Selective resistance to the PARP inhibitor olaparib in a mouse model for BRCA1-deficient metaplastic breast cancer

Metaplastic breast carcinoma (MBC) is a rare histological breast cancer subtype characterized by mesenchymal elements and poor clinical outcome. A large fraction of MBCs harbor defects in breast cancer 1 (BRCA1). As BRCA1 deficiency sensitizes tumors to DNA cross-linking agents and poly(ADP-ribose) polymerase (PARP) inhibitors, we sought to investigate the response of BRCA1-deficient MBCs to the PARP inhibitor olaparib. To this end, we established a genetically engineered mouse model (GEMM) for BRCA1-deficient MBC by introducing the MET proto-oncogene into a BRCA1-associated breast cancer model, using our novel female GEMM ES cell (ESC) pipeline. In contrast to carcinomas, BRCA1-deficient mouse carcinosarcomas resembling MBC show intrinsic resistance to olaparib caused by increased P-glycoprotein (Pgp) drug efflux transporter expression. Indeed, resistance could be circumvented by using another PARP inhibitor, AZD2461, which is a poor Pgp substrate. These preclinical findings suggest that patients with BRCA1-associated MBC may show poor response to olaparib and illustrate the value of GEMM-ESC models of human cancer for evaluation of novel therapeutics.

PARP Inhibitor Resistance - What Is Beyond BRCA1 or BRCA2 Restoration?

Nearly 10 years ago the usefulness of poly(ADP-ribose) polymerase (PARP) inhibitors to kill BRCA1 or BRCA2-deficient cells was reported, and this finding has served as a prime example of the concept of synthetic lethality in the context of anticancer therapy. The clinical translation of this finding has undergone several ups and downs, however. Despite spectacular responses seen in some patients with BRCA-deficient breast or ovarian cancers, other patients did not show the expected benefit from PARP inhibitor therapy. Thus, like for all novel tailored anti-cancer drugs, upfront and secondary resistance remain major hurdles in the implementation of the initial preclinical finding. We know at least one clinically relevant mechanism of PARP inhibitor resistance: the reversion of BRCA function by secondary mutations. Nevertheless, it is also clear that this mechanism does not explain all cases of resistance. At the moment, we only have a poor understanding of BRCA reversion-independent resistance mechanisms. Preclinical data have pointed in several directions, e.g. increased drug efflux, reduced drug target levels, or alternative DNA repair. Here, we discuss these mechanisms with a focus on potential DNA repair adaptations.

REV7 counteracts DNA double-strand break resection and affects PARP inhibition

Error-free repair of DNA double-strand breaks (DSBs) is achieved by homologous recombination (HR), and BRCA1 is an important factor for this repair pathway. In the absence of BRCA1-mediated HR, the administration of PARP inhibitors induces synthetic lethality of tumour cells of patients with breast or ovarian cancers. Despite the benefit of this tailored therapy, drug resistance can occur by HR restoration. Genetic reversion of BRCA1-inactivating mutations can be the underlying mechanism of drug resistance, but this does not explain resistance in all cases. In particular, little is known about BRCA1-independent restoration of HR. Here we show that loss of REV7 (also known as MAD2L2) in mouse and human cell lines re-establishes CTIP-dependent end resection of DSBs in BRCA1-deficient cells, leading to HR restoration and PARP inhibitor resistance, which is reversed by ATM kinase inhibition. REV7 is recruited to DSBs in a manner dependent on the H2AX-MDC1-RNF8-RNF168-53BP1 chromatin pathway, and seems to block HR and promote end joining in addition to its regulatory role in DNA damage tolerance. Finally, we establish that REV7 blocks DSB resection to promote non-homologous end-joining during immunoglobulin class switch recombination. Our results reveal an unexpected crucial function of REV7 downstream of 53BP1 in coordinating pathological DSB repair pathway choices in BRCA1-deficient cells.

BRCA2-Deficient Sarcomatoid Mammary Tumors Exhibit Multidrug Resistance.

Pan- or multidrug resistance is a central problem in clinical oncology. Here, we use a genetically engineered mouse model of BRCA2-associated hereditary breast cancer to study drug resistance to several types of chemotherapy and PARP inhibition. We found that multidrug resistance was strongly associated with an EMT-like sarcomatoid phenotype and high expression of the Abcb1b gene, which encodes the drug efflux transporter P-glycoprotein. Inhibition of P-glycoprotein could partly resensitize sarcomatoid tumors to the PARP inhibitor olaparib, docetaxel, and doxorubicin. We propose that multidrug resistance is a multifactorial process and that mouse models are useful to unravel this. Cancer Res; 75(4); 732-41. ©2014 AACR.

