Patrizia Lista

Selective growth response to IL‐3 of a human leukaemic cell line with megakaryoblastic features

A new human leukaemic cell line (M‐O7) with the phenotypic characteristics of CFU‐mega is described. Its cells are positive for T200 leucocyte common antigen (LCA) and negative with MAbs recognizing T and B cells and mature myelomonocytic antigens. In contrast, they react with MAbs recognizing antigenic determinants common to multilineage (CD13, CD33, CD34) and to bipotent erythromegakaryoblastic (CD36, H25) haemopoietic precursors, and with MAbs specific for platelet glycoproteins (CD41w, CD42w). A small proportion (10%) of the cells were large and multinucleated, and on electron microscopy examination showed peripheral splitting of platelet‐like cytoplasm particles. When transferred to a serum‐free Iscove modified Dulbeccos medium supplemented with human insulin and transferrin, M‐O7 cells stop proliferating. Of the haemopoietic growth factors tested for their ability to restore the proliferative activity of this quiescent population, only rH IL‐3 proved effective. Moreover, it also increased the cloning efficiency in methylcellulose more than any other CSFs. The M‐O7 cell line may provide a valuable tool for the biological assay of IL‐3, and a model for biochemical studies of the megakaryocytic lineage.

Interleukin 2 does not promote the in vitro and in vivo proliferation and growth of human acute leukaemia cells of myeloid and lymphoid origin

The effect of recombinant interleukin 2 (IL2) on the in vitro and in vivo proliferation and growth of human acute leukaemia cells of both myeloid and lymphoid origin was investigated. In none of the 25 primary samples tested could a continuously in vitro growing cell line be obtained by adding IL2 to the culture medium. Although IL2 induced a proliferative signal in three of the 31 acute leukaemias analysed, the overall 3H‐thymidine uptake of the neoplastic cells was significantly reduced (P<0.05) in the presence of IL2. The unlikelihood of an important proliferative signal triggered by IL2 was confirmed in a semisolid clonogenic assay, which failed to document an increased colony growth in the 26 samples studied. Furthermore, using a colorimetric assay as a test for cell proliferation and survival, in seven of the 11 fresh acute leukaemia samples tested a 22–40% reduction in viability was observed in the presence of IL2, while in the remaining four, IL2 was ineffective. In order to investigate the effect of IL2 in an in vivo setting, an experimental model in heavily immunosuppressed nu/nu mice was established. In no case did IL2 promote the in vivo proliferation and growth of human myeloid and lymphoid acute leukaemia cells injected in the mice. On the contrary, with seven of the eight leukaemic cell lines which gave rise spontaneously to leukaemic masses, this could be prevented when the mice received locally 300 U of IL2 three times daily for 90 d. IL2 also blocked the growth in vivo of three fresh acute leukaemia samples (two myeloid and one lymphoid). Co‐culture experiments using leukaemic cell lines and increasing numbers of normal lymphocytes suggest that the inhibitory effect of IL2 is probably exerted via an indirect mechanism. These findings, coupled to the well‐documented ability of IL2 to generate lymphokine activated killer cells cytolytic against leukaemic blasts, further point to the potential role of immunotherapy with IL2 in the management of patients with haematological malignancies.

Interleukin 3 enhances the cytotoxic activity of 1-β-d-arabinofuranosylcytosine (ara-C) on acute myeloblastic leukaemia (AML) cells

We have evaluated the possibility of enhancing the cell killing effect of ara‐C on AML blasts by increasing their proliferative activity with haemopoietic growth factors. Leukaemic cells from 10 AML patients were incubated for 3 d in liquid culture in the presence or in the absence of the human recombinant growth factors IL‐1β (5 U/ml) and IL‐3 (3 U/ml), and subsequently exposed to ara‐C (3 μ/ml) for the last 24 h. The number of residual leukaemic stem cells was evaluated by a clonogenic assay in semisolid medium. The results showed that ara‐C exposure inhibits the proliferation of a higher proportion of clonogenic cells in cultures pretreated with growth factors than in the controls (mean inhibitory values: in the absence of growth factors = 49.8%: with IL‐1β=58.3%; with IL‐3 78.9%) The effect was statistically significant only when IL‐3 was used as a growth factor.

