BackgroundAcute rheumatic fever (ARF) rates have declined to near zero in nearly all developed countries. However, in New Zealand rates have not declined since the 1980s. Further, ARF diagnoses in New Zealand are inequitably distributed--occurring almost exclusively in Māori (the indigenous population) and Pacific children--with very low rates in the majority New Zealand European population. With ARF diagnosis, secondary prophylaxis is key to prevent recurrence. The purpose of this study was to identify the perceived enablers and barriers to secondary recurrence prophylaxis following ARF for Māori patients aged 14–21.MethodsThis study took a systems approach, was informed by patient voice and used a framework method to explore potential barriers and enablers to ongoing adherence with monthly antibiotic injections for secondary prophylaxis. Qualitative interviews were conducted with 19 Māori ARF patients receiving recurrence prophylaxis in the Waikato District Health Board region. Participants included those fully adherent to treatment, those with intermittent adherence or those who had been “lost to follow-up.”ResultsBarriers and enablers were presented around three factors: system (including access/resources), relational and individual. Access and resources included district nurses coming to patients as an enabler and lack of income and time off work as barriers. Relational characteristics included support from family and friends as enablers and district nurse communication as predominantly a positive although not enabling factor. Individual characteristics included understanding, personal responsibility and fear/pain of injections.ConclusionThis detailed exploration of barriers and enablers for ongoing secondary prophylaxis provides important new information for the prevention of recurrent ARF. Among other considerations, a national register, innovative engagement with youth and their families and a comprehensive pain management programme are likely to improve adherence to ongoing secondary prophylaxis and reduce the burden of RHD for New Zealand individuals, families and health system.

Trastuzumab emtansine (T-DM1, Kadcyla) is an antibody-drug conjugate used in the treatment of HER2-positive breast cancer. However, its use is limited by acquired and intrinsic resistance, the mechanisms of which are not well understood. Further knowledge of T-DM1 resistance may provide new combination strategies or therapeutic targets to overcome resistance or new predictive biomarkers to identify the patients most likely to benefit from T-DM1 therapy. To discover genes responsible for T-DM1 sensitivity and resistance in an unbiased manner, we have conducted CRISPR/Cas9 functional genomics screens by a two-stage process. Firstly, we performed whole genome screens in MDA-MB-361 and MDA-MB-453 cells transduced with Cas9 and the GeCKOv2 lentiviral library that were exposed to T-DM1 and its effector DM1 for 8-13 weeks. Gene knockouts enriched or depleted in response to T-DM1 or DM1 treatment in either cell line were identified by sequencing of genomic DNA and differentially expressed genes by RNA sequencing, revealing 599 candidate genes of T-DM1 sensitivity. For high-throughput validation of the 599 genes, we developed a custom library of 2539 guide RNAs (gRNAs) to target these 599 genes, plus non-targeting controls. Cas9-expressing MDA-MB-361 cells were transduced with the custom library and exposed to T-DM1 for 28 days. MAGeCK analysis of gRNA sequencing revealed 11 genes that were significantly enriched and one gene that was significantly depleted at a false discovery rate (FDR) of <0.1. Two of the top hits in the secondary screen were two genes whose loss is known to promote T-DM1 resistance: ERBB2 (HER2) and SLC46A3 (P<8 × 10−5; FDR <0.007). Other top hits were TSC1 and TSC2 (P<3 × 10−6; FDR= 0.0004); which are both tumor suppressor genes and negative regulators of mTOR complex 1 (mTORC1). For subsequent validation, we have generated TSC2 knockout cell pools, which were more resistant to T-DM1 than wildtype cells in a competition growth assay. Knockout clones have been isolated and are being tested for T-DM1 resistance. Since mTOR inhibitors can phenocopy TSC1 and TSC2 by inhibiting mTORC1 activity, we have also evaluated T-DM1 in combination with the mTOR inhibitor KU-0063794 in a sulforhodamine B assay in MDA-MB-361 and MDA-MB-453 cells. Each agent potently inhibited cell proliferation and demonstrated synergistic anti-proliferative activity in combination. Together, our results suggest that TSC1 and TSC2 knockout may promote T-DM1 resistance and that targeting mTOR may be an effective strategy to overcome T-DM1 resistance. Citation Format: Francis W. Hunter, Barbara A. Lipert, Kyla N. Siemens, Aziza Khan, Hilary R. Barker, Troy W. Ketela, William R. Wilson, Tet-Woo Lee, Stephen M. Jamieson. Identification of TSC1 and TSC2 as potential determinants of sensitivity to trastuzumab emtansine [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5853.

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