Yasufumi Matsuda

Induction of DNA strand breaks is critical to predict the cytotoxicity of gemtuzumab ozogamicin against leukemic cells.

Gemtuzumab ozogamicin (GO) consists of the CD33 antibody linked to calicheamicin. The binding of GO to the CD33 antigen on leukemic cells results in internalization followed by the release of calicheamicin, thereby inducing DNA strand breaks. We hypothesized that the induction of DNA strand breaks would be a surrogate marker of GO cytotoxcity. Here, two GO‐resistant variants (HL/GO‐CSA [225‐fold], HL/GO [200‐fold]) were established by serially incubating human leukemia HL‐60 cells with GO with or without a P‐glycoprotein (P‐gp) inhibitor, cyclosporine A, respectively. The CD33 positivity was reduced in both variants. The HL/GO‐CSA cells showed an increased multidrug resistance protein‐1 (MRP1) transcript, and an MRP1 inhibitor partially reversed GO resistance. The HL/GO cells had neither P‐gp nor MRP1 overexpression. Microarray analysis and Western blotting indicated elevated levels of DNA repair‐associated proteins in both variants. Two other leukemic subclones, showing either P‐gp or MRP1 overexpression, were also GO‐resistant. Using single cell gel electrophoresis analysis, it was determined that GO‐induced DNA strand breaks increased dose‐dependently in HL‐60 cells, whereas the number of breaks was reduced in the GO‐resistant cell lines. The induction of DNA strand breaks was correlated with GO sensitivity among these cell lines. The CD33 positivity and the expression levels of transporters were not proportional to drug sensitivity. Using primary leukemic cells, the induction of DNA strand breaks appeared to be associated with GO sensitivity. Thus, GO‐induced DNA strand breaks as the final output of the mechanism of action would be critical to predict GO cytotoxicity.

Combination of panobinostat with ponatinib synergistically overcomes imatinib‐resistant CML cells

The major mechanism of imatinib (IM) resistance of CML is the reactivation of ABL kinase either through BCR‐ABL gene amplification or mutation. We investigated the cytotoxicity of a pan‐ABL tyrosine kinase inhibitor, ponatinib, and a pan‐histone deacetylase inhibitor, panobinostat, against IM‐resistant CML cells in vitro. Two different IM‐resistant cell lines, K562/IM‐R1 and Ba/F3/T315I were evaluated in comparison with their respective, parental cell lines, K562 and Ba/F3. K562/IM‐R1 overexpressed BCR‐ABL due to gene amplification. Ba/F3/T315I was transfected with a BCR‐ABL gene encoding T315I‐mutated BCR‐ABL. Ponatinib inhibited the growth of both K562/IM‐R1 and Ba/F3/T315I as potently as it inhibited their parental cells with an IC50 of 2–30 nM. Panobinostat also similarly inhibited the growth of all of the cell lines with an IC50 of 40–51 nM. This was accompanied by reduced histone deacetylase activity, induced histone H3 acetylation, and an increased protein level of heat shock protein 70, which suggested disruption of heat shock protein 90 chaperone function for BCR‐ABL and its degradation. Importantly, the combination of ponatinib with panobinostat showed synergistic growth inhibition and induced a higher level of apoptosis than the sum of the apoptosis induced by each agent alone in all of the cell lines. Ponatinib inhibited phosphorylation not only of BCR‐ABL but also of downstream signal transducer and activator of transcription 5, protein kinase B, and ERK1/2 in both K562/IM‐R1 and Ba/F3/T315I, and the addition of panobinostat to ponatinib further inhibited these phosphorylations. In conclusion, panobinostat enhanced the cytotoxicity of ponatinib towards IM‐resistant CML cells including those with T315I‐mutated BCR‐ABL.

