Over a period of 28 months, we observed five cases of osteonecrosis of the jaw (ONJ) in cancer patients treated with bisphosphonates (BP) at our institution. This prompted us to undertake a ...retrospective, multicenter study to analyse the characteristics of patients who exhibited ONJ and to define the frequency of ONJ in multiple myeloma (MM). We identified 35 cases in Gruppo Italiano Studio Linfomi centers during the period 2002 - 05. The median time from cancer diagnosis to the clinical onset of ONJ was 70 months. In these 35 cases of ONJ, 24 appeared 20 - 60 months after starting BP treatment. The time for the onset of ONJ was significantly shorter for patients treated with zoledronic acid alone than for those treated with pamidronate followed by zoledronic acid. The frequency of ONJ in the MM group during the study period was 1.9%, although the nature of the present study may have resulted in an underestimation of ONJ cases. Our analysis strongly suggested an association between the use of BP and the occurrence of ONJ, although we were unable to identify any definite risk factors with a retrospective study. The most frequently ONJ-associated clinical characteristics were chemotherapy treatment, steroid treatment, advanced age, female sex, anemia, parodonthopaties dental procedures and thalidomide (in the case of MM patients).
Abstract 2837
Rai and Binet staging systems are not devoid of some limitations, including the lack of evaluation of thoracic and abdominal lymphadenopathies. The widely-used IWCLL guidelines do not ...incorporate use of TB-CT scan in the diagnostic algorithm. In the present study, we investigated whether TB-CT scans could up-stage Binet stage A CLL patients included in the prospective multicenter O-CLL01 GISL study (clinicaltrial.gov ID: NCT00917549), and whether this subgroup presented differences in prognostic markers and in progression-free survival (PFS). To date, 454 patients have been enrolled and TB-CT scans were available in 238 patients. The median age was 60 years (range, 33–71) and 136 (57%) were male. According to Rai, 180 patients were at low risk (stage 0) and 58 at intermediate risk (stages I-II). b2-microglobulin was elevated in 35.5% of cases. Seventy-eight patients (32%) were IgVH unmutated, 108 patients (45%) had a high ZAP-70 expression, 45 patients (19%) were CD38 positive (>30%). FISH data were available in 226/238 cases; the most frequent abnormality was del(13)(q14) (105 pts, 46.5%), followed by trisomy 12 (24 pts, 10.6%), del(11q22.3) (13 pts 5.5%), del(17p13) (4 pts 1.8%) while 80 cases (35.4%) cytogenetics were normal. Cytogenetic abnormalities were clustered in 3 risk groups i.e. low (del(13q14) and normal), intermediate (trisomy 12) and high risk (del(11q22) and del(17p13). Two hundred six out of 238 patients had a minimum follow-up of 6 months and were evaluable for PFS. Considering TB-CT scan, 54 out of 238 analyzed (22.7%) patients converted into Binet stage B. Notably, 63% were male, b2-microglobulin was elevated in 50% of cases, 42.6% were IgVH unmutated, 48.1% had a high ZAP-70 expression, 27.8% were CD38 positive, and 17.6% showed a high-risk FISH. Binet B patients showed a statistically higher rate of cases with high risk cytogenetic abnormalities than Binet A patients (17.6% vs 4.6%; p=0.032). While, no statistically different distribution of gender, age, B2-microglobulin, IgVH mutational status, CD38 or ZAP-70 expression were observed between the two subgroups. After a median follow-up of 24 months 46/206 (22%) evaluable cases showed disease progression. Binet B patients showed a PFS significantly shorter than those with a normal TB-CT (2-years PFS probability, 85.6% vs 68.5%; p<0.0001). According to the Rai classification 102/180 (56.7%) low risk patients were re-defined as intermediate risk with the integration of TB-CT scan. This subset of patients showed a statistically higher rate of cases with elevated ZAP-70 (51.5% vs 35.9%; p=0.049) and CD38 (22.5% vs 10.3%; p=0.045) than patients at low risk. After a median follow-up of 25 months, 23/154 (15%) of evaluable cases showed disease progression. Patients with an intermediate risk Rai stage showed a PFS significantly shorter than those with a low risk (2-years PFS probability, 82% vs 96%; p=0.002). In this setting 70 cases met the diagnostic criteria of monoclonal B-lymphocytosis (<5 × 109/L B- lymphocytes in the blood). With the integration of TB-CT scan 30/70 (42.9%) monoclonal B-lymphocytosis patients were re-defined as