Immune checkpoint inhibitors for POLE or POLD1 proofreading-deficient metastatic colorectal cancer.

BACKGROUND: POLE and POLD1 proofreading deficiency (POLE/D1pd) define a rare subtype of ultramutated metastatic colorectal cancer (mCRC; over 100 mut/Mb). Disease-specific data about the activity and efficacy of immune checkpoint inhibitors (ICIs) in POLE/D1pd mCRC are lacking and it is unknown whether outcomes may be different from mismatch repair-deficient (dMMR)/microsatellite instability-high (MSI-H) mCRCs treated with ICIs. PATIENTS AND METHODS: In this global study, we collected 27 patients with mCRC harboring POLE/D1 mutations leading to proofreading deficiency and treated with anti-programmed cell death-ligand 1 alone +/- anti-cytotoxic T-lymphocyte antigen-4 agents. We collected clinicopathological and genomic characteristics, response, and survival outcomes after ICIs of POLE/D1pd mCRC and compared them with a cohort of 610 dMMR/MSI-H mCRC patients treated with ICIs. Further genomic analyses were carried out in an independent cohort of 7241 CRCs to define POLE and POLD1pd molecular profiles and mutational signatures. RESULTS: POLE/D1pd was associated with younger age, male sex, fewer RAS/BRAF driver mutations, and predominance of right-sided colon cancers. Patients with POLE/D1pd mCRC showed a significantly higher overall response rate (ORR) compared to dMMR/MSI-H mCRC (89% versus 54%; P = 0.01). After a median follow-up of 24.9 months (interquartile range: 11.3-43.0 months), patients with POLE/D1pd showed a significantly superior progression-free survival (PFS) compared to dMMR/MSI-H mCRC [hazard ratio (HR) = 0.24, 95% confidence interval (CI) 0.08-0.74, P = 0.01] and superior overall survival (OS) (HR = 0.38, 95% CI 0.12-1.18, P = 0.09). In multivariable analyses including the type of DNA repair defect, POLE/D1pd was associated with significantly improved PFS (HR = 0.17, 95% CI 0.04-0.69, P = 0.013) and OS (HR = 0.24, 95% CI 0.06-0.98, P = 0.047). Molecular profiling showed that POLE/D1pd tumors have higher tumor mutational burden (TMB). Responses were observed in both subtypes and were associated with the intensity of POLE/D1pd signature. CONCLUSIONS: Patients with POLE/D1pd mCRC showed more favorable outcomes compared to dMMR/MSI-H mCRC to treatment with ICIs in terms of tumor response and survival.

Primary and metastatic brain cancer genomics and emerging biomarkers for immunomodulatory cancer treatment.

Recent studies with immunomodulatory agents targeting both cytotoxic T-lymphocyte protein 4 (CTLA4) and programmed cell death 1 (PD1)/programmed cell death ligand 1 (PDL1) have shown to be very effective in several cancers revealing an unexpected great activity in patients with both primary and metastatic brain tumors. Combining anti-CTLA4 and anti-PD1 agents as upfront systemic therapy has revealed to further increase the clinical benefit observed with single agent, even at cost of higher toxicity. Since the brain is an immunological specialized area it's crucial to establish the specific composition of the brain tumors' microenvironment in order to predict the potential activity of immunomodulatory agents. This review briefly summarizes the basis of the brain immunogenicity, providing the most updated clinical evidences in terms of immune-checkpoint inhibitors efficacy and toxicity in both primary and metastatic brain tumors with the final aim of defining potential biomarkers for immunomodulatory cancer treatment.

Distribution of SNAP25, VAMP1 and VAMP2 in mature and developing deep cerebellar nuclei after estrogen administration.

