Transarterial Administration of Oncolytic Viruses for Locoregional Therapy of Orthotopic HCC in Rats

The overall goal of this surgical intervention is to deliver locoregional oncolytic virus therapy via hepatic arterial administration to orthotopic hepatocellular carcinoma or HCC lesions. This method can help answer key questions in the oncolytic virus field, such as what is the ethicacy of locoregionally applied therapy to treat orthotopic HCC? The main advantage of this technique is that it allows the delivery of high titers of oncolytic virus to multiple HCC lesions.

A limitation of intratumoral or systemic administration. This technique mimics the application of transarterial chemoembolization. The standard of care to many HCC patients allowing for the excellent translation of pre-clinical data.

Before beginning the surgery, don the appropriate personal protective equipment and disinfect the surgical area and equipment with 70%ethanol. Dilute the virus to the appropriate concentration, according to the toxicity of the virus, and the susceptibility of the strain of rat being used. Then pre-fill one milliliter syringes with the diluted virus, and equip the syringes with 30 Gauge needles, taking care to remove any bubbles.

Confirm that the rat is fully sedated by a lack of response to foot pad pinch. Using a small, curved scalpel, make a small, vertical incision in the skin, directly along the midline, and extend the opening up to the xiphoid process. Next, incise the muscle layer in the same way, taking care not to puncture the organs of the abdominal cavity.

Place a retractor across the abdomen to spread the muscle and skin, exposing the entire abdominal area. Then moisten two 7.5 by 7.5 centimeter gauze swabs with physiological saline and spread one gauze across the top of the surgical opening, and one across the bottom. To isolate the hepatic artery, first use two moistened cotton applicator swabs to gently transfer the intestines and cecum out of the abdominal cavity onto the lower gauze swab, folding the gauze over the organs to keep them moist.

Next, gently lift and flip the left lateral liver lobe up onto the upper gauze and use a small pair of spring scissors to cut through the fibrous membrane on the under side of the lobe. Fold the gauze over the lobe to hold it in place. Then, using a dissecting microscope and fine forceps, dissect the thin membrane surrounding the interior caudate lobe, and flip the lobe onto the upper gauze.

Place the interior caudate lobe under the gauze fold with the left lateral lobe, and then dissect the posterior caudate lobe in the same way. After placing the posterior caudate lobe under the gauze, locate the common hepatic artery by its strong pulsation. Following the artery to the anatomical right, use fine forceps to remove the thick membranous tissue obscuring the gastroduodenal and proper hepatic arteries.

Stop any bleeding with a cotton applicator. Then use two fine atraumatic forceps to carefully dissect the common hepatic gastroduodenal and proper hepatic arteries from the surrounding membranes. To inject the hepatic artery, first, pass a 7-0 prolene suture under the gastroduodenal artery, and use a micro needle holder to tie a permanent ligature at the distal end of the artery just above the bifurcation, leaving approximately two inches of suture material on each end.

Next, place a small clamp on the common hepatic artery to temporarily block the blood flow. Within five minutes of placing the clamp, increase the magnification until the gastroduodenal artery is in sharp focus and appears large enough to accurately insert the needle. Then place the injection syringe needle into the lumen of the gastroduodenal artery, and slowly depress the plunger to maximize the transduction efficiency.

When the entire injection volume has been administered, slowly retract the needle from the artery, and pass a second 7-0 prolene suture beneath the gastroduodenal artery. Tie a second permanent ligature above the injection site and remove the clamp from the common hepatic artery. Confirm that the blood flow through the proper hepatic artery towards the liver has been restored, and cut the loose ends of the ligatures.

Confirm that there is no internal bleeding. Then return the externalized tissues to the abdominal cavity. Using 4-0 vicryl and a curved needle, suture the muscle and skin in separate layers with continuous sutures approximately one millimeter apart.

Then monitor the animal until it is fully recovered. The kinetics of intratumoral virus replication is virus-dependent. For example, vesicular stomatitis virus replication peaks within one day of injection.

With numerous viral foci, a parent exclusively within the hepatocellular carcinoma nodules of the injected animals. In contrast, Newcastle disease virus exhibits a prolonged viral replication kinetic, with peak virus titers on day five post-injection. Due to the inherent tumor specificity of the viruses, the pathogen is rapidly cleared from the normal surrounding liver tissue.

Additionally, morphometric analysis of randomly selected vesicular stomatitis virus vector treated tumor nodules demonstrates that tumor transduction efficiency does not correlate with tumor size. Transhepatic arterial administration of a degradable starch microsphere, concomitantly with the viral solution, results in a nearly complete embolization of the tumor nodules, while imposing a minimal effect on the profusion of the normal liver tissue. As further demonstrated by these dynamic contrast-enhanced magnetic resonance images using a gadolinium contrast agent.

Once mastered, this technique can be completed in 20 minutes, if it is performed properly. While attempting this procedure, it’s important to remember to have everything prepared prior to administering the anthesia, to minimize the time required for the intervention. After watching this video, you should have a good understanding of how to dissect the hepatic artery and to prepare it for the injection of oncolytic virus solutions for locoregional therapy of HCC.

Don’t forget that working with viruses can be extremely hazardous and that precautions, such as wearing personal protective clothing and properly disinfecting the work area and instruments should always be taken while performing this procedure.