Smoking habits and osteoporosis in community-dwelling men subjected to dual-X-ray absorptiometry: a cross-sectional study.

BACKGROUND: Active and Environmental Tobacco Smoke (ETS) are a global cause of death. Osteoporosis (Op) is the most common metabolic bone disorder worldwide, impacting on mortality and disability, with high health and welfare costs. Active smoking is a known risk factor for Op, but there is few information regarding Op and ETS in men. PURPOSE: The study aim is to evaluate the association between smoking habits and Op in community-dwelling men that have been subjected to Dual-X-ray Absorptiometry and completed a questionnaire about their own and cohabiter's smoking habits. METHODS: We performed a cross-sectional study based on administrative data. This study is part of the SIMON protocol. The binary logistic regression analysis was used to estimate the role of ETS on the risk of Op, adjusting for age, body mass index (BMI), type 2 diabetes mellitus (T2DM) and eGFR. RESULTS: Four hundred sixteen men were selected and, based on questionnaire replies, 167 were classified as current smokers (CS), 93 as passive smokers (PS) and 156 as never smokers (NS). NS showed a lower prevalence of past fragility fracture, radiological features of osteoporosis and hypovitaminosis D compared to PS and CS (p < 0.05). NS showed a lower prevalence of Op compared to PS and CS, also after correction for age, BMI, T2DM and eGFR (p < 0.05). CONCLUSION: The study results demonstrate that PS and CS have a higher risk of Op, fragility fractures and vitamin D deficiency compared to NS.

Role of active and environmental tobacco smoke on susceptibility to osteoporosis in women undergoing dual-X-ray absorptiometry.

PURPOSE: Current smoking is a risk factor for osteoporosis (Op), but few data are available regarding the passive smoke impact on Op susceptibility. This cross-sectional study aimed to evaluate the association between the smoking habits and Op in community-dwelling women undergoing dual-energy X-ray absorptiometry (DXA). METHODS: On 01/06/2018, general practitioners from "COMEGEN" Medical Cooperative, Naples, Italy, selected the medical records from the last 10 years of women who had a measurement of bone mineral density performed and simultaneously completed a questionnaire about their smoking behaviour and their cohabiters'. The binary logistic regression analysis was used to estimate the role of passive smoke on the risk of Op, adjusting for age and body mass index (BMI). RESULTS: Among 10,616 subjects, 3942 were currently smokers [CS; mean age 69.4 ± 10.4 years; BMI 27.0 ± 4.9 kg/m2], 873 were passive smokers (PS; mean age 67.8 ± 11.6 years; BMI 27.0 ± 4.9 kg/m2) and 5781 were never smokers (NS; mean age 67.8 ± 11.6 years; body mass index (BMI) 27.0 ± 4.9 kg/m2). Of all, 8562 women (mean age 70.3 ± 10.2 yrs; BMI 27.0 ± 4.9 kg/m2) received the Op diagnosis. PS showed an increased Op risk compared to NS [odds ratio (OR) 1.38 (1.14-1.67)] and comparable to CS [OR 1.02 (0.84-1.24)]. CONCLUSION: The study results demonstrate an association between passive smoke and Op in community-dwelling women already presenting with susceptibility to Op according to Italian essential assistance levels, suggesting that passive and active smoke are equivalent Op risk factors in women.

Breast surgeons updating on the thresholds of COVID-19 era: results of a multicenter collaborative study evaluating the role of online videos and multimedia sources on breast surgeons education and training.

OBJECTIVE: Current trends show a rise of attention given to breast cancer patients' quality of life and the surgical reconstructive result. Along with this trend, surgical training quality and efficacy are gaining importance and innovative training methods such as online videos shared on social media portals, are becoming main updating tools. In hazardous times like COVID-19 pandemic nowadays, online communication becomes of vital importance and adaptation and innovation are fundamental to keep research and education alive. The authors aimed to investigate the role of video and multimedia sources on the daily activity and surgical training of a representative group of surgeons specifically dedicated to oncologic, oncoplastic and reconstructive breast surgeries. MATERIALS AND METHODS: A survey was produced and administered to 20 major Italian Breast Centers. Collected data were analyzed with Fisher's Exact Test. RESULTS: From October 2019 to March 2020, a total of 320 surveys were collected. Among the responders, there were 188 trainees (intern medical doctors and residents) and 110 faculty, 72% of them belonged to a plastic surgery environment, while 28% to general surgery environment. Almost all respondents have ever watched videos concerning breast surgery. CONCLUSIONS: The results of the study show how breast surgeons rely on videos and web platforms, mostly YouTube, when searching for training info about surgical procedures. Social media offer great opportunities for sharing knowledge and diffusion of new ideas but greater attention to their reliability is mandatory.