intermediate risk according the Rai classification. No statistically different distribution of clinical and biological parameters were observed between cases who remained in the low risk stage and those who became at intermediate risk. After a median follow-up of 28 months 4/57 cases evaluable for PFS showed a disease progression (2 cases for each subgroup). Considering low risk Rai stage, no statistical difference in PFS was observed among nonCT-upstaged MBL, CT-upstaged MBL, nonCT-upstaged Rai 0, while CT-upstaged Rai 0 cases showed a statistically shorter PFS (p<.0001) than the other groups (Figure 1). Finally, TB-CT scan allowed the early identification of a second neoplasia in 2 cases (lung cancer 1 pt, renal cell carcinoma 1 pt). Our preliminary data indicate that the integration of TB-CT scans in the clinical staging allows for an effective clinical discrimination of Binet A CLL cases in approximately 23% of cases at more advanced stages, predicting a worse clinical outcome. However, the use of TB-CT scanning for upstaging is not beneficial for predicting PFS in MBL cases. A longer follow-up will demonstrate whether the inclusion of TB-CT scan in the initial work-up of patients with early-stage CLL will provide clinically relevant prognostic information. Display omitted
Di Raimondo:celgene: Honoraria. Foà:Bristol-Myers Squibb: Consultancy, Membership on an entity’s Board of Directors or advisory committees.
Abstract 1360
The arbitrary cut-off of 5000/μL chronic lymphocytic leukemia (CLL)-phenotype cells in peripheral blood is generally used to separate monoclonal B-cell lymphocytosis (MBL) from CLL. ...However, a major concern is the biological differentiation, if any, between MBL and CLL. We tried to address the issue therefore analyzing 261 Rai stage 0 patients enrolled in a Gruppo Italiano Studio Linfomi (GISL) prospective multicentre trial designed to validate biological parameters in early CLL as well as to assess the impact on clinical outcome of an early versus delayed policy of treatment with subcutaneous alemtuzumab in the high biological risk. In this cohort, biological characteristics of 105 (40.2%) patients who would be reclassified as MBL using the 2008 CLL diagnostic criteria were compared with those of the remaining 156 patients who had more than 5000/μL CLL-phenotype cells in peripheral blood and fulfilled diagnostic criteria of CLL.
Male to female ratio was similar for MBL and CLL (54/53 vs. 92/66, P=0.21) as was median age (58.18 vs 58.18, P=0.98). Median absolute number of cells with CLL phenotype in peripheral blood was 3120/μL (range,400-4959) in MBL and 9925/μL (range, 5020–110000) in CLL (P<0.0001). No difference in the CD38 status (P=0.48),ZAP-70 expression (P=0.29) or cytogenetic abnormalities as detected by FISH trisomy 12 (P=0.24); deletion 11q (P=0.68); del17p (P=0.09) was found between patients with MBL and CLL. The only feature differentiating CLL from MBL was represented by an excess of patients with unmutated IgVH disease in the former group (CLL,69.2% vs. MBL, 30.8%: P=0.04). In addition, patients with CLL had an about 2-fold risk of having IgVH germline status in comparison to patients with MBL (OR,1.80; 95% CI, 1.02–3.13; P=0.04). Since the arbitrary cut-off of 5000/μL CLL-phenotype cells in peripheral blood failed to identify a peculiar biological profile for either MBL or CLL, we wondered whether a different B-cell threshold based on disease clinical outcome better stratified patients according to biological risk. In an independent cohort including 818 Rai stage 0 patients registered in a GIMEMA (Gruppo Italiano Malattie EMatologiche Maligne dell'Adulto) database, we demonstrated that a count of 10000/ μL B-cells is the best lymphocyte threshold to predict time to first therapy (TFT). When this cut-off was applied to the GISL series we found that the distribution of main high-risk features CD38, P=0.83; trisomy 12,P=0.36; del11q,P=0.85; del17,P=0.37) was similar between patients with B-cell lymphocytes higher and lower than 10000/ μL. Only an excess of cases with unmutated IgVH (P=0.04) and slightly increase of ZAP-70 (P=0.06) characterized patients B-cell higher than 10000 μL. In conclusion, present data obtained from a prospective multicentre study indicate that biological characteristics of CLL are found also in MBL and there is no general predominance of good risk variables in MBL in comparison to CLL. This implies that MBL may not be considered a distinct disease but as an early stage of CLL.
