DeepNEU: cellular reprogramming comes of age – a machine learning platform with application to rare diseases research

Abstract Background Conversion of human somatic cells into induced pluripotent stem cells (iPSCs) is often an inefficient, time consuming and expensive process. Also, the tendency of iPSCs to revert to their original somatic cell type over time continues to be problematic. A computational model of iPSCs identifying genes/molecules necessary for iPSC generation and maintenance could represent a crucial step forward for improved stem cell research. The combination of substantial genetic relationship data, advanced computing hardware and powerful nonlinear modeling software could make the possibility of artificially-induced pluripotent stem cells (aiPSC) a reality. We have developed an unsupervised deep machine learning technology, called DeepNEU that is based on a fully-connected recurrent neural network architecture with one network processing layer for each input. DeepNEU was used to simulate aiPSC systems using a defined set of reprogramming transcription factors. Genes/proteins that were reported to be essential in human pluripotent stem cells (hPSC) were used for system modelling. Results The Mean Squared Error (MSE) function was used to assess system learning. System convergence was defined at MSE < 0.001. The markers of human iPSC pluripotency (N = 15) were all upregulated in the aiPSC final model. These upregulated/expressed genes in the aiPSC system were entirely consistent with results obtained for iPSCs. Conclusion This research introduces and validates the potential use of aiPSCs as computer models of human pluripotent stem cell systems. Disease-specific aiPSCs have the potential to improve disease modeling, prototyping of wet lab experiments, and prediction of genes relevant and necessary for aiPSC production and maintenance for both common and rare diseases in a cost-effective manner

Novel anti-cancer drug COTI-2 synergizes with therapeutic agents and does not induce resistance or exhibit cross-resistance in human cancer cell lines

<div><p>Emerging drug-resistance and drug-associated toxicities are two major factors limiting successful cancer therapy. Combinations of chemotherapeutic drugs have been used in the clinic to improve patient outcome. However, cancer cells can acquire resistance to drugs, alone or in combination. Resistant tumors can also exhibit cross-resistance to other chemotherapeutic agents, resulting in sub-optimal treatment and/or treatment failure. Therefore, developing novel oncology drugs that induce no or little acquired resistance and with a favorable safety profile is essential. We show here that combining COTI-2, a novel clinical stage agent, with multiple chemotherapeutic and targeted agents enhances the activity of these drugs <i>in vitro</i> and <i>in vivo</i>. Importantly, no overt toxicity was observed in the combination treatment groups <i>in vivo</i>. Furthermore, unlike the tested chemotherapeutic drugs, cancer cells did not develop resistance to COTI-2. Finally, some chemo-resistant tumor cell lines only showed mild cross-resistance to COTI-2 while most remained sensitive to it.</p></div

Chemo-resistant cancer cell lines often do not show cross-resistance to COTI-2.

<p>Paclitaxel-resistant (A) and cisplatin-resistant (B) A549, DMS-153, and SHP-77 cells were exposed to IC₅₀ concentrations of COTI-2 and tumor cell proliferation was measured after approximately 4 doublings of control cells. 5FUdR-resistant HeLa cells (C) and vincristine-resistant HN-5a cells (D) were exposed to IC₅₀ concentrations of COTI-2 as described in (A) and (B). Significant differences were assessed by Student’s <i>t</i>-test (<i>p</i><0.05). Data indicates mean values derived from 3 independent experiments ± SEM.</p

Cancer cells do not develop acquired resistance to COTI-2 unlike treatment with paclitaxel and cisplatin.

<p>A549 NSCLC (A), DMS-153 SCLC (B) and SHP-77 SCLC (C) cells were cultured in IC₅₀ concentrations of COTI-2, paclitaxel, or cisplatin for 4 rounds of treatment (5 generations of cells including the parental cells). The surviving 50% of cells from the initial IC₅₀ tested were harvested and cultured for 5 days, after which time this new generation of cells was re-treated with the same agent and a new IC₅₀ value was established. Emerging resistance was identified by increasing IC₅₀ values in successive generations. Significant differences were assessed by Student’s <i>t</i>-test (<i>p</i><0.05). *Significantly different from parental cells treated with the mentioned drug. Data points indicate the mean from 3 independent experiments ± SEM.</p

COTI-2 enhances the cytotoxic activity of paclitaxel and cisplatin.