Brugada syndrome ECG provoked by the selective serotonin reuptake inhibitor fluvoxamine

A patient with an SCN5A p.W822X nonsense mutation, localized in the transmembrane region DII-S4 of the Na(v)1.5 sodium channel and leading to a non-expression of the mutant allele, was prescribed the selective serotonin reuptake inhibitor (SSRI) fluvoxamine (Floxyfral), 100 mg per day. His normal baseline ECG changed to a characteristic Brugada-Type-1-ECG pattern. To investigate whether fluvoxamine may reduce the cardiac sodium current, the effect of this drug was studied on the wild-type voltage-gated cardiac sodium channel Na(v)1.5 stably expressed in HEK293 cells. Patch-clamp recording showed a 50% inhibition of the current at a concentration of 57.3 microM. In our patient, no arrhythmia occurred but the proarrhythmic potential of SSRI in patients with SCN5A mutations cannot be excluded. Therefore, we advise 12-lead ECG control after administering SSRI in these patients.

Differential inhibition sensitivities of MET mutants to the small molecule inhibitor SU11274

Point mutations emerge as one of the rate-limiting steps in tumor response to small molecule inhibitors of protein kinases. Here we characterized the response of the MET mutated variants, V1110I, V1238I, V1206L and H1112L to the small molecule SU11274. Our results reveal a distinct inhibition pattern of the four mutations with IC(50) values for autophosphorylation inhibition ranging between 0.15 and 1.5muM. Differences were further seen on the ability of SU11274 to inhibit phosphorylation of downstream MET transducers such as AKT, ERK, PLCgamma and STAT3 and a variety of MET-dependent biological endpoints. In all the assays, H1112L was the most sensitive to SU11274, while V1206L was less affected under the used concentration range. The differences in responses to SU11274 are discussed based on a structural model of the MET kinase domain.

Reversal of established lupus nephritis and prolonged survival of New Zealand black x New Zealand white mice treated with the topoisomerase I inhibitor irinotecan

Systemic lupus erythematosus is a chronic autoimmune disorder that predominantly affects women of childbearing age. Lupus-associated glomerulonephritis is a major cause of mortality in these patients. Current treatment protocols for systemic lupus erythematosus include cyclophosphamide, prednisolone, azathioprine, and mycophenolate mofetil. However, in mice none of these agents alone or in combination were shown to reverse established proteinuria. Using New Zealand Black x New Zealand White F1 mice, we report that administration of the topoisomerase I inhibitor irinotecan from week 13 completely prevented the onset of proteinuria and prolonged survival up to at least 90 wk without detectable side effects. Furthermore, application of irinotecan to mice with established lupus nephritis, as indicated by grade 3+ (> or =300 mg/dl) and grade 4+ (> or =2000 mg/dl) proteinuria and, according to a median age of 35 wk, resulted in remission rates of 75% and 55%, respectively. Survival was significantly prolonged with 73 wk (grade 3+ and 4+ combined) versus 40 wk for control animals. Although total IgG and anti-dsDNA Abs in the serum and mesangial IgG deposits in the kidneys were not reduced in irinotecan-treated mice, subendothelial immune deposits were considerably diminished, suggesting a prevention of glomerular basement membrane disruption. This effect was accompanied by increased rates of ssDNA breaks and inhibition of renal cell apoptosis being different to what is known about irinotecan in anticancer therapy. In conclusion, our data provide evidence that irinotecan might represent an entirely new strategy for the treatment of systemic lupus erythematosus.

The stem cell gene "inhibitor of differentiation 1" (ID1) is frequently expressed in non-small cell lung cancer

Inhibitor of differentiation 1 (ID1) plays a role in cellular differentiation, proliferation, angiogenesis and tumor invasion. As shown recently, ID1 is positively regulated by the tyrosine kinase SRC in lung carcinoma cell lines and with that appears as a potential new therapeutic target in non-small cell carcinoma (NSCLC). To substantiate this hypothesis we examined ID1, SRC and matrix metalloproteinase-9 (MMP-9) immunohistochemically in human NSCLC specimens.