Hypoxanthine Guanine Phosphoribosyltransferase (HPRT) Deficiencies: HPRT1 Mutations in New Japanese Families and PRPP Concentration

Mutation of hypoxanthine guanine phosphoribosyltransferase (HPRT) gives rise to Lesch–Nyhan syndrome, which is characterized by hyperuricemia, severe motor disability, and self-injurious behavior, or HPRT-related gout with hyperuricemia. Four mutations were detected in two Lesch–Nyhan families and two families with partial deficiency since our last report. A new mutation of G to TT (c.456delGinsTT) resulting in a frameshift (p.Q152Hfs*3) in exon 3 has been identified in one Lesch–Nyhan family. In the other Lesch–Nyhan family, a new point mutation in intron 7 (c.532 + 5G > T) causing splicing error (exon 7 excluded, p.L163Cfs*4) was detected. In the two partial deficiency cases with hyperuricemia, two missense mutations of p.D20V (c.59A > T) and p.H60R (c.179A >G) were found. An increase of erythrocyte PRPP concentration was observed in the respective phenotypes and seems to be correlated with disease severity.

3A Comparison between R-THP-COP and R-CHOP Regimens for the Treatment of Diffuse Large B-cell Lymphoma in Old Patients: A Single-institution Analysis

Objective We retrospectively compared the clinical efficacy and toxicity of rituximab (R)-THP-COP (pirarubicin, cyclophosphamide, vincristine, and prednisolone) with that of R-CHOP (rituximab, adriamicin, cyclophosphamide, vincristine, and prednisolone) in previously untreated old patients with diffuse large B-cell lymphoma (DLBCL). Patients and Methods Patients admitted to our institution between 2004 and 2013 were examined. The patients received either R(375 mg/m2, day 1)-THP-COP (pirarubicin 50 mg/m2 day 1, cyclophosphamide 750 mg/m2 day 1, vincristine 1.4 mg/m2 day 1, and prednisolone 100 mg day 1-5) or R-CHOP (adriamicin 50 mg/m2 day 1, cyclophosphamide 750 mg/m2 day 1, vincristine 1.4 mg/m2 day 1, and prednisolone 100 mg day 1-5). The doses of chemotherapeutic agents were adjusted depending on the patients age and associated complications. The treatment was performed for 6 to 8 cycles. Results Among 74 patients with DLBCL (median 76, range 65-90 years; male 39, female 35), 29 received R-THP-COP, while 45 received R-CHOP. The overall response rates were 94.6% (complete response 86.4%, partial response 8.1%). The 2-year overall and progression-free survival rates were 77.6% and 68.5% for the R-THP-COP regimen and 79.2% and 78.9% for R-CHOP, respectively. No significant differences were found between these two regimens regarding the clinical efficacies. The most frequent adverse event was neutropenia (72.4% for the R-THP-COP regimen, 88.9% for the R-CHOP regimen). The cardiac function as evaluated by ejection fraction values was not impaired in either regimen. Conclusion R-THP-COP was effective and safe as an alternative to R-CHOP.

Efficacy of aprepitant for CHOP chemotherapy-induced nausea, vomiting, and anorexia

The objective of this study was to evaluate whether aprepitant in addition to 5-HT3 receptor antagonist is useful for preventing chemotherapy-induced nausea and vomiting (CINV) and anorexia in patients receiving CHOP therapy, and to evaluate the relationship between in vivo kinetics of plasma substance P and these adverse events. Patients with malignant lymphoma who received CHOP chemotherapy or THP (THP-ADR)-COP therapy were investigated for CINV and anorexia for 5 days after the start of chemotherapy. With the first course of chemotherapy, all patients received only granisetron on day1 as an antiemetic. Patients who experienced nausea, vomiting, or anorexia exceeding grade 1 in the first course received aprepitant for 3 days in addition to granisetron with the second course of CHOP chemotherapy. Plasma substance P concentrations at 24 and 72 hours after chemotherapy were measured. Nineteen patients were evaluated. Nausea, vomiting, or anorexia was observed with the first course in 7 of 19 patients. During the second course with aprepitant, no patients experienced vomiting, and the toxicity grade of nausea, vomiting, or anorexia was decreased compared with those in the first course. Substance P concentrations showed no differences after chemotherapy, in patients with nausea, vomiting, or anorexia and in patients without. The addition of aprepitant to 5-HT3 receptor antagonist appears effective for CINV or anorexia for patients who received CHOP chemotherapy.