Correction: Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph+ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Standardisation and consensus guidelines for minimal residual disease assessment in Philadelphia-positive acute lymphoblastic leukemia (Ph + ALL) by real-time quantitative reverse transcriptase PCR of e1a2 BCR-ABL1.

Minimal residual disease (MRD) is a powerful prognostic factor in acute lymphoblastic leukemia (ALL) and is used for patient stratification and treatment decisions, but its precise role in Philadelphia chromosome positive ALL is less clear. This uncertainty results largely from methodological differences relating to the use of real-time quantitative PCR (qRT-PCR) to measure BCR-ABL1 transcript levels for MRD analysis. We here describe the first results by the EURO-MRD consortium on standardization of qRT-PCR for the e1a2 BCR-ABL1 transcript in Ph + ALL, designed to overcome the lack of standardisation of laboratory procedures and data interpretation. Standardised use of EAC primer/probe sets and of centrally prepared plasmid standards had the greatest impact on reducing interlaboratory variability. In QC1 the proportion of analyses with BCR-ABL1/ABL1 ratios within half a log difference were 40/67 (60%) and 52/67 (78%) at 10-3 and 36/67 (53%) and 53/67 (79%) at 10-4BCR-ABL1/ABL1. Standardized RNA extraction, cDNA synthesis and cycler platforms did not improve results further, whereas stringent application of technical criteria for assay quality and uniform criteria for data interpretation and reporting were essential. We provide detailed laboratory recommendations for the standardized MRD analysis in routine diagnostic settings and in multicenter clinical trials for Ph + ALL.

Interface morphology and gas production by a refractive 347 nm ultraviolet femtosecond laser: Comparison with established laser systems.

PURPOSE: To evaluate gas production and interface quality after refractive lenticule creation. A new ultraviolet (UV) 347 nm femtosecond laser was compared with the FS200 and the Visumax lasers. SETTING: Department of Anatomy II, University of Erlangen-Nürnberg, Erlangen, Germany. DESIGN: Experimental study. METHODS: Refractive lenticules were created in 52 porcine corneas with the UV laser (n = 20), the Visumax laser (n = 20), and a modified FS200 laser (n = 12). Twelve corneas in each laser group were fixed immediately after lenticule creation to entrap the gas produced. Lenticule extraction was omitted. From 16 other corneas (UV and Visumax lasers; n = 8 each) -5.0 diopter (D) and -10.0 D lenticules were extracted. On lenticule extraction, a flap was created and lifted. In 12 additional corneas, a flap was cut using the FS200 laser and lifted afterward; laser in situ keratomileusis was performed on 8 of these samples. The corneas were processed for light microscopy (gas bubble 2-dimensional morphometry) and scanning electron microscopy (interface quality). RESULTS: The Visumax generated approximately one half as much gas as the FS200. The UV laser undercut the Visumax by another 50%. The interfaces created by the FS200 appeared smooth and without irregularities. The optical zone margin was always conspicuous in the Visumax samples but hardly discernible in the UV specimens. Slight irregularities and very small lenticule pieces were found occasionally in both groups. The UV interfaces appeared slightly rougher than the Visumax interfaces. CONCLUSIONS: Ultraviolet laser lenticule extraction seems promising. The interface quality was similar to that of the clinical Visumax system while gas production was reduced significantly.

CD4+CD25+ regulatory T-cell-inactivation in combination with adenovirus vaccines enhances T-cell responses and protects mice from tumor challenge.