Synaptosomal-associated protein of 25kDa (SNAP25), vesicle-associated membrane protein 1 (VAMP1) and 2 (VAMP2) are components of soluble N-ethylmaleimide-sensitive fusion attachment protein receptors (SNARE) complex which is involved in synaptic vesicle exocytosis, a fundamental step in neurotransmitter release. SNARE expression in cerebellum correlates with specific neurotransmitter pathways underlying synaptic diversification and defined synaptic properties. In this study we firstly characterized the distribution of SNAP25, VAMP1 and VAMP2 in the nerve terminals of a defined cerebellar region, the deep cerebellar nuclei (DCN), of adult and newborn rats. Then, given the pivotal role of estradiol (E2) in the synaptic organization of the cerebellar circuitry in early postnatal life, we examined whether administration of E2 in the newborn DCN affected synaptic density and changed the distribution of the presynaptic proteins SNAP25, VAMP1 and VAMP2, together with post synaptic density protein 95 (PSD95). Results showed that: (1) distribution of SNAP25, VAMP1 and VAMP2 in adult DCN differs significantly from that found in newborn DCN; (2) administration of E2 in the newborn DCN affected synaptic density and also changed the distribution of the pre- and postsynaptic proteins. The differential distribution of SNAP25, VAMP1 and VAMP2 in nerve terminals of adult and newborn rats may correlate with specific stages of neuronal phenotypic differentiation. The effects of E2 on SNAP25, VAMP1, VAMP2, PDS95 and synaptic density suggest that pre- and postsynaptic proteins are under estrogenic control during development and that synaptic maturation can also be related with the activity of this steroid.

Immunohistochemical localisation and molecular expression of the steroidogenic enzyme cytochrome P450 17α-hydroxylase /C(17,20)-lyase in the vestibular nuclei of adult male rats.

Many biologically active neurosteroids, including dehydroepiandrosterone (DHEA), are synthesised in the brain. DHEA is a potent endogenous modulator of several neuronal functions, and alterations of DHEA are correlated with various neurobiological deficits. The cytochrome P450 17α-hydroxylase/C(17,20)-lyase (P450C(17) ) plays a pivotal role in the synthesis of DHEA from pregnenolone and progesterone. We investigated the immunohistochemical localisation and molecular expression of P450C(17) in the superior, lateral, medial and inferior vestibular nuclei (VCN) of adult male rats by western blotting and indirect immunofluorescence analysis. Immunoreactive P450C(17) was widely distributed in all VCN and the expression of P450C(17) was confirmed by western blot analysis. The present study demonstrates, for the first time, the presence and anatomical distribution of P450C(17) in the VCN. Given that neurosteroids can modulate neuronal activities in the medial vestibular nucleus, DHEA synthesised in the VCN may play an important role in the control of specific activities at this level.

2-methoxyestradiol induces morpho-functional changes and impairs the microtubular system in mouse neuroblastoma and rat glioma cells.

2-Methoxyestradiol (2ME), a metabolite deriving from 17-beta estradiol, is a well-established antiangiogenic, apoptotic and antiproliferative agent in cell cultures and animal models. 2ME may also exert its cytotoxic activity by interacting with tubulin and by causing an impairment of the microtubular system. The aim of this study was to investigate the relative effectiveness of 2ME on mouse neuroblastoma (C1300) and rat glioma (C6) cell lines in inducing morpho-functional changes and alteration of the microtubular system physiology. Cells, cultured in a medium supplemented with increasing 2ME micromolar concentrations, were submitted to morphological investigations, MTT assay and western blot analysis. 2ME-exposed cell lines displayed in comparison with control cells, morpho-functional changes such as reduction in cell number, a globular/shrunken shape, retraction or absence of cytoplasmic processes, inhibition of cell growth and cell decreased viability. Interestingly, all changes detected were more evident in C1300 cells than in C6 cells. Western blot analysis showed that the total and the tyrosinated a-tubulin expression was reduced more intensely in the C1300 than in C6 cells; whereas the acetylated a-tubulin expression did not significantly decrease in either cell lines. Results demonstrate that 2ME is more effective in neural cells than in glial cells. The alteration of total and tyrosinated a-tubulin expression suggests that 2ME effectiveness could be strictly related to an impairment of microtubule system physiology resulting in morpho-functional changes, block of mitosis and cell death.

D-glucose induces microtubular changes in C1300 neuroblastoma cell line through the incorporation of 3-nitro-L-tyrosine into tubulin.

