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Papers by Andrea Gallotta

Journal of Hepatology, 2011

Cancer biomarkers : section A of Disease markers, 2013

Prostate cancer (PCa) represents the most common solid tumor affecting men and its early detectio... more Prostate cancer (PCa) represents the most common solid tumor affecting men and its early detection remains the best approach to improve survival rates. The assessment of serum levels of PSA is currently used for PCa screening but the low specificity of the test results in a high number of false positives. Other forms of PSA may be detected in the bloodstream including PSA associated with immunoglobulin M (PSA-IgM) which, alone or combined with PSA, has shown diagnostic accuracy for PCa. The aim of the study is to improve the diagnostic accuracy of PSA-IgM by developing a multivariable model which includes serum biomarkers and routine diagnostic parameters to obtain a predictive index useful in the post-screening clinical practice. One hundred sixty male patients with clinical suspect of PCa underwent a trans-rectal ultrasound guided first prostate biopsy with a standardized sampling scheme. To generate the model, we assessed the presence of PSA and PSA-IgM complexes in sera of patie...

Clinics in Laboratory Medicine, 2012

Journal of Molecular Recognition, 2011

A novel dendrimeric peptide ligand dubbed D-PAM-Φ was designed to achieve a high capacity for hum... more A novel dendrimeric peptide ligand dubbed D-PAM-Φ was designed to achieve a high capacity for human IgG through the decoration of the D-PAM scaffold. The design criteria based on the introduction of small hydrophobic groups on the D-PAM structure were supported by the recently published solid-state structure of D-PAM complexed to the Fc fragment of a recombinant human IgG1 and by molecular dynamic simulations that provided information on the mode of binding of phenylacetyl-D-PAM (D-PAM-Φ). D-PAM-Φ was immobilised on an activated solid support and compared with the parent D-PAM affinity matrix. The newly obtained affinity sorbent was evaluated for its capacity to selectively capture polyclonal human IgG; the binding capacity was approximately 10 mg/ml, an almost 10-fold enhancement with respect to the D-PAM-functionalised matrices without the specificity of binding being reduced. The new ligand was also effective in the capturing of recombinant humanised IgG1 from a clarified cell culture supernatant. Under a typical laboratory-scale affinity chromatography assembly and preliminarily optimised binding conditions, the affinity purification of humanised IgG1 from culture supernatants rendered the desired product, with purity higher than 90%. The results suggest that the application of the computational approach on the structure of the D-PAM-Fc complex may be very valuable in the development of novel lead molecules for the downstream processing of human or humanised antibodies used in therapy.

Journal of Molecular Recognition, 2010

Journal of Hepatology, 2011

Journal of Hepatology, 2006

Journal of Enzyme Inhibition and Medicinal Chemistry, 2012

Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibit... more Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibitors. Validate the use of recombinant neuraminidase expressed in baculovirus located on the viral surface capsule to develop a neuraminidase inhibitor screening assay. Recombinant baculovirus particles displaying neuraminidase N1 and N3 were used as enzyme sources. The assay set-up required the use of 2'-(4-methylumbelliferyl)-α-D-acetyl neuraminic acid as substrate and oseltamivir carboxylate as benchmark inhibitor. The assay was set up in a standard 96-well plate. The within- and between-assay coefficients of variation were, on average, less than 10%. The 50% inhibitory concentration values of the inhibitor were in good agreement with those determined by independent kinetic experiments. The assay showed satisfactory within- and between-assay repeatability. The obtained results suggest that recombinant baculovirus expressing neuraminidase located on the virus membrane capsule can be used to set up affordable and reliable neuraminidase inhibitors screening assays.