Cancer vaccines are a promising approach to treating tumors or preventing tumor relapse through induction of an immune response against tumor-associated antigens (TAA). One major obstacle to successful therapy is the immunological tolerance against self-antigens which limits an effective antitumor immune response. As a transient reduction of immunological tolerance may enable more effective vaccination against self-tumor antigens, we explored this hypothesis in a CEA tolerant animal model with an adenovirus expressing CEA vaccine in conjunction with inactivation of CD4(+)CD25(+) regulatory T cells. This vaccination modality resulted in increased CEA-specific CD8(+), CD4(+) T cells and antibody response. The appearance of a CD4(+) T-cell response correlated with a stronger memory response. The combined CD25(+) inactivation and genetic vaccination resulted in significant tumor protection in a metastatic tumor model. Non-invasive tumor visualization showed that not only primary tumors were reduced, but also hepatic metastases. Our results support the viability of this cancer vaccine strategy as an adjuvant treatment to prevent tumor relapse in cancer patients.

Retrospective comparison of qualitative and quantitative reverse transcriptase polymerase chain reaction in diagnosing and monitoring the ALL1-AF4 fusion transcript in patients with acute lymphoblastic leukaemia.

We compared quantitative reverse transcriptase polymerase chain reaction (Q-RT-PCR) to qualitative RT-PCR in determining response to therapy and predicting clinical outcome in 18 retrospectively selected patients with ALL positive for the ALL1-AF4 fusion and with frozen RNA samples collected at diagnosis and during follow-up (96 samples analysed). The ALL1-AF4 junction was detected by qualitative RT-PCR in 18 patients and by Q-RT-PCR in 17 patients (one patient harboured the rare e10-e6 ALL1-AF4 junction, which falls outside of the primer and probe location designed for the Q-RT-PCR). In three of the 12 patients negative to qualitative RT-PCR after induction therapy, a small number of ALL1-AF4 copies was detected by Q-RT-PCR. Thus nine patients were negative and eight positive. Seven of the eight positive patients suffered a relapse, including two of the three patients positive to Q-RT-PCR yet negative to qualitative RT-PCR. Moreover, we found two (5%) discordant results among the 39 follow-up tests of the nine patients who converted to a negative qualitative-quantitative PCR status. The results suggest that qualitative RT-PCR is more appropriate for the routine diagnosis of this genetic alteration. However, Q-RT-PCR is more accurate in assessing the molecular response after induction treatment and could be more useful in clinical decision-making in ALL1-AF4-positive ALL patients.

Partial deletions of long arm of chromosome 6: biologic and clinical implications in adult acute lymphoblastic leukemia.

Within 285 adult acute lymphoblastic leukemias (ALL) included in the multicenter GIMEMA 0496 trial and prospectively studied by conventional cytogenetics, 18 cases (6%) with long arm deletion of chromosome 6 (6q) were identified. These cases were divided into: (i) del(6q) only (n = 6); (ii) del(6q) plus other numerical and/or structural abnormalities (n = 8); (iii) del(6q) and other 'specific' translocations (n = 4). The biologic and clinical features of the patients carrying this anomaly, as well as their outcome, were compared with those of 267 patients without del(6q). A T cell phenotype was more frequently associated with del(6q) cases in general (P = 0.001) and particularly with cases presenting del(6q) as the isolated abnormality (P = 0.0027). No significant difference with respect to multidrug resistance (MDR)/P glycoprotein expression was observed between the two groups of patients (21% vs 28% of MDR-positive cases, respectively). A BCR-ABL fusion transcript was less frequently detected in cases with del(6q) (11%) compared with those without the anomaly (29%). p15 and p16 deletions were identified by Southern blot analysis in 21% of cases with del(6q) and in 26% of cases without del(6q). In this latter group, a T cell phenotype was less frequently associated with p15 and/or p16 deletion than in the group carrying del(6q) (36% vs 100% of cases, P = 0.011). Overall, patients with ALL and del(6q) had a high complete remission (CR) rate (83%); however, they had a lower 18 month event-free survival (31% vs 41%) and a higher relapse rate (70% vs 37%, P = 0.02) compared with patients without del(6q). To date, this is the largest series of adult ALL cases reported with del(6q) homogeneously treated, which have also been prospectively studied for MDR expression and for the detection of known fusion genes. This anomaly, as an isolated change, identifies a subset of cases with hyperleukocytosis (median WBC count 52 x 10(9)/l) and a strict correlation with a T cell phenotype. Overall, del(6q) seems to be associated with an unfavorable clinical outcome, although this finding will need to be confirmed by extended FISH analysis.