Musto:Celgene: Honoraria; Janssen Cilag: Honoraria.
Abstract 4613
Biologic risk factors such as immunoglobulin variable heavy chain (IgVH) gene mutation status and CD38 and ZAP-70 expression levels, along with genomic aberrations, have been integrated ...in clinical prognostic evaluation of CLL. Additionaly, CLL subsets expressing a certain stereotyped B-cell receptors have also been indicated to share biological and clinical features.
We investigated, by FISH, the incidence of the major cytogenetic alterations (+12 and 13q14, 17p13, 11q23 deletions), their clinical implication and their relationship with prognostic biomarkers in 344 out of 384 Binet A CLLs enrolled in the prospective multicenter O-CLL1 GISL trial. Stereotypy subsets identification have been performed in 324 patients.
Molecular markers characterization and FISH protocols were previously reported (Cutrona et al. Haematologica, 2008; Fabris et al. Genes Chromosomes Cancer, 2008), while stereotyped subsets were defined according to Stamatopoulos et al (Blood, 2007) and Murray et al (Blood, 2008).
At least one abnormality was found in 225/344 (65.4%) cases. The most frequent abnormality was del(13q14), detected in 173 CLLs (50.3%) followed by +12 (44/344;12.8%) (one case harboring 17p13 deletion), del(17p13) (9/344, 2.6%) and del(11q23) (18/344, 5.2%). 13q14 deletion was found as a sole abnormality in 155 (45%) patients; in the remaining cases, it was combined with +12 (3 pts) and 17p13 (4 pts) or 11q23 deletions (11 pts). The 13q deletion was found as a monoallelic deletion in 139/173 (80.3%); the presence of a biallelic deletion (> 20% of interphase nuclei) was found in the remaining 34 cases. No acquisition of new cytogenetic aberrations was evidenced among the 13 CLLs developing progressive disease (range, 6 to 32 months; median, 20 months); in only one case, the proportion of nuclei with 17p13 and 13q14 deletions increased from the time of diagnosis (from 33% to 92%). Biomarkers data were available in all of the patients. CD38 percentages (mean value ± sem) were 7.9±1.3, 15.1±1.9, 51.7±5.5, 22.0±7.8,40.8±13.2, 39.8±7.3 for del(13q14), normal karyotype, +12, del(11q23), del(17p13) and multiple alterations, respectively (p<0.0001). The percentages of IgVH mutations significantly correlated with cytogenetic alterations; namely, 5.7±0.2 for cases with del(13q14), 4.7±0.4 for normal karyotype, 2.3±0.5 for +12, 0.05±0.05 for del(11q23), 2.0±1.1 for del(17p13) and 1.0±0.4 for multiple alterations (p<0.0001). Similarly, a significant correlation was found for ZAP-70 expression: namely 32.9±1.6 for cases with del(13q14), 38.5±2.1 for normal karyotype, 46.4±3.6 for +12, 67.0±8.3 for del(11q22), 41.0±12.8 for del(17p13) and 50.7±5.4 multiple alterations (p<0.0001). Cytogenetic abnormalities were clustered in 3 risk groups i.e. low del(13q14) and normal; intermediate (+12); and high risk 17p13 and 11q23 deletions and correlated with a scoring system in which patients were stratified in 4 different groups according to the absence or presence of 1, 2 or 3 biomarkers (Morabito et al., Br. J. Haematol., 2009). Notably, 166/175 cases scoring 0, gathered in the low FISH group, whereas 21/26 high FISH risk cases clustered in scoring 2–3 (p<0.0001). A significantly higher risk of starting treatment was found in high vs. intermediate (p=0.024) and low FISH risk (p=0.001) CLLs. Finally, stereotyped IgVH sequences were found in 108/324 (33%). Unfavorable stereotyped subsets (#1, #2, #3, #7 and #9) were significantly more frequent in CLLs with poor-prognostic aberrations (p=0.0203; RR=3.589).