The microtubular network of neurons is involved in several functions such as formation and tropism of cellular processes, cell division and intracellular transport. A lot of evidences testify that the microtubular network of neurons can be impaired by oxidative stress. A condition of oxidative stress is often possible when D-glucose overloads its metabolic pathway, resulting in an increase in reactive oxygen species and subsequent neurological disorders. The aim of this work was to check in undifferentiated mouse neuroblastoma cells (C1300) the possible oxidative effects of D-glucose on microtubules. Using a concentration of 110mM D-glucose, cell morphology, growth rate, viability and catalase activity were seriously altered. Noteworthy, an increase in 3-nitro-L-tyrosine and a downregulation of tubulins was found in D-glucose-exposed cells, whereas another cytoskeletal proteins, namely actin, did not show any changes. In conclusion, microtubular network can be impaired by D-glucose through specific nitrosative effects, suggesting a possible mechanism at the basis of hyperglycemia-induced neuronal damage.

Comparative bone histology of adult horses (Equus caballus) and cows (Bos taurus).

Bone microstructure of domestic herbivores is still not completely understood. Indeed, works focused on the bone histology of numerous Mammalian species frequently led to misunderstandings because of the high number of variations such as the kind of bone, section orientation, species, breed and age. Moreover, attempts to identify the species in archaeozoological studies by a mere qualitative approach have not been encouraging and in recent years quantitative methods, based on image processing and statistical analysis, have appeared. The present study was undertaken to determine whether morphometrical and morphological differences exist in the compact bone structure of the femur and humerus between horses and cows. Measurements such as area, perimeter, minimum and maximum diameter of osteons and Haversian canals as well as the osteonal density were carried out on cross sections of eight humeri and eight femurs of the two herbivores investigated. In agreement with other authors, the qualitative investigation confirmed that the compact bone of horses and cows can be classified as dense Haversian tissue. Osteons of the horse were more numerous and composed of a higher number of well-defined lamellae when compared with the cow. Diameter, perimeter and area of osteons and Haversian canals were always higher in horses than in cows and this pattern could be related to the different locomotor behaviour of these animals.

Ancient pompeian dogs--morphological and morphometric evidence for different canine populations.

This article examines the morphological features of the dog during the Roman Age on the basis of osseous and dental remains dug up in Pompeii. The material, consisting of 113 canine bones and teeth, was subjected to both morphological and morphometrical analyses and was compared with modern canine breeds. In most cases, the age at death, shoulder height and other phenotypic features were ascertained. The examined Pompeian canine population fell mainly into two categories: small- and large-sized animals. Among the former, one brachycephalic and two dolichocephalic subjects were included. Such morphological features agree with what is described in numerous texts and appears in mosaics, bas-reliefs and frescoes of the Roman Age. As small-sized dogs cannot be classified as Canes Venatici (sporting dogs), Canes Villatici (watch dogs) and Canes Pastorales (shepherd dogs) according to Columella's De re rustica, these animals may be considered as lapdogs.

Testosterone induces neuroprotection from oxidative stress. Effects on catalase activity and 3-nitro-L-tyrosine incorporation into alpha-tubulin in a mouse neuroblastoma cell line.

3-nitro-L-tyrosine is formed by nitric oxide following different pathways such as NADPH oxidase, xanthine oxidase or glutamate NMDA receptor activation and is involved in the pathology of different neurological disorders. Unlike estradiol, a neuroprotective role of androgens against oxidative cell injury has not been fully investigated. This work targets the possible effects of testosterone on neuroblastoma cells exposed to 3-nitro-L-tyrosine. C1300 mouse undifferentiated neuroblastoma cells exposed to 3-nitro-L-tyrosine were cultured in the presence of testosterone. Morphological examination, proliferation and nuclear viability assays were performed. The expression of tyrosinated alpha-tubulin and incorporation of 3-nitro-L-tyrosine into protein were also estimated. Cells exposed to 3-nitro-L-tyrosine showed globular shape, reduced cytoplasmic processes and growth inhibition in comparison with controls. When testosterone was added to the medium, these changes were not evident. In addition, testosterone induced an upregulation of tyrosinated alpha-tubulin, a marker of neuronal plasticity, and a decrease in 3-nitro-L-tyrosine incorporation into tubulin. Our results suggest that testosterone exposure can diminish 3-nitro-L-tyrosine toxic effects on the morphology and growth rate of neuroblastoma cells. The upregulation of tyrosinated alpha-tubulin in testosterone-exposed cells would be consistent with concurrent plasticity events. Failure in alpha-tubulin nitration detected in cells exposed to both 3-nitro-L-tyrosine and testosterone, may support the idea that testosterone interferes with 3-nitro-L-tyrosine protein incorporation. Moreover, testosterone-induced neuroprotection likely entails a linkage with the androgen receptor as is suggested by the flutamide-induced inhibition of the hormone activity. Finally, the neuroprotective effects of testosterone in neuroblastoma cells could deal with the cellular antioxidant defence system, as shown by testosterone-induced increase in catalase activity.