Myeloperoxidase gene expression in non-infant pro-B acute lymphoblastic leukaemia with or without ALL1/AF4 transcript.

In this study, we examined myeloperoxidase (MPO) gene expression in a series of 31 non-infant pro-B acute lymphoblastic leukaemia (ALL) patients that included 16 cases with the t(4;11) translocation and/or the resultant ALL1/AF4 chimaeric gene. Sixteen out of 31 cases (51%) were MPO mRNA positive/enzyme negative. MPO mRNA was detected in nine out of 16 (56%) and seven out of 15 (47%) patients with and without the ALL1/AF4 fusion transcript respectively. The comparative study between MPO mRNA positive and negative cases showed statistically significant differences with regard to age and white blood cell (WBC) count, and was 39.5 years vs. 26.3 years (P = 0.016) and 71.4 x 10(9)/l vs. 157.8 x 10(9)/l (P = 0.046) in the MPO mRNA positive and negative groups respectively. The correlation analysis between MPO mRNA expression, age, WBC count and leukaemic relapse according to the presence/absence of the ALL1/AF4 fusion showed that the statistically significant differences observed in the whole group were related mostly to the ALL1/AF4-positive ALL patients. In fact, in this latter group, the mean WBC count and patients' age were 85 +/- 79 x 10(9)/l vs. 289.8 +/- 102 x 10(9)/l (P = 0.0005) and 44.8 +/- 15.3 years vs. 26.7 +/- 13.7 years (P = 0.01) in patients with and without MPO mRNA expression respectively. It appears, therefore, that the assessment of MPO mRNA expression enables a further dissection of leukaemia heterogeneity in apparently homogeneous genetic/immunophenotypic ALL subsets.

A prospective study of residual-disease monitoring of the ALL1/AF4 transcript in patients with t(4;11) acute lymphoblastic leukemia.

Twenty-five patients (22 adults and 3 infants) with ALL1/AF4-positive acute lymphoblastic leukemia (ALL) were prospectively monitored by reverse transcriptase-polymerase chain reaction (RT-PCR) between January 1992 and July 1999. After high-dose induction and consolidation chemotherapy without bone marrow transplantation, all patients had a complete hematologic remission. Using nested RT-PCR (sensitivity 10(-4)), we observed conversion to PCR negativity in 11 (44%) of the patients. Thirteen of the 14 patients who did not have a molecular remission had a relapse at a median time of 4 months (range, 1 - 20 months). Of the 11 patients who had a conversion to PCR negativity, 5 reconverted to PCR positivity within 1 to 14 months. These 5 patients all progressed to hematologic relapse after 2, 3, 4, 4, and 7 months, respectively. Of the remaining 6 patients, 4 are in persistent hematologic and molecular remission at 12, 14, 88, and 96 months, whereas 2 are early in their follow-up. Actuarial probabilities of relapse and overall survival were 100% and 0% at 14 and 24 months and 67% and 43% at 96 and 100 months, respectively, in patients who had persistent RT-PCR positivity and in those who had a molecular remission. For both relapse and survival, the differences observed between the two groups were significant (P =.003 and P <.005, respectively). This study, which represents the first prospective analysis of residual-disease monitoring carried out in a substantial series of patients with t(4;11)-positive ALL, emphasizes the clinical relevance of RT-PCR-based methods to monitor minimal residual disease in this leukemia subset. (Blood. 2000;95:96-101)

HLA-C and HLA-DQB1 compatibility in unrelated cord blood transplants.