Journal of Clinical Virology, 2006

Journal of Biomolecular Screening, 2011

European Journal of Clinical Investigation, 2008

Digestive and Liver Disease, 2011

Digestive and Liver Disease, 2008

Digestive and Liver Disease, 2009

Digestive and Liver Disease, 2009

Clinical Chemistry and Laboratory Medicine, 2000

Cancer Detection and Prevention, 2007

Bioorganic & Medicinal Chemistry, 2012

Evidences of oseltamivir resistant influenza patients raised the need of novel neuraminidase inhi... more Evidences of oseltamivir resistant influenza patients raised the need of novel neuraminidase inhibitors. In this study, five oseltamivir analogs PMC-31-PMC-36, synthesised according to the outcomes of a rational design analysis aimed to investigate the effects of substitution at the 5-amino and 4-amido groups of oseltamivir on its antiviral activity, were screened for their inhibition against neuraminidase N1 and N3. The enzymes used as models were from the avian influenza A H7N1 and H7N3 viruses. The neuraminidase inhibition assay was carried out by using recombinant species obtained from a baculovirus expression system and the fluorogenic substrate MUNANA. The assay was validated by using oseltamivir carboxylate as a reference inhibitor. Among the tested compounds, PMC-36 showed the highest inhibition on N1 with an IC(50) of 14.6±3.0nM (oseltamivir 25±4nM), while PMC-35 showed a significant inhibitory effect on N3 with an IC(50) of 0.1±0.03nM (oseltamivir 0.2±0.02nM). The analysis of the inhibitory properties of this panel of compounds allowed a preliminary assessment of a structure-activity relationship for the modification of the 4-amido and 5-amino groups of oseltamivir carboxylate. The substitution of the acetamido group in the oseltamivir structure with a 2-butenylamido moiety reduced the observed activity, while the introduction of a propenylamido group was well tolerated. Substitution of the free 5-amino group of oseltamivir carboxylate with an azide, decreased the activity against both N1 and N3. When these structural changes were both introduced, a dramatic reduction of activity was observed for both N1 and N3. The alkylation of the free 5-amino group in oseltamivir carboxylate introducing an isopropyl group seemed to increase the inhibitory effect for both N1 and N3 neuraminidases, displaying a more pronounced effect against N1.

Journal of Hepatology, 2011

Cancer biomarkers : section A of Disease markers, 2013

Prostate cancer (PCa) represents the most common solid tumor affecting men and its early detectio... more Prostate cancer (PCa) represents the most common solid tumor affecting men and its early detection remains the best approach to improve survival rates. The assessment of serum levels of PSA is currently used for PCa screening but the low specificity of the test results in a high number of false positives. Other forms of PSA may be detected in the bloodstream including PSA associated with immunoglobulin M (PSA-IgM) which, alone or combined with PSA, has shown diagnostic accuracy for PCa. The aim of the study is to improve the diagnostic accuracy of PSA-IgM by developing a multivariable model which includes serum biomarkers and routine diagnostic parameters to obtain a predictive index useful in the post-screening clinical practice. One hundred sixty male patients with clinical suspect of PCa underwent a trans-rectal ultrasound guided first prostate biopsy with a standardized sampling scheme. To generate the model, we assessed the presence of PSA and PSA-IgM complexes in sera of patie...

Clinics in Laboratory Medicine, 2012

Journal of Molecular Recognition, 2011

A novel dendrimeric peptide ligand dubbed D-PAM-Φ was designed to achieve a high capacity for hum... more A novel dendrimeric peptide ligand dubbed D-PAM-Φ was designed to achieve a high capacity for human IgG through the decoration of the D-PAM scaffold. The design criteria based on the introduction of small hydrophobic groups on the D-PAM structure were supported by the recently published solid-state structure of D-PAM complexed to the Fc fragment of a recombinant human IgG1 and by molecular dynamic simulations that provided information on the mode of binding of phenylacetyl-D-PAM (D-PAM-Φ). D-PAM-Φ was immobilised on an activated solid support and compared with the parent D-PAM affinity matrix. The newly obtained affinity sorbent was evaluated for its capacity to selectively capture polyclonal human IgG; the binding capacity was approximately 10 mg/ml, an almost 10-fold enhancement with respect to the D-PAM-functionalised matrices without the specificity of binding being reduced. The new ligand was also effective in the capturing of recombinant humanised IgG1 from a clarified cell culture supernatant. Under a typical laboratory-scale affinity chromatography assembly and preliminarily optimised binding conditions, the affinity purification of humanised IgG1 from culture supernatants rendered the desired product, with purity higher than 90%. The results suggest that the application of the computational approach on the structure of the D-PAM-Fc complex may be very valuable in the development of novel lead molecules for the downstream processing of human or humanised antibodies used in therapy.

