) and intermediate-grade (9) NHL patients. Fifteen patients, who had received up to 4 regimens prior to therapy with FAMP-Cy were treated with fludarabine (25 mg/m2) and cyclophosphamide (300 mg/m2) for 3 consecutive days followed by G-CSF (5 microg/kg). The overall response was 74%, 4 achieving complete responses (CR) and 7 partial responses (PR). All patients with low-grade NHL responded (4 CR, 2 PR); 5 patients with intermediate-grade NHL achieved PR lasting for a median of 5 months. The main toxicity encountered was moderate myelosuppression. Three patients had febrile neutropenia, one had drug-induced fever and a single patient developed severe neurotoxicity. Opportunistic infections due to lymphopenia were not seen. The combination of fludarabine and cyclophosphamide used as a salvage regimen showed an impressive response in a small group of heavily pretreated low-grade NHL patients who had previously received a large number of prior regimens. FAMP-Cy had limited effect in a similar group of intermediate-grade NHL patients. Results with this "failure" regimen are encouraging, however further studies are needed in order to confirm these observations in a larger series of patients.

A polymorphism in the BCL-6 gene is associated with follicle center lymphoma.

Follicle center lymphoma (FCL) accounts for approximately 40% of all non-Hodgkin's lymphomas (NHL). The genetic-environmental interactions involved in the etiology and pathogenesis of this disease are unknown. In our previous study a single nucleotide polymorphism (SNP) (397C) in the regulatory untranslated first intron region of the BCL-6 gene was found in four of the eight FCL patients but in none of the 10 healthy controls. To further evaluate the potential association between the 397C allele of the BCL-6 gene and FCL, we performed a case-control study. Genomic DNA was isolated from 85 FCL patients, from 98 control cases without a previous history of malignancy, treated at Stanford University Medical Center for non-malignant disorders and from 90 samples from the DNA Polymorphism Discovery Resource. The 397G and the 397C polymorphic alleles were identified by a PCR-RFLP method. To evaluate the possible effect of this polymorphism on gene expression, BCL-6 mRNA levels in nine FCL tumors with the 397G-G genotype and in nine FCL tumors with the 397G-C genotype were measured by quantitative real-time RT-PCR. The 397C polymorphic allele was found in 32 FCL cases (37.6%), in 20 controls (20.4%) and in 17 (18.9%) samples from the DNA Polymorphism Discovery Resource. The prevalence of the 397G-C and 397C-C genotypes was significantly higher in FCL cases than in control group (p = 0.01). No difference in BCL-6 gene expression was observed between FCL cases with 397G-G and 397G-C genotypes. The present study demonstrates a possible association between the 397C allele of the BCL-6 proto-oncogene and FCL. The similar levels of BCL-6 mRNA expression in 397G-G and in 397G-C FCL cases suggests that any possible oncogenic effect of the polymorphic allele would not simply be related to a direct effect on BCL-6 gene expression and suggests the existence of other FCL susceptibility genes that are in linkage disequilibrium with the 397C allele of the BCL-6 gene.