Our data indicate that cytogenetic lesions predicting unfavorable prognosis show a relatively low incidence in newly diagnosed Binet stage A CLLs and are significantly associated with negative prognostic biomarkers predictive of disease progression. Our prospective study also confirms the prognostic value of risk FISH categories in predicting the time to the first treatment and revealed a higher rate of unfavorable stereotyped IgVH subsets in patients carrying poor-prognostic genomic aberrations. Finally, preliminary evidence in a limited number of cases indicates that the acquisition of new genetic abnormalities seem to be an infrequent event during disease progression.
No relevant conflicts of interest to declare.
Abstract 2423
Chronic lymphocytic leukemia (CLL) is characterized by an extremely variable clinical course. Mutational status of the immunoglobulin heavy-chain variable (IGHV) region defines two ...disease subsets with different prognosis. A fraction of CLL cases carries highly homologous B-cell receptors (BCR), i.e. characterized by non-random combinations of immunoglobulin heavy-chain variable (IGHV) genes and heavy-chain complementarity determining region-3 (HCDR3).
We performed sequence analysis to characterize IGHV regions in a panel of 1133 CLL patients investigated by a multicenter Italian study group. A total of 1148 rearrangements were identified; the analysis of stereotyped subsets was performed based on previously reported criteria (Messmer et al, J Exp Med 2004; Stamatopuolos et al, Blood 2007). Specifically, we compared all our sequences with those found in three different publicly available data sets (Stamatopoulos et al, Blood 2007; Murray et al, Blood 2008 and Rossi et al, 2009 Clin Cancer Res). In addition, a pairwise alignment within all sequences was performed in order to discover novel potential subsets (HCDR3 identity > 60%).
Based on the 2% cut-off used to discriminate between Mutated (M) and Unmutated (UM) cases, 777 sequences (67.59%) were classified as M, while 371 sequences (32.3%) as UM. The most represented IGHV genes within mutated cases were IGHV4-34 (104/118) and IGHV3-23 (85/96), whereas IGHV1-69 (97/112) was the most frequently used in the UM group. Interestingly, the IGHV3-21 gene, reported to be frequently expressed in CLL patients from Northern Europe, was present in only a small fraction of cases (24; 2.07%), confirming a previous finding reported by Ghia et al (Blood 2005) in a smaller panel. In our series, stereotyped HCDR3 sequences were found in 407/1148 (35.45%) patients, 177 of whom were M and 230 were UM cases. Overall, we observed that stereotyped sequences were significantly associated with UM IGHV status (Fisher's exact test, P<0.0001). Among the 407 stereotyped HCDR3 sequences, 345 belong to the clusters reported by Murray et al and 14 to those described by Rossi et al., 2009 Clin Cancer Res. The most frequent stereotyped subsets identified in our panel were #1 (35 cases), #7 (28 cases), #4 (24 cases), #3 and #9 (16 cases), #28 (13 cases), and #2 (12 cases), together with subsets #5, #8, #10, #12, #13, #16 and #22 (all ranging from 6 to 9 cases). Finally, we were able to identify by auto-matching analysis 48 sequences potentially specific for 23 novel putative stereotype subsets.