Journal of Molecular Recognition, 2010

Journal of Hepatology, 2011

Journal of Hepatology, 2006

Journal of Enzyme Inhibition and Medicinal Chemistry, 2012

Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibit... more Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibitors. Validate the use of recombinant neuraminidase expressed in baculovirus located on the viral surface capsule to develop a neuraminidase inhibitor screening assay. Recombinant baculovirus particles displaying neuraminidase N1 and N3 were used as enzyme sources. The assay set-up required the use of 2'-(4-methylumbelliferyl)-α-D-acetyl neuraminic acid as substrate and oseltamivir carboxylate as benchmark inhibitor. The assay was set up in a standard 96-well plate. The within- and between-assay coefficients of variation were, on average, less than 10%. The 50% inhibitory concentration values of the inhibitor were in good agreement with those determined by independent kinetic experiments. The assay showed satisfactory within- and between-assay repeatability. The obtained results suggest that recombinant baculovirus expressing neuraminidase located on the virus membrane capsule can be used to set up affordable and reliable neuraminidase inhibitors screening assays.

Journal of Clinical Virology, 2006

Journal of Biomolecular Screening, 2011

European Journal of Clinical Investigation, 2008

Digestive and Liver Disease, 2011

Digestive and Liver Disease, 2008

Digestive and Liver Disease, 2009

Digestive and Liver Disease, 2009

Clinical Chemistry and Laboratory Medicine, 2000

Cancer Detection and Prevention, 2007

Bioorganic & Medicinal Chemistry, 2012

Evidences of oseltamivir resistant influenza patients raised the need of novel neuraminidase inhi... more Evidences of oseltamivir resistant influenza patients raised the need of novel neuraminidase inhibitors. In this study, five oseltamivir analogs PMC-31-PMC-36, synthesised according to the outcomes of a rational design analysis aimed to investigate the effects of substitution at the 5-amino and 4-amido groups of oseltamivir on its antiviral activity, were screened for their inhibition against neuraminidase N1 and N3. The enzymes used as models were from the avian influenza A H7N1 and H7N3 viruses. The neuraminidase inhibition assay was carried out by using recombinant species obtained from a baculovirus expression system and the fluorogenic substrate MUNANA. The assay was validated by using oseltamivir carboxylate as a reference inhibitor. Among the tested compounds, PMC-36 showed the highest inhibition on N1 with an IC(50) of 14.6±3.0nM (oseltamivir 25±4nM), while PMC-35 showed a significant inhibitory effect on N3 with an IC(50) of 0.1±0.03nM (oseltamivir 0.2±0.02nM). The analysis of the inhibitory properties of this panel of compounds allowed a preliminary assessment of a structure-activity relationship for the modification of the 4-amido and 5-amino groups of oseltamivir carboxylate. The substitution of the acetamido group in the oseltamivir structure with a 2-butenylamido moiety reduced the observed activity, while the introduction of a propenylamido group was well tolerated. Substitution of the free 5-amino group of oseltamivir carboxylate with an azide, decreased the activity against both N1 and N3. When these structural changes were both introduced, a dramatic reduction of activity was observed for both N1 and N3. The alkylation of the free 5-amino group in oseltamivir carboxylate introducing an isopropyl group seemed to increase the inhibitory effect for both N1 and N3 neuraminidases, displaying a more pronounced effect against N1.