<p>DMS-114 (A and C) and SHP-77 cells (B and D) were cultured overnight then exposed to the indicated doses of paclitaxel and cisplatin plus or minus a pre-determined dose of COTI-2 (IC₂₅) for 4 days before cell viability was determined. The asterix (*) indicates a significant greater-than-additive effect in the combination therapy compared to single agent alone, <i>p</i><0.05, Student’s <i>t</i>-test. Data are the average mean of 3 independent experiments ± SEM. (E) AN3-CA human endometrial cells (1 x 10⁷) were injected into the right flanks of athymic nude mice (n = 10 mice per group). Xenografts were grown to an average volume of 170 mm³ before animals received treatment i.v. Vehicle control and COTI-2 (25 mg/kg) were administered 3 times a week on alternate days until study end. The schedule for paclitaxel was daily for 5 days (5 mg/kg). In the combination arm, animals received COTI-2 (25 mg/kg) 3 times a week for the entire study and 5 injections of paclitaxel (5 mg/kg). *Significantly different from the paclitaxel alone treatment group, Student’s <i>t</i>-test, <i>p</i><0.05. Error bars represent SEM.</p

Combining COTI-2 with cetuximab and erlotinib synergistically enhances the efficacy of these drugs against human colorectal cancer cells.

<p>Human colorectal cancer cell lines HCT-15 (A), SW-620 (B), and COLO-205 (C) were treated with varying concentrations of COTI-2, cetuximab, erlotinib, or a combination of COTI-2 and either EGFR inhibitor. Tumor cells were allowed to proliferate for 4 days in the presence of drug(s) before cell viability was determined. All data points indicate the mean of 5 independent measures of viability ± SEM. *Significant difference from cells treated with COTI-2 alone using a Student’s <i>t</i>-test (<i>p</i><0.05).</p

The effect of COTI-2 treatment on U87-MG cells in combination with temsirolimus and rapamycin.

<p>U87-MG human glioma cells were cultured in the presence of various concentrations of temsirolimus plus COTI-2 (A) or rapamycin plus COTI-2 (B) for 4 days before cell viability was determined. Black circles indicate the combination of COTI-2 and temsirolimus (A) or rapamycin (B) and the white circles indicate treatment with COTI-2 alone. Data are the average mean of 6 independent experiments ± SEM.*Significant difference, Student’s <i>t</i>-test, <i>p</i><0.05.</p

Only some chemotherapeutic drugs with similar molecular targets show enhanced activity when combined with COTI-2.

<p>DMS-114 (A and C) and SHP-77 (B and D) SCLC cells were treated with various concentrations of carboplatin (A and B) or vincristine (C and D) in combination with or without an IC₂₅ concentration of COTI-2 for 4 days before cell viability was determined. The asterix (*) indicates a significant greater-than-additive effect in the combination therapy compared to single agent alone, <i>p</i><0.05, Student’s <i>t</i>-test. Data are the average mean of 3 independent experiments ± SEM. (E and F) PANC-1 human pancreatic carcinoma cells (2 x 10⁶) were injected into each flank of NCr-<i>nu</i> mice (n = 12 mice per group). Xenografts were grown to ~100 mm³ before animals received treatment, which consisted of the vehicle control, COTI-2 (125 mg/kg), gemcitabine (100 mg/kg), or the combination (COTI-2 at 125 mg/kg and gemcitabine at 100 mg/kg) (E) or the vehicle control, COTI-2 (125 mg/kg), abraxane (15 mg/kg), or the combination (COTI-2 at 125 mg/kg and abraxane at 15 mg/kg) (F). COTI-2 was delivered <i>p</i>.<i>o</i>. with a schedule of 5 days on treatment and 2 days off weekly. Gemcitabine (100 mg/kg) was administered i.p., every second day, for a total of 6 injections. Abraxane (15 mg/kg) was administered i.v., once per day for 5 consecutive days. The dosing schedule for the combination treatments was identical to that of the single agent treatments for each drug. COTI-2 administration was initiated 1 day after treatment with either gemcitabine or abraxane. *Significantly different from single agent gemcitabine or abraxane treatment groups, Student’s <i>t</i>-test, <i>p</i><0.05.</p