In our series we identified 407/1148 (35.45%) stereotyped HCDR3 sequences. The percentage was higher than that reported by Stamatopoulos et al and Murray et al. This discrepancy may partially be due to the different approach used in our analysis, namely the matching to a general data set including all published stereotyped subsets instead of the auto-matching performed by those Authors. We demonstrated a significant association between IGHV status and stereotyped sequences and confirmed the finding that #1 is the most frequent subset identified so far. Finally, we were able to identify a series of 23 novel putative subsets that will require further confirmation.
No relevant conflicts of interest to declare.
Abstract 2375
Poster Board II-352
A prognostic index based on widely available clinical and laboratory features was recently proposed to predict survival in patients with previously untreated ...patients with chronic lymphocytic leukemia (CLL) by MD Anderson investigators. However, whether proposed clinical risk categories may surrogate new biological variables of prognostic relevance (i.e., mutational status of the IgVH gene regions, ZAP-70 or CD-38 expression, cytogenetic abnormalities) is unclear thus far.
In a series of 160 asymptomatic Binet stage A patients enrolled in a Gruppo Italiano Studio Linfomi (GISL) multicentre trial designed to validate prospectively biological parameters in early CLL as well as to assess the impact on clinical outcome of an early versus delayed policy of treatment with subcutaneous alemtuzumab in the high biological risk, we evaluated whether clinical categories derived from newly proposed prognostic index reflected biological risk.
Since the original prognostic index was derived from a database including cases with more advanced disease we used an optimal cutoff search to determine how to best split Binet stage A patients in different prognostic groups. To this purpose an independent patient cohort consisting of 310 Binet stage A patients included in a GIMEMA (Gruppo Italiano Malattie EMatologiche Maligne dell'Adulto) database was used. According to recursive partitioning (RPART) model, a classification tree was built that identified two subsets of patients who scored respectively: 0-3 (low risk) and 4-7 (high risk). Therefore, by prognostic index, 48.7% and 51.2% of 160 asymptomatic stage A patients, respectively, met criteria of low risk and high risk disease.
In our prospective series high- risk score was more frequently associated with both unmutated IgVH status (P=0.009) and higher CD38-expression (P=0.002); in contrast only a trend towards an increased ZAP-70 expression could be found (P=0.06). As far as cytogenetic abnormalities are concerned, we observed that 11q deletion occurred more frequently among patients belonging to high-risk score (P=0.005), while cases with 13q deletion or trisomy 12 were homogeneously distributed among low- and high-risk patient category(P=0.151 and P=0.452, respectively). We did not consider suitable for correlation analysis 17p deletion since observed only in 2 out of 160 Binet stage A patients.
In conclusion, our results demonstrate in a prospective cohort of patients with early CLL that clinical categories of a revised score index may surrogate biological parameters of prognostic relevance. The observation reinforces the revised IWCLL guidelines recommendations to assess the risk of CLL patients on clinical basis and to deserve biological studies to patients eligible for clinical trials.
No relevant conflicts of interest to declare.
Abstract 2341
Poster Board II-318
The clinical heterogeneity of chronic lymphocytic leukemia (CLL) requires parameters to stratify patients into prognostic subgroups to adapt treatment ranging from ...‘watch and wait' to allogeneic stem cell transplantation. To this end, several parameters such as lymphocyte doubling time, β-2 microglobulin, CD38 and ZAP-70 expression, immunoglobulin variable heavy chain (IgVH) mutation status and genetic abnormalities, as assessed by fluorescence in situ hybridization (FISH), have been integrated in clinical practice.
In the present study, we investigated by FISH the incidence of the known major cytogenetic alterations (+12 and 13q14, 17p13, 11q23 deletions) in a series of Binet A B-CLL patients included in the prospective O-CLL1 GISL study started in April 2007.