BACKGROUND AND OBJECTIVE: Umbilical cord blood (UCB) cells have been definitively proved to be a source of hematopoietic stem cells with repopulating capacity when transplanted into pediatric hosts with neoplastic or non-neoplastic disease. Moreover, due to the immaturity of the UCB lymphoid compartment, these transplants are usually associated with a low incidence and severity of GvHD. This clinical observation and the immaturity of the UCB lymphoid compartment justify the acceptance of UCB units which differ from their recipient by 1 or 2 HLA antigens of the six HLA A, B and DRB1 antigens conventionally typed. Whether the number and type of HLA disparities affect clinical outcome of UCB transplants has not, however, been clearly demonstrated yet. DESIGN AND METHODS: In the present study on 14 pediatric patients with high risk leukemia transplanted with UCB from unrelated donors, evaluation of HLA compatibility was extended to HLA-C and DQB1 genes and correlated to the engraftment rate and occurrence of GvHD. Conditioning regimen and GvHD prophylaxis were identical in all cases. HLA-A and B antigens were typed by serology, whereas DNA based methods were used to define HLA-C gene groups, and HLA-DRB1 and DQB1 alleles. RESULTS: Conventional HLA-A, B and DRB1 typing demonstrated that 12 recipient/donor pairs differed at one HLA locus, while 2 pairs had 2 HLA disparities. The extended HLA-typing showed that only one out of the six pairs with a different HLA-A locus had additional mismatches at HLA-C and DQB1 loci, whereas all the remaining 8 pairs, which already differed at HLA-B and/or DRB1 loci after conventional typing, had additional HLA-C and/or DQB1 mismatches (p = 0.002). By contrast, engraftment rate and occurrence of GvHD did not significantly correlate with level of HLA-mismatches even after extended HLA-typing. INTERPRETATION AND CONCLUSIONS: The present data show that additional mismatched HLA-C and/or DQB1 antigens are significantly more frequent in pairs which after conventional HLA-typing differed at HLA-B and/or DRB1 loci, than in those showing one HLA-A mismatch. This observation provides an additional criterion for selection of UCB donors with the closest HLA-match when more than one unit are available. We did not, however, observe any correlation between engraftment rate, occurrence of GvHD and degree of HLA disparities detected either by standard or extended typing. These data support the notion that certain HLA differences do not affect the clinical outcome of UCB transplants and indicate that the expensive and time consuming molecular typing of HLA-C and DQB1 loci might be avoided for UCB donor selection.

Umbilical cord blood transplant from unrelated HLA-mismatched donors in children with high risk leukemia.

In the last 3 years, 14 children with high-risk leukemia (11 ALL, 2 AML and 1 CML) underwent cord blood transplantation from unrelated HLA-mismatched donors at a median of 99 days from the start of search. Eight patients were transplanted in second CR, one in accelerated phase, three at relapse and two patients in first CR. Conditioning regimen (fractionated TBI, etoposide, CY and anti-lymphocyte serum) and prophylaxis of GVHD (CsA and 6-methylprednisolone) were identical for all patients. Neutrophils >0.5x10(9)/l were reached at a median of 33 days from transplant, but in four cases we observed an autologous hematopoietic reconstitution (three spontaneous, one after autologous BM rescue). Acute and chronic GVHD were observed in 10/14 and 3/8 evaluable cases, respectively. Three patients died of transplant-related toxicity and three patients relapsed. The probabilities of event-free, disease-free and overall survival were 50, 53 and 64%, respectively. Cord blood transplant from HLA-mismatched unrelated donor is a valid option for the treatment of children with high-risk leukemia. With our eligibility criteria, conditioning regimen and prophylaxis of graft-versus-host disease, the main obstacles to successful transplant were represented by graft failure and fatal acute GVHD.

A prospective molecular study of chimaerism in patients with haematological malignancies receiving unrelated cord blood or bone marrow transplants: detection of mixed chimaerism predicts graft failure with or without early autologous reconstitution in cord blood recipients.

We prospectively studied the chimaerism status in the bone marrow (BM) and peripheral blood (PB) of 23 patients receiving umbilical cord (UCB, 14 cases) or BM (nine cases) transplants from unrelated donors by PCR amplification of four individual-specific VNTR genetic loci. Haematological engraftment, with persistent full donor pattern. was observed in 10/14 (72%) patients receiving UCB and in 9/9 (100%) patients transplanted with marrow from an unrelated donor (MUD). In contrast, the remaining four patients converted to an autologous pattern. Three out of these four patients had an early autologous haematological reconstitution reaching a neutrophil level >0.5 x 10(9)/l at days 27, 33 and 37 after transplant, respectively. In all three of these patients, chimaerism analysis demonstrated an early appearance of donor cells (i.e. within 35 d after UCB transplant) showing a transient full donor (one case) or mixed chimaerism condition (two cases). Despite the early autologous haemopoietic reconstitution, one of the three patients died of GVHD at day 60, which was explained by the demonstration of low levels of donor lymphoid cells. In the MUD group all nine patients converted to a persistent full donor pattern with haematological reconstitution, accompanied in two of them by transient mixed chimaerism lasting to days 60 and 270 after transplant. Our data show that monitoring of chimaerism may predict graft failure with or without early autologous haemopoietic reconstitution in patients receiving unrelated UCB transplants. Furthermore, chimaerism analysis may identify, in patients with autologous reconstitution, those at risk of severe GVHD in whom immunosuppressive therapy should not be discontinued.