[Silica, silicosis, and lung cancer: analysis if the literature and mortality studies of minor workers in Sardinia]. / Silice, silicosi e cancro del polmone: analisi della letteratura e studi di mortalità in minatori della Sardegna.

Starting from a short review of the recent epidemiological studies available in the international literature concerning the association between silica, silicosis and lung cancer, the results of two mortality studies performed in Sardinia are reported. The first study concerns a 20-year follow-up of 1741 miners employed in 1973 in two metalliferous Sardinian mines. In the second study the cause specific mortality of 724 patients with silicosis, firstly diagnosed by standard chest x-ray between 1964 and 1970 in our Institute, has been analysed by a cohort study extended to December 31, 1997. The findings indicate that the slight increased lung cancer mortality observed in these cohorts, more than to the severity of radiological silicosis or to the entity of the cumulative exposure to crystalline silica dust in itself, was significantly associated to other risk factors as cigarette smoking, airflow obstruction and radon-daughters exposure in underground mines.

Mortality from lung cancer among silicotic patients in Sardinia: an update study with 10 more years of follow up.

OBJECTIVES: To evaluate the association between silica, silicosis and lung cancer, the mortality of 724 patients with silicosis, first diagnosed by standard chest x ray film between 1964 and 1970, has been analysed by a cohort study extended to 31 December 1997. METHODS: Smoking and detailed occupational histories were available for each member of the cohort as well as the estimated lifetime exposure to respirable silica dust and radon daughters. Two independent readers blindly classified standard radiographs according to the 12 point International Labour Organisation (ILO) scale. Lung function tests meeting the American Thoracic Society's criteria were available for 665 patients. Standardised mortality ratios (SMRs) for selected causes of death were based on the age specific Sardinian regional death rates. RESULTS: The mortality for all causes was significantly higher than expected (SMR 1.35, 95% confidence interval (95% CI) 1.24 to 1.46) mainly due to tuberculosis (SMR 22.0) and to non-malignant chronic respiratory diseases (NMCRD) (SMR 6.03). All cancer deaths were within the expected numbers (SMR 0.93; 95% CI 0.76 to 1.14). The SMR for lung cancer was 1.37 (95% CI 0.98 to 1.91, 34 observed), increasing to 1.65 (95% CI 0.98 to 2.77) allowing for 20 years of latency since the first diagnosis of silicosis. Although mortality from NMCRD was strongly associated to the severity of radiological silicosis and to the extent of the cumulative exposure to silica, SMR for lung cancer was weakly related to the ILO categories and to the cumulative exposure to silica dust only after 20 years of lag interval. A significant excess of deaths from lung cancer (SMR 2.35) was found among silicotic patients previously employed in underground metal mines characterised by a relatively high airborne concentration of radon daughters and among ever smokers who showed an airflow obstruction at the time of the first diagnosis of silicosis (SMR 3.29). Mortality for lung cancer related to exposure was evaluated with both the Cox's proportional hazards modelling within the entire cohort and a nested case-control study (34 cases of lung cancer and 136 matched controls). Both multivariate analyses did not show any significant association with cumulative exposure to silica or severity of silicosis, but confirmed the association between mortality for lung cancer and relatively high exposure to radon, smoking, and airflow obstruction as significant covariates. CONCLUSIONS: The findings indicate that the slightly increased mortality for lung cancer in this cohort of silicotic patients was significantly associated with other risk factors-such as cigarette smoking, airflow obstruction, and estimated exposure to radon daughters in underground mines-rather than to the severity of radiological silicosis or to the cumulative exposure to crystalline silica dust itself.