Molecular markers characterization and FISH analyses were performed as previously reported (Cutrona et al. Haematologica, 2008; Fabris et al. GCC, 2008). A cut-off value of 2% was used to distinguish mutated and unmutated patients. CD38 and ZAP-70 were determined by flow-cytometry and a 30% cut-off was used to distinguish between positive or negative cases.
Up to date, 326 patients have been enrolled in the trial and FISH data concerning trisomy 12 and 13q14, 17p13, 11q23 deletions were available in 305 patients. At least one abnormality was found in 197 (64%) cases. The most frequent was del(13)(q14) (150/305, 49%), followed by +12 (40/303, 13%) (in one and three cases accompanied by 17p13 and 13q14 deletions, respectively), del(17)(p13) (7/305, 2%) and del(11)(q23) (17/305, 5%). 13q14 deletion was found as a sole abnormality in 134 patients; in the remaining cases, it was combined with +12 (3 pts) and 17p13 (3 pts) or 11q23 (10 pts) deletions. Among patients with 13q14 deletions, 99 were monoallelic, 12 biallelic and 39 showed a combination of the two patterns. Biomarkers data were available in all of the patients: 95/305 (31%) cases had unmutated IgVH genes; ZAP-70 and CD38 were positive in 117/305 (38%) and 72/305 (23%) cases, respectively. Concerning the distribution of cytogenetic aberrations, the unmutated IgVH group included 29/150 (19%) 13q14 deleted cases, 23/40 (57%) cases with trisomy 12 and 4/7 (57%) and 16/17 (94%) with 17p13 and 11q23 deletions, respectively. ZAP-70-positive groups included 43/150 (28%) 13q14 deleted cases, 26/40 (65%) cases showing trisomy 12 and 5/7 (71%) and 12/17 (70%) with 17p13 and 11q23 deletions, respectively. Finally, CD38-positive cases included 18/150 (12%) 13q14 deleted cases, 26/40 (65%) cases carrying trisomy 12 and 5/7 (71%) and 7/17 (41%) with 17p13 and 11q23 deletions, respectively. The percentages of IgVH mutations significantly correlated with cytogenetic alterations; namely, 5.8±0.3 for cases with del(13)(q14), 4.6±0.4 for normal karyotype, 2.6±0.5 in +12, 0.3±0.2 in del(11)(q23), and 1.7±0.9 in del(17)(p13) cases (p for trend <0.0001). A significant correlation was also found for ZAP-70 expression: namely 32±1.8 for cases with del(13)(q14), 38.6±2.2 for normal karyotype, 47.6±3.7 for +12, 55.8±7.0 for del(11)(q22) and 42.4±11.7 for del(17)(p13) (p<0.0001). Similarly, CD38 percentages were (mean value ± sem) 9.3±1.7, 16.9±2.1, 52.9±5.7, 26.8±6.2, 37.0±12.7 for del(13)(q14), normal karyotype, +12, del(11)(q23) and del(17)(p13) alterations, respectively (p for trend <0.0001). Finally, cytogenetic abnormalities were clustered in 3 risk groups i.e. low del(13)(q14) and normal; intermediate (+12); and high risk del(11)(q23) and del(17)(p13) and significantly correlated (p<0.0001) with a scoring system in which cases were stratified in 4 different groups according to the absence (group 0) or presence of 1 (group 1), 2 (group 2) or 3 (group 3) biomarkers (Morabito et al., BJH, 2009, voce). Interestingly, 147/154 cases scoring 0, gathered in the low FISH group, whereas 17/22 high FISH risk cases clustered in scoring 2-3.
Our preliminary results indicate that in Binet stage A B-CLL patients at diagnosis cytogenetic abnormalities with an expected negative clinical impact are relatively few (7.2%) but significantly associated with prognostic biomarkers which negatively predict the clinical outcome in B-CLL.
No relevant conflicts of interest to declare.
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