Umbilical cord blood (UCB) transplant from unrelated mismatched donor in patients with high risk (HR) leukemia.

Ten consecutive children with high risk leukemia have been submitted to UCB transplant from unrelated HLA mismatched donors. All patients received an identical regimen for conditioning and GVHD prophylaxis. The median dose of viable nucleated cells infused was 2.6 x 10(6)/kg b.w. Among the nine patients evaluable for engraftment the hematopoiesis was of full donor origin in six patients and autologous in three. At a median follow-up of 9 months, six of nine (67%) patients are alive in CR.

Umbilical cord blood transplant from HLA-mismatched unrelated donor in high-risk leukemia.

Twelve consecutive children with high-risk leukemia have been submitted to UCB transplant from unrelated 1 or 2 loci HLA-mismatched donor. All patients received an identical regimen for conditioning and GVHD prophylaxis. The median dose of viable nucleated cells infused was 2.8 x 10(7)/kg bw (range 1.4-7.9). Of 11 patients evaluable for engraftment, the hematopoiesis was of full donor origin in seven patients and autologous in four. The probability of disease-free survival at 1 and 2 years from UCB transplant is 60 and 42%, respectively.

ALL1 gene alterations in acute leukemia: biological and clinical aspects.

BACKGROUND AND OBJECTIVE: The ALL1 gene, also referred to as MLL, HRX or Htrx1, is interrupted in the vast majority of translocations involving the chromosome band 11q23. Alterations in this gene are reported in approximately 5-10% of acute leukemias (AL) and characterize different leukemic subtypes such as infant (< 12 months of age) AL, topoisomerase II inhibitors-related (TR) AL and a small subset of de novo AML and ALL. Distinguishing features of ALL1 alterations include the striking heterogeneity of its recombinations, i.e., more than 30 chromosome partners have been described in ALL1 rearrangements, and the lack of association with a definite lineage. The objective of this article is to review the biological and structural properties of ALL1 gene and its various fusion proteins, and to discuss the clinical relevance of these lesions with special emphasis on their role in molecular diagnosis and monitoring of minimal residual disease. EVIDENCE AND INFORMATION SOURCES: The material examined in the present review includes data published by the authors in this field, articles and abstracts published in journals covered by the Science Citation Index and Medline, as well as some more recent personal unpublished observations. STATE OF THE ART: The ALL1 gene spans approximately 90 kb of DNA in length, and consists of 36 exons, ranging in size from 65 bp to 4249 bp. ALL1 codifies for a major transcript of approximately or equal to 15 kb. It encodes a protein of more than 3910 amino acids, containing three regions sharing sequence homology with the Drosophila trithorax gene. These homologies suggest that ALL1 is a transcription factor controlling development and/or differentiation of human cells. To date, twelve ALL1 partner genes have been characterized which are involved in the following translocations: t(4;11), t(9;11), t(6;11), t(11;19), t(1;11) t(10;11), t(11;16), t(11;17) and t(X;11). Since all these genes do not share relevant homologies among each other, their putative role in ALL1 activation still remains to be clarified. The analysis of ALL1 breakpoint cluster region (bcr) shows that several DNA motifs implicated in illegitimate recombination events are located within the bcr. Thus, mapping of breakpoints in the different subtypes of ALL1 +ve leukemia may help in understanding the events leading to translocations in human ALs. In this respect, data on ALL1 breakpoint localization suggest that similar pathogenetic mechanisms may underlie infant and TR AL and that these events might differ from those occurring in de novo AL. The availability of this molecular marker provides a new tool for diagnostic purposes and characterization of ALs and for monitoring of minimal residual disease. To date, the prognostic value of ALL1 rearrangements has been clearly demonstrated for infant ALs only, whereas the clinical relevance of ALL1 rearrangements in the other leukemic subtypes needs further evaluation by future prospective studies on a larger number of patients homogeneously treated. As concerning studies on minimal residual disease, data on PCR monitoring of the ALL1/AF4 fusion transcript, resulting from the t(4;11) translocation, show the clinical relevance of this molecular test in predicting outcome and, as a consequence, in designing individual post-remission therapies. PERSPECTIVES: It is expected that future studies will provide more detailed information regarding either the normal ALL1 function and/or the leukemogenic effect of ALL1 alterations, together with a better definition of the prognostic relevance of the hybrid proteins formed by this gene at diagnosis and during remission of disease.