Thymoma: inter-relationships among World Health Organization histology, Masaoka staging and myasthenia gravis and their independent prognostic significance: a single-centre experience §

OBJECTIVE In thymomas, World Health Organization (WHO) histology, Masaoka stage and myasthenia gravis (MG) have long been considered important for patient management and outcome. Their role has been independently investigated in the past. Few studies, however, focussed on the correlations among these variables. The aim of the present study was to retrospectively evaluate, in our patient population of resected thymomas, the inter-relationships among MG, WHO histology and Masaoka stage, and to look at how and to what extent one variable is associated with the other two in terms of clinical presentation and survival. METHODS From January 1990 to October 2008, 255 patients received resection of thymoma. MG was present in 105 cases (41%). Histology by WHO was: 25 A (10%), 72 AB (28%), 65 B1 (25%), 69 B2 (27%) and 24 B (9%). Masaoka staging was stage I, 54 cases (21%), stage II, 86(34%), stage III 79 (31%), and stage IVA 36 (14%). Ordinal and logistic regression models were undertaken to analyse correlations among ordinal (WHO histology and Masaoka stage) and categorical (MG, A vs B WHO types) variables. Univariate and multivariate survival analysis were also performed using the same covariates. Overall survival (OS) and disease-free survival (DFS) were calculated. RESULTS MG was associated with early Masaoka stages (odds ratio (OR) 0.45, 95% confidence interval (CI) 0.33-0.62) and B-type thymomas (OR 1.59, 95% CI 1.23-2.05). B-type thymomas were associated with high Masaoka stage (OR 0.46, 95% CI 0.36-0.60). High Masaoka stage was associated with non-MG (OR 3.27; 95% CI 2.00-5.34). In univariate survival analysis, MG (p = 0.01) and Masaoka stage (p = 0.0001) were significant prognostic indicators using OS. Using DFS, WHO histology (A/AB vs B1/B2/B3 types) (p = 0.05) and Masaoka stage (p = 0.0001) had a prognostic significance. In multivariate analysis, only Masaoka stage was an independent prognostic covariate using OS (hazard ratio (HR) 2.57, 95% CI 1.46-4.52, p = 0.001) and DFS (HR 3.18, 95% CI 1.56-6.52, p = 0.001). CONCLUSIONS In thymomas, MG, WHO histology and Masaoka stage are inter-related. MG has an influence on histology and stage at presentation, while two clinical/histologic patterns are more likely: early Masaoka stage A/AB WHO type and high Masaoka stage/B WHO type. Among the three factors, only Masaoka stage had a prognostic significance on OS and DFS. Our results suggest that a consistent staging system for thymomas should take into account all three variables.

Different sensitivity of normal and leukaemic progenitor cells to Ara-C and IL-3 combined treatment.

Summary. In the present study the effects of a combined treatment with cytosine‐arabinoside (Ara‐C) and inter‐leukin‐3 (IL‐3) on acute myeloblastic leukaemia clonogenic cells and on normal haemopoietic progenitors was investigated, with the aim of improving the tumoricidal effect of cycle specific drugs.

Lymphokine-activated killer (LAK) cells inhibit the clonogenic growth of human leukemic stem cells

The effect of lymphokine‐activated killer (LAK) cells on the in vitro clonogenic capacity of acute myeloid leukemia (AML) blasts was investigated in a semisolid medium assay. The leukemic clonogenic capacity of 11 AML cases, selected on the basis of their ability to grow in vitro, was highly reduced following overnight preincubation with LAK effectors. The degree of colony inhibition, which ranged between 66% and 98% (mean 83.8% ± 11.4 SD), was quantitatively greater than by 51Cr release, which gave rise to lytic values between 5% and 65% (mean 43.2% ± 19.2 SD). The demonstration that the clonogenic inhibition was still induced following a shorter pre‐incubation period (4 hours) suggests that the effect is unlikely to be due only to the generation of cytotoxic activity during the incubation time. The possibility that LAK cells may be employed in the management of residual disease is strengthened by the evidence that the clonogenic potential of samples containing as few as 20% and 14.3% leukemic cells could be almost completely abolished by LAK effectors. These findings further point the possible role of adoptive immunotherapy with interleukin 2/LAK cells in the treatment of patients with acute leukemia.

