Activity of KRM 1648 alone or in combination with ethambutol or clarithromycin against Mycobacterium avium in beige mouse model of disseminated infection.

Rifamycins are active against slowly growing mycobacteria, such as Mycobacterium tuberculosis and Mycobacterium kansasii, but the majority of rifamycins thus far investigated both in vitro and in vivo are inactive or have only modest activity against the Mycobacterium avium complex (MAC). We investigated the activity of three doses of the semisynthetic benzoxazinorifamycin KRM 1648, alone or in combination with ethambutol or clarithromycin, in beige mice challenged with the MAC strain 101. Our results show the following. (i) KRM 1648 was significantly effective against MAC infection as determined by the reduction of the number of bacteria in the blood, liver, and spleen when administered at doses of 20 and 40 mg/kg of body weight per day but not at 10 mg/kg/day, compared with untreated controls. (ii) KRM 1648 (40 mg/kg/day) administered in combination with ethambutol (100 mg/kg/day) resulted in significant reduction in bacteremia compared with values for untreated controls (P 0.001), KRM 1648 alone (P = 0.019), and ethambutol alone (P = 0.003). Furthermore, the combination of KRM 1648 and ethambutol was associated with a significant decrease of the number of bacteria in the spleen and the liver compared with values for both untreated controls and each drug alone (P < 0.001 for all comparisons). (iii) KRM 1648 (40 mg/kg/day) administered in combination with clarithromycin (200 mg/kg/day) resulted in a significant decrease of the number of bacteria in the blood and the spleen compared with the number for untreated controls (P < 0.001 for all comparisons). In our experience, using MAC 101 as the challenging organism, KRM 1648 is the first the number of bacteria in the blood and spleen compared with the number for untreated controls (P >0.001 for all comparions). In our experience, using MAC 101 as the challenging organism, KRM 1648 is the first rifamycin with significant activity in vivo against MAC infection in beige mice.     

Telomerase confers resistance to caspase-mediated apoptosis

There is growing evidence that accelerated telomeric attrition and/or aberrant telomerase activity contributes to pathogenesis in a number of diseases. Likewise, there is increasing interest to develop new therapies to restore or replace dysfunctional cells characterized by short telomeric length using telomerase-positive counterparts or stem cells. While telomerase adds telomeric repeats de novo contributing to enhanced proliferative capacity and lifespan, it may also increase cellular survival by conferring resistance to apoptosis. Consequently, we sought to determine the involvement of telomerase for reduced apoptosis using ovarian surface epithelial cells. We found that expression of hTERT, the catalytic component of telomerase, was sufficient and specific to reduce caspase-mediated cellular apoptosis. Further, hTERT expression reduced activation of caspases 3, 8, and 9, reduced expression of pro-apoptotic mitochondrial proteins t-BID, BAD, and BAX and increased expression of the anti-apoptotic mitochondrial protein, Bcl-2. The ability of telomerase to suppress caspase-mediated apoptosis was p-jnk dependent since abrogation of jnk expression with jip abolished resistance to apoptosis. Consequently, these findings indicate that telomerase may promote cellular survival in epithelial cells by suppressing jnk-dependent caspase-mediated apoptosis.     

Human SHPRH is a ubiquitin ligase for Mms2-Ubc13-dependent polyubiquitylation of proliferating cell nuclear antigen

Human SHPRH gene is located at the 6q24 chromosomal region, and loss of heterozygosity in this region is seen in a wide variety of cancers. SHPRH is a member of the SWI/SNF family of ATPases/helicases, and it possesses a C(3)HC(4) RING motif characteristic of ubiquitin ligase proteins. In both of these features, SHPRH resembles the yeast Rad5 protein, which, together with Mms2-Ubc13, promotes replication through DNA lesions via an error-free postreplicational repair pathway. Genetic evidence in yeast has indicated a role for Rad5 as a ubiquitin ligase in mediating the Mms2-Ubc13-dependent polyubiquitylation of proliferating cell nuclear antigen. Here we show that SHPRH is a functional homolog of Rad5. Similar to Rad5, SHPRH physically interacts with the Rad6-Rad18 and Mms2-Ubc13 complexes, and we show that SHPRH protein is a ubiquitin ligase indispensable for Mms2-Ubc13-dependent polyubiquitylation of proliferating cell nuclear antigen. Based on these observations, we predict a role for SHPRH in promoting error-free replication through DNA lesions. Such a role for SHPRH is consistent with the observation that this gene is mutated in a number of cancer cell lines, including those from melanomas and ovarian cancers, which raises the strong possibility that SHPRH function is an important deterrent to mutagenesis and carcinogenesis in humans.     