Infant acute leukemias show the same biased distribution of ALL1 gene breaks as topoisomerase II related secondary acute leukemias.

The ALL1 gene (also called MLL, HRX, or Htrx1) at the cytogenetic band 11q23 is consistently altered by chromosome rearrangements in acute leukemias (ALs) of early infancy, in ALs developed after exposure to topoisomerase (topo) II-inhibitory drugs, and in a small subset of de novo ALs in children and adults. Because exposure to natural or medicinal substances blocking topo II during pregnancy have been proposed as etiological agents for infant leukemia, we have compared the distribution of ALL1 gene breakpoints in infant leukemias with an altered ALL1 gene configuration to those in secondary leukemia associated with prior exposure to topo II targeting drugs and in reference to the major topo consensus binding site in exon 9. ALL1 gene breakpoint distribution was determined by Southern blot hybridization and/or reverse transcription-PCR of the ALL1/AF4 fusion cDNA in 70 patients. Using restriction enzyme analysis, the 8.3-kb ALL1 breakpoint cluster region was divided in a centromeric portion of 3.5 kb (region A) and telomeric portion of a 4.8 kb (region B). ALL1 breakpoint were located in region A in 8 of 28 (28.5%) cases of infant ALs, 16 of 24 (66%) cases of de novo ALs, and 0 of 5 cases of therapy-related (TR) ALs. Conversely, ALL1 breakpoints in region B were detected in 20 of 28 (71.5%) cases of infant AL, 8 of 24 (33%) cases of de novo AL, and 5 of 5 (100%) cases of TR AL (P = 0.002). These results were confirmed by direct sequencing of the ALL1/AF4 fusion transcript in 30 cases (19 infants and 11 child and adult de novo cases). The analysis of ALL1/AF4 junction types showed that children and adults with de novo leukemia had ALL1 breakpoints in intron 6 (9 cases) or intron 7 (2 cases), whereas breakpoints in infant cases were mainly located in intron 8 (14 cases) and less frequently in intron 6 (4 cases) and intron 7 (1 case). The difference in ALL1 breakpoint location between infant and noninfant AL patients with ALL1/AF4 fusion was statistically significant (P = 0.00005). These data demonstrated that infant and TR ALs share a similar biased clustering of ALL1 gene breakpoints, which supports the possibility that topo II inhibitors may also operate in utero and play a crucial role in the etiology of infant leukemia.

Multigenetic lesions in infant acute leukaemias: correlations with ALL-1 gene status.

In this study we investigated the presence of structural lesions in the ALL-1, p53 and p16 (cyclin-dependent kinase 4 inhibitor) genes in leukaemic cells obtained from 22 patients with infant acute leukaemia (aged < 18 months). Of these, 18 cases were classified as acute lymphoblastic leukaemia (ALL) and four as acute myeloid leukaemia (AML). Tumour DNAs were analysed by a combination of Southern blot. polymerase chain reaction (PCR), single-strand conformation polymorphism (SSCP), and direct sequence analyses. The results showed ALL-1 gene rearrangements in 15/22 (68%) cases, p53 gene mutations in 5/22 (26%), and a homozygous deletion of p16 in a single T-ALL case. p53 and p16 alterations were all found in the group of patients with ALL-1 gene rearrangements. p53 mutations were more often associated with a myeloid phenotype (3/5). In summary, multiple molecular alterations were found in 6/15 (40%) infant acute leukaemias with ALL-1 rearrangements. As to the clinical course, patients with additional lesions had similar clinical outcome with respect to patients with ALL-1 gene rearrangement as the sole genetic aberration. This may support the hypothesis that ALL-1 alterations are genetic events per se sufficient to confer a fully malignant phenotype to the leukaemic clone.