Bilateral total knee arthroplasty in a patient with hemophilia A, high inhibitor titre and aneurysma spurium of the popliteal artery. A case report

The authors report on bilateral simultaneous knee arthroplasty in a 40-year-old male patient with haemophilia A, high inhibitor titre and an aneurysma spurium of the right popliteal artery. Both knees showed a fixed flexion deformity of 20 degrees. To build up haemostasis, treatment with activated prothrombin complex concentrate (APCC) and recombinant activated factor seven (rFVIIa) was initiated preoperatively. A tourniquet was used on both sides during the operation and factor VIII (FVIII) was administered to further correct coagulopathy. On the eleventh postoperative day the patient complained of increasing pain and pressure in the right knee. An ultrasound suggested aneurysm, which was confirmed by substraction angiography. Under the protection of rFVIIa the aneurysm could be coiled and further rehabilitation was uneventful. At one year post-op the patient presented a range of motion of 90/5/0 degrees for both knees and had returned to full time office work. This case indicates that haemophiliacs with high antibody titre and destruction of both knees can be operated on in one session in order to diminish the operative risk of two consecutive surgical procedures, thus allowing an effective rehabilitation programme. Because of the significant frequency of popliteal aneurysms, preoperative angiography is recommended.

A novel mode of action of the putative sphingosine kinase inhibitor 2-(p-hydroxyanilino)-4-(p-chlorophenyl) thiazole (SKI II): induction of lysosomal sphingosine kinase 1 degradation

Sphingosine kinase 1 (SK1) is a key enzyme in the generation of sphingosine 1-phosphate (S1P) which critically regulates a variety of important cell responses such as proliferation and migration. Therefore, inhibition of SK-1 has been suggested to be an attractive approach to treat tumor growth and metastasis formation.

Recurrent non-melanoma skin cancer: remission of field cancerization after conversion from calcineurin inhibitor- to proliferation signal inhibitor-based immunosuppression in a cardiac transplant recipient

Non-melanoma skin cancers (NMSCs) are the most common malignancies after solid organ transplantation. Their incidence increases with time after transplantation. Calcineurin-inhibitors (CNIs) and azathioprine are known as skin neoplasia-initiating and -enhancing immunosuppressants. In contrast, increasing clinical experience suggests a relevant antiproliferative effect of mammalian target of rapamycin inhibitors, also named proliferation signal inhibitors (PSIs). We report the case of a cardiac allograft recipient with an impressive and consolidated reduction of recurrent NMSC, observed after conversion from CNI-therapy to a PSI-based protocol.

The protease inhibitor alpha-2-macroglobulin-like-1 is the p170 antigen recognized by paraneoplastic pemphigus autoantibodies in human

Paraneoplastic pemphigus (PNP) is a devastating autoimmune blistering disease, involving mucocutaneous and internal organs, and associated with underlying neoplasms. PNP is characterized by the production of autoantibodies targeting proteins of the plakin and cadherin families involved in maintenance of cell architecture and tissue cohesion. Nevertheless, the identity of an antigen of Mr 170,000 (p170), thought to be critical in PNP pathogenesis, has remained unknown.

The effects of PTK787/ZK222584, an inhibitor of VEGFR and PDGFR? pathways, on intussusceptive angiogenesis and glomerular recovery from Thy1.1 nephritis

The aim of our study was to investigate the phenomenon of intussusceptive angiogenesis with a focus on its molecular regulation by vascular endothelial growth factor receptor (VEGFR)/platelet-derived growth factor receptor β (PDGFRβ) pathways and biological significance for glomerular recovery after acute injury. Glomerular healing by intussusception was examined in a particular setting of Thy1.1 nephritis, where the lysis of mesangial cells results in an initial collapse and successive rebuilding of glomerular capillary structure. Restoration of capillary structure after induction of Thy1.1 nephritis occurred by intussusceptive angiogenesis resulting in i) rapid expansion of the capillary plexus with reinstatement of the glomerular filtration surface and ii) restoration of the archetypical glomerular vascular pattern. Glomerular capillaries of nephritic rats after combined VEGFR2 and PDGFRβ inhibition by PTK787/ZK222584 (PTK/ZK) were tortuous and irregular. However, the onset of intussusceptive angiogenesis was influenced only after long-term PTK/ZK treatment, providing an important insight into differential molecular regulation between sprouting and intussusceptive angiogenesis. PTK/ZK treatment abolished α-smooth muscle actin and tensin expression by injured mesangial cells, impaired glomerular filtration of microspheres, and led to the reduction of glomerular volume and the presence of multiple hemorrhages detectable in the tubular system. Collectively, treatment of nephritic patients with PTK/ZK compound is not recommended.