Complete response of metastatic cutaneous squamous cell carcinoma to cetuximab plus paclitaxel

BACKGROUND AND METHODS Cetuximab therapy results strongly active in advanced cutaneous squamous cell carcinoma (cSCC). A patient affected by a rapidly progressing, already irradiated and cisplatin-refractory cSCC, with lung, pleura and thoracic lymph nodes metastasis, was treated with weekly cetuximab and paclitaxel. RESULTS Treatment was well tolerated and a partial response was obtained after four months of cetuximab plus paclitaxel therapy. Then we continued maintenance cetuximab for another seven months with tumor shrinkage until complete response, maintained after six months. CONCLUSIONS Cetuximab was safely associated with paclitaxel, obtaining a rapid tumor response in cisplatin-refractory metastatic cSCC. Single-agent cetuximab maintenance sustained tumor shrinkage until complete response.

Induction of proliferation of acute myeloblastic leukemia (AML) cells with hemopoietic growth factors

Like their normal counterparts, leukemic blasts have recently been shown to respond to hemopoietic growth factors in both suspension culture and in semisolid media. In the present study, we have evaluated the proliferative response of 35 AML cases to colony-stimulating factors (CSFs) containing conditioned media derived from the human cell lines GCT, 5637, MO and MG U87, and to human recombinant IL-1 (rh-IL1), IL-3 (rhIL-3), GM-CSF (rhGM-CSF) and G-CSF (rhG-CSF). In the great majority of cases, an increase of 3H-thymidine (3H-TdR) uptake was obtained in response to at least one conditioned medium. The labeling index (LI) and the growth fraction (GF), evaluated in a restricted group of cases, were also increased by the growth factors, suggesting that they act by recruiting leukemic cells in cycle from the resting compartment. The ability of blast populations to form colonies was also studied. Conditioned media were found to induce or significantly increase the clonogenic capacity in 20 cases out of 22. The response of leukemic cells to human recombinant CSFs and rhIL-1, used alone or in combination, was also assayed. The results, in agreement with those obtained with conditioned media, show that each leukemic case displays a different pattern of response to CSFs, and that optimal growth conditions must be individually assessed. The possibility of increasing the fraction of cycling cells in AML populations may represent a way to render them more sensitive to cytostatic agents, with a view to new therapeutic strategies.

Lifetime growth and risk of testicular cancer

Adult height is associated with testicular cancer risk. We studied to what extent this association is explained by parental height, childhood height and age at puberty. We conducted a case–control study on germ‐cell testicular cancer patients diagnosed in 1997–2008 and resident in the Province of Turin. Information was collected using mailed questionnaires in 2008–2011. Specifically, we asked for adult height (in cm), height at age 9 and 13 (compared to peers) and age at puberty (compared to peers). We also asked for paternal and maternal height (in cm) as indicators of genetic components of adult height. The analysis included 255 cases and 459 controls. Odds ratios (ORs) of testicular cancer were estimated for the different anthropometric variables. Adult height was associated with testicular cancer risk [OR: 1.16, 95% confidence interval (CI): 1.03–1.31 per 5‐cm increase]. The risk of testicular cancer was only slightly increased for being taller vs. shorter than peers at age 9 (OR: 1.55, 95% CI: 0.91–2.64) or age 13 (OR: 1.26, 95% CI: 0.78–2.01), and parental height was not associated with testicular cancer risk. The OR for adult height was 1.32 (95% CI: 1.12–1.56) after adjustment for parental height. Among participants with small average parental height (<167 cm or less), the OR of testicular cancer for tall (>180 cm) vs. short (<174 cm) subjects was 3.47 (95% CI: 1.60–7.51). These results suggest that the association between height and testicular cancer is likely to be explained by environmental factors affecting growth in early life, childhood and adolescence.

Baldness and testicular cancer: the EPSAM case-control study.

The etiology of testicular cancer is largely unexplained. Research has mainly focused on prenatal exposures, especially to sex hormones, while less attention has been paid to exposures that may act also postnatally. As baldness has been previously associated with testicular cancer risk we focused on baldness and body hairiness, which are both associated with androgen activity. We used data of the Postnatal Exposures and Male Health (EPSAM) study, a case–control study on testicular cancer conducted in the Province of Turin, Italy, involving cases diagnosed between 1997 and 2008. Information was collected using mailed questionnaires. Analyses included 255 cases and 459 controls. We calculated ORs and 95% CIs to estimate testicular cancer risk among those who developed baldness and among those with body hairiness. We found an inverse association between testicular cancer and baldness (OR: 0.67, 95% CI: 0.46–0.98) and body hairiness (OR: 0.78, 95% CI: 0.53–1.16), although the latter had wider CIs. The inverse association between baldness and testicular cancer is consistent with the results from previous studies. These results suggest that androgens activity may influence testicular cancer risk.