The alternative Ctf18-Dcc1-Ctf8-replication factor C complex required for sister chromatid cohesion loads proliferating cell nuclear antigen onto DNA

The linkage of sister chromatids after DNA replication ensures the faithful inheritance of chromosomes by daughter cells. In budding yeast, the establishment of sister chromatid cohesion requires Ctf8, Dcc1, and Ctf18, a homologue of the p140 subunit of the replication factor C (RFC). In this report we demonstrate that in 293T cells, Flag-tagged Ctf18 forms a seven-subunit cohesion-RFC complex comprised of Ctf18, Dcc1, Ctf8, RFCp40, RFCp38, RFCp37, and RFCp36 (Ctf18-RFC). We demonstrate that a stoichiometric heteroheptameric Ctf18-RFC complex can be assembled by coexpressing the seven proteins in baculovirus-infected insect cells. In addition, the two other stable subcomplexes were formed, which include a pentameric complex comprised of Ctf18, RFCp40, RFCp38, RFCp37, and RFCp36 and a dimeric Dcc1-Ctf8. Both the five- and seven-subunit Ctf18-RFC complexes bind to single-stranded and primed DNAs and possess weak ATPase activity that is stimulated by the addition of primed DNA and proliferating cell nuclear antigen (PCNA). These complexes catalyzed the ATP-dependent loading of PCNA onto primed and gapped DNA but not onto double-stranded nicked or single-stranded circular DNAs. Consistent with these observations, both Ctf18-RFC complexes substituted for the replicative RFC in the PCNA-dependent DNA polymerase delta-catalyzed DNA replication reaction. These results support a model in which sister chromatid cohesion is linked to DNA replication.     

Refined mapping of genomic rearrangements involving the short arm of chromosome 9 in acute lymphoblastic leukemias and other hematologic malignancies

Deletions of chromosomal band 9p21 have been detected in various tumor types as well as in more than 20% of acute lymphoblastic leukemia (ALL). These deletions frequently include the entire interferon (IFN) gene cluster as well as the methylthioadenosine phosphorylase (MTAP) gene. Recently, the CDKN2 gene (p16INK4A, MTS I, CDK41) was proposed as a candidate tumor-suppressor gene on 9p21 because it is frequently deleted in cell lines derived from multiple tumor types. To determine if CDKN2 or another closely related gene on 9p is the target of 9p deletions in ALL and other hematologic malignancies, we analyzed 20 primary patient samples (13 ALL, 2 acute myeloid leukemias [AML], and 5 non-Hodgkin's lymphomas [NHL]) with 9p rearrangements using Southern blot analysis, fluorescence in situ hybridization (FISH), and single- strand conformation polymorphism (SSCP) for alterations of CDKN2. Homozygous deletions of the CDKN2/CDKN2B (p15) region were detected in 10 cases (50%; 6 ALL, 2 AML, and 2 NHL). In 1 additional case, the intensity of the Southern blot band was significantly reduced, suggesting a CDKN2 deletion in a subpopulation of the malignant cells. No CDKN2 or CDKN2B rearrangements were seen. The IFN gene cluster was homozygously deleted in 2 of 15 (13%) analyzed cases, whereas the MTAP gene was deleted in 6 of 15 cases (40%). In addition, hemizygous deletions of the CDKN2 region were identified in 6 ALL cases using interphase FISH. No point mutation of the coding region of CDKN2 was detected by SSCP in these cases. We conclude that CDKN2 is the most frequently homozygously deleted marker on 9p. The absence of point mutations in the coding region of CDKN2 in cases with hemizygous 9p deletions and the frequent codeletion of MTAP, CDKN2B, and other yet unidentified neighboring genes suggest that the simultaneous deletion of these genes may be necessary for the selective growth advantage of malignant cells.     

Computed tomography findings of Crouzon syndrome: A case report

Crouzon syndrome is a genetic condition characterized by a premature fusion of skull sutures resulting in head and facial deformities. Crouzon syndrome is usually suspected at birth through physical examination or in the antenatal period via ultrasonographic assessment. Once Crouzon syndrome is suspected, advanced imaging methods such as three-dimensional computed tomography must be requested, showing early signs of cranial sutures fusion. In this paper, we present a case of a six-year-old girl who was taken to a pediatrician control appointment due to abnormal facies. During the physical examination, a suspicion of Crouzon syndrome was raised. Therefore, a head computed tomography was requested, showing asymmetrical calvarium thickening, diffuse indentation of the inner table of the skull, and moderate hydrocephalus with a big cyst in the posterior fossa. Due to these findings, the patient was remitted to maxillofacial surgery for further evaluation; however, the medical appointment could not be achieved as a consequence of the poor medical insurance of the girl. This paper aims to describe and discuss the computed tomography findings of Crouzon syndrome.     

Liposarcoma of the chest wall mimicking a breast mass in a man: a case report

We report a rare case of dedifferentiated liposarcoma in a man. A 60-year-old male patient presented with a left mass involving the whole breast area, with no lymph node enlargement, growing during a one-year-period. Imaging studies revealed a fat-containing mixed-density mass apparently associated with the pectoralis major muscle. A core biopsy was performed that yielded a diagnosis of a well-differentiated liposarcoma. Further tests to check for metastases were ordered and no distant disease was found. Left mastectomy with en bloc resection of the pectoralis major muscle was performed. The pathologic diagnosis revealed a high-grade dedifferentiated liposarcoma with extensive necrosis. This tumor type is primarily described in the retroperitoneum and extremities. We report an unusual presentation of a liposarcoma mimicking a breast mass.