PNB 001 (GPP- Balacovin)

PNB 001 / GPP- Balacovin for the treatment of Covid 19, Inflammatory Bowel Disease, Inflammatory Pain, and Small Cell Lung Cancer

Background about target The peptide cholecystokinin (CCK) was originally discovered in the gastrointestinal tract and has been shown to mediate pancreatic secretion and contraction of the gallbladder. CCK was then described in the mammalian central nervous system (CNS) as a gastrin-like immunoreactive material. Two types of CCK receptors (type A, “alimentary”, and type B, “brain”) have been distinguished. CCK-A and CCK-B receptor types have been shown to differ by their relative affinity for the natural ligands, their differential distribution, and their molecular structure. Gastrin, a peptide hormone secreted by the parietal cells of the stomach, binds to the CCK-B receptor and elicits its function. Upon binding to CCK2, it releases histamine, which in turn plays a role in releasing hydrochloric acid/gastric acid. In addition, gastrin also plays important role in cell proliferation and maturation of the GI tract. Gastrin is also produced in excess in gastrinoma or gastric cancers. Inflammatory Bowel Syndrome patients had higher secretion and sustained secretion of CCK after fat meals. CCK is also involved in GI motor response after meals. Hence, CCK receptor antagonists are recommended for IBD.

Preclinical studies: PNB-001 is an isoform-selective antagonist that binds to CCK2 at 20 nM. In an isolated tissue assay, using CCK-5 as an agonist the antagonizing properties were confirmed. L-365,260 was used as the best CCK2 gastrin antagonist standard and PNB-001 was 10 times more potent and of much higher magnitude.
PNB-001 was tested in a rat model of indomethacin-induced IBD and was compared to the positive control, prednisolone. PNB-001, at 5 mg/kg and 20 mg/kg p.o., was extremely effective in reducing the inflammation- and IBD-dependent damage to various gastrointestinal tissues and organs. Gross pathological changes and histopathological observations demonstrated IBD and Crohn’s disease in this animal model were completely reversed by PNB-001. This reversal effect was dose-dependent and highly consistent with CCK literature.

In the tail-flick assay in mice, 0.5 mg/kg PNB001 was found to be analogue to 40 mg/kg tramadol about the latency period. Subsequent experiments were performed to determine the dose and time response of PNB001 to understand the longevity of the effect. In rats, PNB-001 was administered orally and it demonstrated a sustained pain tolerance and this effect was either comparable or superior to the effect observed with tramadol.
Subsequently, the efficacy of PNB001 was evaluated in a more complex model of pain. Rats were grouped and a dose-response curve of PNB001 (p.o or i.p.) was performed and compared to morphine in formalin-induced neuropathic (phase I) and inflammation pain (phase II) models. PNB-001 was extremely effective in this model of pain.

Preclinical development studies: PNB001 has a short half-life of 1.20 min in rat liver microsome and approximately 12 min in dogs and human liver microsome. Subsequent studies were performed to determine the pharmacokinetic (PK) properties of PNB-001. Rats were administered orally with 20 mg/kg PNB001 to determine the circulating concentration. PNB001 peak concentration was achieved at 40 min. PNB001 had a half-life of 9 h and had a relatively low bioavailability. Considering the bioavailability in a species with rapid liver metabolism, we expected PNB001 to have a manifold bioavailability in humans.

PNB-001 was evaluated in various ADME studies, including cytochrome P450 enzyme inhibition, plasma protein binding, and CaCo2 permeability. PNB-001 did not inhibit Cyp3A4, Cyp2C9, and Cyp1A2 up to 10 µM and Cyp2C19 until 3 µM. With 97% of PNB001 bound to rat and human plasma, the amount of free unbound PNB-001 is comparable to some of the marketed drugs. CaCo2 permeability studies indicated that PNB001 has high permeability and is not effluxed in the B>A direction. Regulatory toxicology and safety pharmacology studies (acute toxicity studies in rats and mice, 7-day dose range-finding studies in rats and dogs, 28 days toxicology studies in rats and dogs, PK studies in rats and dogs, safety pharmacology studies (cardiac, renal, respiratory, and neuro), chromosomal aberration and mutagenicity studies) conducted with PNB001 under GLP environment and in accordance with ICH guidelines demonstrate extremely high levels of safety and tolerability. The doses selected were 50 to 100 fold above the efficacy dose.

Synthetic chemistry: PNB-001 synthetic scheme has been established both in non-GMP and GMP conditions. Moreover, synthetic chemistry efforts were optimized to synthesize PNB-001 on a multi-kg scale.
Formulation studies: Formulation development, stability, and physicochemical characterization of PNB001 were conducted. The studies demonstrated that PNB001 was highly stable with ideal physicochemical properties.

Phase I clinical trials: Phase I SAD clinical trial was completed and PNB001 was found extremely safe over the dose range from 25-1500 mg. A MAD phase 1 clinical trial was also completed at low, medium, and high doses (50, 100, and 200 mg), and in total PNB001 was tested now in 74 healthy subjects. Overall PNB-001 was tested very safe and the target plasma concentrations were reached. AEs shown by the subjects were very mild, such as vomiting in a female subject and at a high dose, an increase in ALT was identified in one subject.

Phase II clinical trials: A multi-centre, randomized, parallel-group, comparative, open-label study to assess efficacy and safety of PNB-001 in patients with moderate COVID-19 infection was conducted. Patients were randomly assigned to receive PNB-001 100 mg orally with Best Care (BC) (PNB-001 + BC) or only Best Care (BC). A total of 40 patients were randomized into two arms (20 in PNB-001 + BC arm and 20 in BC arm) and received the respective treatments. The primary endpoint, change in the 8-point WHO Ordinal Scale score for COVID-19 showed significant clinical improvement from baseline to day 15 with PNB-001 + BC (P=0.042). One patient on PNB-001+BC and two patients on BC arm died (1 Vs 2; HR: 2.0 [95%CI=0.18, 22.05]; P=0.56) by Day 28. Mean Chest Xray score showed significant improvement (2.05 Vs 1.16; P=0.032), as well as more patients, quickly showed complete improvement. Patients needed a shorter duration of hospitalization and on day 15, 1 patient remained in the hospital on PNB-001 + BC compared to 5 on BC (P=0.048), thus giving an 80% of reduction in the duration of hospitalization. The mean duration of supplemental oxygen requirement was shorter in the PNB-001 + BC arm. In the PNB-001 + BC arm, 50% of patients were off oxygen on day 6, compared to day 8 in the BC arm. Exploratory analysis was done for ESR, CRP, IL-6, and N/L ratio, and the immune parameters showed a statistically significant reduction by Day 15. Lymphocytes were increased into the reference range (P=0.032) and neutrophils were reduced (P=0.013). The role of PNB-001 as an immune modulator was established. NLR was reduced significantly for PNB-001 + BC. A total of 24 (11 vs 13) (PNB-001 + BC vs BC) adverse events were reported in 18 (8 vs 10) (PNB-001 + BC vs BC) patients and none of the 11 adverse effects, was related to PNB-001. The overall safety profile was better in the PNB-001 + BC arm than in the BC arm. PNB-001 when combined with BC improved the clinical status of patients with moderate COVID-19 infection compared to BC alone. PNB-001 was well tolerated by patients with moderate COVID-19 and acted by stimulation of the immune system.