Mortality in a cohort of men expressing the glucose-6-phosphate dehydrogenase deficiency.

The objective of this study was to test the hypothesis of a lower mortality from cancer and cardiovascular diseases among men expressing glucose-6-phosphate dehydrogenase (G6PD) deficiency. We designed a mortality study based on death certificates from January 1, 1982 through December 31, 1992 in a cohort of G6PD-deficient men. Cohort members were 1,756 men, identified as expressing the G6PD-deficient phenotype during a 1981 population screening of the G6PD polymorphism. The setting was the island of Sardinia, Italy. Outcome measures were cause-specific standardized mortality ratios (SMRs), which were computed as 100 times the observed/expected ratio, with the general Sardinian male population as the reference. Deaths from all causes were significantly less than expected due to decreased SMRs for ischemic heart disease (SMR, 28; 95% confidence interval [CI], 10 to 62), cerebrovascular disease (SMR, 22; 95% CI, 6 to 55), and liver cirrhosis (SMR, 12; 95% CI, 0 to 66), which explained 95.6% of the deficit in total mortality. All cancer mortality was close to the expectation, with a significant increase in the SMR for non-Hodgkin's lymphoma (SMR, 545; 95% CI, 147 to 1,395). A decrease in mortality from cardiovascular diseases was one of the study hypotheses, based on an earlier human report and experimental evidence. However, selection bias is also a likely explanation. Further analytic studies are warranted to confirm whether subjects expressing the G6PD-deficient phenotype are protected against ischemic heart disease and cerebrovascular disease. This cohort study is consistent with more recent case-control studies in rejecting the hypothesis of a decreased cancer risk among G6PD-deficient subjects. The observed increase in mortality from non-Hodgkin's lymphoma and decrease in mortality from liver cirrhosis were not previously reported.

Occupational exposures as risk factors for gastric cancer in Italy.

Occupational associations with gastric cancer were investigated in a multicenter case-control study in Italy involving interviews with 640 histologically confirmed male cases and 959 controls, randomly selected from the resident populations of the study areas. From information on the three jobs each person held the longest, risks were evaluated according to employment in 35 occupations (ever or 21+ years) and to estimated exposure (ever or 21+ years) to six chemicals using a job-exposure matrix. All risk estimates were adjusted by personal, demographic, and dietary variables identified as gastric-cancer risk factors in previous analyses. The only significantly increased risk was observed for sailors, seamen, and allied groups (ever employed: odds ratio [OR] = 2.9; 95 percent confidence interval [CI] = 1.1-8.0; 21+ years: OR = 3.1, CI = 0.8-13). Nonsignificant increases after 21+ years of employment were observed for forestry workers, miners, and janitors and cleaners. Crude ORs were elevated significantly among farmers, but adjusting for demographic and lifestyle factors largely eliminated the association: a nonsignificant 30 percent excess risk remained for farm laborers, but there was no rise in risk among long-term farm laborers and no excess among farm owners. Application of the job-exposure matrix revealed excess risks of borderline significance associated with potential exposure to mineral dusts and nitrogen oxides. For subjects with 21+ years of potential exposure, nonsignificantly increased risks were related to mineral dusts, asbestos, fertilizers, and nitrosamines.(ABSTRACT TRUNCATED AT 250 WORDS)

Helicobacter pylori antibodies in areas of Italy at varying gastric cancer risk.

In a survey of 930 adults aged 35-74 years randomly sampled from the general population of four areas of Italy, two at low and two at high risk for gastric cancer, plasma levels of Helicobacter pylori IgG antibodies were assayed in order to investigate associations with the geographical distribution of gastric cancer and other dietary and life-style factors, as assessed by personal interview. H. pylori positivity (antibody titer above or equal to 10 micrograms/ml), 45% overall, increased with age and was inversely associated with social class but showed little geographical variation or association with dietary variables and blood nutrients. H. pylori positivity was also associated with increased blood levels of pepsinogens, particularly pepsinogen II. The authors discuss these findings in relation to those from a previous case-control study of gastric cancer in the same areas.

Antispasmodic activity of tert-aminoalkylderivatives of 3-benzyl-, 3-benzylaza- and 3-benzyldiazaquinoxalinones.

Tert-aminoalkylderivatives of quinoxalin-2-ones, aza- and diazaquinoxalin-2-ones bearing in position 3 a benzyl group were assayed to evaluate the antispasmodic activity. The tested compounds exhibited moderate aspecific antispastic properties that do not warrant further investigation.

Nitrate and N-nitrosoproline excretion in two Italian regions with contrasting rates of gastric cancer: the role of nitrate and other factors in endogenous nitrosation.

Exposure to nitrate and propensity for endogenous nitrosation were examined in 80 healthy males, aged 25-40 years, residing in areas of Italy with long-standing high (Florence) and low (Cagliari) rates of gastric cancer. Nitrate exposure was assessed by measurement of urinary nitrate excretion over 12 hr, and endogenous nitrosation was assessed using the N-nitrosoproline test (NPRO-test). Our hypothesis was whether the geographic variation in cancer rate correlated with nitrate exposure or nitrosating ability. Exposure to background sources of NPRO was significantly higher in the high-risk subjects (phi = 0.04) whereas no differences were found in exposure to nitrate or in urinary NPRO levels after L-proline loading (test NPRO levels). The regional difference in test NPRO was almost completely accounted for by background NPRO exposure. Examination of individual rather than grouped data revealed that exposure to nitrate was a major factor in NPRO formation. No other factors studied (age, dietary-questionnaire-assessed intake of anti-oxidant vitamins) had a significant effect. Geographical variation in gastric cancer risk did not, therefore, correlate with either nitrate exposure or propensity for endogenous nitrosation of L-proline.

Synthesis and choleretic activity of 3-[2-(3-R', 4-R'', 5-R'''-benzyl)-5-R-benzimidazol-1-yl]-butanoic acids.

On the ground of the evidentiated choleretic activity of 3-[2-benzylbenzimidazol-1-yl]butanoic acid, 28 new acids were prepared in order to evaluate the influence of suitable substitutions in either C5 of heteroring or C3', C4', C5' of benzyl group in position 2 on the choleretic activity. Pharmacological results after i.v. administration of 0.5 mmol/Kg in rats confirmed a general high choleretic activity that in eleven cases showed during the first 4 hours an increase of bile volume higher than 80%, that is superior to that produced by dehydrocholic acid. Only in a few cases the bile volume increase was less than 37% of basal value.

Plasma pepsinogens, nutrients, and diet in areas of Italy at varying gastric cancer risk.

In a survey of 930 adults aged 35-74 years randomly sampled from the general population of four areas of Italy at different risks for gastric cancer (GC), plasma levels of pepsinogens (PGI and PGII) and fat-soluble vitamins were assayed. Pepsinogen levels were used to identify individuals with chronic atrophic gastritis (CAG). Severe CAG (PGI < or = 20 pg/liter) affected 5.8% of the population, but the prevalence rose with increasing age and declining social class. Severe CAG was 5 times more common in areas with high compared to low rates of GC. Risk also rose with increasing consumption of salted/dried fish but was inversely related to dietary intake of beta-carotene and to plasma retinol and cholesterol levels. The prevalence of moderate CAG (PGI > 20 pg/liter, but PGI/PGII < or = 2.9) was 6.3%. Moderate CAG was also related to age and social class and increased 1.8-fold in areas where GC rates were high, but was not strongly associated with diet or plasma nutrients. The authors discuss these findings in relation to those from a previous case-control study of GC in the same areas.

[Tert-aminoalkyl derivatives of quinoxalinones, aza- and diazaquinoxalinones with analgesic activity]. / Terz-amminoalchilderivati di chinossalinoni, aza e diazachinossalinoni ad attività analgesica.

A series of tert-aminoalkyl-derivatives of quinoxalin-2-one, aza- and diazaquinoxalin-2-one bearing in position 3 a benzyl group was prepared in order to compare with analogous 3-methyl derivatives as regards analgesic activity. The substitution causes various effects. In compounds (I) and (VI-IX) is found the expected increase in analgesic activity but with contemporaneous rise in toxicity. The compounds (IV) and (V) are of interest due to the presence of a strong separation of DL50 from DE50.